Glomerulosclerosis is one of the complications of diabetes that occurs after many years of uncontrolled hyperglycemia. Mesangial cells (MCs) exposed to high glucose (HG) for short periods have shown that transforming growth factor-beta (TGF-beta) and activated diacylglycerol-dependent protein kinase C (PKC) mediate increased collagen formation. Our study examined collagen formation by MCs exposed to HG for 8 weeks. Exposure to HG in overnight culture resulted in the activation of all PKC isoforms. In contrast, 8-week exposure to HG resulted in the persistent activation of PKC-delta, did not change PKC-alpha or -beta activity, and decreased PKC-varepsilon activity while increasing collagen I and IV gene and protein expression. Collagen IV accumulation was reversed by specific PKC-delta inhibition. Collagen IV gene expression was completely normalized by TGF-beta neutralization; however, this was associated with plasminogen activator inhibitor-1 (PAI-1) overexpression and a modest reduction in collagen protein. Our studies suggest that prolonged exposure to HG results in PKC-delta-driven collagen accumulation by MCs mediated by PAI-1 but independent of TGF-beta.Kidney International advance online publication, 29 August 2007; doi:10.1038/sj.ki.5002517.