Johnson TR, etal., J Immunol. 2003 Feb 15;170(4):2037-45.
Although well studied in settings of helminth infection and allergen sensitization, the combined contributions of IL-4 and IL-13 and their signaling pathways in models of viral pathogenesis have not been reported. Using a murine model of respiratory syncytial vi
rus (RSV) infection, we evaluated the contribution of IL-13, alone and in conjunction with IL-4, during immunization with recombinant vaccinia virus expressing RSV G glycoprotein (vvGs) or with formalin-inactivated RSV (FI-RSV). We showed that both IL-4 and IL-13 activity must be inhibited to modulate G-specific responses resulting in severe RSV-induced disease. Inhibition of IL-4 or IL-13 activity alone had minimal impact on disease in vvGs-immunized mice. However, treatment of IL-4-deficient mice with IL-13Ra during vvGs immunization reduced IL-5, IL-13, and eotaxin production and pulmonary eosinophilia after RSV challenge. In contrast, FI-RSV-induced immune responses were diminished when either IL-4 or IL-13 activity was blocked. After RSV challenge, these type 2 T cell responses were also diminished in vvGs-primed IL-4Ralpha-deficient mice. Our data suggest that secreted vvGs uses mechanisms requiring signaling through the IL-4Ralpha-chain by either IL-4 or IL-13 for induction of eosinophilia and is the first description of the relative contributions of IL-4, IL-13, and their receptors in viral pathogenesis.
Cell adhesion molecules have been implicated as key organizers of synaptic structures, but there is still a need to determine how these molecules facilitate neurotransmitter receptor recruitment to developing synapses. Here, we identify erythrocyte protein band 4.1-like 3 (protein 4.1B) as an intrac
ellular effector molecule of Synaptic Cell Adhesion Molecule 1 (SynCAM1) that is sufficient to recruit NMDA-type receptors (NMDARs) to SynCAM1 adhesion sites in COS7 cells. Protein 4.1B in conjunction with SynCAM1 also increased the frequency of NMDAR-mediated mEPSCs and area of presynaptic contact in an HEK293 cell/ neuron co-culture assay. Studies in cultured hippocampal neurons reveal that manipulation of protein 4.1B expression levels specifically affects NMDAR-mediated activity and localization. Finally, further experimentation in COS7 cells show that SynCAM1 may also interact with protein 4.1N to specifically effect AMPA type receptor (AMPAR) recruitment. Thus, SynCAM1 may recruit both AMPARs and NMDARs by independent mechanisms during synapse formation.
Fogel AI, etal., J Neurosci. 2007 Nov 14;27(46):12516-30.
Synapses are asymmetric cell junctions with precisely juxtaposed presynaptic and postsynaptic sides. Transsynaptic adhesion complexes are thought to organize developing synapses. The molecular composition of these complexes, however, remains incompletely understood, precluding us from understanding
how adhesion across the synaptic cleft guides synapse development. Here, we define two immunoglobulin superfamily members, SynCAM 1 and 2, that are expressed in neurons in the developing brain and localize to excitatory and inhibitory synapses. They function as cell adhesion molecules and assemble with each other across the synaptic cleft into a specific, transsynaptic SynCAM 1/2 complex. Additionally, SynCAM 1 and 2 promote functional synapses as they increase the number of active presynaptic terminals and enhance excitatory neurotransmission. The interaction of SynCAM 1 and 2 is affected by glycosylation, indicating regulation of this adhesion complex by posttranslational modification. The SynCAM 1/2 complex is representative for the highly defined adhesive patterns of this protein family, the four members of which are expressed in neurons in divergent expression profiles. SynCAMs 1, 2, and 3 each can bind themselves, yet preferentially assemble into specific, heterophilic complexes as shown for the synaptic SynCAM 1/2 interaction and a second complex comprising SynCAM 3 and 4. Our results define SynCAM proteins as components of novel heterophilic transsynaptic adhesion complexes that set up asymmetric interactions, with SynCAM proteins contributing to synapse organization and function.
Respiratory syncytial virus (RSV) requires protein biosynthesis machinery to generate progeny. There is evidence that RSV might alter some translation components since stress granules are formed in their host cells. Consistent with these observations, we found
that RSV induces dephosphorylation of 4EBP1 (eIF4E-binding protein), an important cellular translation factor. Our results show no correlation between the 4EBP1 dephosphorylation time and the decrease in the global rate of protein synthesis. Interestingly, treatment with rapamycin stimulates virus generation. The results suggest that RSV is a virus that still contains unknown mechanisms involved in the translation of their mRNAs through the alteration or modification of some translation factors, such as 4EBP1, possibly to favor its replicative cycle.
Lindemans CA, etal., Clin Exp Immunol. 2006 Jun;144(3):409-17.
Respiratory syncytial virus (RSV) is a common cause of lower respiratory tract disease (LRTD) in infants. Eosinophils have been suggested to play a role in the disease pathogenesis of LRTD. Inflammation can induce functional and morphological alterations of peri
pheral blood granulocytes. In patients with RSV LRTD, we aimed to investigate the eosinophil activation status by analysing surface markers. In vitro stimulation of eosinophils with cytokines leads to up-regulation of CD11b and priming markers recognized by the recently developed priming markers A17 and A27, whereas interleukin (IL)-5Ralpha is being down-regulated. In 51 patients and 10 controls we examined the expression of these surface markers on eosinophils in moderate to severe RSV-induced LRTD patients at the time of admission and 6 weeks later during the convalescence phase. RSV-patients were characterized by a higher eosinophil CD11b expression compared to controls. Although basal A17 and A27 expression was not increased, we observed a significantly higher expression of these priming epitopes on N-formyl-methionyl-leucyl-phenylalanine (fMLP)-stimulated cells of RSV patients compared with cells of controls, indicative of prior in vivo priming. Furthermore, IL-5Ralpha expression was down-regulated on peripheral blood eosinophils of these patients. Follow-up blood samples showed normalization of all markers but CD11b, which was persistently increased. Utilizing cellular markers, we observed that peripheral blood eosinophils from infants with RSV LRTD are in a more activated state compared to eosinophils of controls, which normalizes only partially during convalescence.
Respiratory syncytial virus (RSV) is the most common cause of infant hospitalizations and severe RSV infections are a significant risk factor for childhood asthma. The pathogenic mechanisms responsible for RSV induced immunopathophysiology remain elusive. Using
an age-appropriate mouse model of RSV, we show that IL-33 plays a critical role in the immunopathogenesis of severe RSV, which is associated with higher group 2 innate lymphoid cells (ILC2s) specifically in neonates. Infection with RSV induced rapid IL-33 expression and an increase in ILC2 numbers in the lungs of neonatal mice; this was not observed in adult mice. Blocking IL-33 with antibodies or using an IL-33 receptor knockout mouse during infection was sufficient to inhibit RSV immunopathogenesis (i.e., airway hyperresponsiveness, Th2 inflammation, eosinophilia, and mucus hyperproduction); whereas administration of IL-33 to adult mice during RSV infection was sufficient to induce RSV disease. Additionally, elevated IL-33 and IL-13 were observed in nasal aspirates from infants hospitalized with RSV; these cytokines declined during convalescence. In summary, IL-33 is necessary, either directly or indirectly, to induce ILC2s and the Th2 biased immunopathophysiology observed following neonatal RSV infection. This study provides a mechanism involving IL-33 and ILC2s in RSV mediated human asthma.
Respiratory syncytial virus (RSV) infection may have an effect on the development of T cell memory responses. RSV bronchiolitis in infants is associated with a transient decline in circulating lymphocytes. We hypothesized that the mechanism underlying this lymph
openia is apoptosis. Blood was taken from 32 infants during primary RSV bronchiolitis and three months later. Using flow cytometry, we found that absolute numbers of both CD3+/CD4+ T-helper lymphocytes (P = 0.029) and CD3+/CD8+ cytotoxic lymphocytes (CTL) (P = 0.043) were significantly reduced during acute infection. Up-regulated expression both of Fas (P < 0.001) and tumour necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor (P < 0.001) was found during acute illness on both CD3+/CD4+ and CD3+/CD8+ lymphocytes, when compared with convalescent samples. Expression of Fas on CD4+ lymphocytes was inversely related to CD4+ number (P = 0.03). Plasma levels of soluble Fas ligand (P = 0.028) and caspase-1 (P = 0.037), determined by enzyme-linked immunosorbent assay, were increased during bronchiolitis. Plasma interleukin-18, a product of caspase-1 activity, was not raised. Taken together, these data suggest that in acute RSV infection, CD4+ helper lymphocytes and CD8+ cytotoxic lymphocytes are primed to undergo apoptosis. This is a mechanism through which lymphopenia may occur and T cell memory may be altered.
Rash A, etal., Eur J Clin Invest. 2016 Feb;46(2):141-5. doi: 10.1111/eci.12576. Epub 2016 Jan 18.
BACKGROUND: The diagnosis of vasovagal syncope continues to be difficult despite the use of accurate histories, tilt testing and implantable loop recorders. A circulating biomarker might be useful to facilitate diagnoses. Both endothelin-1 and vasopressin are
increased during positive tilt tests resulting in syncope. Copeptin is a stable cleavage product of vasopressin formation. We conducted a pilot study to assess the utility of endothelin-1 and copeptin as circulating biomarkers of vasovagal syncope. METHODS: Three populations were studied: syncope patients, epilepsy patients and controls. Vasovagal syncope diagnosis was ascertained with the Calgary Syncope Score and epilepsy diagnosis was confirmed with EEG. Plasma levels of endothelin-1 were measured using by ELISA and copeptin levels were determined using an EIA kit. RESULTS: Asymptomatic control subjects had mean age 35 +/- 11 years (7/22 male); epileptic subjects had mean age 32 +/- 7 years (4/15 male); and syncope subjects had mean age 33 +/- 16 years (4 of 21 male). Circulating plasma levels of endothelin-1 and copeptin were no different among the three groups. Mean concentrations of endothelin-1 were as follows: syncope, 23 +/- 32 pg/mL; controls, 21 +/- 17 pg/mL; and epileptics, 18 +/- 12 pg/mL. Mean concentrations of copeptin were as follows: syncope, 1.29 +/- 0.79 ng/mL; controls, 1.25 +/- 0.79 ng/mL; and seizures, 1.23 +/- 0.45 ng/mL. There were no significant correlations between syncope frequency and copeptin or endothelin-1 levels. CONCLUSION: Circulating plasma endothelin-1 and copeptin levels are not significantly different among populations of controls, syncope patients and seizure patients.
Tang QQ, etal., Proc Natl Acad Sci U S A 2003 Jan 7;100(1):44-9. Epub 2002 Dec 26.
When induced to differentiate, growth-arrested 3T3-L1 preadipocytes synchronously reenter the cell cycle and undergo mitotic clonal expansion (MCE) followed by expression of genes that produce the adipocyte phenotype. The preadipocytes traverse the G(1)S checkpo
int synchronously as evidenced by the expressionactivation of cdk2-cyclin-EA, turnover of p27kip1, hyperphosphorylation of Rb, translocation of cyclin D(1) from nuclei to cytoplasm and GSK-3beta from cytoplasm to nuclei, and incorporation of [(3)H]thymidine into DNA. As the cells cross the G(1)S checkpoint, CEBPbeta acquires DNA-binding activity, initiating a cascade of transcriptional activation that culminates in the expression of adipocyte proteins. The mitogen-activated protein kinaseextracellular signal-regulated kinase kinase (MEK) inhibitor PD98059 delays, but does not block, MCE and differentiation, the extent of the delay causing a comparable delay in the expression of cell-cycle markers, MCE, and adipogenesis. The more potent and specific MEK inhibitor UO126 and the cyclin-dependent kinase inhibitor roscovitine, which inhibit the cell cycle at different points, block MCE, expression of cell cycle and adipocyte markers, as well as adipogenesis. These results show that MCE is a prerequisite for differentiation of 3T3-L1 preadipocytes into adipocytes.
Kerr MH and Paton JY, Am J Respir Crit Care Med. 1999 Apr;159(4 Pt 1):1115-8.
Infection with respiratory syncytial virus (RSV) is a common cause of respiratory disease in infancy. Surfactant phospholipids have been shown to be reduced in severe RSV infection. Reduction in surfactant proteins might also contribute to the pathogenesis of th
is disease. We investigated daily levels of surfactant proteins in bronchoalveolar lavage (BAL) fluid from 18 ventilated infants with RSV infection (median age 3.1 mo) and in a control group of 16 ventilated surgical patients (median age 0.4 mo). Surfactant proteins were measured by ELISA, total protein by the Lowry method. Surfactant protein A (SP-A) was reduced in BAL fluid from children with RSV infection (median 5.6 micrograms/ml; range 0.6 to 151.9 micrograms/ml) compared with control samples (median 9.0 micrograms/ml; range 0.5 to 139.6 micrograms/ml, p = 0.0368). Surfactant protein B (SP-B) was lower in the RSV group (median 12.0 ng/ml; range 0 to 60. 8 ng/ml) than in control patients (median 118.1 ng/ml; range 0 to 778.2 ng/ml, p < 0.0000). Surfactant protein D (SP-D) was also reduced in the RSV group (median 130.3 ng/ml; range 0 to 1,486.0 ng/ml) versus median 600.4 ng/ml; range 0 to 1,869.0 ng/ml, p < 0. 0000. Total protein levels were higher in the RSV group (median 0.49 mg/ml; range 0.13 to 2.46 mg/ml versus median 0.36 mg/ml; range 0.07 to 1.65 mg/ml, p = 0.0079). The median value of SP-A was significantly lower in the initial sample (2.3 micrograms/ml) than in the final one (6.0 micrograms/ml). However, no significant correlation was found between surfactant protein concentrations and disease severity measured by arterial alveolar oxygen ratio. We conclude that alterations in surfactant protein concentrations are present in severe RSV infection and speculate that these may contribute to the abnormalities of lung function seen in this condition.
Soygur B and Sati L, Reproduction. 2016 Nov;152(5):R167-78. doi: 10.1530/REP-16-0031. Epub 2016 Aug 2.
Human life begins with sperm and oocyte fusion. After fertilization, various fusion events occur during human embryogenesis and morphogenesis. For example, the fusion of trophoblastic cells constitutes a key process for normal placental development. Fusion in the placenta is facilitated by sync
yle='font-weight:700;'>syncytin 1 and syncytin 2. These syncytins arose from retroviral sequences that entered the primate genome 25 million and more than 40 million years ago respectively. About 8% of the human genome consists of similar human endogenous retroviral (HERVs) sequences. Many are inactive because of mutations or deletions. However, the role of the few that remain transcriptionally active has not been fully elucidated. Syncytin proteins maintain cell-cell fusogenic activity based on ENV: gene-mediated viral cell entry. In this review, we summarize how syncytins and their receptors are involved in fusion events during human reproduction. The significance of syncytins in tumorigenesis is also discussed.
Respiratory syncytial virus (RSV) is the leading cause of bronchiolitis and viral pneumonia in infants and young children. Administration of a formalin inactivated vaccine against RSV to children in the 1960s resulted in increased morbidity and mortality in vacc
ine recipients who subsequently contracted RSV. This incident precluded development of subunit RSV vaccines for infants for over 30 years, because the mechanism of illness was never clarified. An RSV vaccine for infants is still not available. Here, we demonstrate that enhanced RSV disease is mediated by immune complexes and abrogated in complement component C3 and B cell-deficient mice but not in controls. Further, we show correlation with the enhanced disease observed in children by providing evidence of complement activation in postmortem lung sections from children with enhanced RSV disease.
Respiratory syncytial virus (RSV) is the most common viral respiratory pathogen during infancy world wide. It induces innate and adaptive immune response in host cells. The toll like receptor 4 (TLR4)/CD14 complex is particularly important for the initiation of
an innate immune response to RSV. Thus we were interested whether an association exists between severe RSV associated diseases and polymorphisms within TLR4 and CD14. We genotyped the CD14 promotor polymorphism -C159T and the two common TLR4 amino acid variants (D259G, and T359I) in 131 infants with severe RSV associated diseases and 270 controls. Statistical analyses of single polymorphisms made use of the Armitage's trend test, haplotypes were calculated by FAMHAP, FASTEHPLUS and Arlequin. All polymorphisms were in Hardy Weinberg Equilibrium. We found marginal association between amino acid exchange D259G in TLR4 with RSV infection p=0.0545). Furthermore, haplotypes analysis of the two TLR4 polymorphisms by three independent programs revealed association of haplotypes with severe RSV infection (p
Stark JM, etal., J Virol. 2010 Mar;84(5):2257-69. Epub 2009 Dec 16.
Respiratory syncytial virus (RSV) is the major cause of lower respiratory tract infection in infants, with about half being infected in their first year of life. Yet only 2 to 3% of infants are hospitalized for RSV infection, suggesting that individual susceptib
ility contributes to disease severity. Previously, we determined that AKR/J (susceptible) mice developed high lung RSV titers and showed delayed weight recovery, whereas C57BL/6J (resistant) mice demonstrated low lung RSV titers and rapid weight recovery. In addition, we have reported that gene-targeted mice lacking the cystic fibrosis transmembrane conductance regulator (Cftr; ATP-binding cassette subfamily C, member 7) are susceptible to RSV infection. For this report, recombinant backcross and F2 progeny derived from C57BL/6J and AKR/J mice were infected with RSV, their lung titers were measured, and quantitative trait locus (QTL) analysis was performed. A major QTL, designated Rsvs1, was identified on proximal mouse chromosome 6 in both recombinant populations. Microarray analysis comparing lung transcripts of the parental strains during infection identified several candidate genes that mapped to the Rsvs1 interval, including Cftr. These findings add to our understanding of individual RSV susceptibility and strongly support a modifier role for CFTR in RSV infection, a significant cause of respiratory morbidity in infants with cystic fibrosis.
McNamara PS, etal., Eur Respir J. 2004 Jan;23(1):106-12.
Respiratory syncytial virus (RSV) bronchiolitis is an important cause of severe respiratory disease in infants. This study aimed to characterise changes in pulmonary pro- and anti-inflammatory responses in infants with RSV bronchiolitis over the course of the il
lness. On the day of intubation (Day 1) and the day of extubation (Day X), nonbronchoscopic bronchoalveolar lavage was performed on term and preterm infants ventilated for RSV bronchiolitis and on control infants on Day 1. Tumour necrosis factor (TNF)-alpha, soluble TNF receptor (sTNFR) and interleukin (IL)-6 messenger ribonucleic acid (mRNA) and protein were measured. Twenty-four infants, born at term and 23 infants born preterm with RSV bronchiolitis and 10 controls were recruited. TNF-alpha and IL-6 mRNA and protein in infants with bronchiolitis were greater than the control group on Day 1. In preterm infants, who were ventilated for longer than term infants, TNF-alpha and IL-6 proteins decreased between Day 1 and Day X. Concentrations of sTNFRs differed between groups on Day 1, but levels did not change between Day 1 and Day X. Large amounts of tumour necrosis factor-alpha and interleukin-6 in the respiratory syncytial virus-infected lung suggest important roles for these cytokines in the pathogenesis of respiratory syncytial virus bronchiolitis. The decrease in tumour necrosis factor-alpha and interleukin-6 protein in preterm infants may reflect the prolonged clinical course seen in these infants.
Respiratory syncytial virus (RSV) is a common cause of infection that is associated with a range of respiratory illnesses, from common cold-like symptoms to serious lower respiratory tract illnesses such as pneumonia and bronchiolitis. RSV is the single most imp
ortant cause of serious lower respiratory tract illness in children <1 year of age. Host innate and acquired immune responses activated following RSV infection have been suspected to contribute to RSV disease. Toll-like receptors (TLRs) activate innate and acquired immunity and are candidates for playing key roles in the host immune response to RSV. Leukocytes express TLRs, including TLR2, TLR6, TLR3, TLR4, and TLR7, that can interact with RSV and promote immune responses following infection. Using knockout mice, we have demonstrated that TLR2 and TLR6 signaling in leukocytes can activate innate immunity against RSV by promoting tumor necrosis factor alpha, interleukin-6, CCL2 (monocyte chemoattractant protein 1), and CCL5 (RANTES). As previously noted, TLR4 also contributes to cytokine activation (L. M. Haynes, D. D. Moore, E. A. Kurt-Jones, R. W. Finberg, L. J. Anderson, and R. A. Tripp, J. Virol. 75:10730-10737, 2001, and E. A. Kurt-Jones, L. Popova, L. Kwinn, L. M. Haynes, L. P. Jones, R. A. Tripp, E. E. Walsh, M. W. Freeman, D. T. Golenbock, L. J. Anderson, and R. W. Finberg, Nat. Immunol. 1:398-401, 2000). Furthermore, we demonstrated that signals generated following TLR2 and TLR6 activation were important for controlling viral replication in vivo. Additionally, TLR2 interactions with RSV promoted neutrophil migration and dendritic cell activation within the lung. Collectively, these studies indicate that TLR2 is involved in RSV recognition and subsequent innate immune activation.
Doengi M, etal., Hippocampus. 2016 Mar;26(3):319-28. doi: 10.1002/hipo.22524. Epub 2015 Oct 1.
The survival of adult-born dentate gyrus granule cells critically depends on their synaptic integration into the existing neuronal network. Excitatory inputs are thought to increase the survival rate of adult born neurons. Therefore, whether enhancing the stability of newly formed excitatory synaps
es by overexpressing the synaptic cell adhesion molecule SynCAM 1 improves the survival of adult-born neurons was tested. Here it is shown that overexpression of SynCAM 1 improves survival of adult-born neurons, but has no effect on the proliferation rate of precursor cells. As expected, overexpression of SynCAM 1 increased the synapse density in adult-born granule neurons. While adult-born granule neurons have very few functional synapses 15 days after birth, it was found that at this age adult-born neurons in SynCAM 1 overexpressing mice exhibited around three times more excitatory synapses, which were stronger than synapses of adult-born neurons of control littermates. In summary, the data indicated that additional SynCAM 1 accelerated synapse maturation, which improved the stability of newly formed synapses and in turn increased the likelihood of survival of adult-born neurons.
Hashimoto K, etal., Am J Respir Crit Care Med. 2004 Aug 1;170(3):306-12. Epub 2004 May 6.
Respiratory syncytial virus (RSV) is associated with wheezing and childhood asthma. We previously reported that RSV infection prolongs methacholine-induced airway hyperresponsiveness in ovalbumin (OVA)-sensitized mice. In addition, allergically sensitized RSV-in
fected (OVA/RSV) mice had more abundant airway epithelial mucus production compared with OVA mice 14 days after infection, whereas there was almost no mucus in mice that were only RSV infected. We hypothesized that this increased mucus was associated with mucosal expression of Muc5ac, a mucus gene expression in airways, and gob-5, a member of the Ca(2)(+)-activated chloride channel family. By histochemical analysis, we found that there was significantly increased staining for gob-5 and Muc5ac in the airways of OVA/RSV mice compared with either OVA mice or allergically sensitized mice that were challenged with inactivated RSV, and virtually no detectable staining in the RSV group. These findings were confirmed by Western blot analysis. The increased mucus expression in the OVA/RSV group was associated with increased lung levels of interleukin-17, a factor known to stimulate airway mucin gene expression. The impact of virus infection combined with allergic inflammation on mucus production may partially explain the more severe disease and airway hyperresponsiveness associated with RSV in the setting of atopy.
Liao S and Ruddle NH, J Immunol. 2006 Sep 1;177(5):3369-79.
The mature phenotype of peripheral lymph node (LN) high endothelial venules (HEVs), defined as MAdCAM-1(low) PNAd(high) LTbetaR(high) HEC-6ST(high), is dependent on signaling through the lymphotoxin-beta receptor (LTbetaR). Plasticity of PLN HEVs during immunization with oxazolone was apparent as a
reversion to an immature phenotype (MAdCAM-1(high) PNAd(low) LTbetaR(low) HEC-6ST(low)) followed by recovery to the mature phenotype. The recovery was dependent on B cells and was inhibited by LTbetaR-Ig treatment. Concurrent with HEV reversion, at day 4 following oxazolone or OVA immunization, reduced accumulation of Evans blue dye and newly activated DCs in the draining LNs revealed a temporary afferent lymphatic vessel (LV) functional insufficiency. T cell priming to a second Ag was temporarily inhibited. At day 7, lymphangiogenesis peaked in both the skin and draining LN, and afferent LV function was restored at the same time as HEV phenotype recovery. This process was delayed in the absence of B cells. LV and HEV both express the LTbetaR. During lymphangiogenesis in the draining LN, HEV, and LV were directly apposed; some vessels appeared to express both PNAd and LYVE-1. Pretreatment with LTbetaR-Ig drastically reduced the number of PNAd+ LYVE-1+ vessels, suggesting a reduction in LV and HEV cross-talk. The concordance in time and function and the close physical contact between LVs and HEVs in the remodeling process after immunization indicate that the two vascular systems are in synchrony and engage in cross-talk through B cells and LTbetaR.
The membrane proteins synaptobrevin, syntaxin, and SNAP-25 form the core of a ubiquitous fusion machine that interacts with the soluble proteins NSF and alpha-SNAP. During regulated exocytosis, membrane fusion is usually strictly controlled by Ca2+ ions. However, the mechanism by which Ca2+ regulate
s exocytosis is still unclear. Here we show that the membranes of exocrine secretory granules contain an 18-kDa protein, syncollin, that binds to syntaxin at low Ca2+ concentrations and dissociates at concentrations known to stimulate exocytosis. Syncollin has a single hydrophobic domain at its N-terminus and shows no significant homology with any known protein. Recombinant syncollin inhibits fusion in vitro between zymogen granules and pancreatic plasma membranes, and its potency falls as Ca2+ concentration rises. We suggest that syncollin acts as a Ca2(+)-sensitive regulator of exocytosis in exocrine tissues.
Respiratory syncytial virus (RSV) is a major viral pathogen of infants and the elderly. Significant morbidity is caused by an overexuberant mixed lung cell infiltrate, which is thought to be driven by chemokines. One of the main chemotactic mediators responsible
for the movement of eosinophils is CCL11 (eotaxin). Using a mouse model of eosinophilic bronchiolitis induced by RSV, we show here that treatment in vivo with a blocking antibody to CCL11 greatly reduces lung eosinophilia and disease severity. In addition, anti-CCL11 caused a striking inhibition of CD4-T-cell influx and shifted cytokine production away from interleukin-5 without reducing the resistance to viral replication. These results suggest that in addition to influencing eosinophil diapedesis and survival, anti-CCL11 has an action on T cells. These studies strengthen the case for anti-CCL11 treatment of Th2-driven diseases.
Li W and Shen HH, Chin Med J (Engl). 2007 Jan 5;120(1):5-11.
BACKGROUND: Respiratory syncytial virus (RSV) is a common pathogen in the lower respiratory tract of infants and children. Recent studies have shown that in adults, especially in the elderly population who have relatively weak immunity, lower respiratory tract i
nfection is not uncommon. RSV was detected in 22% hospitalized patients with acute exacerbation of chronic obstructive pulmonary disease (COPD), and the detection rate was only next to that of parvovirus and influenza virus respectively. Further studies revealed that lung infection of RSV could lead to inflammatory destruction and structural remodeling. This study was undertaken to examine the effect of infection with RSV on matrix metalloproteinase (MMPs) in mice, and to explore the role of RSV in the pathogenesis of pulmonary diseases. METHODS: Twenty BALB/c mice were divided randomly into an RSV infection group and a blank control group. The mice in the RSV infection group were given 100 microl liquid containing 10(6) PFU of RSV by intranasal instillation. Three days after the infection, the bronchoalveolar lavage fluid (BALF) was harvested and RT-PCR and Western blotting were used to detect MMP-9 and the expression of tissue inhibitors of matrix metalloproteinase (TIMP)-1 mRNA in lung tissues. Gelatin zymography was employed to detect the activities of MMP-9 and MMP-2 in BALF. Immunohistochemistry was used to determine the expressions of E-cadherin (E-cd) and proliferating cell nuclear antigen (PCNA) in the lung tissues. RESULTS: In the BALF of the mice infected with RSV, the activities of MMP-9 and MMP-2 were significantly increased (t = 6.08, P < 0.01 and t = 5.68, P < 0.01). The levels of mRNA and proteins of MMP-9 in the lung tissues of the mice infected with RSV were significantly elevated, and the mRNA and protein levels were significantly higher than those of the blank controls. Though the ratio of MMP-9/TIMP-1 mRNA in the lung tissues of the infected mice was not significantly different from that of the normal controls, the ratio of the MMP-9/TIMP-1 protein in the RSV infection group was significantly higher than that in the control group. Moreover, the number of cells with positive E-cd expression was decreased and the number of cells positive for PCNA was increased, with an enhanced expression. CONCLUSIONS: In mice, infection with RSV can significantly increase the activities of MMP-9 and MMP-2, and conspicuously elevate the mRNA transcription of MMP-9. RSV infection can increase the activity of gelatinase while up-regulating its inhibitory factors but increase its protein ratio of MMP-9/TIMP-1 in lung tissues, thereby facilitating fibrosis after structural destruction of the airway. The resultant status might be an important factor causing chronic reconstruction of the airway.
Grilo LS, etal., Ann Noninvasive Electrocardiol. 2011 Apr;16(2):213-8. doi: 10.1111/j.1542-474X.2011.00419.x.
We report the case of a woman with syncope and persistently prolonged QTc interval. Screening of congenital long QT syndrome (LQTS) genes revealed that she was a heterozygous carrier of a novel KCNH2 mutation, c.G238C. Electrophysiological and biochemical charac
terizations unveiled the pathogenicity of this new mutation, displaying a 2-fold reduction in protein expression and current density due to a maturation/trafficking-deficient mechanism. The patient's phenotype can be fully explained by this observation. This study illustrates the importance of performing genetic analyses and mutation characterization when there is a suspicion of congenital LQTS. Identifying mutations in the PAS domain or other domains of the hERG1 channel and understanding their effect may provide more focused and mutation-specific risk assessment in this population.
Wang SZ, etal., Eur Respir J. 2000 Feb;15(2):358-66.
Respiratory epithelium is both a target and an effector of airway inflammation. Adhesion molecules on epithelium play an important role in a variety of airway diseases. Respiratory syncytial virus (RSV) is the most important pathogen for airway diseases in infan
ts. The expression of adhesion molecules on epithelium in RSV infection, however, is unclear. The expression of selected adhesion molecules and major histocompatibility complex (MHC) class I and II antigens on a human alveolar type II epithelial cell line (A549) infected with RSV was investigated by means of flow cytometry and immunocytochemistry. The results showed that intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) were expressed on A549 cells at a low level. E-cadherin and MHC class I antigen were constitutively expressed on the cells. RSV infection of A549 cells significantly upregulated the expression of ICAM-1, VCAM-1 and MHC class I and II antigens on these cells. RSV infection also altered the expression of E-cadherin on A549 cells. Immunostaining showed that E-cadherin was mainly upregulated around or in RSV-induced giant cells. These data suggest that respiratory syncytial virus infection of respiratory epithelial cells enhances the expression of adhesion molecules and major histocompatibility complex antigens. These changes may play an important role in the pathophysiology of respiratory syncytial virus disease.
Zhang Q, etal., Acta Paediatr. 2013 May;102(5):e210-4. doi: 10.1111/apa.12182. Epub 2013 Feb 21.
AIM: This study was designed to evaluate the diagnostic value of B-type natriuretic peptide (BNP) in syncope in children and adolescents. METHODS: Serum BNP concentration was measured by electrochemiluminescence assay in 62 consecutive children and adolescents
hospitalized for syncope. RESULTS: Of the 62 children and adolescents hospitalized for syncope, 39 had noncardiac syncope, of whom 37 (59.7%) had autonomic-mediated reflex syncope and two (3.2%) had syncope of unknown cause. Twenty-three patients (37.1%) had cardiac syncope: 11 of these had cardiac arrhythmias and 12 had structural cardiac/cardiopulmonary disease. Patients with cardiac syncope had significantly higher serum BNP than those with non-cardiac syncope (958.78 +/- 2443.41 pg/mL vs 31.05 +/- 22.64 pg/mL, p < 0.05). Logistic multivariate regression analysis revealed that urinary incontinence during syncopal episodes, ECG abnormalities and increased serum BNP levels were independent predictors of cardiac syncope. At a cut-off value of 40.65 pg/mL, serum BNP was associated with significant risk of a cardiac cause of syncope, with sensitivity of 73.9% and specificity of 70.0% for distinguishing cardiac syncope from noncardiac syncope. CONCLUSION: Serum BNP was helpful in differentiating cardiac syncope from noncardiac syncope in children and adolescents.
Reflex syncope is defined by a self-terminating transient loss of consciousness associated with an exaggerated response of the vagal reflexes upon orthostatic challenges. A hereditary component has previously been suggested. We hypothesized that variations in ge
nes encoding proteins mediating the vagal signaling in the heart may be involved in reflex syncope pathogenesis. We systematically resequenced the entire coding regions and flanking intron sequences in 5 genes in the cardiac post-synaptic parasympathetic signaling pathway [muscarinic acetylcholine receptor M2 (CHRM2); G-protein beta-1 subunit (GNB1); G-protein gamma-2 subunit (GNG2); potassium inwardly rectifying channel, subfamily J, member 3 (KCNJ3); and potassium inwardly rectifying channel, subfamily J, member 5 (KCNJ5)] in 74 patients with well-characterized reflex syncope of either cardioinhibitory [Vasovagal Syncope International Study (VASIS-IIB), N = 38] or vasodepressor (VASIS-III, N = 36) type. We identified 2 novel genetic variants (CHRM2 c.1114C>G and GNG2 c.87+34G>A) and several known variants (GNB1: c.267+14G>A, c.267+19C>T, and c.738C>T; KCNJ3: c.119A>G, c.591C>T, c.1038T>C, and c.1494T>C; KCNJ5: c. 171T>C, c.810T>G, c.834T>C, c.844C>G, c.938+7C>T, and c.938-10G>A). The minor allele frequency of the KCNJ5 c.938+7C>T variant was significantly lower in patients than in the control group (0.014 versus 0.089, P = 0.001), and the frequency of heterozygosity and homozygosity was lower in cardioinhibitory patients compared to controls. Genetic variations in genes responsible for the vagal signaling in the heart, including CHRM2, GNB1, GNG2, KCNJ3, and KCNJ5, are not major contributors to the pathogenesis of reflex syncope of vasodepressor or cardioinhibitory types.
Respiratory syncytial virus (RSV) is the leading cause of lower respiratory tract illnesses in infants worldwide. TLR4 signal pathway plays a critical role in regulating immune response against RSV infection. However, the activation of TLR4 in RSV infection is s
till unclear. In present study, the expression levels of miR-26b and TLR4 mRNA were detected in peripheral blood mononuclear cells (PBMCs) of children with or without RSV infected bronchiolitis. The expression levels of TLR4 and its downstream genes IFNbeta and CCL5 were also quantified in PBMCs infected with RSVDeltaG or RSV A2 in vitro. The results showed that children with RSV infection had higher miR-26b level and lower TLR4 mRNA level in PBMCs. miR-26b was predicted to target TLR4. In vitro, miR-26b mimic markedly down-regulated TLR4 mRNA/protein expression and IFNbeta/CCL5 concentrations while miR-26b inhibitor up-regulated these levels. This study reveals that RSV infection inhibits TLR4 signaling via up-regulation of miR-26b, which provides a potential therapeutic target for preventing and treating RSV infection.
Gu H, etal., Sci Rep. 2016 Jan 27;6:19840. doi: 10.1038/srep19840.
Respiratory syncytial virus (RSV) infection is a major cause of severe lower respiratory illness in infants and young children, but the underlying mechanisms responsible for viral pathogenesis have not been fully elucidated. To date, no drugs or vaccines have be
en employed to improve clinical outcomes for RSV-infected patients. In this paper, we report that angiotensin-converting enzyme-2 (ACE2) protected against severe lung injury induced by RSV infection in an experimental mouse model and in pediatric patients. Moreover, ACE2 deficiency aggravated RSV-associated disease pathogenesis, mainly by its action on the angiotensin II type 1 receptor (AT1R). Furthermore, administration of a recombinant ACE2 protein alleviated the severity of RSV-induced lung injury. These findings demonstrate that ACE2 plays a critical role in preventing RSV-induced lung injury, and suggest that ACE2 is a promising potential therapeutic target in the management of RSV-induced lung disease.
The innate immune system contributes to the earliest phase of the host defense against foreign organisms and has both soluble and cellular pattern recognition receptors for microbial products. Two important members of this receptor group, CD14 and the Toll-like receptor (TLR) pattern recognition rec
eptors, are essential for the innate immune response to components of Gram-negative and Gram-positive bacteria, mycobacteria, spirochetes and yeast. We now find that these receptors function in an antiviral response as well. The innate immune response to the fusion protein of an important respiratory pathogen of humans, respiratory syncytial virus (RSV), was mediated by TLR4 and CD14. RSV persisted longer in the lungs of infected TLR4-deficient mice compared to normal mice. Thus, a common receptor activation pathway can initiate innate immune responses to both bacterial and viral pathogens.
Piedimonte G, etal., Am J Physiol. 1999 Oct;277(4 Pt 1):L831-40.
Respiratory syncytial virus (RSV) is a major respiratory pathogen in infants. The first goal of this study was to determine whether the infection following endotracheal inoculation of RSV in Fischer 344 rats results in increased inflammatory responses to substan
ce P (SP) either released by capsaicin from sensory nerves or injected into the circulation. Five days after inoculation, the extravasation of Evans blue-labeled albumin after capsaicin or SP was significantly greater in RSV-infected airways than in pathogen-free controls. The peptide-degrading activity of the regulatory enzyme neutral endopeptidase was unaffected by RSV. However, SP(NK(1)) receptor mRNA levels increased fivefold in RSV-infected lungs, and the density of SP binding sites in the bronchial mucosa increased threefold. These data suggest that RSV makes the airways abnormally susceptible to the proinflammatory effects of SP by upregulating SP(NK(1)) receptor gene expression, thereby increasing the density of these receptors on target cells. This effect may contribute to the inflammatory reaction to the virus and could be a target for the therapy of RSV disease and its sequelae.
Respiratory syncytial virus (RSV) is an important viral pathogen of otitis media, bronchiolitis, and pneumonia. As infection of the upper airways is a precondition for the development of these diseases, understanding RSV pathogenesis and the host response induce
d by RSV in this niche may enable the development of novel therapeutic strategies against this virus. We have used a microarray approach and showed that expression of the gene that encodes the antiviral protein viperin was significantly upregulated in the chinchilla nasopharynx up to 1 week after RSV challenge. Overexpression of human viperin in vitro diminished the ability of RSV to infect HeLa or A549 cells. Furthermore, transduction of the chinchilla airways with a recombinant adeno-associated virus vector that encodes viperin resulted in reduced titers of RSV in the nasopharyngeal lavage fluid. Collectively, these data indicated that viperin plays a significant role in the innate immune defense against RSV.
Kawasaki Y, etal., J Med Virol. 2009 Dec;81(12):2104-8.
To evaluate whether KL-6 concentration is a useful biomarker of the severity of respiratory syncytial virus (RSV) bronchiolitis, we determined KL-6 concentrations in patients with RSV bronchiolitis with or without chronic heart disease (CHD). We enrolled 52 pati
ents who had been diagnosed with RSV bronchiolitis and required admission to the hospital at the Department of Pediatrics of Fukushima Medical University School of Medicine from 2004 to 2005. These patients were divided into two groups: Group 1 consisted of patients without any underlying disease, and Group 2 consisted of patients with CHD. These patients were assigned to three categories. Stage A consisted of patients without oxygen dosage, stage B of patients who required oxygen dosage, and stage C of patients required artificial respiration. We evaluated baseline characteristics, clinical features, and serum KL-6 concentration in Group 1, Group 2, and a control group (healthy infants without infection). Mean serum KL-6 concentrations in patients with RSV bronchiolitis were higher than those in the control group (471.8 +/- 236.9 and 127.1 +/- 69.1 U/ml, respectively). Mean serum KL-6 concentration was higher in Group 2 than in Group 1 (692.8 +/- 313.1 and 390.4 +/- 132.7 U/ml, respectively). Mean serum KL-6 concentrations were higher in stage C than in stages A and B, and mean serum KL-6 concentrations were higher in stage B than in stage A. These findings suggest that serum KL-6 is associated with the severity of RSV bronchiolitis and that it may be a useful biomarker for the severity of RSV bronchiolitis.
Glasser SW, etal., Am J Physiol Lung Cell Mol Physiol. 2009 Jul;297(1):L64-72. Epub 2009 Mar 20.
Patients with mutations in the pulmonary surfactant protein C (SP-C) gene develop interstitial lung disease and pulmonary exacerbations associated with viral infections including respiratory syncytial virus (RSV). Pulmonary infection with RSV caused more severe
interstitial thickening, air space consolidation, and goblet cell hyperplasia in SP-C-deficient (Sftpc(-/-)) mice compared with SP-C replete mice. The RSV-induced pathology resolved more slowly in Sftpc(-/-) mice with lung inflammation persistent up to 30 days postinfection. Polymorphonuclear leukocyte and macrophage counts were increased in the bronchoalveolar lavage (BAL) fluid of Sftpc(-/-) mice. Viral titers and viral F and G protein mRNA were significantly increased in both Sftpc(-/-) and heterozygous Sftpc(+/-) mice compared with controls. Expression of Toll-like receptor 3 (TLR3) mRNA was increased in the lungs of Sftpc(-/-) mice relative to Sftpc(+/+) mice before and after RSV infection. Consistent with the increased TLR3 expression, BAL inflammatory cells were increased in the Sftpc(-/-) mice after exposure to a TLR3-specific ligand, poly(I:C). Preparations of purified SP-C and synthetic phospholipids blocked poly(I:C)-induced TLR3 signaling in vitro. SP-C deficiency increases the severity of RSV-induced pulmonary inflammation through regulation of TLR3 signaling.
Devincenzo JP, etal., Am J Respir Crit Care Med. 2010 Jul 9.
Rationale and objective: Respiratory syncytial virus (RSV) is the leading cause of childhood lower respiratory infection, yet viable therapies are lacking. Two major challenges have stalled antiviral development: ethical difficulties performing pediatric proof-o
f-concept studies and the prevailing concept that disease is immune-mediated rather than being driven by viral load. We developed a human experimental wild-type RSV infection model to address these challenges. METHODS: Healthy volunteers (n = 35), in 5 cohorts, received increasing quantities (3.0 - 5.4 log PFU/person) of wild-type RSV-A intranasally. RESULTS: Overall, 77% of volunteers consistently shed virus. Infection rate, viral loads, disease severity and safety were similar between cohorts and were unrelated to quantity of RSV received. Symptoms began near the time of initial viral detection, peaked in severity near when viral load peaked, and subsided as viral loads (measured by RT-PCR) slowly declined. Viral loads correlated significantly with intranasal proinflammatory cytokine concentrations (IL-6, IL-8). Increased viral load correlated consistently with increases in multiple different disease measurements (symptoms, physical exam, amount of nasal mucus). CONCLUSION: Viral load appears to drive disease manifestations in humans with RSV infection. The observed parallel viral and disease kinetics support a potential clinical benefit of RSV antivirals. This reproducible model facilitates development of future RSV therapeutics.
Respiratory Syncytial Virus (RSV) infection is an important cause of severe infant bronchiolitis, partly due to lower airway inflammation orchestrated by virus-induced chemokine secretion. Chemokine receptors may therefore be therapeutic targets. We investigated
RSV-induced chemokine receptor (CCR) 1, 2 and 5 surface expressions in a cellular model and in infants. RSV infection increased human monocytic CCR1, 2 and 5 expression, as assessed by FACS, via replication-dependent mechanisms. CCR1 and CCR5 levels peaked at 36 h and CCR2 levels at 48 h. Monocytes from infants with RSV-bronchiolitis significantly increased CCR1 expression after ex vivo RSV infection compared to controls. Expression of CCR5 also increased, and correlated with CCR1 expression (r=0.78, p<0.0001). CCR1 upregulation correlated with disease severity markers. Monocyte CCR1 receptors were functionally active as stimulation resulted in calcium influx. CCR1/5 blocking strategies may be useful in decreasing cellular inflammation in RSV infection.
Tumor necrosis factor (TNF-)alpha is a proinflammatory cytokine that is important in the innate host defence and thus in the defence of infectious agents. However, in excess it provokes the development of chronic inflammatory diseases. The aim of this study was to test association of TNF with severe
RSV bronchiolitis as example of an infectious disease and asthma as representative for a chronic inflammatory condition. The following study populations were genotyped for 4 polymorphisms within TNF-beta (rs909253) and TNF-alpha (rs1799964, rs1799724, rs1800629): 322 asthmatic children, 151 children with severe respiratory syncytial virus (RSV) bronchiolitis and 270 controls. Furthermore, serum TNF-alpha levels were measured by a FlowCytomix Assay. Asthma showed association with two TNF-alpha polymorphisms as well as with TNF haplotypes (p = 0.0050). In contrast, RSV bronchiolitis was associated with TNF haplotypes (p < 0.00001) but not with any single polymorphism. In addition, TNF-alpha serum levels correlated with rs1799724 (p = 0.034). A genetically mediated up-regulation of TNF-alpha expression might provoke a pronounced inflammation of the airways and thus a more severe course of RSV infection as well as the onset of asthma. It remains to be elucidated whether severe RSV bronchiolitis starts TNF-alpha upregulation and is one first step in the direction to asthma later in life, or whether both diseases are independent from each other and supported by TNF-alpha upregulation.
Zhang M, etal., J Paediatr Child Health. 2016 Jan;52(1):25-9. doi: 10.1111/jpc.12985. Epub 2015 Aug 19.
AIM: Respiratory syncytial virus (RSV) infection is a major health problem in young children. This study aimed to investigate whether the interleukin (IL)-4 -590C/T and -33 C/T polymorphisms were associated with the susceptibility to RSV infection in Chinese Ha
n children and with the severity of the infection. METHODS: The IL-4 -590C/T and -33 C/T SNPs were evaluated in 218 inpatients with RSV bronchiolitis and 303 healthy controls. The severity of RSV bronchiolitis was evaluated using a respiratory scoring system. RESULTS: The frequencies of IL-4 -590 CC, CT and TT in the 218 RSV bronchiolitis patients versus the 303 controls were 1.4% versus 1.7%, 21.1% versus 34% and 77.5% versus 64.4%, respectively. The frequencies of IL-4 -33 CC, CT and TT were 1.4%, 20.6% and 78%, respectively, in RSV bronchiolitis patients and 2.3%, 35.6% and 62%, respectively, in the controls. The frequencies of the IL-4 -590C/T and -33 C/T polymorphisms were significantly different between the two groups. There was a significant difference in the frequency of the T-T haplotype formed by -590C/T and -33C/T between the two groups. The respiratory score of the RSV bronchiolitis cases with -590TT was 3.02 +/- 0.44, which was significantly higher than those with -590CT (2.80 +/- 0.44). The score for patients with -33TT was 3.03 +/- 0.43, which was significantly higher than the score of 2.78 +/- 0.46 for patients with -33CT. CONCLUSIONS: In Chinese Han children, the IL-4 -590C/T and -33 C/T polymorphisms were associated with the susceptibility to RSV and the severity of the infection.
Monick MM, etal., J Immunol. 2007 Aug 1;179(3):1648-58.
Respiratory syncytial virus (RSV) is a ubiquitous virus that preferentially infects airway epithelial cells, causing asthma exacerbations and severe disease in immunocompromised hosts. Acute RSV infection induces inflammation in the lung. Thymus- and activation-
regulated chemokine (TARC) recruits Th2 cells to sites of inflammation. We found that acute RSV infection of BALB/c mice increased TARC production in the lung. Immunization of BALB/c mice with individual RSV proteins can lead to the development of Th1- or Th2-biased T cell responses in the lung after RSV infection. We primed animals with a recombinant vaccinia virus expressing either the RSV fusion (F) protein or the RSV attachment (G) protein, inducing Th1- and Th2-biased pulmonary memory T cell responses, respectively. After RSV infection, TARC production significantly increased in the vaccinia virus G-primed animals only. These data suggest a positive feedback loop for TARC production between RSV infection and Th2 cytokines. RSV-infected lung epithelial cells cultured with IL-4 or IL-13 demonstrated a marked increase in the production of TARC. The synergistic effect of RSV and IL-4/IL-13 on TARC production reflected differential induction of NF kappa B and STAT6 by the two stimuli (both are in the TARC promoter). These findings demonstrate that RSV induces a chemokine TARC that has the potential to recruit Th2 cells to the lung.
Cheng X, etal., Am J Med Genet A. 2005 Nov 15;139(1):13-8.
Genetic and auditory studies of 731 children with severe-to-profound hearing loss in US schools for the deaf and 46 additional children receiving clinical services for hearing loss ranging from moderate to profound demonstrated that mutations in the connexin 26 (GJB2) and connexin 30 (GJB6) genes ex
plain at least 12% of those with nonsyndromic sensorineural deafness. Otoacoustic emissions (OAEs) testing to detect functional outer hair cells indicated that 76 of the children had emissions and therefore may have (as yet unconfirmed) auditory neuropathy/dys-synchrony (AN/AD). Five of these children with OAEs were GJB2 homozygotes or compound heterozygotes with the genotypes 35delG/35delG, W77X/W77X, 35delG/360delGAG, 35delG/V95M, and V84M/M34T. In particular, unilateral AN/AD was confirmed in a child with moderate hearing loss and the 35delG/V95M genotype. Detecting OAEs in individuals with GJB2 mutations suggests that lack of functional gap junctions as a result of GJB2 mutations does not necessarily destroy all outer hair cell function.
Respiratory syncytial virus (RSV) glycoprotein G mimics fractalkine, a CX(3)C chemokine, which mediates chemotaxis of leukocytes expressing its receptor, CX(3)CR1. The aim of this study was to examine the relationship between RSV infection and expression of perf
orin and IFN-gamma in CX(3)CR1-expressing peripheral blood CD8(+) T cells. Samples were collected from infants with RSV bronchiolitis, both in the acute and convalescence phase (n = 12), and from their age- and sex-matched healthy controls (n = 15). Perforin expression and IFN-gamma secretion in CX(3)CR1(+) CD8(+) T cells were assessed by four-color flow cytometry. The NF-kappaB p50 and p65 subunit levels were also determined as markers of RSV-induced inflammation. Study results showed perforin and CX(3)CR1 expression to be significantly lower in the convalescent phase of infected infants than in healthy controls. There was no significant difference in IFN-gamma secretion and NF-kappaB binding activity between two time-points in RSV-infected infants, or when compared with healthy controls. Infants with prolonged wheezing had lower acute-phase CX(3)CR1 levels in peripheral blood. These data indicate existence of an event persisting after acute RSV infection that is able to modulate effector functions of cytotoxic T cells, and also link disease severity with CX(3)CR1 expression.
Zhao DY, etal., Zhonghua Er Ke Za Zhi. 2008 Feb;46(2):89-93.
OBJECTIVE: Respiratory syncytial virus (RSV) infects nearly all children under two years of age. It is poorly understood why a few children who were infected with RSV develop bronchiolitis that require hospital admission while most have a relatively minor illnes
s. Several recent studies have obtained some indications for the involvement of genetic heterogeneity in RSV bronchiolitis, implying that the clinical outcome of RSV infection perhaps is determined by genetic factors. Regulated on activation, normal T cell expressed and secreted RANTES plays a key role in the pathophysiology of RSV bronchiolitis. The purpose of this study was to explore the genetic association between the RANTES gene promoter -28C/G polymorphism and RSV bronchiolitis in Chinese Han ethnic group population. METHODS: The study recruited 238 hospitalized patients (186 male and 52 female) under 12 months of age, with a clinical diagnosis of bronchiolitis due to RSV, the sex, age, hospital stay, SaO2 at the time of admission, personal and family history of atopy were recorded. The 288 healthy control subjects (206 male and 82 female), who had no evidence of personal or familial history of atopy and no history of wheezing, were chosen at the same time. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was used to identify the polymorphism at position -28C/G of the RANTES promoter. The total IgE concentrations in serum samples were measured by enzyme-linked immunosorbent assay (ELISA). The absolute peripheral blood eosinophil counts were measured by using an automated hematology analyzer. RESULTS: The distribution of RANTES -28C/G gene polymorphism was in accordance with Hardy-Weinberg equilibrium. Compared to control subjects, significant difference was demonstrated for genotypes and allele frequencies of the RANTES -28C/G polymorphism in patients with RSV bronchiolitis (G = 10.22, P < 0.01; chi2 = 9.708, P < 0.01). Compared with the wild type CC, the -28G allele carriers demonstrated a 2.09-fold increased risk of RSV bronchiolitis (OR = 2.09, 95% CI = 1.32 - 3.30, P < 0.01). Interestingly, both the percentage of personal history of atopy and the percentage of family history of atopy for the -28G allele carriers were significantly higher (P < 0.05) than that for those CC homozygotes carriers in RSV bronchiolitis. Compared with the wild type CC, the -28G allele carriers demonstrated a 1.85-fold increased risk of the personal history of atopy (OR = 1.85, 95% CI = 1.01 - 3.38, P = 0.045) and a 1.91-fold increased risk of the family history of atopy (OR = 1.91, 95% CI = 1.03 - 3.54, P = 0.037), and the absolute peripheral blood eosinophil counts for the -28G allele carriers were significantly higher (P < 0.05). CONCLUSION: The RANTES gene promoter -28C/G polymorphism is associated with the susceptibility to RSV bronchiolitis, and the -28G allele is an important predisposing factor for the personal history of atopy and the family history of atopy in RSV bronchiolitis.
Zhang YL, etal., Zhongguo Dang Dai Er Ke Za Zhi. 2009 Oct;11(10):821-4.
OBJECTIVE: To study the association of tumor necrosis factor (TNF)-alpha-308G/A and interleukin (IL)-6-174G/C gene polymorphisms with the susceptibility of respiratory syncytial virus (RSV) bronchiolitis. METHODS: Polymerase chain reaction-restriction fragment l
ength polymorphism ( PCR-RFLP) was used to identify the polymorphisms at position 308 of the TNF-alpha promoter and -174 of the IL-6 promoter in 150 children with RSV bronchiolitis and 120 healthy children (control group). RESULTS: The frequency of GG, GA and AA genotypes of TNF-alpha-308 was 68.0%, 28.0% and 4.0% respectively in the RSV group, while that was 80.8%, 19.2% and 0 respectively in the control group. There were significant differences in the genotype frequencies between the two groups (chi2=5.665, p<0.05). The frequency of G and A alleles in the RSV group was 82.0% and 18.0% respectively, while that was 90.4% and 9.6% respectively in the control group. The frequency of A allele in the RSV group was significantly higher than that in the control group (chi2=7.726, p<0.05). Compared to the children carrying G allele, the children with A allele demonstrated a 2.071-fold increased risk of RSV bronchiolitis (OR=2.071, p<0.05). Only GG genotype was detected at position -174 of IL-6 in both groups. CONCLUSIONS: TNF-alpha-308A is associated with the susceptibility of RSV bronchiolitis and it might be an important candidate gene for the development of RSV bronchiolits. IL-6-174G/C gene polymorphism is not found in children in Wenzhou area.
Xia H, etal., Nan Fang Yi Ke Da Xue Xue Bao. 2009 Apr;29(4):724-8.
OBJECTIVE: To investigate the effect of respiratory syncytial virus (RSV) infection on the production of thymic stromal lymphopoietin (TSLP) and Th1/Th2 balance in asthmatic mice. METHODS: Thirty-two female BALB/c mice were randomly divided
into 4 groups, namely the PBS group, ovalbumin (OVA) group, RSV group and OVA/RSV group. The mice were sensitized by OVA and then stimulated with nebulized OVA, and RSV was inoculated into the nasal cavity of the mice. BUXCO noninvasive lung function detection was performed to examine the airway response to metacholine, and enzyme-linked immunosorbent assay (ELISA) was used to detect the serum levels of IL-4, IL-5, IL-13, and IFN-gamma in the mice. The cells in the bronchoalveolar lavage fluid (BALF) were counted and classified, and the supernatants of the BALF were used for the detection of TSLP. Histopathological changes in the lung tissues of the mice were examined using HE staining, and immunohistochemistry using anti-mouse TSLP antibody was performed to examine TSLP expressions in the airway epithelial cells. RESULTS: RSV infection promoted the production of TSLP in the asthmatic mice, and the concentration of TSLP in OVA/RSV group (2.13-/+0.05 ng/ml) was significantly higher than that in the other groups (P<0.01). RSV infection increased the serum levels of IL-4, IL-5, IL-13, and IFN-gamma in the mice. The total BALF cells, eosinophils, lymphocytes and neutrophils in OVA/RSV group were significantly higher than those in the other groups; noninvasive lung function examination showed higher Penh value in OVA/RSV group (318.66-/+50.87) than in the other groups when the inhaled metacholine increased to 6.25 mg/ml (P<0.01). More obvious and extensive airway inflammatory cell infiltration in OVA/RSV group were observed, and immunohistochemical staining also showed higher expression of TSLP in the airway epithelial cells of OVA/RSV group. CONCLUSIONS: RSV infection promotes the production of TSLP in the airway epithelial cells and increases the level of Th2 cytokines in asthmatic mice. Concurrent RSV infection can exacerbate Th2 inflammatory reaction in asthmatic mice.
Tan H, etal., Zhong Nan Da Xue Xue Bao Yi Xue Ban. 2009 Dec;34(12):1189-95.
OBJECTIVE: To elucidate intracellular transcription factor activation of C(7) approximately T(5) dorsal root ganglia in rats recurrently infected with respiratory syncytial virus (RSV). METHODS: Eighty 1 approximately 2 weeks old Sprague-Daw
ley rats were randomly divided into 2 groups: a control group and a RSV-infection group. The rats in the RSV-infection group were infected with 5 x 10(5) U/mL RSV once a week and the rats in the control group were treated with culture medium without RSV. Airway response was measured after 8 weeks. Lung tissue was submitted for HE staining and in situ hybridization. The C(7) approximately T(5) dorsal root ganglia were obtained for the preliminary screening of the intracellular transcription factors by TranSignal protein/DNA combo array. Nuclear protein of C(7) approximately T(5) dorsal root ganglia were extracted and submitted to Western blot. RESULTS: Airway response in the RSV-infection group was higher than that in the control group (P<0.05). HE staining showed inflammatory cell infiltration, and in situ hybridization demonstrated positive RSV RNA in the RSV-infection rat lung which was not present in the control group, thus validating the efficacy of our model. TranSignal protein/DNA combo array screening showed that 55 transcription factors increased by at least 2 folds in the C(7) approximately T(5) DRG cells of the RSV-infection group. The transcription factors Smad and interferon regulatory factor (1 or 2) were the 2 most upregulated transcription factors identified by combo array screening (59 and 43 fold increase compared with the control, respectively). Western blot confirmed Smad(1/2/3) and IRF-1 upregulate while IRF-2 remained unchanged. CONCLUSION: Respiratory syncytial virus infection results in airway hyperresponsiveness and transcription factor activation in C(7) approximately T(5) spinal adorsal root ganglia in rats, which may contribute to airway nerve network dysfunction and airway hyperresponsiveness.
Gutierrez-Roelens I, etal., Eur J Hum Genet. 2006 Dec;14(12):1313-6. Epub 2006 Aug 9.
The prevalence of congenital heart defects is approximately 1% of all live births. Identifying the genes responsible for cardiac malformation is the first step to understand pathogenesis. Heterozygous mutations in the CSX/NKX2-5 (NKX2E) gene have been identified to cause atrial septal defect (ASD) a
nd/or atrioventricular (AV) conduction disturbance in some families. However, there is great variability in expressivity of the phenotype between the patients with a CSX/NKX2-5 mutation. We screened four sporadic patients and three index cases of families with ASD and/or conduction defects. In one of them, a CSX/NKX2-5 mutation was identified. This novel mutation (p.Tyr256X) was inherited in a three-generation family causing five individuals to have cardiac anomalies ranging from ASD to arrhythmias. Interestingly, all the observed AV conduction disturbances were at the nodal level, manifesting first as an AV block of the first degree and evolving toward a second-degree block. Atrial fibrillation, previously reported in three individuals with CSX/NKX2-5 mutations, was observed in three patients.
Patoine D, etal., Can J Cardiol. 2011 Mar-Apr;27(2):263.e5-12. doi: 10.1016/j.cjca.2010.12.045.
BACKGROUND: A 43-year-old woman suffering from Steinert syndrome was admitted after experiencing multiple episodes of torsades de pointes-related syncope. OBJECTIVES: To elucidate the pathophysiology of these arrhythmic events. METHODS AND RESULTS: We obtained
DNA from the patient and sequenced the coding region of KCNQ1, KCNH2, SCN5A, KCNE1, and KCNE2 genes. A single nucleotide change was identified in the KCNQ1 gene at position 608 (T608C), resulting in a substitution from leucine to proline at position 203 (L203P). CHO cells were used to express either wild-type KCNQ1, wild-type KCNQ1+L203P KCNQ1 (50:50), or L203P KCNQ1, along with KCNE1 to recapitulate the slow cardiac delayed rectifier potassium current (I(Ks)). Patch-clamp experiments showed that the variant L203P causes a dominant negative effect on I(Ks). Coexpression of wild-type KCNQ1 and L203P KCNQ1 (50:50) caused a ~75% reduction in current amplitude when compared to wild-type KCNQ1 alone (131.40 +/- 23.27 vs 567.25 +/- 100.65 pA/pF, P < .001). Moreover, when compared with wild-type KCNQ1 alone, the coexpression of wild-type KCNQ1 and L203P KCNQ1 (50:50) caused a 7.5-mV positive shift of midpoints of activation (from 27.5 +/- 2.4 to 35.1 +/- 1.2 mV, P < .05). The wild-type KCNQ1 and L203P KCNQ1 (50:50) coexpression also caused alteration of I(Ks) kinetics. The activation kinetics of the L203P variant (50:50) were slowed compared with wild-type KCNQ1, while the deactivation kinetics of L203P (50:50) were accelerated compared with wild type, all these further contributing to the "loss-of-function" phenotype of I(Ks) associated with the variant L203P. CONCLUSION: Torsades de pointes and episodes of syncope are very likely to be due to the KCNQ1 variant L203P found in this patient.
Zheng J, etal., Virol Sin. 2015 Jun;30(3):190-9. doi: 10.1007/s12250-015-3597-0. Epub 2015 Jun 19.
HEp-2 cells persistently infected with respiratory syncytial virus (RSV) are a heterogeneous mixture of viral antigen-positive and -negative variants; however, the mechanism through which viral replication becomes latent remains unclear. In this study, we inve
stigated the potential mechanism by which RSV escapes from innate immune surveillance. Persistent-infected RSV HEp-2 cells were isolated and cell clones were passaged. The RSV-persistent cells produced viruses at a lower titer, resisted wild-type RSV re-infection, and secreted high levels of interferon-ss (IFN-ss), macrophage inflammatory protein-1alpha (Mip-1alpha), interleukin-8 (IL-8), and Rantes. Toll-like receptor 3 (TLR3), retinoic acid inducible gene-I (RIG-I), and suppressor of cytokine signaling 1 (SOCS1) levels were upregulated in these cells. The silencing of TLR3 mRNA decreased the expression of SOCS1 protein and the secretion of cytokines. RSV-persistent cells are in an inflammatory state; upregulation of SOCS1 is related to the TLR3 signaling pathway, which could be associated with the mechanism of viral persistence.
Respiratory syncytial virus (RSV) causes serious respiratory illness in infants and elderly. RSV infection induces short-lived immunity, which leaves people prone to re-infection. In contrast, the RSV fusion (F) protein formulated with a novel adjuvant (∆F/TriAd
j) elicits long term protective immunity. A comparison of RSV-immunized mice to mice vaccinated with a single dose of ∆F/TriAdj showed no difference in IgG1 and IgG2a production; however, local IgA secreting memory B cell development and B cell IgA production were significantly lower in RSV vaccinated mice than in ∆F/TriAdj-immunized mice. This indicates a potential reason as to why long-term immunity is not induced by RSV infection. The comparison also revealed that germinal center lymphocyte populations were higher in ∆F/TriAdj-vaccinated mice. Furthermore, ∆F/TriAdj induced higher gene expression of activation-induced cytidine deaminase (AID), as well as IL-6, IL-21, TGF-β cytokines, which are key players in IgA class switch recombination, ultimately leading to a sustained long-term memory response.
In mammalian testes, "A-single" spermatogonia function as stem cells that sustain sperm production for fertilizing eggs. Yet, it is not understood how cellular niches regulate the developmental fate of A-single spermatogonia. Here, immunolabeling studies in rat testes define a novel population of ER
BB3(+) germ cells as approximately 5% of total SNAP91(+) A-single spermatogonia along a spermatogenic wave. As a function of time, ERBB3(+) A-single spermatogonia are detected during a 1- to 2-day period each 12.9-day sperm cycle, representing 35%-40% of SNAP91(+) A-single spermatogonia in stages VIII-IX of the seminiferous epithelium. Local concentrations of ERBB3(+) A-single spermatogonia are maintained under the mean density measured for neighboring SNAP91(+) A-single spermatogonia, potentially indicative of niche saturation. ERBB3(+) spermatogonia also synchronize their cell cycles with epithelium stages VIII-IX, where they form physical associations with preleptotene spermatocytes transiting the blood-testis barrier and Sertoli cells undergoing sperm release. Thus, A-single spermatogonia heterogeneity within this short-lived and reoccurring microenvironment invokes novel theories on how cellular niches integrate with testicular physiology to orchestrate sperm development in mammals.
Gutierrez RC, etal., Neuroscience. 2009 Aug 4;162(1):208-21. doi: 10.1016/j.neuroscience.2009.04.062. Epub 2009 May 3.
The neuroligin (NL) gene family codes for brain specific cell adhesion molecules that play an important role in synaptic connectivity. Recent studies have identified NL mutations linked to patients with autism spectrum disorders (ASD). Cognitive deficits seen in autistic patients are hypothesized
to arise from altered synchronicity both within and between brain regions. Here we show how the expression of autism-associated neuroligin mutation R471C-NL3 affects synchrony in dissociated cultures of rat hippocampal neurons. Spontaneous network activity patterns of cultures expressing wild type and mutant NL3 were measured by optical techniques. Firing events were quantified and compared by cross-correlation analysis. Our results suggest that NL3 overexpression enhances synchrony of spontaneous activity patterns, however, this ability is reduced with the R471C-NL3 mutation. We investigated the structural basis of this phenomenon using fractal dimension analysis to characterize the arrangement of axon trajectories. R471C-NL3 cultures were associated with lower fractal dimensions and higher lacunarity values, indicating a decrease in the complexity of axonal architecture. Transfection of R471C-NL3 into a subpopulation of cells in a network resulted in neuronal degeneration. This degeneration likely affected the inhibitory population of neurons, as there were half as many (P<0.01, n=12) glutamate decarboxylase (GAD) 65 expressing cells in R471C-NL3 cultures compared to wild type NL3 and control cultures. Electrophysiological recordings showed a reduction of inhibitory activity in networks carrying the mutation in comparison to networks overexpressing wild-type NL3. Together, these data support the hypothesis that the autism-associated NL3 mutation affects information processing in neuronal networks by altering network architecture and synchrony.
Bannai H, etal., J Biol Chem. 2004 Dec 17;279(51):53427-34. Epub 2004 Oct 8.
mRNA transport and local translation in the neuronal dendrite is implicated in the induction of synaptic plasticity. Recently, we cloned an RNA-interacting protein, SYNCRIP (heterogeneous nuclear ribonuclear protein Q1/NSAP1), that is suggested to be important f
or the stabilization of mRNA. We report here that SYNCRIP is a component of mRNA granules in rat hippocampal neurons. SYNCRIP was mainly found at cell bodies, but punctate expression patterns in the proximal dendrite were also seen. Time-lapse analysis in living neurons revealed that the granules labeled with fluorescent protein-tagged SYNCRIP were transported bi-directionally within the dendrite at approximately 0.05 microm/s. Treatment of neurons with nocodazole significantly inhibited the movement of green fluorescent protein-SYNCRIP-positive granules, indicating that the transport of SYNCRIP-containing granules is dependent on microtubules. The distribution of SYNCRIP-containing granules overlapped with that of dendritic RNAs and elongation factor 1alpha. SYNCRIP was also found to be co-transported with green fluorescent protein-tagged human staufen1 and the 3'-untranslated region of inositol 1,4,5-trisphosphate receptor type 1 mRNA. These results suggest that SYNCRIP is transported within the dendrite as a component of mRNA granules and raise the possibility that mRNA turnover in mRNA granules and the regulation of local protein synthesis in neuronal dendrites may involve SYNCRIP.
van der Wal GE, etal., Ann Surg. 2012 Jan;255(1):86-94. doi: 10.1097/SLA.0b013e318238346a.
OBJECTIVE: Resection of a primary colorectal carcinoma (CRC) can be accompanied by rapid outgrowth of liver metastases, suggesting a role for angiogenesis. The aim of this study is to investigate whether the presence of a primary CRC is associated with changes in angiogenic status and pro
liferation/apoptotic rate in synchronous liver metastases and/or adjacent liver parenchyma. METHODS: Gene expression and localization of CD31, HIF-1α, members of the vascular endothelial growth factor (VEGF) and Angiopoietin (Ang) system were studied using qRT-PCR and immunohistochemistry in colorectal liver metastases and nontumorous-adjacent liver parenchyma. Proliferation and apoptotic rate were quantified. Three groups of patients were included: (1) simultaneous resection of synchronous liver metastases and primary tumor (SS-group), (2) resection of synchronous liver metastases 3 to 12 months after resection of the primary tumor [late synchronous (LS-group)], and (3) resection of metachronous metastases >14 months after resection of the primary tumor (M-group). RESULTS: In all 3 groups a higher expression of the angiogenic factors was encountered in adjacent liver parenchyma as compared to the metastases. VEGFR-2 gene expression was abundant in adjacent liver parenchyma in all 3 groups. VEGF-A and VEGFR-1 were prominent in adjacent parenchyma in the SS-group. The SS-group showed the highest Ang-2/Ang-1 ratio both in the metastases and the adjacent liver. This was accompanied by a high turnover of tumor cells. CONCLUSION: In the presence of the primary tumor, the liver parenchyma adjacent to the synchronous liver metastases provides an angiogenic prosperous environment for metastatic tumor growth.
Tal G, etal., J Infect Dis. 2004 Jun 1;189(11):2057-63. Epub 2004 May 12.
BACKGROUND: The clinical spectrum of respiratory syncytial virus (RSV) bronchiolitis in previously healthy infants is extremely variable. Thus, it is likely that factors such as genetic heterogeneity contribute to disease severity. Toll-like receptor 4 (TLR4) an
d CD14 are part of a receptor complex involved in the innate immune response to RSV. METHODS: The association of the TLR4 mutations (Asp299Gly and Thr399Ile) and the CD14/-159 polymorphism were analyzed in 99 infants hospitalized with severe RSV bronchiolitis (group I). Eighty-two ambulatory infants with mild RSV bronchiolitis (group II) and 90 healthy adults (group III) composed the 2 control groups. The TLR4 mutations and the CD14/-159 polymorphism were genotyped by use of reverse-transcriptase polymerase chain reaction and restriction fragment-length polymorphism analysis, respectively. RESULTS: Each of the TLR4 mutations, either alone or in cosegregation, were associated with severe RSV bronchiolitis: the Asp299Gly and Thr399Ile mutations were significantly overrepresented in group I, compared with groups II and III. No association between the CD14/-159 polymorphism and RSV bronchiolitis was found. CONCLUSIONS: These findings suggest that TLR4 mutations, but not the CD14/-159 polymorphism, are associated with an increased risk of severe RSV bronchiolitis in previously healthy infants.
Gulec C, etal., Anadolu Kardiyol Derg. 2014 Nov;14(7):617-24. doi: 10.5152/akd.2014.4972. Epub 2014 Feb 10.
OBJECTIVE: Homeodomain Only Protein X (HOPX) is an unusual homeodomain protein which regulates Serum Response Factor (SRF) dependent gene expression. Due to the regulatory role of HOPX on SRF activity and the regulatory role of SRF on cardiac hypertrophy, we aimed to investigate the relationship bet
ween HOPX gene variations and hypertrophic cardiomyopathy (HCM). METHODS: In this study, designed as a case-control study, we analyzed coding and flanking non-coding regions of the HOPX gene through 67 patients with HCM and 31 healty subjects. Certain regions of the gene were investigated by Single Stranded Conformation Polymorphism (SSCP) and Restriction Fragment Length Polymorphism (RFLP). Statistical analyses of genotypes and their relationship with clinical parameters were performed by chi-square, Kruskal-Wallis and the Fisher's exact test. RESULTS: In 5' Untranslated Region (UTR) and intronic region of the HOPX gene, we found a C>T substitution and an 8-bp insertion/deletion (In/Del) polymorphism, respectively. These two polymorphisms seemed to constitute an haplotype. While the frequency of homozygous genotypes of In/Del and C/T polymorphisms were found significantly lower in the patients with syncope (p=0.014 and p=0.017, respectively), frequency of their heterozygous genotypes were found significantly higher in the patients with syncope (p=0.048 and p=0.030, respectively). CONCLUSION: Though there was not found any mutation in coding sequence of HOPX gene, two non-coding polymorphisms were found related to syncope in HCM patients. While homozygous status of these polymorphisms was found to be protective against the syncope, their heterozygous status seemed to be a risk factor for syncope in HCM patients. Our results suggest that HOPX may contribute to pathogenesis or manifestation of HCM as a modifier gene.
Hoebee B, etal., J Infect Dis. 2003 Jan 1;187(1):2-11. Epub 2002 Dec 13.
The association of variants of genes encoding interleukin (IL)-4 and the IL-4 receptor alpha chain (IL-4Ralpha) with respiratory syncytial virus (RSV) bronchiolitis was examined in hospitalized infants. Polymorphisms in IL-4 (C-590T) and IL-4Ralpha (I50V and Q55
1R) were genotyped by restriction fragment-length polymorphism analysis. Control subjects included parents of the hospitalized children (for the transmission/disequilibrium test), and a random population sample (for the case-control study). Results were also analyzed in a combination of these 2 tests, using Fisher's method. The IL-4 590T allele was found more frequently among children hospitalized with RSV than expected in the case-control (odds ratio [OR], 1.43; P=.04) and combination (OR, 1.41; P=.02) tests. Among children who were >6 months old when they were hospitalized, compared with the control group or with the <6 months old who were hospitalized for RSV infection, higher frequencies of both the IL-4 590T allele and the IL-4Ralpha R551 allele were found. These results indicate that gain-of-function variants of T helper type 2 cytokine genes may play a role in increasing the severity of RSV disease, which appears more pronounced after the first half-year of life.
Respiratory syncytial virus (RSV) is a leading cause of infant mortality worldwide. Although anti-RSV Ab prophylaxis has greatly reduced infant mortality in the United States, there is currently no vaccine or effective antiviral therapy. RSV fusion (F) protein a
ctivates cells through TLR4. Two single nucleotide polymorphisms (SNPs) encoding Asp299Gly and Thr399Ile substitutions in the TLR4 ectodomain were previously associated with TLR4 hyporesponsiveness and increased susceptibility to bacterial infection. Prevalence of these SNPs was analyzed in a case series of 105 DNA samples extracted from archived nasal lavage samples from high-risk infants/young children with confirmed RSV disease who participated in two seminal clinical trials for anti-RSV prophylaxis. Frequencies of TLR4 SNPs in the case series were compared with those of literature controls, healthy adults, infants, and young children who presented with symptoms of respiratory infections (but not preselected for high risk for RSV). Both SNPs were highly associated with symptomatic RSV disease in this largely premature population (p < 0.0001), with 89.5% and 87.6% of cases being heterozygous for Asp299Gly and Thr399Ile polymorphisms versus published control frequencies of 10.5% and 6.5%, respectively. The other two control groups had similarly low frequencies. Our data suggest that heterozygosity of these two extracellular TLR4 polymorphisms is highly associated with symptomatic RSV disease in high-risk infants and support a dual role for TLR4 SNPs in prematurity and increased susceptibility to RSV not revealed by analysis of either alone.
Evin L, etal., Bratisl Lek Listy. 2016;117(3):142-7.
OBJECTIVES: The aim of this prospective study was to investigate the impact of genetic polymorphisms of beta3 subunit of G-protein on the occurrence of vasovagal syncope, hemodynamic parameters and heart rate variability during head-up tilt test (HUT). BACKGROUN
D: G-proteins play an important role in the intracellular transmission of impulses in cardiovascular autonomic reflexes. METHODS: In 157 patients with suspected vasovagal syncope HUT was performed. Ninety-one patients (38 men, 53 women, mean age 48 +/- 17 years) had positive HUT. Control group consisted of 109 subjects (69 men, 40 women, mean age 37 +/- 16 years) with no history of syncope. Results of HUT, hemodynamic parameters and LF, HF, LF/HF, SDNN, RMSSD parameters of heart rate variability were compared in patients with different genotypes. C825T polymorphism of beta3 subunit of G-protein was determined in the study subjects. RESULTS: There was no significant difference in the distribution of genotypes between patients and control group. Also, there was no significant difference in hemodynamic parameters. A statistically significant difference was found between genotypes in LF/HF in the early HUT (mean rank CC: 48.68 vs CT: 35.51 vs TT: 34.14; p = 0.039) and at RMSSD at the time of syncope (mean rank CC: 32.38 vs CT: 42.74 vs TT: 18.50; p = 0.026). CONCLUSIONS: In this study, the relation of C825T polymorphism of beta3 subunit of G-protein to vasovagal syncope was not documented (Tab. 2, Fig. 4, Ref. 37).
Bone morphogenetic protein-1 (BMP-1)/tolloid-like proteinases, here called BTPs, include the proteases originally identified for their roles in the C-terminal maturation of fibrillar procollagens ("procollagen C-proteinase"). Though numerous other substrates have since been discovered, the BTPs rem
ain the main proteases involved in extracellular matrix assembly with little or no implication in matrix degradation. During the same period however, the BTPs have also become established as important proteases in the activation of growth factors, including TGF-beta1, BMP-2/-4, GDF-8/-11 and IGFs, as well as the release of anti-angiogenic fragments from parent proteins. The BTPs are therefore key players in many pathophysiological processes such as morphogenesis, tissue repair and tumor progression. This mini-review summarizes our current knowledge of the functions of BTPs, their substrates and unusual mechanisms of regulation, and discusses their potential as new targets for future therapies.
Rossi CR and Kiesel GK, Am J Vet Res. 1977 Nov;38(11):1901-4.
Development of fluorescence in bovine embryonic lung cells infected with bovine respiratory syncytial virus (BRSV) was studied by the fluorescent antibody (FA) test. Similar patterns of fluorescence were seen with the direct FA test, in which the immunoglobulin
G fraction of antiserum to BRSV was conjugated with fluorescein isothiocyanate and used; and the indirect test, in which antiserum to the Long strain of respiratory syncytial virus and fluorescein isothiocyanate-conjugated anti-rabbit immunoglobulin G were used. In different trials, fluorescence was first detected between 16 and 18 hours after inoculation with BRSV. Fluorescence always was confined to the cytoplasm. Before 24 hours, fluorescence consisted of fine fibrils, usually parallel to the long axis of the cell, and cytoplasmic granules. After 24 hours, coincident with rounding of the cells, fluorescence slowly moved to the periphery of the cytoplasm. Under the growth conditions used, syncytia did not develop. By the FA test and as determined by the release of BRSV into the supernatant fluid, the minimal time for a single cycle of infection was between 24 and 26 hours.
Enhanced respiratory syncytial virus disease, a serious pulmonary disorder that affected recipients of an inactivated vaccine against respiratory syncytial virus in the 1960s, has delayed the development of vaccines against
the virus. The enhanced disease was characterized by immune complex-mediated airway hyperreactivity and a severe pneumonia associated with pulmonary eosinophilia. In this paper, we show that complement factors contribute to enhanced-disease phenotypes. Mice with a targeted disruption of complement component C5 affected by the enhanced disease displayed enhanced airway reactivity, lung eosinophilia, and mucus production compared to wild-type mice and C5-deficient mice reconstituted with C5. C3aR expression in bronchial epithelial and smooth muscle cells in the lungs of C5-deficient mice was enhanced compared to that in wild-type and reconstituted rodents. Treatment of C5-deficient mice with a C3aR antagonist significantly attenuated airway reactivity, eosinophilia, and mucus production. These results indicate that C5 plays a crucial role in modulating the enhanced-disease phenotype, by affecting expression of C3aR in the lungs. These findings reveal a novel autoregulatory mechanism for the complement cascade that affects the innate and adaptive immune responses.
Currie SM, etal., J Immunol. 2016 Mar 15;196(6):2699-710. doi: 10.4049/jimmunol.1502478. Epub 2016 Feb 12.
Respiratory syncytial virus (RSV) is a leading cause of respiratory tract infection in infants, causing significant morbidity and mortality. No vaccine or specific, effective treatment is currently available. A more complete understanding of the key components
of effective host response to RSV and novel preventative and therapeutic interventions are urgently required. Cathelicidins are host defense peptides, expressed in the inflamed lung, with key microbicidal and modulatory roles in innate host defense against infection. In this article, we demonstrate that the human cathelicidin LL-37 mediates an antiviral effect on RSV by inducing direct damage to the viral envelope, disrupting viral particles and decreasing virus binding to, and infection of, human epithelial cells in vitro. In addition, exogenously applied LL-37 is protective against RSV-mediated disease in vivo, in a murine model of pulmonary RSV infection, demonstrating maximal efficacy when applied concomitantly with virus. Furthermore, endogenous murine cathelicidin, induced by infection, has a fundamental role in protection against disease in vivo postinfection with RSV. Finally, higher nasal levels of LL-37 are associated with protection in a healthy human adult RSV infection model. These data lead us to propose that cathelicidins are a key, nonredundant component of host defense against pulmonary infection with RSV, functioning as a first point of contact antiviral shield and having additional later-phase roles in minimizing the severity of disease outcome. Consequently, cathelicidins represent an inducible target for preventative strategies against RSV infection and may inform the design of novel therapeutic analogs for use in established infection.
Inoue Y, etal., J Infect Dis. 2007 Jun 1;195(11):1618-24. Epub 2007 Apr 23.
BACKGROUND: The contribution that genetic polymorphisms of Toll-like receptor (TLR) 4 and of CD14--both of which recognize respiratory syncytial virus (RSV) in the innate immune response--make to RSV bronchiolitis in the Japanese population has not yet been clar
ified. METHODS: This study genotyped 2 TLR4 mutations, Asp299Gly and Thr399Ile, and 2 single-nucleotide polymorphisms (SNPs) of CD14, -550 C/T and -159 C/T, in 54 children with RSV bronchiolitis and in 203 control subjects. CD14 SNPs and the serum level of soluble CD14 (sCD14) also were examined in 67 cord-blood specimens and in serum samples from 73 6-year-old children. RESULTS: No TLR4 mutations were found. The frequencies of both the CC genotype and the C allele of CD14 -550 C/T were significantly higher in children with RSV bronchiolitis than in the control subjects. The serum level of sCD14 was significantly higher in children with the CC genotype of CD14 -550 C/T than in those with the CT and TT genotypes. CONCLUSIONS: CD14 -550 C/T, which is related to the serum level of sCD14, is associated with the development of RSV bronchiolitis in the Japanese population. This study's results indicate that, in different ethnic groups, different genetic factors contribute to the development of RSV bronchiolitis.
Heidema J, etal., J Immunol. 2007 Dec 15;179(12):8410-7.
A protective role for CD8+ T cells during viral infections is generally accepted, but little is known about how CD8+ T cell responses develop during primary infections in infants, their efficacy, and how memory is established after viral clearance. We studied CD8+ T cell responses in bronchoalveolar
lavage (BAL) samples and blood of infants with a severe primary respiratory syncytial virus (RSV) infection. RSV-specific CD8+ T cells with an activated effector cell phenotype: CD27+CD28+CD45RO+CCR7-CD38+HLA-DR+Granzyme B+CD127- could be identified in BAL and blood. A high proportion of these CD8+ T cells proliferated and functionally responded upon in vitro stimulation with RSV Ag. Thus, despite the very young age of the patients, a robust systemic virus-specific CD8+ T cell response was elicited against a localized respiratory infection. RSV-specific T cell numbers as well as the total number of activated effector type CD8+ T cells peaked in blood around day 9-12 after the onset of primary symptoms, i.e., at the time of recovery. The lack of a correlation between RSV-specific T cell numbers and parameters of disease severity make a prominent role in immune pathology unlikely, in contrast the T cells might be involved in the recovery process.
It is known that lymphopenia caused by apoptosis may occur during severe respiratory syncytial virus (RSV) infection. However, further evidence about how T-cell subsets may be affected in infants during severe RSV bronchiolitis is needed to understand the mechan
isms through which immunological memory may be altered. There is increasingly convincing evidence that RSV may be associated with the development of atopy and asthma. Surrogates of Th1, Th2 and regulatory T-lymphocyte populations were measured in blood from children with acute RSV bronchiolitis and in convalescence using the cell surface receptors CXCR3, CCR4 and CD25, respectively. Samples were also obtained from healthy age-matched controls. Plasma levels of the chemokines interferon-gamma inducible protein-10 (IP-10) and thymus and activation-regulated chemokine (TARC), which are known ligands for CXCR3 and CCR4, were also measured. Free plasma DNA was measured using quantitative PCR. CXCR3-positive cells were significantly decreased during acute infection (p = 0.013), while CCR4 and CD25 T-cell populations were unchanged. Plasma levels of IP-10 were markedly elevated in acute infection (p = 0.001). Convalescent samples were not significantly different to control samples for lymphocyte phenotypes or plasma chemokines. Elevated free plasma DNA was detected during acute infection compared with convalescence and controls. A profound reduction in the Th1, but not Th2, and CD25-positive lymphocyte populations associated with exaggerated IP-10 production occurs in severe RSV bronchiolitis. Free DNA is detectable in plasma. This may allow significant alterations in the generation of T-cell memory.
Grundschober C, etal., J Biol Chem 2001 Dec 14;276(50):46751-8.
Genes under a 24-h regulation period may represent drug targets relevant to diseases involving circadian dysfunctions. As a testing model of the circadian clock system, we have used synchronized rat fibroblasts that are known to express at least six genes in a c
ircadian fashion. We have determined the expression patterns of 9957 transcripts every 4 h over a total period of 76 h using high density oligonucleotide microarrays. The spectral analysis of our mRNA profiling data indicated that approximately 2% (85 genes) of all expressed genes followed a robust circadian pattern. We have confirmed the circadian expression of previously known clock or clock-driven genes, and we identified 81 novel circadian genes. The majority of the circadian-regulated gene products are known and are involved in diverse cellular functions. We have classified these circadian genes in seven clusters according to their phase of cycling. Our pathway analysis of the mRNA profiling data strongly suggests a direct link between circadian rhythm and cell cycle.
Wang SZ, etal., J Immunol. 2003 Jul 15;171(2):1051-60.
Clara cell secretory protein (CCSP) has been shown to have anti-inflammatory and immunomodulatory functions in the lung. Respiratory syncytial virus (RSV) is the most common cause of respiratory infection in infants and young children. RSV usually infects small
airways and likely interacts with the Clara cells of bronchioles. To determine a possible role for CCSP during acute RSV infection, CCSP-deficient (CCSP(-/-)) and wild-type (WT) mice were intratracheally infected with RSV and the lung inflammatory and immune responses to RSV infection were assessed. RSV-F gene expression was increased in the lungs of CCSP(-/-) mice as compared with WT mice following RSV infection, consistent with increased viral persistence. Lung inflammation was significantly increased in CCSP(-/-) mice as compared with WT mice after infection. Moreover, although the levels of Th1 cytokines were similar, the levels of Th2 cytokines and neutrophil chemokines were increased in the lungs of CCSP(-/-) mice following infection. Physiologic endpoints of exacerbated lung disease, specifically airway reactivity and mucus production, were increased in CCSP(-/-) mice after RSV infection. Importantly, restoration of CCSP in the airways of CCSP(-/-) mice abrogated the increased viral persistence, lung inflammation, and airway reactivity. These findings suggest a role for CCSP and Clara cells in regulating lung inflammatory and immune responses to RSV infection.
Evidence suggests that Toll-like receptor 4 (TLR4) contributes to immune recognition of respiratory syncytial virus (RSV). The TLR4 gene harbours a polymorphism-Asp299Gly-previously associated with reduced TLR4 signalling. To understand of how host genetic varia
tion influences the outcome of RSV infection in children, we examined the association between the TLR4 299Gly allele and severe RSV disease. By genotyping 236 children with RSV infection and 219 healthy controls we found no association between the risk of severe RSV infection and Asp299Gly polymorphisms (P>0.05), and we demonstrate that the TLR4 Asp299Gly genotype does not influence susceptibility to either RSV serotype A or B (P>0.05). Finally, examining the functional impact of the TLR4 Asp299Gly polymorphism (n=58), we demonstrate that proinflammatory cytokine production following TLR4 activation was indistinguishable between homozygous (Asp/Asp) and heterozygous (Asp/Gly) subjects. We conclude that the Asp299Gly TLR4 polymorphism does not alter receptor function and does not influence the risk of severe RSV infection.
Human respiratory syncytial virus (hRSV) is the leading cause of infant hospitalization related to respiratory disease. Infection with hRSV produces abundant infiltration of immune cells into the airways, which combined with an exacerbated pro-inflammatory immun
e response can lead to significant damage to the lungs. Human RSV re-infection is extremely frequent, suggesting that this virus may have evolved molecular mechanisms that interfere with host adaptive immunity. Infection with hRSV can be reduced by administering a humanized neutralizing antibody against the virus fusion protein in high-risk infants. Although neutralizing antibodies against hRSV effectively block the infection of airway epithelial cells, here we show that both, bone marrow-derived dendritic cells (DCs) and lung DCs undergo infection with IgG-coated virus (hRSV-IC), albeit abortive. Yet, this is enough to negatively modulate DC function. We observed that such a process is mediated by Fcgamma receptors (FcgammaRs) expressed on the surface of DCs. Remarkably, we also observed that in the absence of hRSV-specific antibodies FcgammaRIII knockout mice displayed significantly less cellular infiltration in the lungs after hRSV infection, compared with wild-type mice, suggesting a potentially harmful, IgG-independent role for this receptor in hRSV disease. Our findings support the notion that FcgammaRs can contribute significantly to the modulation of DC function by hRSV and hRSV-IC. Further, we provide evidence for an involvement of FcgammaRIII in the development of hRSV pathogenesis.
Yao S, etal., Mucosal Immunol. 2015 Jul;8(4):746-59. doi: 10.1038/mi.2014.106. Epub 2014 Dec 3.
Respiratory syncytial virus (RSV) infection is a leading cause of severe lower respiratory tract illness in young infants, the elderly and immunocompromised individuals. We demonstrate here that the co-inhibitory molecule programmed cell death 1 (PD-1) is selec
tively upregulated on T cells within the respiratory tract during both murine and human RSV infection. Importantly, the interaction of PD-1 with its ligand PD-L1 is vital to restrict the pro-inflammatory activities of lung effector T cells in situ, thereby inhibiting the development of excessive pulmonary inflammation and injury during RSV infection. We further identify that PD-L1 expression on lung inflammatory dendritic cells is critical to suppress inflammatory T-cell activities, and an interferon-STAT1-IRF1 axis is responsible for increased PD-L1 expression on lung inflammatory dendritic cells. Our findings suggest a potentially critical role of PD-L1 and PD-1 interactions in the lung for controlling host inflammatory responses and disease progression in clinical RSV infection.
Vieira RA, etal., J Bras Pneumol. 2010 Feb;36(1):59-66.
OBJECTIVE: To determine whether the concentrations of inflammatory mediators (CCL5, soluble intercellular adhesion molecule type 1 [sICAM-1], TNF-alpha, IL-6 and IL-10) in the nasopharyngeal secretion and in the serum of children with lower respiratory tract infection (LRTI) caused by respiratory ... (more)
pan style='font-weight:700;'>syncytial virus (RSV) correlate with the clinical markers of disease severity. METHODS: Between July of 2004 and December of 2005, 30 children less than three months of age, diagnosed with LRTI caused by RSV and admitted to a neonatal ICU, were included in this study. RESULTS: The severity of disease at hospital admission, as determined with a modified clinical scoring system, presented a significant positive correlation with sICAM-1 and IL-10 concentrations in the nasopharyngeal secretion, as well as with IL-6 concentrations in the serum, of the patients. In addition, serum IL-6 concentrations presented a significant positive correlation with the duration of oxygen therapy and with the length of hospital stay. CONCLUSIONS: At hospital admission, the concentrations of sICAM-1 and IL-10 in the nasopharyngeal secretion, as well as the concentration of IL-6 in the serum, could be used as markers of severity in patients with LRTI caused by RSV. The serum levels of IL-6 determined at admission could also be used to predict prolonged oxygen supplementation and hospital stay.
Chirkova T, etal., J Gen Virol. 2015 Sep;96(9):2543-56. doi: 10.1099/vir.0.000218. Epub 2015 Jun 25.
Respiratory syncytial virus (RSV) is a major cause of severe pneumonia and bronchiolitis in infants and young children, and causes disease throughout life. Understanding the biology of infection, including virus binding to the cell surface, should help develop
antiviral drugs or vaccines. The RSV F and G glycoproteins bind cell surface heparin sulfate proteoglycans (HSPGs) through heparin-binding domains. The G protein also has a CX3C chemokine motif which binds to the fractalkine receptor CX3CR1. G protein binding to CX3CR1 is not important for infection of immortalized cell lines, but reportedly is so for primary human airway epithelial cells (HAECs), the primary site for human infection. We studied the role of CX3CR1 in RSV infection with CX3CR1-transfected cell lines and HAECs with variable percentages of CX3CR1-expressing cells, and the effect of anti-CX3CR1 antibodies or a mutation in the RSV CX3C motif. Immortalized cells lacking HSPGs had low RSV binding and infection, which was increased markedly by CX3CR1 transfection. CX3CR1 was expressed primarily on ciliated cells, and approximately 50 % of RSV-infected cells in HAECs were CX3CR1+. HAECs with more CX3CR1-expressing cells had a proportional increase in RSV infection. Blocking G binding to CX3CR1 with anti-CX3CR1 antibody or a mutation in the CX3C motif significantly decreased RSV infection in HAECs. The kinetics of cytokine production suggested that the RSV/CX3CR1 interaction induced RANTES (regulated on activation normal T-cell expressed and secreted protein), IL-8 and fractalkine production, whilst it downregulated IL-15, IL1-RA and monocyte chemotactic protein-1. Thus, the RSV G protein/CX3CR1 interaction is likely important in infection and infection-induced responses of the airway epithelium, the primary site of human infection.
Jeong KI, etal., PLoS One. 2015 Jun 24;10(6):e0130517. doi: 10.1371/journal.pone.0130517. eCollection 2015.
Respiratory syncytial virus (RSV) is the principal cause of bronchiolitis in infants and a significant healthcare problem. The RSV Glycoprotein (G) mediates attachment of the virus to the cell membrane, which facilitates interaction of the RSV Fusion (F) protein
with nucleolin, thereby triggering fusion of the viral and cellular membranes. However, a host protein ligand for G has not yet been identified. Here we show that CX3CR1 is expressed in the motile cilia of differentiated human airway epithelial (HAE) cells, and that CX3CR1 co-localizes with RSV particles. Upon infection, the distribution of CX3CR1 in these cells is significantly altered. Complete or partial deletion of RSV G results in viruses binding at least 72-fold less efficiently to cells, and reduces virus replication. Moreover, an antibody targeting an epitope near the G protein's CX3CR1-binding motif significantly inhibits binding of the virus to airway cells. Given previously published evidence of the interaction of G with CX3CR1 in human lymphocytes, these findings suggest a role for G in the interaction of RSV with ciliated lung cells. This interpretation is consistent with past studies showing a protective benefit in immunizing against G in animal models of RSV infection, and would support targeting the CX3CR1-G protein interaction for prophylaxis or therapy. CX3CR1 expression in lung epithelial cells may also have implications for other respiratory diseases such as asthma.
The induction of inflammatory cytokines during respiratory viral infections contributes to both disease pathogenesis and resolution. The present studies investigated the role of the chemokine CXCL10 and its specific receptor, CXCR3, in the host response to pulmonary respiratory sync
ght:700;'>syncytial virus (RSV) infection. Antibody-mediated neutralization of CXCL10 resulted in a significant increase in disease pathogenesis, including airway hyperresponsiveness (AHR), mucus gene expression, and impaired viral clearance. When the pulmonary cytokine levels were examined, only type I IFN and IL-12p70 were significantly reduced. These latter observations were reflected in reduced dendritic cell (DC) numbers and DC maturation in the lungs of RSV-infected mice treated with anti-CXCL10. Neutralization of the only known receptor for CXCL10, CXCR3, resulted in similar increases in pathogenic responses. When bone marrow-derived DC were incubated with CXCL10 and RSV, an up-regulation of type I IFN was observed. In addition, T lymphocytes were also examined and a significant decrease in the number of RSV M2 peptide-specific CD8(+) T cells was identified. These findings highlight a previously unappreciated role for the CXCL10:CXCR3 signaling axis in RSV-infected animals by recruiting virus-specific T cells into the lung and promoting viral clearance.
Rudd BD, etal., J Immunol. 2006 Feb 1;176(3):1937-42.
The detection of a viral infection by pattern recognition receptors (PAMPs) is an integral part of antiviral immunity. In these studies we have investigated the role of TLR3, which recognizes dsRNA, in Respiratory Syncytial virus (RSV) infection using B6 backgro
und mice with a TLR3 deletion. Although we observed no changes in viral growth, we did find that TLR3-/- mice demonstrated significant increases in mucus production in the airways of RSV-infected mice. The qualitative assessment was observed by examining differentially stained lungs, followed by immunohistochemical staining for gob5, a mucus-associated protein. The histopathologic observations were verified using quantitative gene expression analyses examining gob5 gene expression. Changes in pulmonary mucus production were accompanied by an increase in pulmonary IL-13 as well as IL-5 expression and eosinophils in the airways of TLR3-/- mice. Examining leukocytes in the airway indicated an accumulation of eosinophils in TLR3-/- mice, but not wild-type mice, after RSV infection. Isolated lung draining lymph node cells from TLR3-/- mice produced significant increases in Th2-type cytokines, IL-5, and IL-13, compared with wild-type TLR3+/+ mice only after RSV infection. To demonstrate a causative link, we depleted TLR3-/- mice of IL-13 during RSV infection and found that mucus and gob5 expression in the lungs was attenuated. Together, these studies highlight that although TLR3 may not be required for viral clearance, it is necessary to maintain the proper immune environment in the lung to avoid developing pathologic symptoms of disease.
The interaction between mite allergen sensitization and respiratory syncytial virus (RSV) infection at the level of cytokine mRNA expression was examined in a murine model in the present study. Primary RSV infection enhances expression of inflammatory cytokines
such as IL-6, IFN-gamma, and eotaxin in the lung and upregulates the expression of Th2-like cytokines IL-10 and IL-13 in the spleen in BALB/c mice. Mite antigen-sensitized and RSV-infected (ASRSV) mice show enhanced (P < 0.05) total serum IgE compared to antigen-sensitized mice. However, the levels of viral mRNA in the lung tissues are comparable between RSV-infected and ASRSV mice. It is concluded that compartmentalization of cytokine expression following RSV infection plays a role in the augmentation of Th2-like and IgE antibody response to RSV.
After peripheral axotomy, synapses are eliminated from the somata of spinal motoneurons. Recent evidence indicates that synaptic adhesion molecules play a role in maintenance of synaptic contacts, but so far such molecules have not been investigated in the context of synapse elimination after injury
. In vitro, the neuroligins (NLGs) and SynCAM1 drive formation of synapses, and RNAi of NLGs results in decreased synaptic input, indicating an important role for these molecules in synaptic biology. To address potential involvement of NLGs and SynCAMs in postinjury synapse elimination, we investigated the mRNA expression of NLG1, -2, and -3; SynCAM1 and -3; and PSD-95--an intracellular NLG-binding scaffolding protein--in rat spinal motoneurons in control animals and after sciatic nerve transection (SNT). mRNA signals for NLG2, NLG3, SynCAM1, and SynCAM3, but not NLG1, were seen in uninjured motoneurons. Immunoreactivity for SynCAM was seen in close relation to synaptophysin immunoreactivity on the surface of motoneurons and in close relation to neurofilament immunoreactivity in the sciatic nerve. After axotomy, the signals for NLG2, NLG3, and SynCAM1 mRNAs decreased, whereas the signal for NLG1 mRNA remained undetectable and that for SynCAM3 remained at control levels. The signal for PSD-95 mRNA decreased gradually and reached approximately 50% of control values 2 weeks after axotomy. Thus the retrograde response to axotomy of spinal motoneurons involves a rapid down-regulation of NLG2, NLG3, and SynCAM1 mRNAs and a gradual decrease in PSD-95 mRNA. This indicates that down-regulation of synaptic adhesion molecules plays a role in postinjury synapse elimination.
Wu X, etal., Mol Med Rep. 2016 Jan;13(1):167-73. doi: 10.3892/mmr.2015.4518. Epub 2015 Nov 6.
The present study was designed to examine the expression and function of matrix metalloproteinase19 (MMP19), which is downregulated following respiratory syncytial virus (RSV) infection. The diverse expression levels of MMP were examined using a designed cDNA ex
pression array. The expression and secretion of MMP19 was examined using reverse transcriptionquantitative polymerase chain reaction (RTqPCR) analysis and ELISA, respectively. The proliferation of epithelial cells and lung fibroblasts were examined using flow cytometry. The epithelialmesenchymal transition (EMT) was also examined by performing western blot and RTqPCR analyses. The results of the cDNA assay showed that infection with RSV resulted in the abnormal expression of certain metalloproteinases. Among these, the expression of MMP19 decreased 3 and 7 days following infection. By using flow cytometric, western blot and RTqPCR analyses, the present study demonstrated that the downregulation of MMP19 inhibited the proliferation of epithelial cells, promoted the EMT and induced the proliferation of lung fibroblasts. Taken together, the findings of the present study suggested that the downregulation of MMP19 following RSV infection may be associated with the development of airway hyperresponsiveness.
Su YC, etal., J Virol. 2015 Feb;89(3):1564-78. doi: 10.1128/JVI.01536-14. Epub 2014 Nov 19.
Human respiratory syncytial virus (RSV) is a major cause of morbidity and severe lower respiratory tract disease in the elderly and very young, with some infants developing bronchiolitis, recurrent wheezing, and asthma following infection. Previous studies in
humans and animal models have shown that vaccination with formalin-inactivated RSV (FI-RSV) leads to prominent airway eosinophilic inflammation following RSV challenge; however, the roles of pulmonary eosinophilia in the antiviral response and in disease pathogenesis are inadequately understood. In vivo studies in mice with eotaxin and/or interleukin 5 (IL-5) deficiency showed that FI-RSV vaccination did not lead to enhanced pulmonary disease, where following challenge there were reduced pulmonary eosinophilia, inflammation, Th2-type cytokine responses, and altered chemokine (TARC and CCL17) responses. In contrast to wild-type mice, RSV was recovered at high titers from the lungs of eotaxin- and/or IL-5-deficient mice. Adoptive transfer of eosinophils to FI-RSV-immunized eotaxin- and IL-5-deficient (double-deficient) mice challenged with RSV was associated with potent viral clearance that was mediated at least partly through nitric oxide. These studies show that pulmonary eosinophilia has dual outcomes: one linked to RSV-induced airway inflammation and pulmonary pathology and one with innate features that contribute to a reduction in the viral load. IMPORTANCE: This study is critical to understanding the mechanisms attributable to RSV vaccine-enhanced disease. This study addresses the hypothesis that IL-5 and eotaxin are critical in pulmonary eosinophil response related to FI-RSV vaccine-enhanced disease. The findings suggest that in addition to mediating tissue pathology, eosinophils within a Th2 environment also have antiviral activity.
Bouchet A, etal., PLoS One. 2013 Dec 31;8(12):e81874. doi: 10.1371/journal.pone.0081874. eCollection 2013.
Synchrotron Microbeam Radiation Therapy (MRT) relies on the spatial fractionation of the synchrotron photon beam into parallel micro-beams applying several hundred of grays in their paths. Several works have reported the the
rapeutic interest of the radiotherapy modality at preclinical level, but biological mechanisms responsible for the described efficacy are not fully understood to date. The aim of this study was to identify the early transcriptomic responses of normal brain and glioma tissue in rats after MRT irradiation (400Gy). The transcriptomic analysis of similarly irradiated normal brain and tumor tissues was performed 6 hours after irradiation of 9 L orthotopically tumor-bearing rats. Pangenomic analysis revealed 1012 overexpressed and 497 repressed genes in the irradiated contralateral normal tissue and 344 induced and 210 repressed genes in tumor tissue. These genes were grouped in a total of 135 canonical pathways. More than half were common to both tissues with a predominance for immunity or inflammation (64 and 67% of genes for normal and tumor tissues, respectively). Several pathways involving HMGB1, toll-like receptors, C-type lectins and CD36 may serve as a link between biochemical changes triggered by irradiation and inflammation and immunological challenge. Most immune cell populations were involved: macrophages, dendritic cells, natural killer, T and B lymphocytes. Among them, our results highlighted the involvement of Th17 cell population, recently described in tumor. The immune response was regulated by a large network of mediators comprising growth factors, cytokines, lymphokines. In conclusion, early response to MRT is mainly based on inflammation and immunity which appear therefore as major contributors to MRT efficacy.
Vaccination with formalin-inactivated respiratory syncytial virus (FI-RSV) vaccine or RSV G glycoprotein results in enhanced pulmonary disease after live RSV infection. Enhanced pulmonary disease is characterized by pulmonary eosinophilia and is associated with
a substantial inflammatory response. We show that the absence of the G glycoprotein or G glycoprotein CX3C motif during FI-RSV vaccination or RSV challenge of FI-RSV-vaccinated mice, or treatment with anti-substance P or anti-CX3CR1 antibodies, reduces or eliminates enhanced pulmonary disease, modifies T-cell receptor Vbeta usage, and alters CC and CXC chemokine expression. These data suggest that the G glycoprotein, and in particular the G glycoprotein CX3C motif, is key in the enhanced inflammatory response to FI-RSV vaccination, possibly through the induction of substance P.
Kim HH, etal., J Korean Med Sci. 2007 Feb;22(1):37-42.
Bronchiolitis is a risk factor for the development of childhood asthma. Eosinophilic inflammation in airways plays an important role in the pathophysiology of both bronchiolitis and asthma. To investigate this inflammation, we measured the eosinophil cationic protein (ECP), regulated on activation n
ormal T-cell expressed and secreted (RANTES) and eotaxin levels in nasopharyngeal secretions (NPS). Twenty-eight patients with RSV bronchiolitis (RSV group), 11 patients with non-RSV bronchiolitis (non-RSV group) and 7 controls were enrolled in this study. ECP, RANTES, and eotaxin levels were measured by enzyme immunoassays. The ECP level in the NPS of the RSV group was significantly higher than that in the NPS of the non-RSV group and controls. RANTES and eotaxin levels in infants with bronchiolitis were significantly higher than those in the controls, but there was no significant difference between the RSV and non-RSV groups. In conclusion, with regard to eosinophilic airway inflammation, as compared with non-RSV bronchiolitis, RSV bronchiolitis may be more similar to childhood asthma.
Sigurs N, etal., Acta Paediatr. 1994 Nov;83(11):1151-5.
Eosinophil cationic protein (ECP) in nasal secretions was determined in 34 infants with respiratory syncytial virus (RSV) bronchiolitis during the acute infection stage and one and six months later. ECP in serum was determined in 19 of these children at the same
time. Myeloperoxidase (MPO) was determined in the same 19 children at the acute infection stage and after one month. All children were followed prospectively for two years after the infection with regard to the development of bronchial obstructive symptoms. Asthma, defined as three or more episodes of bronchial obstruction verified by a physician, developed in 18% of children and less severe obstructive symptoms in 29%. A screening test for food IgE antibodies in serum was performed six months and a skin prick test two years after the acute infection. Nasal ECP/albumin ratios after six months were significantly higher than during the acute RSV infection. MPO, but not ECP, levels in serum were significantly elevated at the time of acute infection compared with levels after one month. Nasal ECP/albumin ratios at the acute infection were compared to a control group of 27 infants with non-RSV upper respiratory tract infections and did not differ. It was not possible to predict, either from ECP/albumin ratios in nasal secretion or from ECP and MPO in serum, which children would develop asthma, other bronchial obstructive symptoms or positive IgE tests.
Kadri Z, etal., Genes Dev. 2015 Dec 15;29(24):2603-16. doi: 10.1101/gad.267633.115.
Tight coordination of cell proliferation and differentiation is central to red blood cell formation. Erythropoietin controls the proliferation and survival of red blood cell precursors, while variations in GATA-1/FOG-1 complex composition and concentrations drive their maturation. However, clear evi
dence of cross-talk between molecular pathways is lacking. Here, we show that erythropoietin activates AKT, which phosphorylates GATA-1 at Ser310, thereby increasing GATA-1 affinity for FOG-1. In turn, FOG-1 displaces pRb/E2F-2 from GATA-1, ultimately releasing free, proproliferative E2F-2. Mice bearing a Gata-1(S310A) mutation suffer from fatal anemia when a compensatory pathway for E2F-2 production involving insulin-like growth factor-1 (IGF-1) signaling is simultaneously abolished. In the context of the GATA-1(V205G) mutation resulting in lethal anemia, we show that the Ser310 cannot be phosphorylated and that constitutive phosphorylation at this position restores partial erythroid differentiation. This study sheds light on the GATA-1 pathways that synchronize cell proliferation and differentiation for tissue homeostasis.
The effects of exogenous hormones, used for estrus synchronization and ovarian hyper stimulation, on cumulus oocyte complexes (COCs) gene expression in sexually mature rats were determined using microarrays. Gene expression in COCs collected from GnRH (G(trt)),
GnRH + eCG (G + E(trt)), and GnRH + eCG + hCG (G + E + H(trt)) treatments were compared to COCs from naturally cycling (NC) rats before the preovulatory luteninizing hormone surge. There was no significant difference in gene expression among NC, G(trt), and G + E(trt); however, over 2,600 genes were significantly different between NC and G + E + H(trt) (P < 0.05). Genes upregulated in G + E + H(trt) encode for: proteins that are involved in prostaglandin synthesis (Ptgs2, Pla2g4a, and Runx1) and cholesterol biosynthesis (Hmgcr, Sc4mol, and Dhcr24); receptors that allow cholesterol uptake (Ldlr and Scarb1), regulate progesterone synthesis (Star), and inactivate estrogen (Sult1e1); and downstream effectors of LH signal (Pgr, Cebpb, Creb3l1, Areg, Ereg, and Adamts1). Conversely, G + E + H(trt) downregulated genes encoding proteins involved in: DNA replication and cell cycle progression (Ccne2, Orc5l, Rad50, and Mcm6); reproductive developmental process; and granulosa cell expansion (Gdf9, Bmp15, Amh, Amhr2, Bmpr1b, Tgfb2, Foxl2, Pde3a, Esr2, Fshr, Ybx2, Ccnd2, Ccnb1ip1, and Zp3); maternal effect genes required for embryo development (Zar1, Npm2, Nlrp5, Dnmt1, H1foo, and Zfp57); amino acid degradation; and ketogenesis (Hmgcs2, and Cpt1b). These results from the rat show that hormones used for estrus synchronization (G(trt)) and ovarian hyper stimulation (G + E(trt)) had minimal effects on gene expression, whereas induction of ovulation (G + E + H(trt)) caused major changes in gene expression of rat COCs. This study provides comprehensive information about regulated genes during late follicle development and ovulation induction.
Cleal JK, etal., J Physiol. 2011 Feb 15;589(Pt 4):987-97. doi: 10.1113/jphysiol.2010.198549. Epub 2011 Jan 4.
Fetal growth depends on placental transfer of amino acids from maternal to fetal blood. The mechanisms of net amino acid efflux across the basal membrane (BM) of the placental syncytiotrophoblast to the fetus, although vital for amino acid transport, are poorl
y understood. We examined the hypothesis that facilitated diffusion by the amino acid transporters TAT1, LAT3 and LAT4 plays an important role in this process, with possible effects on fetal growth. Amino acid transfer was measured in isolated perfused human placental cotyledons (n = 5 per experiment) using techniques which distinguish between different transport processes. Placental TAT1, LAT3 and LAT4 proteins were measured, and mRNA expression levels (measured using real-time quantitative-PCR) were related to fetal and neonatal anthropometry and dual-energy X-ray absorptiometry measurements of neonatal lean mass in 102 Southampton Women's Survey (SWS) infants. Under conditions preventing transport by amino acid exchangers, all amino acids appearing in the fetal circulation were substrates of TAT1, LAT3 or LAT4. Western blots demonstrated the presence of TAT1, LAT3 and LAT4 in placental BM preparations. Placental TAT1 and LAT3 mRNA expression were positively associated with measures of fetal growth in SWS infants (P < 0.05). We provide evidence that the efflux transporters TAT1, LAT3 and LAT4 are present in the human placental BM, and may play an important role in the net efflux of amino acids to the fetus. Unlike other transporters they can increase fetal amino acid concentrations. Consistent with a role in placental amino acid transfer capacity and fetal growth TAT1 and LAT3 mRNA expression showed positive associations with infant size at birth.
Jewett KA, etal., Mol Brain. 2016 Mar 22;9:32. doi: 10.1186/s13041-016-0214-6.
BACKGROUND: Neural network synchrony is a critical factor in regulating information transmission through the nervous system. Improperly regulated neural network synchrony is implicated in pathophysiological conditions such
as epilepsy. Despite the awareness of its importance, the molecular signaling underlying the regulation of neural network synchrony, especially after stimulation, remains largely unknown. RESULTS: In this study, we show that elevation of neuronal activity by the GABA(A) receptor antagonist, Picrotoxin, increases neural network synchrony in primary mouse cortical neuron cultures. The elevation of neuronal activity triggers Mdm2-dependent degradation of the tumor suppressor p53. We show here that blocking the degradation of p53 further enhances Picrotoxin-induced neural network synchrony, while promoting the inhibition of p53 with a p53 inhibitor reduces Picrotoxin-induced neural network synchrony. These data suggest that Mdm2-p53 signaling mediates a feedback mechanism to fine-tune neural network synchrony after activity stimulation. Furthermore, genetically reducing the expression of a direct target gene of p53, Nedd4-2, elevates neural network synchrony basally and occludes the effect of Picrotoxin. Finally, using a kainic acid-induced seizure model in mice, we show that alterations of Mdm2-p53-Nedd4-2 signaling affect seizure susceptibility. CONCLUSION: Together, our findings elucidate a critical role of Mdm2-p53-Nedd4-2 signaling underlying the regulation of neural network synchrony and seizure susceptibility and reveal potential therapeutic targets for hyperexcitability-associated neurological disorders.
Regulatory Foxp3(+) CD4 T cells (Tregs) prevent spontaneous inflammation in the lungs, inhibit allergic and asthmatic responses, and contribute to tolerance to inhaled allergens. Additionally, Tregs have previously been shown to suppress the CD8 T cell response during persistent virus infections. Ho
wever, little is known concerning the role that Tregs play in modulating the adaptive immune response during acute respiratory virus infections. We show following acute respiratory syncytial virus (RSV) infection that Foxp3(+) CD4 Tregs rapidly accumulate in the lung-draining mediastinal lymph nodes and lungs. BrdU incorporation studies indicate that Tregs undergo proliferation that contributes to their accumulation in the lymph nodes and lungs. Following an acute RSV infection, pulmonary Tregs modulate CD25 expression and acquire an activated phenotype characterized as CD11a(high), CD44(high), CD43(glyco+), ICOS(+), and CTLA-4(+). Surprisingly, in vivo depletion of Tregs prior to RSV infection results in delayed virus clearance concomitant with an early lag in the recruitment of RSV-specific CD8 T cells into the lungs. Additionally, Treg depletion results in exacerbated disease severity, including increased weight loss, morbidity, and enhanced airway restriction. In Treg-depleted mice there is an increase in the frequency of RSV-specific CD8 T cells that coproduce IFN-gamma and TNF-alpha, which may contribute to enhanced disease severity. These results indicate that pulmonary Tregs play a critical role in limiting immunopathology during an acute pulmonary virus infection by influencing the trafficking and effector function of virus-specific CD8 T cells in the lungs and draining lymph nodes.
Given the critical role of pattern recognition receptors (PRRs) in acid nucleic recognition in the initiation of innate immunity and the orchestration of adaptive immunity, the aim of this study was to determine whether any heterogeneity of PRR expression in the airway tracts of infants with respira
tory syncytial virus (RSV) infection might explain the broad clinical spectrum of RSV-associated bronchiolitis in infants. For this purpose, the levels of melanoma differentiation-associated protein-5 (MDA-5), retinoic acid inducible gene-1 (RIG-1), and Toll-like receptor 3 (TLR-3), TLR-7, TLR-8, and TLR-9 mRNAs were evaluated, using TaqMan quantitative reverse transcription-PCR, in cells from nasopharyngeal washes collected from 157 infants suffering from acute bronchiolitis whether or not they were associated with respiratory viruses. High interindividual variability was observed in both virus-positive and -negative infants; however, the relative gene expression levels of MDA-5, RIG-1, TLR-7, and TLR-8 were significantly higher in the virus-infected group, whereas the expression levels of TLR-3 and TLR-9 were not significantly different. The differences in the gene expression of MDA-5, RIG-1, TLR-7, and TLR-8 were more evident in infants with RSV infection than in those with bocavirus or rhinovirus infection. In RSV-infected infants, PRR-mRNA levels also were analyzed in relation to interferon protein levels, viral load, clinical severity, days of hospitalization, age, and body weight. A significant positive correlation was observed only between RSV viral load and RIG-1 mRNA levels. These findings provide the first direct evidence that, in infants with respiratory virus-associated bronchiolitis, especially RSV, there are substantial changes in PRR gene expression; this likely is an important determinant of the clinical outcome of bronchiolitis.
Syncytins are envelope genes from endogenous retroviruses, "captured" for a role in placentation. They mediate cell-cell fusion, resulting in the formation of a syncytium (the syncytiot
rophoblast) at the fetomaternal interface. These genes have been found in all placental mammals in which they have been searched for. Cell-cell fusion is also pivotal for muscle fiber formation and repair, where the myotubes are formed from the fusion of mononucleated myoblasts into large multinucleated structures. Here we show, taking advantage of mice knocked out for syncytins, that these captured genes contribute to myoblast fusion, with a >20% reduction in muscle mass, mean muscle fiber area and number of nuclei per fiber in knocked out mice for one of the two murine syncytin genes. Remarkably, this reduction is only observed in males, which subsequently show muscle quantitative traits more similar to those of females. In addition, we show that syncytins also contribute to muscle repair after cardiotoxin-induced injury, with again a male-specific effect on the rate and extent of regeneration. Finally, ex vivo experiments carried out on murine myoblasts demonstrate the direct involvement of syncytins in fusion, with a >40% reduction in fusion index upon addition of siRNA against both syncytins. Importantly, similar effects are observed with primary myoblasts from sheep, dog and human, with a 20-40% reduction upon addition of siRNA against the corresponding syncytins. Altogether, these results show a direct contribution of the fusogenic syncytins to myogenesis, with a demonstrated male-dependence of the effect in mice, suggesting that these captured genes could be responsible for the muscle sexual dimorphism observed in placental mammals.
Jesinghaus M, etal., Genes Chromosomes Cancer. 2016 Mar;55(3):268-77. doi: 10.1002/gcc.22330. Epub 2015 Dec 9.
Synchronous colorectal carcinomas (sCRC) are clinically challenging neoplasms. Although the epidemiological characteristics are quite well established, their biological basis is still poorly understood. Hence, we performed comprehensive molecular profiling of 23
sCRC cases comprising 50 synchronous primary tumors, 5 metastases, and corresponding normal tissue by targeted deep sequencing of 30 CRC-related genes, microsatellite analysis and analysis for methylated MLH1. We identified a striking inter- and intratumoral genetic heterogeneity of sCRC. Twenty (87%) cases showed genetic heterogeneity leaving only three cases with tumors that had an identical genetic make-up. Intertumoral heterogeneity was frequently observed for clinically actionable genes, including KRAS. Specifically, 44% of the cases harbored tumors of which at least one was KRAS mutated and the other KRAS wildtype. Moreover, 48% of the cases had at least double, sometimes even triple or quadruple mutations in KRAS, APC, TP53, PIK3CA, and TGFBR2, most of them being subclonal events. Lastly, we detected four cases (17%) with microsatellite instable (MSI) tumors with one case harboring one MSI- and a distinct microsatellite stable carcinoma. Our data demonstrate a striking genetic heterogeneity not only between different sCRC of a single case but also within a single tumor. These results contribute to the biological understanding of sCRC and directly impact genotyping strategies and oncological decision making. Testing one tumor or a single metastasis may not suffice in the sCRC setting as clinically relevant and tumor-specific genetic information may be left undetected compromising optimal oncological therapy. (c) 2015 Wiley Periodicals, Inc.
Siezen CL, etal., Pediatr Infect Dis J. 2009 Apr;28(4):333-5.
Prematurity is a risk factor for severe respiratory syncytial virus bronchiolitis. We show that genetic factors in innate immune genes (IFNA13, IFNAR2, STAT2, IL27, NFKBIA, C3, IL1RN, TLR5), in innate and adaptive immunity (IFNG), and in airway remodeling genes
BACKGROUND: Respiratory syncytial virus (RSV) is a common cause of severe lower respiratory tract infection in infants. Only a proportion of children infected with RSV require hospitalization. Because known risk factors for severe disease, such as premature birt
h, cannot fully explain differences in disease severity, genetic factors have been implicated. METHODS: To study the complexity of RSV susceptibility and to identify the genes and biological pathways involved in its development, we performed a genetic association study involving 470 children hospitalized for RSV bronchiolitis, their parents, and 1008 random, population controls. We analyzed 384 single-nucleotide polymorphisms (SNPs) in 220 candidate genes involved in airway mucosal responses, innate immunity, chemotaxis, adaptive immunity, and allergic asthma. RESULTS: SNPs in the innate immune genes VDR (rs10735810; P=.0017), JUN (rs11688; P=.0093), IFNA5 (rs10757212; P=.0093), and NOS2 (rs1060826; P=.0031) demonstrated the strongest association with bronchiolitis. Apart from association at the allele level, these 4 SNPs also demonstrated association at the genotype level (P=.0056, P=.0285, P=.0372, and P=.0117 for the SNPs in VDR, JUN, IFNA5, and NOS2, respectively). The role of innate immunity as a process was reinforced by association of the whole group of innate immune SNPs when the global test for groups of genes was applied (P=.046). CONCLUSION: SNPs in innate immune genes are important in determining susceptibility to RSV bronchiolitis.
Blaise S, etal., Proc Natl Acad Sci U S A. 2003 Oct 28;100(22):13013-8. Epub 2003 Oct 13.
Screening human sequence databases for endogenous retroviral elements with coding envelope genes has revealed 16 candidate genes that we assayed for their fusogenic properties. All 16 genes were cloned in a eukaryotic expression vector and assayed for cell-cell fusion by using a large panel of mamm
alian cells in transient transfection assays. Fusion was observed for two human endogenous retrovirus (HERV) envelopes, the previously characterized HERV-W envelope, also called syncytin, and a previously uncharacterized gene from the HERV-FRD family. Cells prone to env-mediated fusion were different for the two envelopes, indicating different receptor usage. A search for the FRDenv gene in primates indicated that the corresponding proviral element is present in all simians, from New World monkeys to humans, being absent only in prosimians. Cloning of the corresponding env genes in simians disclosed conservation of the fully coding status of the gene, and most remarkably, conservation of its fusogenic property. Finally, a Northern blot analysis for the expression of the FRD family among a series of human tissues demonstrated specific expression in the placenta, as previously demonstrated for the other fusogenic human envelope of the HERV-W family. Altogether, the present data have identified a previously uncharacterized envelope (that we propose to name syncytin 2 after renaming syncytin as syncytin 1) with a potential role in placenta formation, and the identification of the complete set of retroviral envelopes with fusogenic properties now allows a definite analysis of the possible role of HERV in this physiological process, via classical genetic approaches.
Bakre A, etal., J Gen Virol. 2015 Nov;96(11):3179-91.
Human respiratory syncytial virus (RSV) is a major health challenge in the young and elderly owing to the lack of a safe and effective vaccine and proven antiviral drugs. Understanding the mechanisms by which viral genes and proteins modulate the host response
to infection is critical for identifying novel disease intervention strategies. In this study, the RSV non-structural protein NS1 was shown to suppress miR-24 expression during infection. Lack of NS1 was linked to increased expression of miR-24, whilst NS1 overexpression suppressed miR-24 expression. NS1 was found to induce Kruppel-like factor 6 (KLF6), a transcription factor that positively regulates the transforming growth factor (TGF)-b pathway to induce cell cycle arrest. Silencing of KLF6 led to increased miR-24 expression via downregulation of TGF-beta. Treatment with exogenous TGF-beta suppressed miR-24 expression and induced KLF6. Confocal microscopy showed co-localization of KLF6 and RSV NS1. These findings indicated that RSV NS1 interacts with KLF6 and modulates miR-24 expression and TGF-beta, which facilitates RSV replication.
QT prolongation, a risk factor for arrhythmias, can result from genetic variants in one (or more) of the genes governing cardiac repolarization as well as intake of drugs known to affect a cardiac K(+) channel encoded by human ether-a-go-go-related gene (HERG). In this paper, we will report a case
of drug-induced long QT syndrome associated with an H(1)-receptor antagonist, hydroxyzine, in which a mutation was identified in the HERG gene. After taking 75 mg of hydroxyzine for several days, a 34-year-old female began to experience repetitive syncope. The deleterious effect of hydroxyzine was suspected because QTc interval shortened from 630 to 464 ms after cessation of the drug. Later on, the patient was found to harbor an A614V-HERG mutation. By using the patch-clamp technique in the heterologous expression system, we examined the functional outcome of the A614V mutation and confirmed a dominant-negative effect on HERG expression. Hydroxyzine concentration-dependently inhibited both wild-type (WT) and WT/A614V-HERG K(+) currents. Half-maximum block concentrations of WT and WT/A614V-HERG K(+) currents were 0.62 and 0.52 microM, respectively. Thus, accidental combination of genetic mutation and intake of hydroxyzine appeared to have led to a severe phenotype, probably, syncope due to torsade de pointes.
Shirato K, etal., Virus Res. 2015 Dec 2;210:213-26. doi: 10.1016/j.virusres.2015.08.006. Epub 2015 Aug 12.
Human respiratory syncytial virus (RSV) is a major causative agent of respiratory tract infections in children worldwide. Preterm children or those with underlying cardiopulmonary disorders are at particularly high risk of developing severe and lethal RSV respi
ratory tract infections; however, there are currently no effective vaccines or anti-viral drugs. To identify targets for the development of drugs to treat RSV infections, we investigated host cell factors involved in the replication of RSV. To this end, MDCK cells with low susceptibility to RSV were transfected with cDNA libraries derived from RSV-susceptible human lung or HeLa cells. A microarray analysis was subsequently performed on parental MDCK cells and MDCK cells that were converted to an RSV-susceptible form. Among the genes identified, chemokine (C-C motif) ligand 2 (CCL2), retinoic acid receptor responder protein 2 (RARRES2) and ephrin-B2 (EFNB2) had a positive effect on RSV replication. Expression of these genes in MDCK cells resulted in a 10- to 100-fold increase in RSV replication. CCL2 expression also disrupted the distribution of claudin-1, a tight junction protein, suggesting that CCL2 plays a role in claudin-based tight junction formation during RSV replication. The knockdown of EFNB2 and RARRES2 by siRNA in RSV-susceptible cell lines (HEp-2 and A549) resulted in reduced RSV replication, suggesting that EFNB2 and RARRES2 participate in RSV replication. Together, our findings suggest that CCL2, RARRES2 and EFNB2 are host cell factors involved in RSV replication.
Luk JM, etal., Biochem Biophys Res Commun 2003 Aug 8;307(4):782-90.
The molecular mechanism regulating spermatogenesis at different developmental stages remains largely unknown. In a vitamin A-deficiency (VAD) rat model, five distinct histologically defined, stage-synchronized testes: (i) resting spermatogonia and preleptotene s
permatocytes at Day 0 of post-vitamin A treatment (PVA); (ii) early pachytene spermatocytes at Day 7 PVA; (iii) late pachytene at Day 15 PVA; (iv) round spermatids at Day 25 PVA; and (v) elongated spermatids at Day 35 PVA were used to study gene expression profiles by mRNA differential display. Twenty-four differentially expressed cDNA fragments were identified and cloned; oligonucleotide sequence analyses indicated that there are 12 novel gene sequences, half of which share no apparent match in current GenBank/EMBL databases. Other 12 VAD clones share sequence homology to membrane channel and transport, transcription and translation, cell cycle and morphogenesis, inducer and transducer, surface or secreted glycoproteins or enzymes, and other miscellaneous molecules. Semi-quantitative RT-PCR analyses against different stages of VAD testes demonstrated: (i) restricted expression of VAD1.2 and 1.3 (novel) on Day 25 PVA when round spermatids form; (ii) escalating pattern of VAD12 (Cx43) in Sertoli cells; and (iii) relative constant levels of VAD4 (A5D3), VAD26.1 (ribonuclease), and VAD27 (GRP8) in spermatogenesis.
Castilow EM, etal., J Immunol. 2008 Feb 15;180(4):2376-84.
Respiratory syncytial virus (RSV) is the leading cause of lower respiratory tract disease in children. Children previously vaccinated with a formalin-inactivated RSV vaccine experienced enhanced morbidity and mortality upon natural RSV infection. Histological an
alysis revealed the presence of eosinophils in the pulmonary infiltrate of the vaccinated children. Eosinophils are characteristic of Th2 responses, and Th2 cells are known to be necessary to induce pulmonary eosinophilia in RSV-infected BALB/c mice previously immunized with a recombinant vaccinia virus (vv) expressing the RSV G protein (vvG). Using IL-13-deficient mice, we find that IL-13 is necessary for eosinophils to reach the lung parenchyma and airways of vvG-immunized mice undergoing RSV challenge infection. IL-13 acts specifically on eosinophils as the magnitude of pulmonary inflammation, RSV G protein-specific CD4 T cell responses, and virus clearance were not altered in IL-13-deficient mice. After RSV challenge, eosinophils were readily detectable in the blood and bone marrow of vvG-immunized IL-13-deficient mice, suggesting that IL-13 is required for eosinophils to transit from the blood into the lung. Pulmonary levels of CCL11 and CCL22 protein were significantly reduced in IL-13-deficient mice indicating that IL-13 mediates the recruitment of eosinophils into the lungs by inducing the production of chemokines important in Th2 cell and eosinophil chemotaxis.
Dodd JS, etal., J Immunol. 2009 Dec 1;183(11):7006-13. Epub 2009 Nov 13.
IL-9 is a cytokine of great current interest associated with allergic/Th2 responses. High levels of IL-9 are present in bronchial secretions from infants with respiratory syncytial virus (RSV) bronchiolitis. To test its effects in RSV disease with a Th2 profile,
BALB/c mice were vaccinated with recombinant vaccinia virus expressing the RSV G protein. On RSV challenge, immunized mice developed augmented disease characterized by enhanced pulmonary Th2 and local IL-9 production peaking on days 7-10 of RSV infection. Depletion with anti-IL-9 Ab at vaccination or RSV challenge enhanced viral clearance. Depletion only at challenge had no effect on disease severity, whereas depletion at immunization and challenge enhanced Th1 responses, inhibited virus-specific IgG1 production, and enhanced disease severity. By contrast, depletion of IL-9 at immunization boosted IgG2a and inhibited the Th2 response and disease during subsequent infection without a concomitant increase in type 1 cytokines. Adoptive transfer of secondary memory CD4 T cells from the spleens of IL-9-depleted mice into naive recipients replicated many of the effects of depletion, indicating that IL-9 acts via CD4 T cells. Therefore, IL-9 is a previously unknown but key modulator of antiviral immunity, regulating T and B cell responses and having potent and specific effects on viral lung disease.
Respiratory syncytial virus (RSV) causes significant morbidity and mortality in infants worldwide. Severe RSV infections in infants cause bronchiolitis, wheeze, and/or cough and significantly increase the risk for developing asthma. RSV pathogenesis is thought t
o be due to a Th2-type immune response initiated in response to RSV infection, specifically in the infant. Using a neonatal mouse system as an appropriate model for human infants, we sought to determine whether local inhibition of IL-4Ralpha expression during primary RSV infection in the neonate would prevent Th2-skewed responses to secondary RSV infection and improve long-term pulmonary function. To reduce IL-4Ralpha expression, antisense oligonucleotides (ASOs) specific for IL-4Ralpha were administered intranasally to neonatal mice at the time of primary infection. Mice were initially infected with RSV at 1 wk of age and were reinfected at 6 wk of age. Administration of IL-4Ralpha ASOs during primary RSV infection in neonatal mice abolished the pulmonary dysfunction normally observed following reinfection in the adult. This ablation of pulmonary dysfunction correlated with a persistent rebalancing of the Th cell compartment with decreased Th2 responses (i.e., reduced goblet cell hyperplasia, Th2 cells, and cytokine secretion) and increased Th1 responses (i.e., elevated Th1 cell numbers and type I Abs and cytokines). Our data support our hypothesis that a reduction in the Th2 immune response during primary infection in neonates prevents Th2-mediated pulmonary pathology initially and upon reinfection and further suggest that vaccine strategies incorporating IL-4Ralpha ASOs may be of significant benefit to infants.
Sun Y, etal., PLoS Pathog. 2015 Sep 3;11(9):e1005122. doi: 10.1371/journal.ppat.1005122. eCollection 2015 Sep.
Human respiratory syncytial virus (RSV) is a major cause of severe respiratory illness in children and susceptible adults. RSV blocks the development of the innate antiviral immune response and can grow to high titers in the respiratory tract. Here we demonstrat
e that immunostimulatory defective viral genomes (iDVGs) that are naturally generated during RSV replication are strong inducers of the innate antiviral response to RSV in mice and humans. In mice, RSV iDVGs stimulated the expression of antiviral genes, restricted viral replication, and prevented weight loss and lung inflammation. In human cells, the antiviral response to RSV iDVGs was dominated by the expression of IFN-lambda1 over IFN-beta and was driven by rapid intranuclear accumulation of the transcription factor IRF1. RSV iDVGs were detected in respiratory secretions of hospitalized patients, and their amount positively correlated with the level of expression of antiviral genes in the samples. Infection of explanted human lung tissue from different donors revealed that most humans can respond to RSV iDVGs and that the rate of accumulation of iDVGs during infection directly correlates with the quality of the antiviral response. Taken together, our data establish iDVGs as primary triggers of robust antiviral responses to RSV and provide the first evidence for an important biological role for naturally occurring iDVGs during a paramyxovirus infection in humans.
Larranaga CL, etal., Pediatr Infect Dis J. 2009 Oct;28(10):867-73.
BACKGROUND: Respiratory syncytial virus (RSV) is a major cause of acute lower respiratory infection in infants. The immune response plays a leading role in the severity of the disease. We hypothesized that severe RSV disease is associated with an impaired immune
response characterized by low circulating T lymphocytes and plasma cytokine concentrations. METHODS: We evaluate the in vivo immune responses of previously healthy infants with their first proven RSV-acute lower respiratory infection that required hospitalization. According to the clinical severity, defined by using a strict scoring system, the in vivo immune response was compared through the analysis of plasma cytokine values and the phenotyping of peripheral blood lymphocyte and natural killer (NK) cells. RESULTS: Absolute blood cell counts of CD4+, CD8+, and CD19+ lymphocytes and NK cells were lower in subjects with RSV than in control infants. Lowest cell counts were observed in more severe RSV-infected infants. Significant low values were obtained in CD8+ lymphocytes (P = 0.03) and nonactive NK cells, that express CD94 antigen (P = 0.046). In contrast, activated NK cells that do not express CD94 molecules were significantly higher in RSV infected infants than in healthy controls (% of cells: P = 0.004). The interferon-gamma and tumor necrosis factor-alpha values in RSV infected patients were lower than in controls subjects. Interleukin-17 cytokine was not detected in healthy infants and the largest concentration was found in moderately ill patients as compared with severe cases (P = 0.033). RSV infection showed significantly higher interleukin-8 chemokine than in control infants (P = 0.024). CONCLUSION: We propose that severe RSV infection in very young infants is associated with poor blood proinflammatory cytokine production, low counts of CD8+ T cells and with a greater activity of a group of NK cells, that are independent of the major histocompatibility complex class Ib recognition system.
Gagro A, etal., Clin Exp Immunol. 2004 Feb;135(2):267-72.
The fusion protein of the respiratory syncytial virus (RSV) binds to the pattern recognition receptors, TLR4 and CD14, and initiates innate immunity response to the virus. The aim of the study was to investigate the expression of TLR4 on peripheral blood lymphoc
ytes and monocytes in peripheral blood of infants in both acute and convalescent phase of RSV bronchiolitis (n = 26). In addition, TNF-alpha expression in lipopolysaccharide-stimulated monocytes was also assessed. The results showed TLR4 to be expressed predominantly by monocytes in both sick infants and controls. During the acute phase of infection monocytes up-regulated TLR4 in eight infants, which returned to the levels recorded in controls 4-6 weeks from infection. There was no difference in the percentage of TNF-alpha secreting monocytes. Of the clinical parameters tested, minimal oxygen saturation was found to correlate negatively with this expression in the group of infants with increased TLR4. Additional studies are under way to correlate this finding with the outcome of the immune response to RSV.
Haeberle HA, etal., J Virol. 2001 Jan;75(2):878-90.
Lower respiratory tract disease caused by respiratory syncytial virus (RSV) is characterized by profound airway mucosa inflammation, both in infants with naturally acquired infection and in experimentally inoculated animal models. Chemokines are central regulato
ry molecules in inflammatory, immune, and infectious processes of the lung. In this study, we demonstrate that intranasal infection of BALB/c mice with RSV A results in inducible expression of lung chemokines belonging to the CXC (MIP-2 and IP-10), CC (RANTES, eotaxin, MIP-1beta, MIP-1alpha, MCP-1, TCA-3) and C (lymphotactin) families. Chemokine mRNA expression occurred as early as 24 h following inoculation and persisted for at least 5 days in mice inoculated with the highest dose of virus (10(7) PFU). In general, levels of chemokine mRNA and protein were dependent on the dose of RSV inoculum and paralleled the intensity of lung cellular inflammation. Immunohisthochemical studies indicated that RSV-induced expression of MIP-1alpha, one of the most abundantly expressed chemokines, was primarily localized in epithelial cells of the alveoli and bronchioles, as well as in adjoining capillary endothelium. Genetically altered mice with a selective deletion of the MIP-1alpha gene (-/- mice) demonstrated a significant reduction in lung inflammation following RSV infection, compared to control littermates (+/+ mice). Despite the paucity of infiltrating cells, the peak RSV titer in the lung of -/- mice was not significantly different from that observed in +/+ mice. These results provide the first direct evidence that RSV infection may induce lung inflammation via the early production of inflammatory chemokines.
Kashino G, etal., Int J Radiat Oncol Biol Phys. 2009 May 1;74(1):229-36. doi: 10.1016/j.ijrobp.2008.09.060.
PURPOSE: To determine whether glioma cells irradiated with a microplanar X-ray beam exert bystander effects. METHODS AND MATERIALS: Microplanar beam irradiation of glioma cells in vitro was done using the SPring-8 synchrotron radiation facility. The amount of DN
A double-strand breaks (dsbs) was measured by the fluorescence intensity of phosphorylated H2AX or the number of 53BP1 foci. The dose distribution in a cell population exposed to a single microplanar beam was determined by the amount of phosphorylated H2AX-positive cells. Bystander effects were determined by counting the number of 53BP1 foci in nonirradiated cells treated with conditioned medium from cultures of irradiated cells. RESULTS: More DNA dsbs were detected in cells adjacent to an area irradiated by the single beam than in cells in distant, nonirradiated areas as a result of bystander effects caused by scattered X-rays and DNA dsbs. In support of this, more 53BP1 foci were observed in nonirradiated, conditioned medium-treated cells than in control cells (i.e., cells not treated with irradiation or conditioned medium). These results suggest that DNA dsbs were induced in nonirradiated cells by soluble factors in the culture medium. In addition, we observed cellular migration into areas irradiated with peak doses, suggesting that irradiated cells send signals that cause nonirradiated cells to migrate toward damaged cells. CONCLUSIONS: Bystander effects are produced by factors secreted as a result of slit-type microplanar X-ray beam irradiation.
T cells secreting interleukin (IL)-4 and IL-5 (T helper cell type 2 [Th2] cells) play a detrimental role in a variety of diseases, but specific methods of regulating their activity remain elusive. T1/ST2 is a surface ligand of the IL-1 receptor family, expressed on Th2- but not on interferon (IFN)-g
amma-producing Th1 cells. Prior exposure of BALB/c mice to the attachment (G) or fusion (F) protein of respiratory syncytial virus (RSV) increases illness severity during intranasal RSV challenge, due to Th2-driven lung eosinophilia and exuberant Th1-driven pulmonary infiltration, respectively. We used these polar models of viral illness to study the recruitment of T1/ST2 cells to the lung and to test the effects of anti-T1/ST2 treatment in vivo. T1/ST2 was present on a subset of CD4(+) cells from mice with eosinophilic lung disease. Monoclonal anti-T1/ST2 treatment reduced lung inflammation and the severity of illness in mice with Th2 (but not Th1) immunopathology. These results show that inhibition of T1/ST2 has a specific effect on virally induced Th2 responses and suggests that therapy targeted at this receptor might be of value in treating Th2-driven illness.
Puthothu B, etal., Pediatr Infect Dis J. 2007 Dec;26(12):1094-8.
BACKGROUND: Respiratory syncytial virus (RSV) is a major cause of bronchiolitis in infants. During the course of RSV infection, predominant T helper cell (TH) 2 response is associated with disease progression, whereas predominant TH1 reaction provides convalesce
nce. Interleukin (IL)-18 plays an important role in adjusting the TH1/TH2 immune response to viral infections. Thus, we tested the hypothesis that polymorphisms in IL-18 were associated with severe RSV-associated diseases. METHODS: We chose to study the promotor polymorphisms -607A/C (rs1946518) and -137G/C (rs187238), the 2 exon polymorphisms 113T/G (rs360718) and 127C/T (rs360717), and 2 intron polymorphisms 5304A/G (rs795467) and 133G/C (rs360721) within the IL-18 gene. Genotyping was performed on 154 children with severe RSV infection as defined by strict clinical criteria and on 270 controls. Statistical analyses of single polymorphisms made use of the Armitage's trend test, haplotypes were calculated with FASTEHPLUS and FAMHAP. RESULTS: -133G/C showed association with severe RSV infection (P = 0.043). The association was further supported by haplotype analyses with all 6 polymorphisms (P < 0.00001 for association with RSV). CONCLUSIONS: This study indicates possible involvement of IL-18 in the determination of severe RSV-associated diseases. Defining the genetic basis of RSV bronchiolitis might help us in identifying new drug targets for a more specific therapy. In addition, it might enable an early identification of children at risk for RSV bronchiolitis and thus make a selective prevention feasible.
Respiratory syncytial virus (RSV) causes bronchiolitis, the main cause of infantile hospitalization. Immunity against reinfection is poor, and there is great interest in boosting vaccine responses using live vectors expressing host cytokines. We therefore constr
ucted a recombinant RSV expressing murine interleukin 18 (RSV/IL-18), a cytokine capable of inducing strong antiviral immune responses. In vitro RSV/IL-18 replicated at wild-type levels and produced soluble IL-18. In naive BALB/c mice, RSV/IL-18 infection significantly increased both IL-18 mRNA and protein and attenuated the peak viral load 3-fold. Despite a reduced viral load, RSV/IL-18 infection caused a biphasic weight loss at days 2 and 6 postinfection that was not seen in wild-type infection. Day 2 disease was associated with enhanced pulmonary natural killer (NK) cell numbers and activity and was prevented by NK cell depletion during infection; day 6 disease was correlated with CD8 T-cell recruitment and was enhanced by NK cell depletion. IL-18 expression during priming also enhanced RSV-specific antibody responses and T-cell responses on secondary RSV infection. Therefore, while IL-18 boosted antiviral immunity and reduced the viral load, its coexpression worsened disease. This is the first recombinant RSV with this property, and these are the first studies to demonstrate that NK cells can induce pathology during pulmonary viral infections.
McNamara PS, etal., Lancet. 2004 Mar 27;363(9414):1031-7.
BACKGROUND: Respiratory syncytial virus (RSV) bronchiolitis is the most prevalent acute wheezing disorder in infants and is associated with recurrent wheeze and asthma in childhood. Interleukin 9, a type 2 cytokine has been proposed as a key cytokine in suscepti
bility to asthma. We aimed to investigate whether interleukin 9 was produced in the lungs of infants with severe RSV disease and if found, from which cells it originated. METHODS: We did 150 non-bronchoscopic bronchoalveolar lavages during the course of ventilation in 24 term infants and 21 preterm infants ventilated for RSV bronchiolitis. We also did 10 bronchoalveolar lavages on the day of intubation in 10 control infants ventilated for non-respiratory causes. We measured pulmonary interleukin 9 mRNA and protein in samples from all groups. We used immunostaining to identify the cells that produce interleukin 9. FINDINGS: Interleukin 9 mRNA expression, which persisted over the course of ventilation, was noted in all infants with bronchiolitis. Three of the control group also showed interleukin 9 mRNA expression. Median interleukin 9 protein concentration on day 1 (1.9 microg/L [range 0.1-36.2]) was significantly greater in term infants with bronchiolitis than either preterm infants (0.4 microg/L [0.1-2.9]; p<0.05) or the control group (0.7 microg/L [0.4-2.5]; p<0.05). There was a trend for interleukin 9 protein concentrations in term, but not preterm infants to decrease over time. Immunostained cell smears showed that most interleukin 9 expression in bronchoalveolar lavage was by neutrophils. INTERPRETATION: In term infants with RSV bronchiolitis, we noted large amounts of interleukin 9 mRNA and interleukin 9 protein. Neutrophils seem to be the main source of this type 2 cytokine. Interleukin 9 production by neutrophils may contribute to the pathogenesis of RSV disease. These findings may be relevant to other disease processes in the lung where neutrophils are the predominant inflammatory cell type.
Respiratory syncytial virus (RSV) is the most important cause of lower respiratory tract disease in young children. In the 1960s, infants vaccinated with formalin-inactivated RSV developed a more severe disease characterized by excessive inflammatory immunopatho
logy in lungs upon natural RSV infection. The fear of causing the vaccine-enhanced disease (VED) is an important obstacle for development of safe and effective RSV vaccines. The recombinant vaccine candidate G1F/M2 immunization also led to VED. It has been proved that cellular memory induced by RSV vaccines contributed to VED. Interleukin-27 (IL-27) and IL-23 regulate Th1, Th17, and/or Th2 cellular immune responses. In this study, mice coimmunized with pcDNA3-IL-27 and G1F/M2 were fully protected and, importantly, did not develop vaccine-enhanced inflammatory responses and immunopathology in lungs after RSV challenge, which was correlated with moderate Th1-, suppressed Th2-, and Th17-like memory responses activated by RSV. In contrast, G1F/M2- or pcDNA3-IL-23+G1F/M2-immunized mice, in which robust Th2- and Th17-like memory responses were induced, developed enhanced pulmonary inflammation and severe immunopathology. Mice coimmunized with G1F/M2 and the two cytokine plasmids exhibited mild inflammatory responses as well as remarkable Th1-, suppressed Th2-, and Th17-like memory responses. These results suggested that Th1-, Th2-, and Th17-like memory responses and, in particular, excessive Th2- and Th17-like memory responses were closely associated with VED; IL-27 may inhibit VED following respiratory syncytial virus infection by regulating cellular memory responses.
Schuurhof A, etal., Pediatr Pulmonol. 2010 Jun;45(6):608-13.
The predominance of severe respiratory syncytial virus (RSV) bronchiolitis in boys compared to girls is well known, but its mechanism is not yet understood. This is the first study focusing on gender-specific genetic factors affecting the risk of severe RSV infe
ction using a previously described cohort. We determined 347 single-nucleotide polymorphisms (SNPs) in 470 children hospitalized for RSV infection, their parents, and 1,008 random population controls. We tested if these SNPs exerted a different effect in boys and girls by performing statistical interaction tests. Only one SNP (rs2069885) had a gender-specific significant association with RSV infection, severe enough to require hospitalization (P-value 0.00057). The major allele of this structural polymorphism in the interleukin (IL)-9 gene is associated with an increased susceptibility to severe RSV infection in boys, while there is a decreased susceptibility in girls. Haplotype analysis of two SNPs in the IL-9 gene (rs2069885 and rs1799962) showed overrepresentation of the TT haplotype in girls with severe RSV bronchiolitis requiring hospitalization indicating that this is the haplotype conferring the highest risk in girls. In conclusion, the IL-9 genetic polymorphism (rs2069885) has an opposite effect on the risk of severe RSV bronchiolitis in boys and girls. Although so far a difference in IL-9 production in boys and girls has not been reported, this study may help in explaining the different risks of severe RSV bronchiolitis in boys and girls.
Gonzalez-Sanz R, etal., J Virol. 2016 Jan 13;90(7):3428-38. doi: 10.1128/JVI.02695-15.
Human respiratory syncytial virus (RSV), for which neither a vaccine nor an effective therapeutic treatment is currently available, is the leading cause of severe lower respiratory tract infections in children. Interferon-stimulated gene 15 (ISG15) is a ubiquiti
n-like protein that is highly increased during viral infections and has been reported to have an antiviral or a proviral activity, depending on the virus. Previous studies from our laboratory demonstrated strong ISG15 upregulation during RSV infection in vitro. In this study, an in-depth analysis of the role of ISG15 in RSV infection is presented. ISG15 overexpression and small interfering RNA (siRNA)-silencing experiments, along with ISG15 knockout (ISG15(-/-)) cells, revealed an anti-RSV effect of the molecule. Conjugation inhibition assays demonstrated that ISG15 exerts its antiviral activity via protein ISGylation. This antiviral activity requires high levels of ISG15 to be present in the cells before RSV infection. Finally, ISG15 is also upregulated in human respiratory pseudostratified epithelia and in nasopharyngeal washes from infants infected with RSV, pointing to a possible antiviral role of the molecule in vivo. These results advance our understanding of the innate immune response elicited by RSV and open new possibilities to control infections by the virus. IMPORTANCE: At present, no vaccine or effective treatment for human respiratory syncytial virus (RSV) is available. This study shows that interferon-stimulated gene 15 (ISG15) lowers RSV growth through protein ISGylation. In addition, ISG15 accumulation highly correlates with the RSV load in nasopharyngeal washes from children, indicating that ISG15 may also have an antiviral role in vivo. These results improve our understanding of the innate immune response to RSV and identify ISG15 as a potential target for virus control.
Respiratory syncytial virus (RSV) infection potentiates neurogenic inflammation in rat airways. Because some vascular effects of sensory nerves are mediated by cysteinyl leukotrienes (cysLTs), we studied whether the receptor antagonist montelukast inhibits neuro
genic plasma extravasation in RSV-infected rats. Pathogen-free rats were inoculated at 2 wk (weanlings) or 12 wk (adults) of age with RSV or virus-free medium and treated with montelukast or its vehicle starting 1 day before inoculation. Five days postinoculation, we measured the extravasation of Evans blue-labeled albumin in the respiratory tract after stimulation of sensory nerves with capsaicin. Montelukast had no effect in the extrapulmonary airways but abolished albumin extravasation in the intrapulmonary airways of RSV-infected rats, with a larger effect in weanlings than in adults. Increased concentrations of 5-lipoxygenase-encoding mRNA and cysLTs, as well as numerous mast cells, were detected in the lung tissues of RSV-infected weanling rats. These observations suggest that the release of neuropeptides from capsaicin-sensitive sensory nerves and nonneuronal cells in the lungs of RSV-infected young rats increases vascular permeability by promoting the release of leukotrienes from mast cells.
Schwickert A, etal., PLoS One. 2015 Dec 10;10(12):e0143993. doi: 10.1371/journal.pone.0143993. eCollection 2015.
MicroRNAs (miRNAs, micro ribonucleic acids) are pivotal post-transcriptional regulators of gene expression. These endogenous small non-coding RNAs play significant roles in tumorigenesis and tumor progression. miR-142-3p expression is dysregulated in several breast cancer subtypes. We aimed at inves
tigating the role of miR-142-3p in breast cancer cell invasiveness. Supported by transcriptomic Affymetrix array analysis and confirmatory investigations at the mRNA and protein level, we demonstrate that overexpression of miR-142-3p in MDA-MB-231, MDA-MB-468 and MCF-7 breast cancer cells leads to downregulation of WASL (Wiskott-Aldrich syndrome-like, protein: N-WASP), Integrin-alphaV, RAC1, and CFL2, molecules implicated in cytoskeletal regulation and cell motility. ROCK2, IL6ST, KLF4, PGRMC2 and ADCY9 were identified as additional targets in a subset of cell lines. Decreased Matrigel invasiveness was associated with the miR-142-3p-induced expression changes. Confocal immunofluorescence microscopy, nanoscale atomic force microscopy and digital holographic microscopy revealed a change in cell morphology as well as a reduced cell volume and size. A more cortical actin distribution and a loss of membrane protrusions were observed in cells overexpressing miR-142-3p. Luciferase activation assays confirmed direct miR-142-3p-dependent regulation of the 3'-untranslated region of ITGAV and WASL. siRNA-mediated depletion of ITGAV and WASL resulted in a significant reduction of cellular invasiveness, highlighting the contribution of these factors to the miRNA-dependent invasion phenotype. While knockdown of WASL significantly reduced the number of membrane protrusions compared to controls, knockdown of ITGAV resulted in a decreased cell volume, indicating differential contributions of these factors to the miR-142-3p-induced phenotype. Our data identify WASL, ITGAV and several additional cytoskeleton-associated molecules as novel invasion-promoting targets of miR-142-3p in breast cancer.
Phenotypic integration patterns in the mammalian skull have long been a focus of intense interest as a result of their suspected influence on the trajectory of hominid evolution. Here we test the hypothesis that perturbation of cartilage growth, which directly affects only the chondrocranium during
development, will produce coordinated shape changes in the adult calvarium and face regardless of mechanism. Using two murine models of cartilage undergrowth that target two very different mechanisms, we show that strong reduction in cartilage growth produces a short, wide, and more flexed cranial base. This in turn produces a short, wide face in both models. Cranial base and face are already correlated early in ontogeny, and the relationship between these modules gains structure through postnatal growth and development. These results provide further evidence that there exist physical interactions between developing parts of the phenotype that produce variation at a distance from the actual locus upon which a particular selective pressure is acting. Phenotypic changes observed over the course of evolution may not all require adaptationist explanations; rather, it is likely that a substantial portion of observed phenotypic variation over the history of a clade is not directly adaptive but rather a secondary consequence of some local response to selection.
Rademacher N, etal., Sci Rep. 2016 Oct 19;6:35283. doi: 10.1038/srep35283.
At neuronal synapses, multiprotein complexes of trans-synaptic adhesion molecules, scaffold proteins and neurotransmitter receptors assemble to essential building blocks required for synapse formation and maintenance. Here we describe a novel role for the membrane-associated guanylate kinase (MAGUK)
protein MPP2 (MAGUK p55 subfamily member 2) at synapses of rat central neurons. Through interactions mediated by its C-terminal SH3-GK domain module, MPP2 binds to the abundant postsynaptic scaffold proteins PSD-95 and GKAP and localises to postsynaptic sites in hippocampal neurons. MPP2 also colocalises with the synaptic adhesion molecule SynCAM1. We demonstrate that the SynCAM1 C-terminus interacts directly with the MPP2 PDZ domain and that MPP2 does not interact in this manner with other highly abundant postsynaptic transmembrane proteins. Our results highlight a previously unexplored role for MPP2 at postsynaptic sites as a scaffold that links SynCAM1 cell adhesion molecules to core proteins of the postsynaptic density.
Liu J, etal., Int Immunopharmacol. 2015 Sep;28(1):337-43. doi: 10.1016/j.intimp.2015.05.035. Epub 2015 Jun 1.
It has been reported that natural helper cells, which are a non-T, non-B innate lymphoid cell type expressing c-Kit and ST2, mediate influenza-induced airway hyper-reactivity by producing substantial IL-13. However, little is known about natural helper cells for the development of RSV-induced airway
inflammation, particularly eosinophilic infiltration. By using BALB/c mice that were infected intranasally with RSV, it became clear that infection with RSV can induce an increase in the absolute number of natural helper cells in the lungs of mice. It seems likely that these natural helper cells contribute to the massive eosinophilic infiltration in an IL-13-dependent manner. In fact, the number of IL-13-producing natural helper cells as well as the expression of IL-13 mRNA in natural helper cells was enhanced significantly during RSV infection, suggesting that natural helper cells might be cellular source of the Th2-type cytokine IL-13. Indeed, adoptive transfer of pulmonary natural helper cells augmented not only the production of IL-13 but also the infiltration of eosinophils in the lungs of transferred mice. Pulmonary natural helper cells can produce IL-13 following response to IL-33, which was increased markedly in the lungs of mice after intranasal RSV infection. The expression of IL-13 mRNA in pulmonary natural helper cells was up-regulated by in vitro IL-33 stimulation. Furthermore, blockade of IL-33 receptor subunit, ST2, diminished the frequency of IL-13-producing natural helper cells. Taken together, these results demonstrate that natural helper cells may play an important role in RSV-induced pulmonary eosinophilia by producing IL-13 via the IL-33/ST2 pathway.
Han X, etal., Immunotherapy. 2017 Aug;9(9):715-722. doi: 10.2217/imt-2017-0037. Epub 2017 Aug 3.
AIM: Type 2 cytokine production during respiratory virus infection is considered to be linked with asthma exacerbation. As potent Th2 cytokine producers, natural helper (NH) cells play a key role in influenza virus-induced airway hyper-responsiveness. However, it is unclear whether NH cel
ls contribute to respiratory syncytial virus (RSV)-induced airway inflammation, and how the cytokine profile in NH cells is changed during RSV infection. METHODS: BALB/c mice were infected intranasally with RSV. The number of NH cells in lungs was detected by flow cytometry. The expression of cytokine mRNAs was performed by real-time RT-PCR. Cytokines levels were determined by ELISA. RESULTS: Following intranasal infection with RSV, BALB/c mice showed an increase in the expression of mRNAs for Th2-like cytokines in NH cells. Furthermore, adoptive transfer of pulmonary NH cells resulted in a massive infiltration of mononuclear cells, in particular eosinophils and neutrophils in lungs, in parallel with an augmented production of Th2-associated cytokines, such as IL-4, IL-5 and IL-10 in bronchoalveolar lavage fluids, providing convincing evidence that NH cells contribute to RSV-induced lung pathogenesis by producing type 2 cytokines. It should be noted that blocking IL-33 with antibody can diminish the absolute number of pulmonary NH cells and the relative expression of mRNAs for type 2 cytokines in pulmonary NH cells, suggesting that IL-33 is necessary for activating Th2-type NH cells. CONCLUSION: These results reveal that pulmonary NH cells might participate in RSV-induced airway inflammation by producing large quality of type 2 cytokines in an IL-33-dependent manner.
Hu C, etal., Am J Physiol Lung Cell Mol Physiol 2002 Aug;283(2):L494-502.
Nerve growth factor (NGF) controls sensorineural development and responsiveness and modulates immunoinflammatory reactions. Respiratory syncytial virus (RSV) potentiates the proinflammatory effects of sensory nerves in rat airways by upregulating the substance P
receptor, neurokinin 1 (NK(1)). We investigated whether the expression of NGF and its trkA and p75 receptors in the lungs is age dependent, whether it is upregulated during RSV infection, and whether it affects neurogenic inflammation. Pathogen-free rats were killed at 2 (weanling) to 12 (adult) wk of age; in addition, subgroups of rats were inoculated with RSV or virus-free medium. In pathogen-free rats, expression of NGF and its receptors in the lungs declined with age, but RSV doubled expression of NGF, trkA, and p75 in weanling and adult rats. Exogenous NGF upregulated NK(1) receptor expression in the lungs. Anti-NGF antibody inhibited NK(1) receptor upregulation and neurogenic inflammation in RSV-infected lungs. These data indicate that expression of NGF and its receptors in the lungs declines physiologically with age but is upregulated by RSV and is a major determinant of neurogenic inflammation.
Yang M, etal., Chin Herb Med. 2024 Jul 29;16(4):638-655. doi: 10.1016/j.chmed.2024.07.007. eCollection 2024 Oct.
OBJECTIVE: Qingfei Tongluo Plaster (QFP), an improved Chinese medicine hospital preparation, is an attractive treatment option due to its well clinical efficacy, convenience, economy, and patient compliance in the treatment of respiratory syncytial vi
rus (RSV) pneumonia. The aim of this study was to investigate the efficacy mechanism of QFP on RSV rats from the perspective of alleviating lung inflammation and further explore the changes of serum metabolites and metabolic pathways in RSV rats under the influence of QFP. METHODS: This study used network pharmacological methods and molecular docking combined with molecular biology and metabolomics from multi-dimensional perspectives to screen and verify the therapeutic targets. Open online databases were used to speculate the gene targets of efficient ingredients and diseases. Then, we used the String database to examine the fundamental interaction of common targets of drugs and diseases. An online enrichment analysis was performed to predict the functional pathways. Molecular docking was applied to discover the binding modes between essential ingredients and crucial gene targets. Finally, we demonstrated the anti-inflammatory ability of QFP in the RSV-evoked pneumonia rat model and explained the mechanism in combination with the metabolomics results. RESULTS: There were 19 critical targets defined as the core targets: tumor necrosis factor (TNF), inducible nitric oxide synthase 2 (NOS2), mitogen-activated protein kinase 14 (MAPK14), g1/S-specific cyclin-D1 (CCND1), signal transducer and activator of transcription 1-alpha/beta (STAT1), proto-oncogene tyrosine-protein kinase Src (SRC), cellular tumor antigen p53 (TP53), interleukin-6 (IL6), hypoxia-inducible factor 1-alpha (HIF1A), RAC-alpha serine/threonine-protein kinase (AKT1), signal transducer and activator of transcription 3 (STAT3), heat shock protein HSP 90-alpha (HSP90AA1), tyrosine-protein kinase JAK2 (JAK2), cyclin-dependent kinase inhibitor 1 (CDKN1A), mitogen-activated protein kinase 3 (MAPK3), epidermal growth factor receptor (EGFR), myc proto-oncogene protein (MYC), protein c-Fos (FOS) and transcription factor p65 (RELA). QFP treated RSV pneumonia mainly through the phosphatidylinositol 3-kinase (PI3K)/RAC AKT pathway, HIF-1 pathway, IL-17 pathway, TNF pathway, and MAPK pathway. Animal experiments proved that QFP could effectively ameliorate RSV-induced pulmonary inflammation. A total of 28 metabolites underwent significant changes in the QFP treatment, and there are four metabolic pathways consistent with the KEGG pathway analyzed by network pharmacology, suggesting that they may be critical processes related to treatment. CONCLUSION: These results provide essential perspicacity into the mechanisms of action of QFP as a promising anti-RSV drug.
BACKGROUND: Respiratory viral infections follow an unpredictable clinical course in young children ranging from a common cold to respiratory failure. The transition from mild to severe disease occurs rapidly and is difficult to predict. The pathophysiology underlying disease severity has remained e
lusive. There is an urgent need to better understand the immune response in this disease to come up with biomarkers that may aid clinical decision making. METHODS: In a prospective study, flow cytometric and genome-wide gene expression analyses were performed on blood samples of 26 children with a diagnosis of severe, moderate or mild Respiratory Syncytial Virus (RSV) infection. Differentially expressed genes were validated using Q-PCR in a second cohort of 80 children during three consecutive winter seasons. FACS analyses were also performed in the second cohort and on recovery samples of severe cases in the first cohort. RESULTS: Severe RSV infection was associated with a transient but marked decrease in CD4+ T, CD8+ T, and NK cells in peripheral blood. Gene expression analyses in both cohorts identified Olfactomedin4 (OLFM4) as a fully discriminative marker between children with mild and severe RSV infection, giving a PAM cross-validation error of 0%. Patients with an OLFM4 gene expression level above -7.5 were 6 times more likely to develop severe disease, after correction for age at hospitalization and gestational age. CONCLUSION: By combining genome-wide expression profiling of blood cell subsets with clinically well-annotated samples, OLFM4 was identified as a biomarker for severity of pediatric RSV infection.
Aroniadou-Anderjaska V, etal., Neuroscience. 2018 Mar 1;373:145-158. doi: 10.1016/j.neuroscience.2018.01.021. Epub 2018 Jan 13.
Synchronous, rhythmic firing of GABAergic interneurons is a fundamental mechanism underlying the generation of brain oscillations, and evidence suggests that NMDA receptors (NMDARs) play a key role in oscillatory activity by regulating the activity of interneuro
ns. Consistent with this, derangement of brain rhythms in certain neuropsychiatric disorders, notably schizophrenia and autism, is associated with NMDAR hypofunction and loss of inhibitory interneurons. In the basolateral amygdala (BLA)-dysfunction of which is involved in a host of neuropsychiatric diseases-, principal neurons display spontaneous, rhythmic "bursts" of inhibitory activity, which could potentially be involved in the orchestration of oscillations in the BLA network; here, we investigated the role of NMDARs in these inhibitory oscillations. Rhythmic bursts of spontaneous IPSCs (0.5 Hz average burst frequency) recorded from rat BLA principal cells were blocked or significantly suppressed by D-AP5, and could be driven by NMDAR activation alone. BLA interneurons generated spontaneous bursts of suprathreshold EPSCs at a similar frequency, which were also blocked or reduced by D-AP5. PEAQX (GluN2A-NMDAR antagonist; 0.4 μM) or Ro-25-6981 (GluN2B-NMDAR antagonist; 5 μM) suppressed the IPSC and EPSC bursts; suppression by PEAQX was significantly greater than that by Ro-25-6981. Immunohistochemical labeling revealed the presence of both GluN2A- and GluN2B-NMDARs on GABAergic BLA interneurons, while, functionally, GluN2A-NMDARs have the dominant role, as suggested by a greater reduction of NMDA-evoked currents by PEAQX versus Ro-25-6981. Entrainment of BLA principal neurons in an oscillatory generation of inhibitory activity depends primarily on activation of GluN2A-NMDARs, and interneuronal GluN2A-NMDARs may play a significant role.
With the emergence of multiple lines of highly effective Human Epidermal Growth Factor Receptor 2 (HER2) directed therapy, accurate identification of HER2 positive tumour has become a critical aspect in the histopathological analysis of breast cancers. Multifocal invasive breast carcinomas are relat
ively common, and given the aggressive inherent biology of HER2 positive disease, identification of even small tumours with HER2 positive status may be of importance for treatment planning. There are currently no clear guidelines as to whether all of these foci should be tested for HER2 status. We reviewed the results of 172 patients in whom HER2 in situ hybridisation (ISH) testing was performed on at least two ipsilateral synchronous invasive carcinomas. Discordant results in different invasive foci were relatively uncommon and occurred in only eight (5%) of the 172 patients. This showed a statistically significant correlation with similarly discordant oestrogen receptor (ER) results. In addition HER2 discordance was more likely amongst different tumour foci if these arose in distinct and separate areas of DCIS. An algorithm based on a combination of College of American Pathologists (CAP) recommendation for HER2 testing, differing ER status and background DCIS profile may be useful in detecting these discordant cases.
BACKGROUND: It is unknown why human metapneumovirus (HMPV) and respiratory syncytial virus (RSV) cause severe respiratory infection in children, particularly in premature infants. Our aim was to investigate if there are defective airway antiviral responses to th
Bolger G, etal., Can J Physiol Pharmacol. 2005 Feb;83(2):198-213.
Intranasal infection of BALB/c mice with respiratory syncytial virus (RSV)-A2 (0.5 x 10(8) - 2.0 x 10(8) plaque-forming units, PFU) produced disease characterized by weight loss (2-3 g) and mortality (60%-100%) with the mean day of death ranging from 6-7 d after
infection. The extent of RSV disease was inoculum titer-dependent and required a replication competent virus. Lung titers of virus peaked at 0.5-1 x 10(6) PFU/g wet weight. Bronchoalveolar lavage fluid (BALF) levels of IL-1beta, TNF-alpha, INF-gamma IL-12, IL-6, MIP-1alpha, RANTES, and protein were elevated, whereas IL-2, IL-4, IL-5, IL-13, and IL-10 were unchanged. Histological assessment of lungs revealed marked inflammatory pathology characterized by bronchiolitis, vasculitis, and interstitial pneumonia. Whole-body plethysmography revealed significant disease-associated deficits of respiratory function. Therapy with ribavirin administered either by the intranasal, subcutaneous, or oral route significantly reduced disease in a dose-dependent manner. Delaying the initiation of therapy resulted in a loss of activity for ribavirin. Synagis administered either intramuscularly as a single dose in prophylaxis or intranasally in prophylaxis, followed by therapy, also significantly reduced disease in a dose-dependent manner. Infection of mice with a high titer inoculum of RSV-A2 resulted in severe and fatal pulmonary disease that was responsive to treatment. This model may be useful to characterize the in vivo activity of experimental therapies for RSV infection.
BACKGROUND: Actin is required for the gene expression and morphogenesis of respiratory syncytial virus (RSV), a clinically important Pneumovirus of the Paramyxoviridae family. In HEp-2 cells, RSV infection also induces actin stress fibers, which may be important
in the immunopathology of the RSV disease. Profilin, a major regulator of actin polymerization, stimulates viral transcription in vitro. Thus, we tested the role of profilin in RSV growth and RSV-actin interactions in cultured cells (ex vivo). RESULTS: We tested three cell lines: HEp-2 (human), A549 (human), and L2 (rat). In all three, RSV grew well and produced fused cells (syncytium), and two RSV proteins, namely, the phosphoprotein P and the nucleocapsid protein N, associated with profilin. In contrast, induction of actin stress fibers by RSV occurred in HEp-2 and L2 cells, but not in A549. Knockdown of profilin by RNA interference had a small effect on viral macromolecule synthesis but strongly inhibited maturation of progeny virions, cell fusion, and induction of stress fibers. CONCLUSIONS: Profilin plays a cardinal role in RSV-mediated cell fusion and viral maturation. In contrast, interaction of profilin with the viral transcriptional proteins P and N may only nominally activate viral RNA-dependent RNA polymerase. Stress fiber formation is a cell-specific response to infection, requiring profilin and perhaps other signaling molecules that are absent in certain cell lines. Stress fibers per se play no role in RSV replication in cell culture. Clearly, the cellular architecture controls multiple steps of host-RSV interaction, some of which are regulated by profilin.
Progesterone pretreatment of ovariectomized rat uteri increases the number of synchronously proliferating stromal cells in response to estradiol 17-beta. To identify the signals involved in stimulating synchronous proliferat
ion, sexually mature ovariectomized rats were injected with progesterone (2 mg) for 3 consecutive days. Estradiol 17-beta (0.2 microg) was administered to initiate cell cycle entry. Uterine samples were removed at various times after hormone administration and changes in wingless (Wnt) pathway effectors and gene targets were identified by microarray. Progesterone pretreatment decreased glycogen synthase kinase-3beta (GSK-3beta) and increased expression of T-cell factor/lymphoid enhancer factor (TCF/LEF). GSK-3beta protein decreased markedly in the uterine stroma of progesterone-pretreated uteri with the concomitant appearance of beta-catenin in these stromal cells. Translocation of beta-catenin from the cytosol to the nuclei in progesterone-pretreated stromal cells was stimulated in response to estradiol. Beta-catenin binding to TCF/LEF increased (P<0.05) in progesterone-pretreated uteri in response to estradiol. Progesterone stimulated the expression of the Wnt target gene urokinase plasminogen activator receptor (uPA-R) in the periluminal uterine stromal cells. The expression of uPA-R increased in progesterone-pretreated stromal cells in response to estradiol administration. Together, the results indicate that progesterone initiates Wnt signaling in the uterine stroma by down-regulating GSK-3beta. However, nuclear translocation of beta-catenin and sufficient complex formation with TCF/LEF to activate stromal cell cycle entry requires estradiol. Stimulation of a uterine stromal cell line to proliferate and differentiate resulted in beta-catenin accumulation, suggesting that endocrine-dependent Wnt signaling controls proliferation and differentiation (decidualization).
Zang N, etal., J Virol. 2015 Dec 16;90(5):2536-43. doi: 10.1128/JVI.02534-15.
Children with acute respiratory syncytial virus (RSV) infection often develop sequelae of persistent airway inflammation and wheezing. Pulmonary C fibers (PCFs) are involved in the generation of airway inflammation and resistance; however, their role in persiste
nt airway diseases after RSV is unexplored. Here, we elucidated the pathogenesis of PCF activation in RSV-induced persistent airway disorders. PCF-degenerated and intact mice were used in the current study. Airway inflammation and airway resistance were evaluated. MMP408 and FSLLRY-NH2 were the selective antagonists for MMP-12 and PAR2, respectively, to investigate the roles of MMP-12 and PAR2 in PCFs mediating airway diseases. As a result, PCF degeneration significantly reduced the following responses to RSV infection: augmenting of inflammatory cells, especially macrophages, and infiltrating of inflammatory cells in lung tissues; specific airway resistance (sRaw) response to methacholine; and upregulation of MMP-12 and PAR2 expression. Moreover, the inhibition of MMP-12 reduced the total number of cells and macrophages in bronchiolar lavage fluid (BALF), as well infiltrating inflammatory cells, and decreased the sRaw response to methacholine. In addition, PAR2 was upregulated especially at the later stage of RSV infection. Downregulation of PAR2 ameliorated airway inflammation and resistance following RSV infection and suppressed the level of MMP-12. In all, the results suggest that PCF involvement in long-term airway inflammation and airway hyperresponsiveness occurred at least partially via modulating MMP-12, and the activation of PAR2 might be related to PCF-modulated MMP-12 production. Our initial findings indicated that the inhibition of PCF activity would be targeted therapeutically for virus infection-induced long-term airway disorders. IMPORTANCE: The current study is critical to understanding that PCFs are involved in long-term airway inflammation and airway resistance after RSV infection through mediating MMP-12 production via PAR2, indicating that the inhibition of PCF activity can be targeted therapeutically for virus infection-induced long-term airway disorders.
Johnson TR, etal., J Leukoc Biol. 2008 Sep;84(3):748-59. Epub 2008 Jun 2.
Severe illness, type 2 cytokine production, and pulmonary eosinophilia are adverse immune responses resulting from respiratory syncytial virus (RSV) challenge of vvGs-immunized mice. We have shown IL-4 and IL-13 activity must be simultaneously inhibited to reduc
e disease severity. We now address the contributions of IL-5, eotaxin-1, and CD4+ and CD8+ T cells to the induction of disease-enhancing immune responses. Depletion of CD4+ T cells during immunization prevented IL-4, IL-13, and eotaxin-1 production, diminished eosinophilia, and reduced weight loss. Conversely, CD8+ T cell depletion did not decrease eosinophilia, weight loss, or type 2 cytokines but did dramatically reduce mucus production and increase eotaxin production. Anti-IL-5 administration at immunization or challenge significantly decreased pulmonary eosinophilia. Strikingly, there were not concomitant decreases in weight loss. Following RSV challenge eotaxin-1-deficient mice immunized with vvGs exhibited significantly less eosinophilia without decreased weight loss or type 2 cytokine production. We conclude CD4+ T cell production of IL-5 and induction of eotaxin-1 are required for vvGs-induced eosinophilia following RSV challenge, while CD8+ T cells appear to down-regulate eotaxin-1 and mucus production. In summary, we demonstrate that pulmonary eosinophilia 1) is a by-product of memory CD4+ T cell activation, 2) does not necessarily correlate with mucus production, and, most importantly, 3) is not required for the RSV G-induced illness in mice. These findings have important implications for the evaluation of candidate RSV vaccines.
Herd KA, etal., J Gen Virol. 2010 May;91(Pt 5):1302-10. Epub 2010 Jan 6.
Human metapneumovirus (hMPV) is a major cause of upper and lower respiratory-tract infection in infants, the elderly and immunocompromised individuals. Virus-directed cellular immunity elicited by hMPV infection is poorly understood, in contrast to the phylogenetically and clinically related pathoge
n human respiratory syncytial virus (hRSV). In a murine model of acute lower respiratory-tract infection with hMPV, we demonstrate the accumulation of gamma interferon (IFN-gamma)-producing CD8+ T cells in the airways and lungs at day 7 post-infection (p.i.), associated with cytotoxic T lymphocytes (CTLs) directed to an epitope of the M2-1 protein. This CTL immunity was accompanied by increased pulmonary expression of Th1 cytokines IFN-gamma and interleukin (IL)-12 and antiviral cytokines (IFN-beta), as well as chemokines Mip-1alpha, Mip-1beta, Mig, IP-10 and CX3CL1. There was also a moderate increase in Th2-type cytokines IL-4 and IL-10 compared with uninfected mice. At 21 days p.i., a strong CTL response could be recalled from the spleen. A similar pattern of CTL induction to the homologous M2-1 CTL epitope of hRSV, and of cytokine/chemokine induction, was observed following infection with hRSV, highlighting similarities in the cellular immune response to the two related pathogens.
More HL, etal., Front Netw Physiol. 2023 Aug 29;3:1208303. doi: 10.3389/fnetp.2023.1208303. eCollection 2023.
Introduction: Tubuloglomerular feedback (TGF) is the negative feedback component of renal blood flow (RBF) autoregulation. Neighbouring nephrons often exhibit spontaneous TGF oscillation and synchronization mediated by endothelial communication, largely via conn
exin40 (Cx40). Methods: We had a knockout (KO) rat made that lacks Cx40. One base pair was altered to create a stop codon in exon 1 of Gja5, the gene that encodes Cx40 (the strain is WKY-Gja55em1Mcwi). Blood pressure (BP)-RBF transfer functions probed RBF dynamics and laser speckle imaging interrogated the dynamics of multiple efferent arterioles that reach the surface (star vessels). Results: The distribution of wild type (WT), heterozygote, and KO pups at weaning approximated the Mendelian ratio of 1:2:1; growth did not differ among the three strains. The KO rats were hypertensive. BP-RBF transfer functions showed low gain of the myogenic mechanism and a smaller TGF resonance peak in KO than in WT rats. Laser speckle imaging showed that myogenic mechanism had higher frequency in KO than in WT rats, but similar maximum spectral power. In contrast, the TGF frequency was similar while peak power of its oscillation was much smaller in KO than in WT rats. In WT rats, plots of instantaneous TGF phase revealed BP-independent TGF synchronization among star vessels. The synchronization could be both prolonged and widespread. In KO rats TGF synchronization was not seen, although BP transients could elicit short-lived TGF entrainment. Discussion: Despite the reduced TGF spectral power in KO rats, there was sufficient TGF gain to induce oscillations and therefore enough gain to be effective locally. We conclude that failure to synchronize is dependent, at least in part, on impaired conducted vasomotor responses.
Qi F, etal., Int Immunopharmacol. 2015 Dec;29(2):408-15. doi: 10.1016/j.intimp.2015.10.022.
Respiratory syncytial virus (RSV) infection can increase the production of IL-33 in lungs of mice. However, little is known about cellular source of IL-33, particularly the types of IL-33-producing cells in innate immune cells during RSV infection. In this stud
y, by using BALB/c mice that were infected intranasally with RSV, it became clear that RSV infection can enhance not only the number of IL-33(+)-alveolar macrophages (AMs) and dendritic cells (DCs), but also the expression of IL-33 mRNA in these cells, suggesting that innate immune cells participate in the production of IL-33. Indeed, in vitro experiments by using murine cell lines found that RSV infection results in more expression of IL-33 mRNA in AMs and DCs, further confirming that these cell types may be an important source of IL-33 during RSV infection. It should be noted that the expression of mRNA for TLR3 and TLR7 was up-regulated in pulmonary AMs during RSV infection. Blockade of TLRs by TLR3 or TLR7 antagonist significantly reduces the levels of IL-33 mRNA in AMs and DCs, suggesting that RSV-induced IL-33 production might be TLRs-dependent manner. Although the expression of TLRs mRNA in pulmonary interstitial macrophages (IMs) was enhanced after RSV infection, stimulation with agonists or inactivated RSV cannot alter the expression of IL-33 mRNA in IMs, suggesting that pulmonary IMs may not be a source of IL-33 during RSV infection. Thus, these results demonstrate that during RSV infection, respiratory macrophages and dendritic cells mediate the production of IL-33 in a TLR-dependent manner.
BACKGROUND: Respiratory Syncytial Virus (RSV) infection is usually restricted to the respiratory epithelium. Few studies have documented the presence of RSV in the systemic circulation, however there is no consistent information whether virus detection in the bl
ood correlates with disease severity. METHODS: Balb/c mice were inoculated with live RSV, heat-inactivated RSV or medium. A subset of RSV-infected mice was treated with anti-RSV antibody 72 h post-inoculation. RSV RNA loads were measured by PCR in peripheral blood from day 1-21 post-inoculation and were correlated with upper and lower respiratory tract viral loads, the systemic cytokine response, lung inflammation and pulmonary function. Immunohistochemical staining was used to define the localization of RSV antigens in the respiratory tract and peripheral blood. RESULTS: RSV RNA loads were detected in peripheral blood from day 1 to 14 post-inoculation, peaked on day 5 and significantly correlated with nasal and lung RSV loads, airway obstruction, and blood CCL2 and CXCL1 expression. Treatment with anti-RSV antibody reduced blood RSV RNA loads and improved airway obstruction. Immunostaining identified RSV antigens in alveolar macrophages and peripheral blood monocytes. CONCLUSIONS: RSV RNA was detected in peripheral blood upon infection with live RSV, followed a time-course parallel to viral loads assessed in the respiratory tract and was significantly correlated with RSV-induced airway disease.
Harcourt J, etal., J Immunol. 2006 Feb 1;176(3):1600-8.
Interactions between fractalkine (CX3CL1) and its receptor, CX3CR1, mediate leukocyte adhesion, activation, and trafficking. The respiratory syncytial virus (RSV) G protein has a CX3C chemokine motif that can bind CX3CR1 and modify CXCL1-mediated responses. In t
his study, we show that expression of the RSV G protein or the G protein CX3C motif during infection is associated with reduced CX3CR1+ T cell trafficking to the lung, reduced frequencies of RSV-specific, MHC class I-restricted IFN-gamma-expressing cells, and lower numbers of IL-4- and CX3CL1-expressing cells. In addition, we show that CX3CR1+ cells constitute a major component of the cytotoxic response to RSV infection. These results suggest that G protein and the G protein CX3C motif reduce the antiviral T cell response to RSV infection.
de Souza AP, etal., Clin Exp Immunol. 2016 Feb;183(2):248-57. doi: 10.1111/cei.12720. Epub 2015 Nov 24.
Respiratory syncytial virus (RSV)-specific CD8(+) T cell responses do not protect against reinfection. Activation of mammalian target of rapamycin (mTOR) impairs memory CD8(+) T cell differentiation. Our hypothesis was that RSV inhibits the formation of CD8(+) T
cells memory responses through mTOR activation. To explore this, human and mouse T cells were used. RSV induced mTOR phosphorylation at Ser2448 in CD8 T cells. mTOR activation by RSV was completely inhibited using rapamycin. RSV-infected children presented higher mTOR gene expression on nasal washes comparing to children infected with metapneumovirus and rhinovirus. In addition, RSV-infected infants presented a higher frequency of CD8(+) pmTORser2448(+) T cells in nasal washes compared to RSV-negative infants. Rapamycin treatment increased the frequency of mouse CD8 RSV-M282-90 pentamer-positive T cells and the frequency of RSV-specific memory T cells precursors. These data demonstrate that RSV is activating mTOR directly in CD8 T cells, indicating a role for mTOR during the course of RSV infection.
BACKGROUND: Respiratory syncytial virus (RSV) is a major health care burden with a particularly high worldwide morbidity and mortality rate among infants. Data suggest that severe RSV-associated illness is in part caused by immunopathology associated
with a robust type 2 response. OBJECTIVE: We sought to determine the capacity of RSV infection to stimulate group 2 innate lymphoid cells (ILC2s) and the associated mechanism in a murine model. METHODS: Wild-type (WT) BALB/c, thymic stromal lymphopoietin receptor (TSLPR) knockout (KO), or WT mice receiving an anti-TSLP neutralizing antibody were infected with the RSV strain 01/2-20. During the first 4 to 6 days of infection, lungs were collected for evaluation of viral load, protein concentration, airway mucus, airway reactivity, or ILC2 numbers. Results were confirmed with 2 additional RSV clinical isolates, 12/11-19 and 12/12-6, with known human pathogenic potential. RESULTS: RSV induced a 3-fold increase in the number of IL-13-producing ILC2s at day 4 after infection, with a concurrent increase in total lung IL-13 levels. Both thymic stromal lymphopoietin (TSLP) and IL-33 levels were increased 12 hours after infection. TSLPR KO mice did not mount an IL-13-producing ILC2 response to RSV infection. Additionally, neutralization of TSLP significantly attenuated the RSV-induced IL-13-producing ILC2 response. TSLPR KO mice displayed reduced lung IL-13 protein levels, decreased airway mucus and reactivity, attenuated weight loss, and similar viral loads as WT mice. Both 12/11-19 and 12/12-6 similarly induced IL-13-producing ILC2s through a TSLP-dependent mechanism. CONCLUSION: These data demonstrate that multiple pathogenic strains of RSV induce IL-13-producing ILC2 proliferation and activation through a TSLP-dependent mechanism in a murine model and suggest the potential therapeutic targeting of TSLP during severe RSV infection.
Respiratory syncytial virus (RSV) nonstructural protein 1(NS1) attenuates type-I interferon (IFN) production during RSV infection; however the precise role of RSV NS1 protein in orchestrating the early host-virus interaction during infection is poorly understoo
d. Since NS1 constitutes the first RSV gene transcribed and the production of IFN depends upon RLR (RIG-I-like receptor) signaling, we reasoned that NS1 may interfere with this signaling. Herein, we report that NS1 is localized to mitochondria and binds to mitochondrial antiviral signaling protein (MAVS). Live-cell imaging of rgRSV-infected A549 human epithelial cells showed that RSV replication and transcription occurs in proximity to mitochondria. NS1 localization to mitochondria was directly visualized by confocal microscopy using a cell-permeable chemical probe for His(6)-NS1. Further, NS1 colocalization with MAVS in A549 cells infected with RSV was shown by confocal laser microscopy and immuno-electron microscopy. NS1 protein is present in the mitochondrial fraction and co-immunoprecipitates with MAVS in total cell lysatesof A549 cells transfected with the plasmid pNS1-Flag. By immunoprecipitation with anti-RIG-I antibody, RSV NS1 was shown to associate with MAVS at an early stage of RSV infection, and to disrupt MAVS interaction with RIG-I (retinoic acid inducible gene) and the downstream IFN antiviral and inflammatory response. Together, these results demonstrate that NS1 binds to MAVS and that this binding inhibits the MAVS-RIG-I interaction required for IFN production.
Rivera-Toledo E, etal., Viruses. 2015 Oct 16;7(10):5361-74. doi: 10.3390/v7102879.
Type-I interferon (IFN-I) production is an early response to viral infection and pathogenic viruses have evolved multiple strategies to evade this cellular defense. Some viruses can establish and maintain persistent infections by altering the IFN-I signaling pathway. Here, we studied IFN-I synthesi
s and response in an in vitro model of persistent infection by respiratory syncytial virus (RSV) in a murine macrophage-like cell line. In this model, interferon regulatory factor 3 was constitutively active and located at nuclei of persistently infected cells, inducing expression of IFN-beta mRNA and protein. However, persistently infected macrophages did not respond in an autocrine manner to the secreted-IFN-beta or to recombinant-IFN-beta, since phosphorylated-STAT1 was not detected by western blot and transcription of the interferon-stimulated genes (ISGs) Mx1 and ISG56 was not induced. Treatment of non-infected macrophages with supernatants from persistently infected cells induced STAT1 phosphorylation and ISGs expression, mediated by the IFN-I present in the supernatants, because blocking the IFN-I receptor inhibited STAT1 phosphorylation. Results suggest that the lack of autocrine response to IFN-I by the host cell may be one mechanism for maintenance of RSV persistence. Furthermore, STAT1 phosphorylation and ISGs expression induced in non-infected cells by supernatants from persistently infected macrophages suggest that RSV persistence may trigger a proinflammatory phenotype in non-infected cells as part of the pathogenesis of RSV infection.
BACKGROUND: Respiratory syncytial virus (RSV) is the leading respiratory viral pathogen in young children worldwide. RSV disease is associated with acute airway obstruction (AO), long-term airway hyperresponsiveness (AHR), and chronic lung inflammation. Using tw
o different mouse strains, this study was designed to determine whether RSV disease patterns are host-dependent. C57BL/6 and BALB/c mice were inoculated with RSV and followed for 77 days. RSV loads were measured by plaque assay and polymerase chain reaction (PCR) in bronchoalveolar lavage (BAL) and whole lung samples; cytokines were measured in BAL samples. Lung inflammation was evaluated with a histopathologic score (HPS), and AO and AHR were determined by plethysmography. RESULTS: Viral load dynamics, histopathologic score (HPS), cytokine concentrations, AO and long-term AHR were similar in both strains of RSV-infected mice, although RSV-infected C57BL/6 mice developed significantly greater AO compared with RSV-infected BALB/c mice on day 5. PCR detected RSV RNA in BAL samples of RSV infected mice until day 42, and in whole lung samples through day 77. BAL concentrations of cytokines TNF-alpha, IFN-gamma, and chemokines MIG, RANTES and MIP-1alpha were significantly elevated in both strains of RSV-infected mice compared with their respective controls. Viral load measured by PCR significantly correlated with disease severity on days 14 and 21. CONCLUSION: RSV-induced acute and chronic airway disease is independent of genetic background.
John AE, etal., Eur J Immunol. 2005 Jan;35(1):108-16.
Severe respiratory syncytial virus (RSV) infection has a significant impact on airway function, and may alter subsequent development of asthma. CCR1 mRNA was significantly up-regulated during primary RSV infection in BALB/c mice, and was also up-regulated during
allergen exposure in sensitized mice. Although CCR1(-/-) mice exhibited similar levels of airway hyperresponsiveness (AHR) as wild-type mice in response to cockroach allergen alone, in animals treated with RSV prior to cockroach antigen (CRA) sensitization and challenge, a significant decrease in exacerbated AHR was observed in the CCR1(-/-) mice. The reduction in AHR after RSV and allergen challenge in CCR1(-/-) mice was not associated with changes in peribronchial eosinophilia, but was accompanied by significantly decreased IL-13 levels in the lungs, as well as an absence of mucus cell staining within the airways. When T lymphocyte numbers were compared in animals receiving CRA to animals receiving a combination of RSV and allergen an increase in both CD4 and CD8 T lymphocytes could be detected in wild-type but not CCR1(-/-) animals. Thus, these data suggest that CCR1-mediated responses have a primary role for inducing severe disease during RSV infection, and may be responsible for altering the lung pathophysiological responses to subsequent allergen challenges via IL-13-mediated mechanisms.
Vasohibin-1 (VASH1) and vasohibin-2 (VASH2), the 2 members of the vasohibin family, have been identified as novel regulators of angiogenesis. VASH1 ceases angiogenesis, whereas VASH2 stimulates sprouting. Here we characterized their functional role in the placenta. Immunohistochemical analysis of hu
man placental tissue clarified their distinctive localization; VASH1 in endothelial cells and VASH2 in trophoblasts. We then used a mouse model to explore their function. Wild-type, Vash1((-/-)), and Vash2((-/-)) mice on a C57BL6 background were used in their first pregnancy. As expected, the fetal vascular area was increased in the Vash1((-/-)) mice, whereas it was decreased in the Vash2((-/-)) mice relative to wild-type. In addition, we noticed that the Vash2((-/-)) mice at 18.5dpc displayed thinner villi of the labyrinth and larger maternal lacunae. Careful observation by an electron microscopy revealed that the syncytiotrophoblast formation was defective in the Vash2((-/-)) mice. To test the possible involvement of VASH2 in the syncytiotrophoblast formation, we examined the fusion of BeWo cells, a human trophoblastoid choriocarcinoma cell line. The forskolin treatment induced the fusion of BeWo cells, and the knockdown of VASH2 expression significantly inhibited this cell fusion. Conversely, the overexpression of VASH2 by the infection with adenovirus vector encoding human VASH2 gene significantly increased the fusion of BeWo cells. Glial cell missing-1 and endogenous retrovirus envelope glycoprotein Syncytin 1 and Syncytin 2 are known to be involved in the fusion of trophoblasts. However, VASH2 did not alter their expression in BeWo cells. These results indicate that VASH1 and VASH2 showed distinctive localization and opposing function on the fetoplacental vascularization. Moreover, our study shows for the first time that VASH2 expressed in trophoblasts is involved in the regulation of cell fusion for syncytiotrophoblast formation.
Bian C, etal., Steroids. 2012 Jan;77(1-2):149-56. Epub 2011 Nov 9.
The structure and function including synaptic plasticity of the hippocampus are deeply affected by steroids in a sex-dependant manner, these processes are believed to be mediated by steroid receptors though their coactivators. Our previous studies have reported the developmental profiles of steroid
receptor coactivator-1 (SRC-1) and PSD-95 in the hippocampus of postnatal female rats and the sex-differences of SRC-1 immunoreactivities in the brain of adult mice. However, whether there are any sex differences about postnatal development of SRC-1 and synaptic proteins in the hippocampus remain unclear. In this study, we investigated the postnatal profile of SRC-1 and key synaptic protein synaptophysin (SYN), PSD-95 and GluR1 in the hippocampus of female and male mice using immunohistochemistry and Western blot. The results showed that in the female hippocampus, the highest levels of SRC-1 were detected at P14, SYN and GluR1 at P30 and PSD-95 at P60; while in the males, the highest levels of SRC-1, SYN and GluR1 were detected at P30, and PSD-95 at P60. Female hippocampus tended to have higher levels of SRC-1, SYN and GluR1 before P30 and PSD-95 before P14; while male hippocampus have higher levels of PSD-95 at P14, P60 and GluR1 at P0. Correlation analysis showed the profiles of SRC-1 were highly correlated with each synaptic protein. The above results showed that in the hippocampus, except some minor sex differences detected at some time-point examined, females and males shared similar postnatal developmental profile and SRC-1 may be deeply involved in the regulation of hippocampal synaptogenesis.
BACKGROUND: Surfactant proteins (SP) are important for the innate host defence and essential for a physiological lung function. Several linkage and association studies have investigated the genes coding for different surfactant proteins in the context of pulmonary diseases such as chronic obstructiv
e pulmonary disease or respiratory distress syndrome of preterm infants. In this study we tested whether SP-B was in association with two further pulmonary diseases in children, i. e. severe infections caused by respiratory syncytial virus and bronchial asthma. METHODS: We chose to study five polymorphisms in SP-B: rs2077079 in the promoter region; rs1130866 leading to the amino acid exchange T131I; rs2040349 in intron 8; rs3024801 leading to L176F and rs3024809 resulting in R272H. Statistical analyses made use of the Armitage's trend test for single polymorphisms and FAMHAP and FASTEHPLUS for haplotype analyses. RESULTS: The polymorphisms rs3024801 and rs3024809 were not present in our study populations. The three other polymorphisms were common and in tight linkage disequilibrium with each other. They did not show association with bronchial asthma or severe RSV infection in the analyses of single polymorphisms. However, haplotypes analyses revealed association of SP-B with severe RSV infection (p = 0.034). CONCLUSION: Thus our results indicate a possible involvement of SP-B in the genetic predisposition to severe RSV infections in the German population. In order to determine which of the three polymorphisms constituting the haplotypes is responsible for the association, further case control studies on large populations are necessary. Furthermore, functional analysis need to be conducted.
Watanabe M, etal., Biochim Biophys Acta. 2008 Jan;1777(1):104-12. Epub 2007 Nov 6.
To identify genes whose expression in brown adipose tissue (BAT) is up- or down-regulated in cold-exposed rats, we performed microarray analysis of RNA samples prepared from the BAT of cold-exposed rats and of rats kept at room temperature. Previously reported elevations of transcript levels of unco
upling protein (UCP1), type II iodothyronine deiodinase (DIO2), and type III adenylate cyclase (AC3) in the BAT of cold-exposed rats over those in that of rats maintained at room temperature were confirmed. In addition to these changes, remarkable elevations of the transcript levels of several genes that seemed to be associated with the processes of cell-cycle regulation and DNA replication were detected in the BAT of cold-exposed rats, possibly reflecting the significant proliferation of brown adipocytes in response to cold exposure. Up-regulation of the gene encoding sarcomeric mitochondrial type creatine kinase in the BAT of cold-exposed rats was also detected by microarray analysis, but subsequent Northern analysis revealed that the expression of not only the sarcomeric mitochondrial type enzyme, but also that of 2 other subtypes, i.e., cytoplasmic brain type and cytoplasmic muscle type, was elevated in the BAT of cold-exposed rats. Microarray analysis also revealed a significant expression of myoglobin in BAT and its elevation in the BAT of cold-exposed rats, and this result was supported by calibrated Northern analysis. On the contrary, several genes such as regulator of G-protein signaling 2 and IMP dehydrogenase 1 were down-regulated in the BAT of cold-exposed rats. The physiological meaning of these changes accompanying cold exposure was discussed.
Despite significant advances in melanoma therapy, melanoma remains the deadliest form of skin cancer, with a 5-year survival rate of only 15%. Thus, novel treatments are required to address this disease. Notch and ERBB are evolutionarily conserved signaling cascades required for the maintenance of
melanocyte precursors. We show that active Notch1 (Notch1(NIC)) and active (phosphorylated) ERBB3 and ERBB2 correlate significantly and are similarly expressed in both mutated and wild-type BRAF melanomas, suggesting these receptors are co-reactivated in melanoma to promote survival. Whereas blocking either pathway triggers modest effects, combining a ?-secretase inhibitor to block Notch activation and a tyrosine kinase inhibitor to inhibit ERBB3/2 elicits synergistic effects, reducing cell viability by 90% and hampering melanoma tumor growth. Specific inhibition of Notch1 and ERBB3 mimics these results, suggesting these are the critical factors triggering melanoma tumor expansion. Notch and ERBB inhibition blunts AKT and NF?B signaling. Constitutive expression of NF?B partially rescues cell death. Blockade of both Notch and ERBB signaling inhibits the slow cycling JARID1B-positive cell population, which is critical for long-term maintenance of melanoma growth. We propose that blocking these pathways is an effective approach to treatment of melanoma patients regardless of whether they carry mutated or wild-type BRAF.
Inhibitory interneurons often generate synchronous activity as an emergent property of their interconnections. To determine the role of electrical synapses in such activity, we constructed mice expressing histochemical reporters in place of the gap junction prot
ein Cx36. Localization of the reporter with somatostatin and parvalbumin suggested that Cx36 was expressed largely by interneurons. Electrical synapses were common among cortical interneurons in controls but were nearly absent in knockouts. A metabotropic glutamate receptor agonist excited LTS interneurons, generating rhythmic inhibitory potentials in surrounding neurons of both wild-type and knockout animals. However, the synchrony of these rhythms was weaker and more spatially restricted in the knockout. We conclude that electrical synapses containing Cx36 are critical for the generation of widespread, synchronous inhibitory activity.
Maruyama R, etal., Am J Physiol Heart Circ Physiol. 2006 Apr;290(4):H1493-502. Epub 2005 Nov 11.
During early apoptosis, adult cardiomyocytes show unusual beating, suggesting possible participation of abnormal Ca(2+) transients in initiation of apoptotic processes in this cell type. Simultaneously with the beating, these cells show dynamic structural alteration resulting from cytoskeletal disin
tegration that is quite rapid. Because of the specialized structure and extensive cytoskeleton of cardiomyocytes, we hypothesized that its degradation in so short a time would require a particularly efficient mechanism. To better understand this mechanism, we used serial video microscopy to observe beta-adrenergic stimulation-induced apoptosis in isolated adult rat cardiomyocytes while simultaneously recording intracellular Ca(2+) concentration and cell length. Trains of Ca(2+) transients and corresponding rhythmic contractions and relaxations (beating) were observed in apoptotic cells. Frequencies of Ca(2+) transients and beating gradually increased with time and were accompanied by cellular shrinkage. As the cells shrank, amplitudes of Ca(2+) transients declined and diastolic intracellular Ca(2+) concentration increased until the transients were lost. Beating and progression of apoptosis were significantly inhibited by antagonists against the L-type Ca(2+) channel (nifedipine), ryanodine receptor (ryanodine), inositol 1,4,5-trisphosphate receptor (heparin), sarco(endo)plasmic Ca(2+)-ATPase (thapsigargin), and Na(+)/Ca(2+) exchanger (KB-R7943). Electron-microscopic examination of beating cardiomyocytes revealed progressive breakdown of Z disks. Immunohistochemical analysis and Western blot confirmed that disappearance of Z disk constituent proteins (alpha-actinin, desmin, and tropomyosin) preceded degradation of other cytoskeletal proteins. It thus appears that, in adult cardiomyocyte apoptosis, Ca(2+) transients mediate apoptotic beating and efficient sarcomere destruction initiated by Z disk breakdown.
BACKGROUND: As a consequence of acute kidney injury (AKI), proximal tubular cells hyperrespond to endotoxin (lipopolysaccharide, LPS) by exaggerated renal Tnf-alpha Production. This LPS hyperresponsiveness is transcriptionally mediated. The epigenetic pathways that control these responses are unknow
n. METHODS/FINDINGS: We applied multiplex chromatin immunoprecipitation platform (Matrix ChIP) to explore epigenetic pathways that underlie endotoxin hyperresponsiveness in the setting of preceding unilateral renal ischemia/reperfusion (I/R) in mouse AKI model. Endotoxin exposure after I/R resulted in enhanced transcription, manifested by hyperresponsive recruitment of RNA polymerase II (Pol II) at the Tnf-alpha gene. At this locus, LPS but not I/R increased levels of Pol II C-terminal domain (CTD) phosho-serine2 &5 and induced dephosphorylation of the transcription-repressive histone H4 phospho-serine-1. In contrast, I/R but not LPS increased the transcription-permissive histone phosphorylation (H3 phospho-serine-10, H3.3 phospho-serine-31) at the Tnf-alpha gene. In agreement with these observations, I/R but not LPS increased activity of cognate kinases (Erk1/2, Msk1/2 and Aurora A) at the Tnf-alpha locus. Cross-talk of histone phosphorylation and acetylation synergize to active gene expression. I/R and LPS increased histone acetylation. (H3K9/14Ac, H4K5/8/12/16Ac, H2KA5Ac, H2BK4/7Ac). Levels of some histone acetyltransferases at this gene (PCAF and MOF) were increased by I/R but not by LPS, while others were induced by either I/R or LPS and exhibited endotoxin hyperresponsive patterns (GCN5, CBP and p300). The adaptor protein 14-3-3 couples histone phosphorylation with acetylation, and tethers chromatin modifiers/transcription elongation factors to target genes. Both I/R and LPS increased levels of 14-3-3 and several chromatin/transcription modifiers (BRD4, BRG1, HP-1gamma and IKKalpha) at the Tnf-alpha gene, all exhibiting endotoxin hyperresponsive recruitment patterns similar to Pol II. CONCLUSIONS: Our results suggest that I/R and LPS differentially trigger phosphorylation (Pol II and histone) and acetylation (histone) epigenetic pathways that interact at the Tnf-alpha gene to generate endotoxin hyperresponse in AKI.
The translocation of tRNAs through the ribosome proceeds through numerous small steps in which tRNAs gradually shift their positions on the small and large ribosomal subunits. The most urgent questions are: (i) whether these intermediates are important; (ii) how the ribosomal translocase, the GTPase
elongation factor G (EF-G), promotes directed movement; and (iii) how the energy of GTP hydrolysis is coupled to movement. In the light of recent advances in biophysical and structural studies, we argue that intermediate states of translocation are snapshots of dynamic fluctuations that guide the movement. In contrast to current models of stepwise translocation, kinetic evidence shows that the tRNAs move synchronously on the two ribosomal subunits in a rapid reaction orchestrated by EF-G and GTP hydrolysis. EF-G combines the energy regimes of a GTPase and a motor protein and facilitates tRNA movement by a combination of directed Brownian ratchet and power stroke mechanisms.
Dupressoir A, etal., Proc Natl Acad Sci U S A. 2005 Jan 18;102(3):725-30. Epub 2005 Jan 11.
Recently, we and others have identified two human endogenous retroviruses that entered the primate lineage 25-40 million years ago and that encode highly fusogenic retroviral envelope proteins (syncytin-1 and -2), possibly involved in the formation of the placen
ta syncytiotrophoblast layer generated by trophoblast cell fusion at the materno-fetal interface. A systematic in silico search throughout mouse genome databases presently identifies two fully coding envelope genes, present as unique copies and unrelated to any known murine endogenous retrovirus, that we named syncytin-A and -B. Quantitative RT-PCR demonstrates placenta-specific expression for both genes, with increasing transcript levels in this organ from 9.5 to 14.5 days postcoitum. In situ hybridization of placenta cryosections further localizes these transcripts in the syncytiotrophoblast-containing labyrinthine zona. Consistently, we show that both genes can trigger cell-cell fusion in ex vivo transfection assays, with distinct cell type specificities suggesting different receptor usage. Genes orthologous to syncytin-A and -B and disclosing a striking conservation of their coding status are found in all Muridae tested (mouse, rat, gerbil, vole, and hamster), dating their entry into the rodent lineage approximately 20 million years ago. Together, these data strongly argue for a critical role of syncytin-A and -B in murine syncytiotrophoblast formation, thus unraveling a rather unique situation where two pairs of endogenous retroviruses, independently acquired by the primate and rodent lineages, would have been positively selected for a convergent physiological role.
Amanatidou V, etal., Pediatr Infect Dis J. 2006 May;25(5):410-4.
BACKGROUND: Recent data suggest that immunologic response during respiratory syncytial virus (RSV) infection is partially modified through interaction of viral G glycoprotein with the host's chemokine receptor, CX3CR1. We hypothesized that two nonsynonymous, sin
gle-nucleotide polymorphisms of the CX3CR1 gene (CX3CR1-V249I and CX3CR1-T280M) that disrupt the affinity of CX3CR1 for its natural ligand (fractalkine) could also affect the G glycoprotein-CX3CR1 pathway. METHODS: To test the hypothesis, DNA samples were obtained from 82 children hospitalized for RSV bronchiolitis in a 1-year period. One hundred twenty sex-matched healthy adults, without a history of severe lower respiratory tract infections, formed the control group. RESULTS: Epidemiologic data showed an increase in the RSV infection rate during the late winter season, with a peak rate in early spring. Genotyping revealed predominance of the 280M-containing genotypes (M/M or T/M) in cases compared with controls (37.8% versus 20.8%, respectively; odds ratio, 2.03; 95% confidence interval, 1.1-3.9; P = 0.025), demonstrating an association between the common CX3CR1-T280M variations and increased risk of severe RSV bronchiolitis. CONCLUSIONS: Our findings support the hypothesis of the pivotal role of the G glycoprotein CX3CR1 pathway in the pathogenesis of RSV bronchiolitis and propose CX3CR1 as a potential therapeutic target.
Pellissier F, etal., BMC Neurosci. 2007 Oct 29;8:90.
BACKGROUND: Cell adhesion molecules are plasma membrane proteins specialized in cell-cell recognition and adhesion. Two related adhesion molecules, Necl-1 and Necl-2/SynCAM, were recently described and shown to fulfill important functions in the central nervous system. The purpose of the work was to
investigate the distribution, and the properties of Necl-3/SynCAM-2, a previously uncharacterized member of the Necl family with which it shares a conserved modular organization and extensive sequence homology. RESULTS: We show that Necl-3/SynCAM-2 is a plasma membrane protein that accumulates in several tissues, including those of the central and peripheral nervous system. There, Necl-3/SynCAM-2 is expressed in ependymal cells and in myelinated axons, and sits at the interface between the axon shaft and the myelin sheath. Several independent assays demonstrate that Necl-3/SynCAM-2 functionally and selectively interacts with oligodendrocytes. We finally prove that Necl-3/SynCAM-2 is a bona fide adhesion molecule that engages in homo- and heterophilic interactions with the other Necl family members, leading to cell aggregation. CONCLUSION: Collectively, our manuscripts and the works on Necl-1 and SynCAM/Necl-2 reveal a complex set of interactions engaged in by the Necl proteins in the nervous system. Our work also support the notion that the family of Necl proteins fulfils key adhesion and recognition functions in the nervous system, in particular between different cell types.
Pakdel A, etal., Cancer Biomark. 2016;16(2):245-52. doi: 10.3233/CBM-150561.
OBJECTIVE: Liver is the primary site for the metastasis from colorectal cancer (CRC). It has been proposed that carcinoembryonic antigen (CEA) might have an instrumental role in the development of hepatic metastasis from human CRC. The aim of the present study was therefore to investigate
the association between preoperative serum CEA concentrations and the incidence of synchronous CRC-liver metastasis. METHODS: A total of 199 patients with CRC who underwent radical surgery at the Shiraz University hospitals were enrolled in this study. The preoperative serum CEA levels of patients without distant metastasis were compared with those with hepatic metastasis and those with other organs metastasis at the time of initial diagnosis. RESULTS: Preoperative serum CEA concentrations were found to be significantly higher in patients with distant metastases compared to patients with local CRC (P < 0.001). Moreover, the mean serum CEA level in patients with liver metastasis was higher than non-metastasis group (42.89 ng/ml vs 9.62 ng/ml) (p < 0.001). We did not find significant differences in serum CEA concentrations between non-metastasis and the other organ metastasis groups (p> 0.05). CONCLUSIONS: We conclude that patients with high preoperative serum CEA levels should have more intensive follow-up for detection of synchronous liver metastasis.
Richman DD and Bozzette SA, J Infect Dis. 1994 May;169(5):968-74.
Many patients infected with human immunodeficiency virus (HIV) yield syncytium-inducing (SI) virus isolates that are cytopathic in cell culture. The presence of SI virus was assessed in 325 persons entering 11 antiretroviral therapy trials and correlated with bo
th CD4 cell declines and clinical end points. Adjusted mean rates of CD4 cell count decline were 40 and 102 cells/year in the non-SI (NSI) and SI groups, respectively (P < .0001). Rates of decline in 16 persons converting from NSI to SI virus averaged 31 cells/year before conversion and 142 cells/year afterward (P = .04). In a nested case-control analysis, persons who experienced surrogate marker end points or opportunistic infections were 2.3-3.5 times more likely to have SI virus than were controls (P = .01-.04) but who died were similar to controls with respect to virus phenotype (P = .70). Presence of the SI phenotype of HIV is a strong predictor of decline in CD4 cell count and progression of disease; however, controlling for the CD4 cell count, the SI phenotype did not increase the immediate risk of death.
Wang Q, etal., PLoS One. 2011;6(9):e24076. doi: 10.1371/journal.pone.0024076. Epub 2011 Sep 2.
For multicellular organisms, different tissues coordinate to integrate physiological functions, although this systematically and gradually declines in the aging process. Therefore, an association exists between tissue coordination and aging, and investigating the evolution of tissue coordination wit
h age is of interest. In the past decade, both common and heterogeneous aging processes among tissues were extensively investigated. The results on spatial association of gene changes that determine lifespan appear complex and paradoxical. To reconcile observed commonality and heterogeneity of gene changes among tissues and to address evolution feature of tissue coordination with age, we introduced a new analytical strategy to systematically analyze genome-wide spatio-temporal gene expression profiles. We first applied the approach to natural aging process in three species (Rat, Mouse and Drosophila) and then to anti-aging process in Mouse. The results demonstrated that temporal gene expression alteration in different tissues experiences a progressive association evolution from spatial synchrony to asynchrony and stochasticity with age. This implies that tissue coordination gradually declines with age. Male mice showed earlier spatial asynchrony in gene expression than females, suggesting that male animals are more prone to aging than females. The confirmed anti-aging interventions (resveratrol and caloric restriction) enhanced tissue coordination, indicating their underlying anti-aging mechanism on multiple tissue levels. Further, functional analysis suggested asynchronous DNA/protein damage accumulation as well as asynchronous repair, modification and degradation of DNA/protein in tissues possibly contributes to asynchronous and stochastic changes of tissue microenvironment. This increased risk for a variety of age-related diseases such as neurodegeneration and cancer that eventually accelerate organismal aging and death. Our study suggests a novel molecular event occurring in aging process of multicellular species that may represent an intrinsic molecular mechanism of aging.
Santos LD, etal., Eur Respir J. 2020 Dec 10. pii: 13993003.03764-2020. doi: 10.1183/13993003.03764-2020.
Respiratory Syncytial Virus (RSV) is the major cause of acute bronchiolitis in infants under 2 years old. Necroptosis has been implicated in the outcomes of respiratory virus infections. Here we report that RSV infection triggers necroptosis in primary mouse mac
rophages and human monocytes in a RIPK1-, RIPK3-, and MLKL-dependent manner. Moreover, necroptosis pathways are harmful to RSV clearance from alveolar macrophages. Additionally, Ripk3-/- mice were protected from RSV-induced weight loss and presented reduced viral loads in the lungs.Alveolar macrophage depletion also protected mice from weight loss and decreased lung RSV virus load. Importantly, alveolar macrophage depletion abolished the upregulation of Ripk3 and Mlkl gene expression induced by RSV infection in the lung tissue.Autocrine TNF mediated RSV-triggered macrophage necroptosis and necroptosis pathways were also involved in TNF secretion even when macrophages were committed to cell death, which can worsen lung injury during RSV infection. In line, Tnfr1-/- mice had a marked decrease in Ripk3 and Mlkl gene expression and a sharp reduction in the numbers of necrotic alveolar macrophages in the lungs. Finally, we provide evidence that elevated nasal levels of TNF are associated with disease severity in infants with RSV bronchiolitis.We propose that targeting TNF and/or the necroptotic machinery may be valuable as therapeutic approaches to reduce the respiratory morbidity caused by RSV infection in young children.
Thomas NJ, etal., Pediatr Res. 2009 Jul;66(1):70-3.
Severity of lung injury with respiratory syncytial virus (RSV) infection is variable and may be related to genetic variations. This preliminary report describes a prospective, family-based association study of children hospitalized secondary to RSV, aimed to det
ermine whether intragenic and other haplotypes of surfactant proteins (SP)-A and SP-D are transmitted disproportionately from parents to offspring with RSV disease. Genomic DNA was genotyped for several SP-A and SP-D single nucleotide polymorphisms (SNPs). Transmission disequilibrium test analysis was used to determine transmission of variants and haplotypes from parents to affected offspring. Three hundred seventy-five individuals were studied, including 148 children with active RSV disease and one or both parents. The SP-A2 intragenic haplotype 1A was found to be protective (p = 0.013). The SP-D SNP DA160_A may possibly be an "at-risk" marker (p = 0.0058). Additional two- and three-marker haplotypes were associated with severe RSV disease, with two being protective (DA11_T/DA160_G and DA160_G/SP-A2 1A/SP-A1 6A). We conclude that there may be associations between SP-A and SP-D and RSV disease. Further study is required to determine whether these variants can be used to target a high-risk patient population in clinical trials aimed at reducing either the symptoms of acute infection or long-term pulmonary sequelae.
Respiratory syncytial virus (RSV) bronchiolitis is characterized by intense inflammation of the airways, and high levels of proinflammatory cytokines and chemokines can be found in respiratory secretions of affected infants. Important among these chemokines are
RANTES (regulated on activation, normal T cell-expressed and -secreted) and macrophage inflammatory-protein alpha, MIP-1alpha, both of which show correlation with severe RSV bronchiolitis. It is not clear whether high levels of these chemokines are important in disease pathogenesis, and this study addresses this question by studying genetic variants of their major receptor, CC chemokine receptor 5. Results from both a case-control and family-based genetic-association analysis show that the -2459G and -2554T variants are associated with severe RSV bronchiolitis (P=.01). It is proposed that these CCR5 variants influence the inflammatory response, and these data provide further evidence of the important role that host genetic variability plays in the determination of disease severity in RSV bronchiolitis.
Hansdottir S, etal., J Immunol. 2010 Jan 15;184(2):965-74. Epub 2009 Dec 11.
Epidemiological studies suggest that low vitamin D levels may increase the risk or severity of respiratory viral infections. In this study, we examined the effect of vitamin D on respiratory syncytial virus (RSV)-infected human airway epithelial cells. Airway ep
ithelium converts 25-hydroxyvitamin D3 (storage form) to 1,25-dihydroxyvitamin D3 (active form). Active vitamin D, generated locally in tissues, is important for the nonskeletal actions of vitamin D, including its effects on immune responses. We found that vitamin D induces IkappaBalpha, an NF-kappaB inhibitor, in airway epithelium and decreases RSV induction of NF-kappaB-driven genes such as IFN-beta and CXCL10. We also found that exposing airway epithelial cells to vitamin D reduced induction of IFN-stimulated proteins with important antiviral activity (e.g., myxovirus resistance A and IFN-stimulated protein of 15 kDa). In contrast to RSV-induced gene expression, vitamin D had no effect on IFN signaling, and isolated IFN induced gene expression. Inhibiting NF-kappaB with an adenovirus vector that expressed a nondegradable form of IkappaBalpha mimicked the effects of vitamin D. When the vitamin D receptor was silenced with small interfering RNA, the vitamin D effects were abolished. Most importantly we found that, despite inducing IkappaBalpha and dampening chemokines and IFN-beta, there was no increase in viral mRNA or protein or in viral replication. We conclude that vitamin D decreases the inflammatory response to viral infections in airway epithelium without jeopardizing viral clearance. This suggests that adequate vitamin D levels would contribute to reduced inflammation and less severe disease in RSV-infected individuals.
