Role of connective tissue growth factor (CTGF) module 4 in regulating epithelial mesenchymal transition (EMT) in HK-2 cells

BackgroundRecent studies have suggested that connective tissue growth factor (CTGF) plays a key role in tissue fibrosis including renal scarring. While studies showed several forms of CTGF with 10–38 kDa in the body fluids, little is known about these small molecule species. We investigated the effect of a 10 kDa CTGF molecule consisting of module 4, on the epithelial mesenchymal transition (EMT) in human proximal tubular cell line (HK-2).MethodsHK2 cells were cultured in DMEM medium. The response of cytokeratin (CK) and vimentin (VIM) mRNA and protein expression to the stimulation of rhCTGF(C) were observed by real-time PCR and immunocytochemistry. At the same time, the morphologic changes were observed by microscopy, and expression of α-smooth muscle actin (α-SMA) and fibronectin (FN) was detected by laser confocal microscope. These effects were compared with CTGF N-terminal [rhCTGF(N)], consisting of module 1–3, and observed in a condition with the addition of anti-CTGF antibody.ResultsRhCTGF(C) induced striking changes in epithelial cells, including changes in cellular morphology, loss of CK, gain VIM and α-SMA, and increased levels of fibronectin. Cocultured with anti-CTGF antibody could abrogate most of these effects, while cells treated with rhCTGF(N) showed no significant phenotypic changes comparing to control group.ConclusionsOur results suggest that module 4 could induce HK-2 cells EMT, whereas the residual fragment has no similar effect in spite of consisting of 3 modules of CTGF molecule.