龙葵碱诱导HepG_2细胞凋亡的观察

Study of solanine on apoptosis in HepG_2 cell

观察龙葵碱对3种消化系统肿瘤细胞株人肝癌细胞HepG2、人胃癌细胞SGC-7901、人大肠癌细胞Ls-174的细胞毒作用.并从细胞凋亡角度揭示龙葵碱对敏感细胞株的作用机制.采用MTT法观察龙葵碱对3种肿瘤细胞株的细胞毒作用;采用AO/EB双染,激光共聚焦扫描显微术(confocal)观察龙葵碱对HepG2肿瘤细胞形态的影响;PI单染,流式细胞仪测定龙葵碱诱导HepG2细胞凋亡的凋亡率及对细胞周期的影响.龙葵碱作用于HepG2、SGC-7901、LS-174的IC50分别为14.47μg/mL、(50μg/mL、(50μg/mL;在形态学观察实验中,阴性对照组细胞形态正常,0.003 2μg/mL、0.016μg/mL龙葵碱使细胞外周呈微弱皱缩状改变,0.08、0.4、2μg/mL龙葵碱使HepG2细胞出现大量碎片及凋亡小体等典型的细胞凋亡形态.凋亡率测定结果表明5个剂量龙葵碱诱导HepG2细胞凋亡率分别为6.0%、14.4%、17.3%、18.9%、32.2%,0.08μg/mL喜树碱诱导HepG2细胞的凋亡率为21.9%.细胞周期观察发现龙葵碱各组G2/M期均消失,S期明显升高.龙葵碱对HepG2人肝癌细胞株比较敏感,能够诱导HepG2细胞凋亡,并将HepG2细胞阻止在S期,影响肿瘤细胞DNA合成.

To study the cytotoxic effect of solanine on human hepatocarcinoma HepG2 cells, human gastrocarcinoma SGC -7901 ceils, and human large intestine cancer Ls -174 ceils, and observe the effect mechanism on apoptosis of sensitive cancer cell. Cytotoxicity is evaluated by MTT assay; HepG2 ceils are stained with AO/EB, and laser confocal scanning microscopy （LCSM） is used to observe the effect of solanine on the morphology of HepG2 ceils ; the cells are stained with PI, and flow cytometry is used to observe the rate of apoptosis and cell Cycle in HepG2 ceils induced by solanine. The average IC50 value is respectively 14.47 μg/ mL, 〉 50 μg/mL, 〉 50 μg/mL; It is found that HepG2 cells in groups treated with 0.003 2 μg/mL and 0.016 μg/mL solanine are wrinkled on the periphery, and that large numbers of ceil fragments and apoptotic bodies, which are a typical sign for apoptosis, has appeared in groups treated with 0.08, 0.4 and 2 μg/mL solanine, but the cell morphology in control group is normal. In the experiment to determine the rate of apoptosis, rates of apoptosis in HepG2 cells induced by 5 different dosages of solanine are found to be 6.0%, 14. 4%,17.3%, 18.9% and 32.2%, respectively, while that of apoptosis induced by 0.08 μg/mL camptothecin is 21.9%. G2/M period disappeare and S period increase in solanine group. Therefore, solanine is sensitive to HepG2 cell. solanine induces the apoptosis in HepG2 cell and block HepG2 in S phase to stop the DNA synthesis.