血小板源性生长因子对血管平滑肌细胞G_0S_2mRNA表达的影响

Effects of PDGF on G_0S_2 mRNA Expression in Vascular Smooth Muscle Cells

目的:研究血小板源性生长因子(PDGF-BB)对血管平滑肌细胞(VSMC)G0S2基因mRNA表达的调控,并初步探讨其机制。方法:体外培养大鼠胸主动脉平滑肌细胞,逆转录实时定量聚合酶联反应(RT-QPCR)法测定PDGF-BBJVSMC中G0S2 mRNA表达影响的时效关系和量效关系;运用不同细胞信号通路阻断剂处理,研究PDGF-BB影响G0S2 mRNA表达的信号途径;转染含有G0S2基因启动子的荧光素酶报告基因质粒pGL3-G0S2-Promotor,检测PDGF-BB对G0S2基因启动子活性的影响。结果:PDGF-BB能够增加平滑肌细胞G0S2 mRNA表达,PDGF-BB诱导VSMC表达G0S2mRNA在12h后达峰值[20ng/ml增加(2.83±0.81)倍;10ng/ml增加(2.99±0.12)倍,P<0.05],随后降低。p38MAPK和PI3K/AKT信号通路阻断剂可显著抑制PDGF-BB诱导的G0S2mRNA表达(P<0.05),并且PI3K/AKT信号通路阻断剂可以明显降低G0S2 mRNA的基础表达水平。荧光素酶活性分析显示,PDGF-BB可增强G0S2基因启动子活性(P<0.05)。结论:PDGF-BB上调VSMC中G0S2基因mRNA的表达,并且可以增强G0S2基因启动子活性,p38MAPK和PI3K/AKT通路可能介导了这种作用。

Objective.To investigate the effect and mechanisms of platelet derived growth factor （PDGFBB） on G0S2mRNA expression in vascular smooth muscle cells （VMSCs）. Methods：Rat VSMCs were isolated and cultured in Dulbecco＇s Modified Eagle Medium supplied with 10% fetal bovine serum. VSMCs were treated with PDGF-BB for different time （2, 6, 12, 24 h）. Total RNA was extracted using TRIzol reagent and reversely transcribed using M-MLV reverse transcriptase. The RT-PCR technique was used to detect the G0S2 gene expression level. The activities of MAPK, ERK, and PI3K/Akt were blocked by SB203580, PD98059, and LY294002 respectively. The luciferase reporter plasmid （pGL3-G0S2-Promoter） was transfected into VSMCs by electro-poration. Results. PDGF-BB can increase the mRNA expression of G0S2 gene in VSMC. G0S2 mRNA expression was induced by PDGF-BB with the gene elevation within 6 h and peaking by 12 h. p38MAPK, and PI3K/AKT signal pathway inhibition can partly inhibit PDGF-BB induced G0S2 mRNA expression. Furthermore, LY294002 can directly inhibit G0S2 mRNA content in VSMCs. Finally, PDGF can increase the activity ofG0S2 gene promoter （P〈0.05）. Conelusion：PDGF-BB can increase the mRNA expression of G0S2 gene and activity of G0S2 gene promoter in VSMCs. p38MAPK and PI3K/AKT signaling pathways mediate PDGF-BB induced G0S2 mRNA expression in VSMCs.