Expression of Smads during in Vitro Transdifferentiation of Hepatic Stellate Cells to Myofibroblasts

Abstract

TGFβ is of crucial importance during transdifferentiation of resting retinoid-storing hepatic stellate cells (HSC) to extracellular matrix producing myofibroblasts (MFB) and consequently, inhibition of TGFβ signal transduction is an effective means for preventing experimental fibrosis. We have shown that isolated HSC lose TGFβ-dependent growth control during in vitro activation and that α2 (I) collagen production in transdifferentiated MFB is TGFβ-independent. Furthermore, Smad complexes with SBE binding activity were only detected in early cultures of HSC, although TGFβ receptor types I and II were significantly expressed in HSC and MFB. In the present report, we compared the expression pattern of TGFβ downstream mediators, i.e., the Smads, in TGFβ responsive HSC versus nonresponding MFB. The transdifferentiation process was monitored by morphology and increasing expression of TGFβ and α-smooth muscle actin, and TGFβ signaling was investigated by (CAGA)9-MLP-Luc. The expression level of all Smads remained essentially unchanged both during the activation process and after TGFβ-treatment. Smad7 was transiently upregulated upon TGFβ stimulation in quiescent HSC, indicating a negative feed back loop in responsive cells. In contrast, MFB neither displayed TGFβ-inducible nor constitutively upregulated Smad7 expression. Instead, Smad3 mRNA was increased in MFB. Our data indicate that abrogation of the TGFβ response in MFB versus HSC is not based on different regulation of Smad expression.