RhoA/Rho-Kinase Contribute to the Pathogenesis of Diabetic Renal Disease
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OBJECTIVE: Accumulation of glomerular matrix proteins is central to the pathogenesis of diabetic nephropathy, with resident mesangial cells (MCs) known to upregulate matrix protein synthesis in response to high glucose. Because activation of the GTPase RhoA has been implicated in matrix upregulation, we studied its role in induction of the matrix protein fibronectin in diabetic MCs and in vivo in diabetic nephropathy. RESEARCH DESIGN AND METHODS: Glucose (30 mmol/l)-induced RhoA/Rho-kinase, AP-1 activation, and fibronectin upregulation were assessed by immunoblotting, luciferase, electrophoretic mobility shift assay, enzyme-linked immunosorbent assay, real-time PCR, Northern blots, and immunofluorescence. Streptozotocin-induced diabetic rats were treated with the rho-kinase inhibitor fasudil, which was compared with enalapril, and functional and pathologic parameters were assessed. RESULTS: Glucose led to RhoA and downstream Rho-kinase activation. Mannitol was without effect. Activity of the transcription factor AP-1, increased in diabetic MCs and kidneys, is important in the profibrotic effects of glucose, and this was dependent on Rho-kinase signaling. Upregulation of fibronectin by glucose, shown to be mediated by activator protein-1 (AP-1), was prevented by Rho-kinase inhibition. RhoA siRNA and dominant-negative RhoA also markedly attenuated fibronectin upregulation by high glucose. Applicability of these findings were tested in vivo. Fasudil prevented glomerular fibronectin upregulation, glomerular sclerosis, and proteinuria in diabetic rats, with effectiveness similar to enalapril. CONCLUSIONS: High glucose activates RhoA/Rho-kinase in MCs, leading to downstream AP-1 activation and fibronectin induction. Inhibition of this pathway in vivo prevents the pathologic changes of diabetic nephropathy, supporting a potential role for inhibitors of RhoA/Rho in the treatment of diabetic renal disease.
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