Differentiation of vascular myofibroblasts induced by transforming growth factor-β1 requires the involvement of protein kinase Cα

Abstract

In response to vascular injury, adventitial fibroblasts can modulate their phenotype to myofibroblasts, cells that participate in arterial remodeling. However, the signaling mechanisms underlying the vascular myofibroblast differentiation remain unknown. Since protein kinase C (PKC) is a key enzyme for cell differentiation, we examined whether PKC isoforms were involved in the vascular myofibroblast differentiation. The association between PKCα and myofibroblast differentiation was investigated in cultured rat aortic fibroblasts treated with transforming growth factor-β1 (TGFβ1). Confocal immunofluorescence microscopy indicated that fibroblasts expressed α-smooth muscle actin (α-SM actin) after TGFβ1 treatment. Moreover, TGFβ1 stimulation increased both PKCα mRNA expression (measured by real-time quantitative RT-PCR) and PKC activity (determined by histone-like pseudosubstrate phosphorylation) in adventitial fibroblasts. Western blot analysis indicated that PKCα protein expression was higher in TGFβ1–treated fibroblasts than in untreated cells. TGFβ1–induced expression of α-SM actin was inhibited in a dose-dependent manner by treating cells with a PKC inhibitor, calphostin C, and was abolished by depleting PKCα with antisense PKCα oligodeoxynucleotides. Our results demonstrate that TGFβ1 induces adventitial myofibroblast differentiation via a PKCα-dependent process.