Overexpression of The Cellular Repressor of E1A-stimulated Genes Inhibits The Apoptosis of Human Vascular Smooth Muscle Cells via Blocking p38/JNK MAP Kinase Activation

The cellular repressor of E1A-stimulated genes (CREG) is a recently described glycoprotein which plays a critical role in keeping cells or tissue in homeostasis states. The current study examined whether and how CREG may regulate VSMC apoptosis. Both loss-of-function (CREG-DW by retrovirus expressing CREG shRNAs) and gain-of-function (CREG-UP by retroviral infection with vector pLNCX-CREG) studies were performed to support this notion. Western blot showed that CREG knockdown stimulated with STS or VP-16 was identified to increase cleaved caspase-3, a marker for apoptosis. It was also observed that the number of cells undergoing apoptosis remarkably increased in CREG-DW cells by annexin V/PI dual-color flow cytometry and TUNE-L assays. Moreover, p38 and JNK mitogen-activated protein kinases were significantly upregulated in CREG-DW and significantly reduced in CREG-UP VSMCs. More importantly, CREG-DW-induced VSMC apoptosis was blocked by a p38-specific inhibitor (SB203580), or by overexpression of a dominant negative p38 alpha (p38 alpha AGF). The data also showed that inactivation of p38 decreased the amount of phosphorylated JNK, indicated that the p38 fusion proteins are functionally active in regulating JNK activity and induced JNK phosphorylation contributes positively to VSMC apoptosis. In addition, CREG-UP increased expression of the VSMC differentiation markers SM alpha-actin and SM-MHC, and reduced cell-associated fibronectin in cultured VSMCs. These results demonstrate for the first time that CREG plays a key role in modulating VSMC apoptosis by p38/JNK signaling transduction pathway in vitro. Accordingly, CREG might have potential applicational perspective in attenuating the progression of atherosclerotic plaques and restenosis after percutaneous coronary intervention.