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目的观察穿山龙提取物(extract from Dioscoreae nipponicae Rhizoma,DNR)联合哈尔满碱(harmane)对人肝癌细胞(HepG2)增殖抑制作用及诱导凋亡的机理。方法采用MTT法检测药物对细胞的生长抑制作用,应用等效曲线-相互指数法(isobole-interaction index method)评价两药的相互作用,倒置显微镜观察细胞形态学变化,流式细胞仪(FCM)检测细胞凋亡率,Western印迹法检测凋亡蛋白procaspase-3的表达。结果穿山龙提取物(DNR)与哈尔满碱质量浓度比为2∶1时,I(相互指数)=0.768<1,联合用药具有协同作用;质量浓度比为1∶2时,I=1.041≈1.0,联合用药具有相加作用。FCM法检测联合用药48、72 h细胞凋亡率与单独用药相比分别增加了10.44%、50.06%,差异具有统计学意义(P<0.05)。Western印迹法检测联合用药诱导HepG2细胞凋亡的机制之一可能与procaspase-3蛋白的表达有关。结论 DNR联合哈尔满碱在体外有明显的协同抗肿瘤的作用,其可能的机制是影响procaspase-3蛋白的表达。
OBJECTIVE: To observe the inhibitory effect of extract from Dioscoreae nipponicae Rhizoma (DNR) combined with harmane on the proliferation of HepG2 cells and the mechanism of apoptosis. Methods MTT assay was used to detect the effect of drugs on cell growth inhibition. The isobole-interaction index method was used to evaluate the interaction between the two drugs. The morphology of the cells was observed by inverted microscope. Flow cytometry (FCM) The apoptosis rate was detected by Western blotting to detect the expression of procaspase-3. Results I (mutual index) = 0.768 <1, the synergistic effect was found when the ratio of DNR to H 2 PO 2 was 2:1: I = 1.041≈ 1.0, combined with additive effect. The FCM assay showed that the apoptotic rates of 48 and 72 h groups were increased by 10.44% and 50.06%, respectively, compared with that of the single drug alone. The difference was statistically significant (P <0.05). One of the mechanisms of Western blotting in detecting the apoptosis of HepG2 cells may be related to the expression of procaspase-3 protein. Conclusions DNR combined with Halhamine has a synergistic antitumor effect in vitro. The possible mechanism is that the expression of procaspase-3 protein is affected by DNR.