Abstract
Transient receptor potential channel C6 (TRPC6) gain-of-function mutations and increased TRPC6 expression in podocytes induce glomerular injury and proteinuria. Sildenafil reduces TRPC6 expression and activity in nonrenal cell types, although the mechanism is unknown. Peroxisome proliferator-activated receptor gamma (PPAR-gamma) is a downstream target of sildenafil in the cyclic guanosine monophosphate (cGMP)-activated protein kinase G (PKG) axis. PPAR-gamma agonists, like pioglitazone, appear antiproteinuric. We hypothesized that sildenafil inhibits TRPC6 expression in podocytes through PPAR-gamma-dependent mechanisms, thereby counteracting podocyte injury and proteinuria. Treatment with sildenafil, the cGMP derivative 8-bromoguanosine 3',5'-cyclic monophosphate sodium salt (8-Br-cGMP), or pioglitazone dose-dependently downregulated podocyte injury-induced TRPC6 expression in vitro Knockdown or application of antagonists of PKG or PPAR-gamma enhanced TRPC6 expression in podocytes and counteracted effects of sildenafil and 8-Br-cGMP. We observed similar effects on TRPC6 promoter activity and TRPC6-dependent calcium influx. Chromatin immunoprecipitation showed PPAR-gamma binding to the TRPC6 promoter. Sildenafil or pioglitazone treatment prevented proteinuria and the increased TRPC6 expression in rats with adriamycin-induced nephropathy and mice with hyperglycemia-induced renal injury. Rats receiving PPAR-gamma antagonists displayed proteinuria and increased podocyte TRPC6 expression, as did podocyte-specific PPAR-gamma knockout mice, which were more sensitive to adriamycin and not protected by sildenafil. Thus, sildenafil ameliorates podocyte injury and prevents proteinuria through cGMP- and PKG-dependent binding of PPAR-gamma to the TRPC6 promoter, which inhibits TRPC6 promoter activity, expression, and activity. Because sildenafil is approved for clinical use, our results suggest that additional clinical study of its antiproteinuric effect in glomerular disease is warranted.
