桃儿七根醇提物对人乳腺癌MCF-7细胞增殖和凋亡的影响

Effects of ethanol extracts from Podophyllum emodi var.chinense on proliferation and apoptosis of breast carcinoma cell line MCF-7

目的:研究桃儿七根醇提物对人乳腺癌MCF-7细胞增殖和凋亡的影响,探讨其抗癌作用的机制。方法:应用MTT法检测桃儿七根醇提物对人乳腺癌MCF-7细胞的生长抑制作用,倒置显徽镜、HE染色及电镜观察细胞形态学改变,流式细胞仪(FCM)分析细胞周期及凋亡率,免疫细胞化学法(ICC)检测细胞增殖、凋亡相关调控蛋白的表达。结果:桃儿七根醇提物可明显抑制MCF-7细胞的增殖,1,2,3 mg．L^(-1)桃儿七根醇提物作用72h后,细胞生长抑制率分别为43．0%,54．9%,78．5%,抑制作用呈时间及浓度依赖性。形态学观察到用药后凋亡细胞典型的形态学特征,伴有多核细胞形成。FCM分析发现桃儿七根醇提物阻断MCF-7细胞生长于细胞周期的G_2/M期,凋亡诱导作用呈时间及浓度依赖性。免疫细胞化学结果显示,2mg·L^(-1)桃儿七根醇提物作用48h后,PCNA,Bcl-2蛋白表达较对照组显著下降;而p21^(WAFI-CIP1)和c-Myc蛋白显著增加(P＜0．01)。结论:桃儿七根醇提物可能通过增加p21^(WAFI-CIP1)蛋白表达及降低PCNA蛋白表达而抑制人乳腺癌MCF-7细胞的增殖,并可能下调Bcl-2蛋白表达和上调c-Myc蛋白表达来促进MCF-7细胞凋亡。

Objective： To investigate the effects and mechanism of ethanol extracts from the roots of Podophyllum emodi var. chinense on proliferation and apoptosis of breast carcinoma cell line MCF-7. Methods： Inhibition of the MCF-7 proliferation by the ethanol extracts from P. emodi var. chinense roots was evaluated using a MTT assay. The morphodifferentiation of the MCF-7 was studied by reversal micro- scope, electron microscope and HE staining. The cellular apoptosis rate and cell cycle phase distribution of the MCF-7 were measured by a flow cytometric assay （FCM）. The expressions of proteins modulating the proliferation and apoptosis were analyzed by an immunocytochemical assay （ICC）. Results： The ethanol extracts suppressed the MCF-7 proliferation in a time-dependent and dose-dependent manner, showing the inhibition rate of 43.0% （1mg·L^-1）, 54.9% （2mg·L^-1） and78.5% （3mg·L^-1） after the culture of the MCF-72 together with the extracts for 72 hours. The morphological study found that the multinucleate giant cells existed in the apoptotic cells after the culture of the MCF-72 with 2 mg·L^-1 of the ethanol extracts. The FCM assay showed that the ethanol extracts induced a G2/M cell cycle arrest in a time- and dose-dependent manner. The ICC assay disclosed that the protein expressions of PCNA, Bcl- 2 significantly decreased （P 〈0.01 ）, and p21^WAF1/CIPI and c-Myc significantly increased （P 〈0.01 ） after the culture of the MCF-7 with the ethanol extracts 2 mg.L^-1 for 48 hours. Conclusion： The ethanol extracts inhibited the MCF-7 proliferation by up-regulation of p21^WAF1/GIP1 level and down-regulation of PCNA level. The apoptosis was produced by down-regulation of Bcl-2 expression and up-regulation of c-Myc proteins expression.