Identification and characterization of transcription factors involved in the myofibroblast formation, and its application to novel therapeutic approach of glomerulonephritis

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Myofibroblasts are recognized as the key to understand the reconstruction and excess matrix formation in injured tissue. We have been investigating the pathophysiological significance of myofibroblasts in renal diseases. We reported that both caldesmon and SMαA are sensitive and useful molecular markers for myofibroblasts in progressive renal disease. In light of the importance of the clinical observation, we started to seek the molecular mechanism of SMαA gene regulation in myofibroblasts to elucidate the underlying mechanisms of myofibroblast formation. Utilizing the in vivo promoter analysis of SMαA gene, we have demonstrated the essential role of intronic CArG motif element for the transcriptional activation of SMαA gene in both smooth muscle cells and in renal myofibroblasts. These results lead us to the further investigation and identification of the transcription factor(s) that bind to this region. We are now on the way of the molecular characterization of the transcription fact … More or that binds to the CArG motif element of the SMαA gene intron 1.The small GTPase Rho is involved in cell-to-substratum adhesion and cell contraction. These actions of Rho mediated by downstream Rho effectors such as Rho-associated coiled-coil forming protein kinase (ROCK) may be partly responsible for the progression of renal interstitial fibrosis. Anti-fibrosis effects of Y-27632, a specific ROCK inhibitor, were studied both in vivo (unilateral ureteral obstruction ; UUO) and in vitro. To investigate the therapeutic efficacy of Y-27632 in UUO kidneys, smooth muscle α actin (SMαA) expression, macrophage infiltration and fibrosis in the obstructed kidneys were studied. SMαA, transforming growth factor β(TGF-β), α1(I) collagen, osteopontin, macrophage chemoattractant peptide-1 (MCP-1), and intercellular adhesion molecule-1 (ICAM-1) gene expression were examined by Northern blotting. To elucidate the mechanism linking the Rho-ROCK pathway with renal fibrosis, the effects of Y-27632 on in vitro cell proliferation and cell migration were studied. In vivo analysis showed that Y-27632 suppressed SMαA expression, macrophage infiltration and interstitial fibrosis, and that Y-27632 suppressed SMαA, TGF-β and α1(I) collagen mRNA expression. In vitro analysis showed that Y-27632 did not suppress proliferation of renal fibroblasts but suppressed migration of macrophages. The Rho-ROCK system may play an important role in the development of tissue fibrosis, and the Rho-ROCK signaling pathway may be a new therapeutic target for preventing interstitial fibrosis in progressive renal disease. Less