Regulation of connective tissue growth factor (CTGF) by hepatocyte growth factor in human tubular epithelial cells

Abstract

Hepatocyte growth factor (HGF) is a pleiotropic protein with renoprotective functions, which have been attributed at least in part to its ability to counteract the profibrotic effects of transforming growth factor beta (TGF-beta). A major downstream mediator of TGF-beta is connective tissue growth factor (CTGF). However, the molecular mechanisms of CTGF regulation by HGF have not yet been investigated. CTGF expression was analysed in human primary tubular epithelial cells (hPTECs) and the cell line HKC-8 by western blotting. Morphological alterations were analysed by immunocytochemistry. HGF induced a transient expression of CTGF, which was maximal after 6 h and returned to baseline after 24 h. Coincubation with TGF-beta increased CTGF protein at 6 h, whereas HGF significantly decreased CTGF induction by TGF-beta after 24 h. Furthermore, HGF induced cell scattering associated with reorganization of focal adhesions and formation of lamellipodia and filopodia. The early induction of CTGF was linked to the HGF-mediated alterations of cell morphology. The PP2 inhibitor of Src-family kinases, which regulate focal adhesion turnover, reduced HGF-mediated upregulation of CTGF. In addition, inhibition of the Rho-kinase, which modulates the actin cytoskeleton, impaired CTGF expression. Combination of both inhibitors further decreased CTGF expression. Comparable inhibitory effects were obtained, when CTGF was induced by the combination of HGF and TGF-beta. We provide evidence for a dual effect of HGF on CTGF regulation in human tubular epithelial cells: transient upregulation of CTGF in the absence of TGF-beta, which was related to alterations of cell morphology, and interference with TGF-beta-mediated CTGF induction after prolonged incubation.