拉马克啉对内皮素-1诱导的大鼠血管平滑肌细胞增殖及蛋白激酶C表达的作用

Effect of levcromakalim on ET-1 induced proliferation and expression of protein kinase C in vascular smooth muscle cells

目的:探讨拉马克啉(levcromakalim)对内皮素-1(ET-1)诱导的大鼠血管平滑肌细胞增殖及蛋白激酶C(protein kinase C,PKC)表达的影响。方法:体外培养Wistar大鼠主动脉血管平滑肌细胞(vascularsmooth muscle cell,VSMCs),用ET-1诱导其增殖,用不同浓度拉马克啉共培养,MTT法评价VSMCs增殖情况,3H-TdR检测DNA合成,流式细胞术检测VSMCs凋亡,逆转录聚合酶链式反应(RT-PCR),Western blot检测PKCαmRNA及蛋白表达。结果:拉马克啉对ET-1所致VSMCs增殖有显著抑制作用,随浓度增加,其MTT活性细胞含量和3H-TdR掺入量都明显减少(P<0.05);拉马克啉呈剂量依赖性地增加G0/G1期VSMCs(P<0.05),促使VSMCs凋亡增多(P<0.05);拉马克啉抑制VSMCs内PKCαmRNA及蛋白表达。结论:拉马克啉抑制ET-1诱导的VSMCs增殖作用,可能的机制是通过下调VSMCs内PKCα表达水平而影响VSMCs增殖和凋亡。

AIM： To observe the effect of levcromakalim （Iev） on endthelin-1 （ ET-1 ） induced proliferation and the expression of protein kinase C （PKC） in cultured rat vascular smooth muscle cells （VSMCs）. METHODS： VSMCs were isolated from rat aorta and were cultured with different concentrations of Lev in vitro after stimulated to proliferation by ET-1 （ 10^-8 mol· L^-1 ）. Vitality of VSMCs was detected by MTT; DNA replication was evaluated by 3H-TdR method. Flow cytometry was used to analyze cell cycle and apoptosis. The expression of PKCα protein and mRNA were determined by Western blot and reverse transcription-polymerase chain reaction （RT-PCR）. RESULTS： Vitality of VSMCs and 3H-TdR intermingle rate were decreased （ P 〈 0.05） in a dose dependent manner which indicated Lev inhibited VSMCs proliferation by inducing ET-1. Levcromakalin increased the ratio of VSMCs apoptosis （ P 〈 0.01 ）. The cell cycle was blocked at G0/G1 phase by Lev in a dose dependent manner （ P 〈 0.05）. Levcromakalim decreased the expression of PKCα protein and mRNA in VSMCs （P 〈 0.05）. CONCLUSION： Levcromakalim inhibits VSMCs proliferation by inducing ET-1 through reducing the expression PKCα.