肝细胞生长因子受体在血管平滑肌细胞增殖凋亡中的表达

目的研究球囊扩张术后血管平滑肌细胞(VSMCs)中肝细胞生长因子(HGF)及其受体(c-Met)mRNA表达的变化,明确HGF受体与VSMCs增殖凋亡分子机制的相关性。方法用逆转录聚合酶链式反应检测36只兔腹主动脉球囊损伤前后不同时间点VSMCs中c-Met的mRNA表达水平;体外实验将选择性c-Met蛋白抑制剂NK-4作用于免VSMCs,运用MTF分别于0、24、48及72h检测细胞增殖状态;流式细胞仪观察NK-4对细胞凋亡的影响,进一步采用Western blot检测药物作用前后细胞周期蛋白(Cyclin D1)、凋亡蛋白(Bcl-2)的表达。结果球囊损伤后VSMCs c-Met mRNA的表达水平明显高于正常VSMCs(P＜0．01),平均为对照组2．42倍;对照组S及G_2/M期DNA百分含量与处理组比值分别为1．31,1．62(P＜0．01),NK-4呈时间、剂量依赖性方式抑制VSMCs增殖,促进其凋亡。72h时对照组与NK-4(1．0mg/L)处理组Cyclin D1、Bcl-2表达水平比值分别为2．37和3．81(P＜0．01),故两者表达水平随NK-4作用时间延长而下降。结论c-Met在球囊扩张术后血管平滑肌细胞中高表达可能在VSMCs过度增殖、凋亡受阻中起重要作用。选择性c-Met蛋白抑制剂NK-4可能通过Cyclin D1,Bcl-2影响VSMCs的增殖与凋亡,提示c-Met抑制剂可作为预防血管成形术后再狭窄新的候选药物。

Expression of HGF/c-Met during proliferation and apoptosis of VSMC in rabbits

Objective To investigate the hepatocyte growth factor(HGF)receptor c-Met mRNA level in vascu- lar smooth muscle cells(VSMCs)after balloon angioplasty in rabbits and reveal the potential effect of c-Met on proliferation and apoptosis of VSMCs.Methods c-Met mRNA in VSMCs and adjacent normal tissue from ab- dominal aorta of 36 tabbits were measured by RT-PCR. Cultured VSMCs of rabbits in vitro were treated with NK-4(a selective c-Met inhibitor)at different time.MTT assay and flow cytometry were used to measure the proliferation and apoptosis.The expression of Cyclin D1 and Bcl-2 were measured by Western blot.Results C-Met mRNA level was increased by 2.42 times in average in VSMCs compared with that in adjacent normal vessel wall(P＜0.01).The ratios of S and G2/M percentages of control group to those of NK-4-treated group were 1.31 and 1.62(P＜0.01)respectively and the ratios of Cyclin D1 and Bcl-2 protein expression were 2.37 and 3.81(P＜0.01),respectively.NK-4 inhibited the cell proliferation and induced apoptosis in a dose-and time-dependent manner and caused significant downregulation of Cychn D1 and Bcl-2.Conclusions The results show that overexpression of c-Met may play a critical role in the proliferation of VSMCs after bal- loon angioplasty.Selective c-Met inhibitor NK-4 may inhibit the proliferation and induce apoptosis of VSMCs through decreasing expression of Cyclin D1 and Bcl-2,indicating that C-Met inhibitor may be a new candidate for prevention of restenosis after angioplasty.