RGD Reference Report - Antisense and short hairpin RNA (shRNA) constructs targeting PIN (Protein Inhibitor of NOS) ameliorate aging-related erectile dysfunction in the rat. - Rat Genome Database

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Antisense and short hairpin RNA (shRNA) constructs targeting PIN (Protein Inhibitor of NOS) ameliorate aging-related erectile dysfunction in the rat.

Authors: Magee, Thomas R  Kovanecz, Istvan  Davila, Hugo H  Ferrini, Monica G  Cantini, Liliana  Vernet, Dolores  Zuniga, Freddi I  Rajfer, Jacob  Gonzalez-Cadavid, Nestor F 
Citation: Magee TR, etal., J Sex Med. 2007 May;4(3):633-43. Epub 2007 Apr 13.
RGD ID: 13207433
Pubmed: PMID:17433082   (View Abstract at PubMed)
DOI: DOI:10.1111/j.1743-6109.2007.00459.x   (Journal Full-text)


INTRODUCTION: Over-expression of penile neuronal nitric oxide synthase (PnNOS) from a plasmid ameliorates aging-related erectile dysfunction (ED), whereas over-expression of the protein inhibitor of NOS (PIN), that binds to nNOS, increases ED.
AIM: To improve this form of gene therapy for ED by comparing the electrical field response of short hairpin RNA (shRNA) for PIN with that of antisense PIN RNA.
MAIN OUTCOME MEASURE: Both shRNA and antisense RNA gene therapy vectors increased intracavernosal pressure in aged rats.
METHODS: PIN small interfering RNA (siRNA), and plasmid constructs for cytomegalovirus promoter plasmid vector (pCMV-PIN), pCMV-PIN antisense RNA, pSilencer2.1-U6-PIN-shRNA; and pSilencer2.1-U6-randomer-shRNA were prepared and validated by transfection into HEK293 cells, determining the effects on PIN expression by Western blot. Plasmid constructs were then injected, followed by electroporation, into the penile corpora cavernosa of aged (20-month-old) Fisher 344 rats and, 1 month later, the erectile response was measured by intracavernosal pressure increase following electrical field stimulation (EFS) of the cavernosal nerve. PIN was estimated in penile tissue by Western blot and real-time reverse transcriptase-polymerase chain reaction. Cyclic guanosine monophosphate (cGMP) measurements were conducted by competitive enzyme immunoassay (EIA). Immunohistofluorescence detected PIN in corporal tissue sections.
RESULTS: In cell culture, PIN siRNA and plasmid-expressed pU6-PIN-shRNA effectively reduced PIN expression from pCMV-PIN. pSilencer2.1-U6-PIN-shRNA corrected the impaired erectile response to EFS in aged rats and raised it above the value for young rats, more efficiently than pCMV-PIN antisense RNA. PIN mRNA expression in the penis was decreased by >70% by the shRNA but remained unaffected by the antisense RNA, whereas PIN protein expression was reduced in both cases, particularly in the dorsal nerve. PIN antisense increased cGMP concentration in treated tissue by twofold.
CONCLUSION: pSilencer2.1-U6-PIN-shRNA gene therapy was more effective than the antisense PIN mRNA in ameliorating ED in the aged rat, thereby suggesting that PIN is indeed a physiological inhibitor of nNOS and nitrergic neurotransmission in the penis.



RGD Manual Disease Annotations    Click to see Annotation Detail View

  
Object SymbolSpeciesTermQualifierEvidenceWithNotesSourceOriginal Reference(s)
DYNLL1Humanimpotence treatmentISODynll1 (Rattus norvegicus) RGD 
Dynll1Ratimpotence treatmentIMP  RGD 
Dynll1Mouseimpotence treatmentISODynll1 (Rattus norvegicus) RGD 

Objects Annotated

Genes (Rattus norvegicus)
Dynll1  (dynein light chain LC8-type 1)

Genes (Mus musculus)
Dynll1  (dynein light chain LC8-type 1)

Genes (Homo sapiens)
DYNLL1  (dynein light chain LC8-type 1)


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