Abstract
Expression of α-smooth muscle actin (αSMA) is constitutive in vascular smooth muscle cells, but is induced in nonmuscle cells such as hepatic stellate cells (HSCs). HSCs play important roles in both physiological homeostasis and pathological response. HSC activation is characterized by αSMA expression, which is regulated by the TGFβ-induced Smad pathway. Recently, protein kinase C (PKC) was identified to regulate αSMA expression. Diacylglycerol kinase (DGK) metabolizes a second-messenger DG, thereby controlling components of DG-mediated signaling, such as PKC. In the present study we aimed to investigate the putative role of DGKα in αSMA expression. Use of a cellular model indicated that the DGK inhibitor R59949 promotes αSMA expression and PKCδ phosphorylation. It also facilitates Smad2 phosphorylation after 30 min of TGFβ stimulation. Furthermore, immunocytochemical analysis revealed that DGK inhibitor pretreatment without TGFβ stimulation engenders αSMA expression in a granular pattern, whereas DGK inhibitor pretreatment plus TGFβ stimulation significantly induces αSMA incorporation in stress fibers. Through animal model experiments, we observed that DGKα-knockout mice exhibit increased expression of αSMA in the liver after 48 h of carbon tetrachloride injection, together with enhanced phosphorylation levels of Smad2 and PKCδ. Together, these findings suggest that DGKα negatively regulates αSMA expression by acting on the Smad and PKCδ signaling pathways, which differentially regulate stress fiber incorporation and protein expression of αSMA, respectively.