雷帕霉素对大鼠主动脉瓣膜间质细胞增殖的影响

目的利用雷帕霉素靶向抑制mTOR并探讨其对大鼠心瓣膜间质细胞增殖的影响。方法体外分离大鼠瓣膜间质细胞后 培养并鉴定；细胞随机分为两组：雷帕霉素组与对照组；MTT法检测并绘制大鼠瓣膜间质细胞用药前后的生长曲线；流式细胞 仪检测雷帕霉素对细胞周期的影响；RT-PCR 检测细胞中S6、P70S6K的mRNA表达水平；Western blotting 检测细胞中S6、 P70S6K、P-S6、P-P70S6K蛋白的表达水平。结果体外分离获得的大鼠瓣膜间质细胞分离培养成功；对照组细胞活性显著高于 加药组细胞（P<0.05），加药组细胞生长变缓。流式细胞术检测发现加药组和对照组细胞的各周期占比均没有明显差别；加药组 细胞的S6蛋白与P70S6K蛋白的磷酸化水平降低（P<0.05）。结论雷帕霉素能抑制瓣膜间质细胞的增殖其原因，可能不是通过 改变细胞周期而是通过抑制mTOR后下调其靶蛋白S6和P70S6K的磷酸化水平引起的。

Effect of rapamycin on proliferation of rat heart valve interstitial cells in vitro

Objective To investigate the effect of rapamycin on the proliferation of rat valvular interstitial cells in primary culture. Methods The interstitial cells isolated from rat aortic valves were cultured and treated with rapamycin, and the cell growth and cell cycle changes were analyzed using MTT assay and flow cytometry, respectively. RT-PCR was used to detect mRNA expression levels of S6 and P70S6K in cells, and the protein expressions level of S6, P70S6K, P-S6, and P-P70S6K were detected using Western blotting. Results Rat aortic valvular interstitial cells was isolated successfully. The rapamycin-treated cells showed a suppressed proliferative activity (P<0.05), but the cell cycle distribution remained unaffected. Rapamycin treatment resulted in significantly decreased S6 and P70S6K protein phosphorylation level in the cells (P<0.05). Conclusion The mechanism by which rapamycin inhibits the proliferation of valvular interstitial cells probably involves suppression of mTOR to lower S6 and P70S6K phosphorylation level but not direct regulation of the cell cycle.