杨梅素抗肝肿瘤细胞HepG2作用及其机制的研究

In vitro research on function and mechanism of myricetin-mediated apoptosis of human liver cancer cell line HepG2

目的探讨杨梅素对肝癌细胞HepG2的生长抑制和诱导凋亡作用及其作用机制。方法应用甲基噻唑蓝(MTT)比色法,测定杨梅素对HepG2细胞株活性的影响,检测其抑制HepG2细胞株的时间效应和剂量效应;应用形态学观察、HE染色和Annexin V-PI流式检测,测定杨梅素对HepG2细胞的凋亡作用,并进行细胞周期的分析,最后对杨梅素对细胞凋亡相关的蛋白p34cdc-2、Cyclin B1、Bcl-2和PARP的影响进行了研究。结果杨梅素对HepG2细胞具有生长抑制及诱导凋亡作用,且呈时间依赖和剂量依赖性,表现为:细胞G2/M期阻滞,出现明显的凋亡峰;Bliss法测定杨梅素对HepG2细胞的IC50值为(32.18±2.31)μmol/L,30、50μmol/L杨梅素对G2/M期细胞比率分别为(41.32±0.90)%和(80.07±0.90)%,差异有统计学意义(P<0.05);p34cdc-2蛋白的表达量未见变化,而CyclinB1蛋白的表达量明显上升,Bcl-2蛋白的表达量锐减,并诱导半胱氨酸蛋白酶Caspase-3的底物PARP蛋白(113 kD)水解为89 kD的特异性水解产物。结论杨梅素通过激活Caspase家族蛋白实现对肝癌细胞HepG2细胞株的增殖抑制及诱导凋亡作用。

Objective To investigate the growth inhibition and apoptosis-inducing effect of myricetin on liver cancer cell line HepG2. Methods Activation of myricetin on cell line HepG2 was determined by MTT test, the suppression of time and dosage effect on cell line HepG2 were detected; myricetin-induced apoptosis was detected by morphology observation, HE staining and Annexin V-PI flow detection, cell cycle was also analyzed. The influences myricetin on Cell apoptosis related protein such as p34^ok-2, Cyclin B1, Bcl-2 and PARP were studied. Results Growth inhibitory and apoptosis-inducing ef- fect were found in myricetin on cell line HepG2, they were time dependent and dose-dependent. Myricetin had a growth inhibitory and apoptosis-inducing effect on HepG2 cell with IC50 （32.18 ± 2.31） μmol/L by Bliss method. Cell proportion of G2/M by 30, 50 μmol/L myricetin was （41.32±0.90）% and （80.07±0.90）%, the difference was statistically significant （P 〈 0.05）; the expression of p34cdc-2 protein was found no change, the expression of Cyclin Blprotein was rised, the expres- sion of Bcl-2 protein was declined sharply, PARP protein with 113 kD was hydrolyzed into 89 kD. Conclusion Myricetin has an growth inhibitory and apoptosis-inducing effect on human liver cancer cell line HepG2 according to the activation of Caspase protein.