槲芪散及槲寄生提取物对肝癌细胞内游离钙离子浓度的影响

摘要

目的 探讨方剂槲芪散及其君药槲寄生提取物多糖、总碱对人肝癌细胞HepG2生长的抑制作用及其对细胞内游离钙离子浓度的影响.方法 采用MTT法测定槲芪散对细胞增生的抑制作用;利用流式细胞仪分析细胞周期、凋亡;利用激光共聚焦显微镜测定细胞内游离钙离子浓度.结果 槲芪散及槲寄生提取物有较好的抑制肿瘤细胞增生的作用,高剂量组除了槲芪散和槲寄生总碱48 h时间点外,OA值均较对照组明显降低(P<0.001),呈现出时间-剂量依赖性;经药物作用48 h后,槲寄生多糖及总碱组G1期细胞比例增加,S期细胞和G2期细胞比例降低,槲芪散组G1期细胞比例无明显变化,S期比例降低,G2期比例增加,3者均使细胞凋亡增加;加入不同浓度的槲芪散、槲寄生多糖及总碱,低浓度时荧光增强不明显,而高浓度均导致细胞内荧光强度瞬间增强,升高到一定水平后,基本维持在一高水平,除槲寄生总碱组有略微下降外,其余2组未见明显下降趋势.结论 槲芪散、槲寄生多糖及总碱均能抑制肿瘤细胞增生,促进细胞凋亡,且这3种药物引起胞内钙超载可能是诱导HepG2凋亡的机制之一.%Objective To probe the inhibitory effecl of human hepalocarcinoma cell line HepG2 and the influence of inlracellular free Ca2+ levels induced by Chinese herbs, Huqi San(HQS) and ils principal drug Ramulus Visci Exlracl. Methods The proliferation of HepG2 were observed by MTT colormelric assay. The cell cycle and the rale of apoplosis of HepG2 were examined by using flow cylomelry. The fluorescenl inlensily of inlracellular free Ca2+ was observed by laser scanning confocal microscopy. Results Both HQS and Ramulus Visci exlracl could inhibil the proliferation of HepG2 in a lime and dose-dependenl manner, the numeric value of OD in high dose group decreased markedly compared lo the conlrol group ( P < 0. 001 ) , expecl the high dose group of HQS and lolal Alkaloid of Viscum al the lime of 48 h. Afler Irealmenl wilh the drugs for 72 h, the cell population in the group of Ramulus Visci Polysaccharides and lolal Alkaloid of Viscum increased in Gl phase and G2 phase and decreased il in S phase; Comparalively, in the group of HQS, the proporlion of the cells in S phase decreased, while cells al G2 phase increased, bul there was no a significant change in the Gl phase. In addilion, all of the three drugs could induce the apoplosis of HepG2. The resulls showed wilh laser scanning confocal microscopy lhal as differenl dose of the three drugs were pul inlo the Pelri dishes respectively, and we found lhat the fluorescenl inlensily did nol change obviously when the cells were Irealed wilh the low dose drugs. Bul as we pul inlo three high dose drugs inlo the Pelri, dishes the fluorescenl inlensily increased inslanlly and mainlained al a high level for periods of lime wilhoul fluclualing nearly excepl a lillle decrease wilh lolal alkaloid of Viscum. ConclusionHuqi San, Ramulus Visci Polysaccharides and lolal alkaloid of Viscum all could inhibil the proliferation of cancer cells and promoled their apoplosis. The calcium overload maybe a possible mechanism of the apoplosis of HepG2 induced by Huqi San and Ramulus Visci Exlracl.