Modulation of transforming growth factorβ response and signaling during transdifferentiation of rat hepatic stellate cells to myofibroblasts

Activation of hepatic stellate cells (HSCs) is the key step in liver fibrogenesis. Increased transforming growth factor β (TGF-β) expression and extracellular matrix production in patients with hepatic fibrosis and experimental models of liver fibrogenesis support implication of TGF-β in the pathogenesis of this disease. However, a causative role for TGF-β during transdifferentiation of HSCs has not been delineated in molecular detail. Using a rat cell culture model of HSC transdifferentiation, we analyzed TGF-β signal transduction and identified changes between stellate cells and their transdifferentiated phenotype. Fully transdifferentiated myofibroblasts, opposed to HSCs, were not inhibited in proliferation activity on treatment with TGF-β1. Furthermore, stimulation of α2 (I) collagen and Smad7 messenger RNA (mRNA) expression by TGF-β1 was achieved in stellate cells but not in myofibroblasts. Northern and Western blot analyses indicated significant expression of TGF-β receptors I and II in both cell types. In contrast, [125I]–TGF-β1 receptor affinity labeling displayed strongly reduced types I, II, and III receptor presentation at the cell surface of myofibroblasts. Moreover, myofibroblasts did not display DNA-binding SMAD proteins in electrophoretic mobility shift assays with a CAGA box. These data indicate that stellate cells are responsive to TGF-β1 treatment and transduce a signal that may play an important role in liver fibrogenesis. Myofibroblasts display decreased availability of surface receptors for TGF-β, which could be based on autocrine stimulation. However, lack of activated SMAD complexes with DNA-binding activity and absence of α2 (I) collagen transcription inhibition by latency-associated peptide (LAP)/anti-TGF-β antibody raise the possibility of TGF-β signaling independent receptor down-regulation in myofibroblasts.