雷公藤内酯醇对牛晶状体上皮细胞增殖及胞浆内游离Ca^(2+)浓度的影响

Effects of triptolide on proliferation of cultured bovine lens epithelial cells and intracellular concentration of Ca^(2+)

目的:探讨雷公藤内酯醇(Tri)对牛晶状体上皮细胞(lens epithelial cell,LECs)增殖的影响及可能的机制。方法:取体外培养的第4代牛LECs,分5组培养。空白对照组:以DMEM培养;增殖对照组:以DMEM+50μg/L rhEGF培养;Tri高、中、低剂量组分别以含50μg/L rhEGF和40μg/L、20μg/L、10μg/L Tri的DMEM培养。分组培养6h、12h、24h、48h、72h后用MTT比色法及流式细胞仪检测细胞增殖抑制率和增殖细胞核抗原(PCNA)阳性表达率;用荧光指示剂Fura-2/AM测定空白对照组、增殖对照组、Tri中剂量组分组培养48h后胞内游离Ca^(2+)浓度。结果:随着作用时间的延长和浓度的增加,Tri对rhEGF诱导的LECs增殖的抑制率逐渐增大;Tri各剂量组PCNA阳性表达率低于增殖对照组;Tri中剂量组胞内Ca^(2+)浓度高于增殖对照组和空白对照组。结论:Tri对LECs的增殖有明显的抑制作用,该作用可能通过影响胞内Ca^(2+)浓度来完成。

Aim ： To explore the effects and the mechanism of triptolide （Tri） on the proliferation of cultured bovine lens epithelial cells （LECs） in vitro. Methods： The fourth generation of bovine LECs were allocated into 5 groups and cultured with DMEM （normal control group）, DMEM ＋50 μg/L rhEGF （proliferation group）,50 μg/L rhEGF ＋40 μg/L Tri ＋ DMEM （high-dose Tri group）,50μg/L rhEGF ＋ 20 μg/L Tri ＋ DMEM （medium-dose Tri group）, 50 μg/L rhEGF ＋ 10 μg/L Tri ＋ DMEM （ high-dose Tri group）. The inhibition rates of LECs and the expression rates of proliferating cell nuclear antigen （PCNA） were measured by MTT （colorimetric assay） and FCM （flow cytometry）at 6 h, 12 h,24 h,48 h, and 72 h after culture, respectively. Cytosolie Ca^2＋ was determined by the fluorescence determination with Fura-2 at 48 h after culture. Results： The inhibition rates of LECs changed in the time-dose-dependent way. In low-dose, medium-dose and high dose Tri groups, the positive rate of PCNA was lower than that of proliferation group. The intracellular Ca^2＋ in medium-dose Tri group was higher than those in proliferation group and normal control group. Conclusion： Tfi could effectively inhibit the proliferation of LECs in vitro, which may be evoked by changing the concentration of the cytosolic Ca^2＋.