益气健脾中药对血管平滑肌细胞凋亡及凋亡相关基因表达的影响

目的：探讨益气健脾中药血清体外诱导血管平滑肌细胞凋亡的作用及其分子机制。方法：采用缺口末端标记（terminal deoxynucleotidyl transferase mediated dUTP nicked labeling,TUNEL)分析、透射电镜、流式细胞仪分析、DNA电泳分析检测细胞凋亡；应用Northern印迹检测bcl-2、c-myc和p53基因表达活性。结果：经中药血清处理的血管平滑肌细胞（vascular smooth muscle cell,VSMC)，可出现典型的凋亡特征，DNA电泳呈梯状图谱；流式细胞仪分析显示G0／G1期细胞阻滞现象，亚二倍体凋亡细胞增多，出现典型的凋亡峰，细胞凋亡率与中药血清浓度具有量效关系。杂交结果提示，30％中药血清可抑制抗凋亡基因bcl-2的表达，上调促凋亡基因c-myc和p3 mRNA水平。结论：益气健脾中药通过影响凋亡相关基因bcl-2、c-myc和p53的表达活性而诱导VSMC凋亡。

Effect of Yiqi Jianpi Drugs on Apoptosis and Relevant Gene Expression in Cultured Vascular Smooth Muscle Cell

OBJECTIVE: To explore the effect of Yiqi Jianpi (YQJP, supplementing Qi and invigorating Spleen) recipe on vascular smooth muscle cells (VSMC) apoptosis and it's mechanisms. METHODS: VSMC apoptosis was detected by terminal deoxynucleotidyl transferase mediated dUTP nicked labeling (TUNEL) analysis, transmission electron microscope (TEM), flow cytometry and DNA agarose electrophoresis. The expression activity of bcl-2, c-myc and p53 genes were determined using Northern blotting. RESULTS: The typical characteristics of apoptotic VSMC was observed following treatment by the YQJP drug serum. The DNA extracted from VSMC revealed a ladder pattern in electrophoresis. Flow cytometric analysis revealed G0/G1 stage cell blocking phenomena, the hypodiploid apoptotic cell increased, displayed typical peak of apoptosis, the rate of cell apoptosis and YQJP serum concentration were in a dose-dependent manner. The result of hybridization showed that 30% of YQJP serum could inhibit apoptotic gene bcl-2 expression, upregulated the level of apoptogenous gene c-myc and p53 mRNA. CONCLUSION: VSMC apoptosis could be induced by YQJP recipe and its mechanism might be through affecting the apoptosis related gene expression activity.