Roles Of SM22α In Transforming Growth Factor β-induced Phenotypic Remodeling Of Vascular Smooth Muscle Cels

Objective: Vascular smooth muscle cells (VSMCs) have the capacity to undergo drastic phenotypic modulation from contractile and differentiated state to proliferative, dedifferentiated, chondrocytic, and osteoblastic phenotypes in arterial diseases such as atherosclerosis. Smooth muscle 22 alpha (SM22α), a 22 kDa protein, is highly expressed in differentiated VSMCs. In this study, we used siRNA to knock down SM22αexpression, and analyzed the roles of SM22αin TGFβ-induced phenotypic remodeling of VSMCs.Methods: Immunoprecipitation assay was used to examine the KLF4 phosphorylation level. TGFβ-induced expression of SM22αand SMα-actin and stress fibres formation were examined by Western blotting and immunofluorescent staining, respectively. Oxidative stress was detected by dihydroethidium (DHE) staining.Results:1 TGFβinduces SM22αexpression in VSMCs TGFβ(2 ng/ml) significantly induced SM22αand SMα-actin expression in VSMCs in time-dependent manner.2 SM22αinhibits TGFβ-induced differentiation transcription regulatory factor KLF4 phosphorylationWestern blot showed that KLF4 phosphorylation level was markedly increased in VSMCs treated with TGFβ. Following knockdown of endogenous SM22αby siRNA, TGFβ-induced KLF4 phosphorylation was further increased. This suggests that SM22αinhibits TGFβ-induced KLF4 phosphorylation. 3 SM22αmediates TGFβ- induced actin stress fibre formationThe expression of endogenous SM22αwas knocked down by specific siRNA in VSMCs. Immunofluorescent staining showed that abundant actin stress fibres co-localized with SM22αin TGFβ-treated cells without SM22αknockdown. However, there was scant actin stress fibre formation after SM22αknockdown. The results suggest that SM22αis involved in TGFβ-induced actin stress fibre formation. 4 SM22αinhibits TGFβ- induced oxidative stressDHE staining showed that TGFβstimulation increased production of O2?- in VSMCs. The O2?- production induced by TGFβwas enhanced after SM22αknockdown. It demonstrated that SM22αmight inhibit TGFβ-induced oxidative stress.Conclusions:1 TGFβinduces SM22αexpression in VSMCs.2 SM22αinhibits TGFβ-induced differentiation transcription regulatory factor KLF4 phosphorylation and oxidative stress.3 SM22αmediates TGFβ- induced actin stress fibre formation.