Connective tissue growth factor regulates the key events in tubular epithelial to myofibroblast transition in vitro

Abstract

Connective tissue growth factor (CTGF) has been reported to play an important role in mediating the profibrotic effects of transforming growth factor-beta (TGF-beta) in various renal diseases. To elucidate the role of CTGF in renal tubular epithelial-myofibroblast transdifferentiation, we examined the expression of alpha-smooth muscle actin (alpha-SMA), vimentin, tenascin-C, and collagen IV expression upon the stimulation of CTGF in cultured human proximal tubular epithelial cell line (HKC), and further investigated the effects of endogenous CTGF blockade on the transdifferentiation process induced by TGF-beta. It is revealed that upon the stimulation of recombinant human CTGF (rhCTGF, 2.5 or 5.0 microg/L), the expression of alpha-SMA and tenascin-C mRNA increased significantly (p<0.01), while collagen IV gene expression decreased significantly (p<0.01), all in a dose-dependent manner. The percentage of alpha-SMA-positive cells was significantly larger in the rhCTGF-stimulated groups than that in negative control (38.9%, 65.5% vs. 2.4%, respectively, p<0.01) as confirmed by flow cytometry. Both cytoplasmic and secretory tenascin-C expression was upregulated by the stimulation of rhCTGF (p<0.01). Under this condition, collagen IV secreted into the culture media was lowered markedly (p<0.01). On RT-PCR analysis, TGF-beta1 upregulated CTGF gene expression, preceding that of alpha-SMA. The alpha-SMA mRNA expression induced by TGF-beta1 was significantly inhibited by CTGF antisense oligodeoxynucleotide (ODN) transfection (p<0.01). With prolonged incubation time, CTGF antisense ODN also inhibited intracellular alpha-SMA protein synthesis, as demonstrated by indirect immuno-fluorescence. So it is concluded that CTGF could promote the transdifferentiation of human renal tubular epithelial cells towards myofibroblasts in vitro, both directly and as a downstream mediator of TGF-beta, and CTGF blockade would be a possible therapeutic target against tubulointerstitial fibrosis.

Proteinase-activated Receptor-2 Transactivation of Epidermal Growth Factor Receptor and Transforming Growth Factor-β Receptor Signaling Pathways Contributes to Renal Fibrosis

“…CTGF, along with TGF␤, promotes epithelial to mesenchymal transition in proximal tubular epithelial cells in vitro (49,50). There is no doubt that Smad-dependent signaling not only regulates epithelial to mesenchymal transition but also the expression of CTGF that contribute to renal fibrosis (51).…”

VEGF ameliorates tubulointerstitial fibrosis in unilateral ureteral obstruction mice via inhibition of epithelial-mesenchymal transition

“…CTGF can promote EMT both directly and as a downstream mediator of TGF-β1 [38] . In our study, VEGF treatment reduced the expression of TGF-β1 and CTGF and upregulated the expression of BMP-7.…”

TGF-β1-Induced Connective Tissue Growth Factor (CCN2) Expression in Human Renal Proximal Tubule Epithelial Cells Requires Ras/MEK/ERK and Smad Signalling

“…We also demonstrated the induction of CCN2 mRNA after 90 min of TGF-β1 treatment, which suggests that CCN2 induction by TGF-β1 precedes mesenchymal transdifferentiation of tubule epithelial cells. The CCN2 protein secreted by PTECs in response to TGF-β probably acts in a paracrine manner to mediate tubular epithelial-mesenchymal transdifferentiation [29] and to stimulate fibroblast proliferation and ECM deposition [14, 17]. …”