Abstract
Introduction: Vascular smooth muscle cell (VSMC) proliferation and apoptosis is associated with the development of atherosclerosis. Although Yes-associated protein (YAP), the nuclear effector of Hippo signaling, is known to regulate cellular growth and survival in cancer cell and heart, the biological role of YAP1 in VSMC is largely unknown. The purpose of this study was to explore the role of YAP1 in VSMC in the cell proliferation and apoptosis.
Methods: VSMC isolated from rat thoracic aorta of sprague-Dawley was used in this study. We tried to gain-or loss-of- function approaches using adenovirus gene transfer or siRNA respectively. Cell proliferation was determined by MTS assay. Cell apoptosis was detected by TUNEL assay. The expression and phosphorylation of signal molecules associated with proliferation or apoptosis were assessed by Western blotting.
Results: Cell proliferation was significantly increased in VSMC infected with wild-type (WT) YAP1 or a constitutive active form YAP1 (YAP1-5SA) compared to GFP-infected cells. Western blot analysis showed an up-regulated expression of HB-EGF and increased phosphorylation of Akt in both WT-YAP1- and YAP1-5SA- infected cells. Interestingly, transfer of culture medium from WT- YAP1- and YAP1-5SA- infected cells to normal VSMC cells significantly increased the phosphorylation of Akt. However, cell proliferation did not observe in cell transferred of the medium. Tunicamycin (ER stress inducer) stimulation caused an apoptosis, accompanied by increased cleaved caspase-3, -8 and CHOP expression and decreased YAP1 and ANKRD1 protein expression. Furthermore, knockdown of ANKRD1 resulted in increased cleaved caspase-3 expression. These phenomena were prevented by overexpression of YAP1- 5SA.
Conclusion: These data demonstrated that YAP1 is a regulator protein involved in cell proliferation and apoptosis in VSMC. Accordingly, YAP1 may be a new promising target for the therapy of cardiovascular disease including atherosclerosis.
