Study On The Mechanisms Underlying The Peroxynitrite-induced Discoidin Domain Receptor-2Expression In Human Brain Vascular Smooth Muscle Cells

Peroxynitrite (ONOO-), a strong biological antioxidant, is highly reactive and unstable. In our preliminary experiments, we demonstrated that, in vitro rat carotid artery ring, ONOO" at concentrations ranged from10-8to10-4mol/L, had a strong vasodilatation effect on the vascular smooth muscle cells.The expression of discoidin domain receptor2(DDR2) in human brain vascular smooth muscle cells (HBVSMCs) subjected in response to ONOO-on human brain vascular smooth muscle cells exhibited a bidirectional manner, which was dependent on concentration of ONOO-. Moreover, research showed that exogenous ONOO "-elicited apoptosis of HBVSMCs apparently involved in activation of DDR2. The precise mechanism, however, by which ONOO-induced the expression of DDR2in HBVSMCs, is still unclear.Objective:The purpose of the present work was major at exploring the possible mechanism underlying the ONOO-induced DDR2expression in HBVSMCs. Methods:In the current study, HBVSMCs were firstly cultured in vitro and pretreated the cells with PD98059, an inhibitor of ERK1/2. Subsequently, the morphological alterations, cell survival rate, expressions of DDR2or matrix metalloproteinases9(MMP9) of HBVSMCs in response to ONOO-were determined by acridine orange staining, MTT assay, Western blot, and Real-time PCR, respectively. Results:The present investigation demonstrated that:1) HBVSMCs pretreated with PD98059still appeared adherent growth and had no obvious morphological changes. Meanwhile, the cell survival rate had no marked statistical difference before and after the pre-intervention;2) At the concentrations of0～10μmol/L, ONOO-significantly increased the DDR2protein expression, but, with the increase in concentration of ONOO-, DDR2protein expression was gradually decreased; and3) When10pμmol/L ONOO-applied to HBVSMCs pretreated with PD98059, the expressions of DDR2at both protein and mRNA level, and MMP9protein level were significantly lowered compared with those in the groups without pretreatment.Conclusion:PD98059suppresses the DDR2expression in HBVSMCs subjected to ONOO-, thereby the mechanism responsible for ONOO--induced DDR2expression is related to the ERK1/2signal transduction pathways. The exact mechanism on this issue needs to be elucidated further.