5-烯丙基-7-二氟亚甲基白杨素诱导人肝癌细胞生长抑制及凋亡.doc

5-烯丙基-7-二氟亚甲基白杨素诱导人肝癌细胞生长抑制及凋亡 　【摘要】 目的：研究5-烯丙基-7-二氟亚甲基白杨素(ADFMChR)对人肝癌(SMMC-7721)细胞生长和凋亡的影响。方法：平皿克隆形成法测定细胞锚定依赖性生长能力；吖啶橙/溴化乙啶(AO/EB)染色荧光显微镜观察凋细胞亡形态,计数细胞凋亡；PI染色流式细胞术(FCM)分析细胞周期和凋亡率。结果：平皿克隆形成法显示：ADFMChR抑制人肝癌(SMMC-7721)细胞生长,且呈浓度依赖性。AO/EB染色荧光显微镜观察发现随着ADFMChR浓度的增加SMMC-7721细胞凋亡率依次增加。PI染色FCM结果表明ADFMChR以浓度依赖方式诱导SMMC-7721细胞凋亡，且使细胞周期阻滞于G1期。结论：ADFMChR具有诱导人肝癌SMMC-7721细胞凋亡的作用，其机制可能与细胞周期G1期阻滞相关。 【关键词】 肝肿瘤 白杨素 细胞周期阻滞 凋亡 Abstrct Objective:To investigate the effect of 5-ally-7-gen-difluormethylene-chrysin (ADFMChR) on the growth and apoptosis of human SMMC- 7721 cells cultured in vitro and its mechanism.Methods:SMMC-7721 cells were treated with various concentrations of ADFMChR.Cell proliferation of SMMC-7721 cells was evaluated by clone formation test.The morphology of cell apoptosis of SMMC-7721 cells were detected by AO/EB double fluorescence stain.The apoptosis rate and cell cyle of SMMC-7721 cells were determined by PI staining flow cytometry.Results:Data of colone formation test indicated that ADFMChR could inhibit the growth of SMMC-7721 cells in a concentration-dependent manner.PI staining flow cytometry and AO/EB double fluorescence stain showed that cell cyle was arrested at G1 phase and the apoptosis of SMMC-7721 cells was induced by ADFMChR in a concentration-dependent manner.Conclusions:ADFMChR posseses induction of apoptosis of SMMC-7721 cells in vitro,and its mechanism might be associated with G1 phase cell cycle arrest. Key words Human liver cancer；Chrysin；Cell-cycle arrest；Apoptosis 白杨素（ChR)具有抗氧化［１］，抗病毒［２］，抗高血压［３］，抗糖尿病［４］，抗基因突变［５］和抗肿瘤等作用［６］。但因其肠道吸收甚少和5,7位羟基被迅速糖基化代谢导致生物利用度和体内活性降低,限制 了它的临床应用。我们以往的实验研究证实：5-烯丙基-7-二氟亚甲基白杨素(ADFMChR)具有显著抑制人胃癌细胞和人结肠癌细胞增殖作用。本实验研究ADFMChR抑制SMMC-7721细胞生长和诱导凋亡作用。 1 材料与方法 1.1 受试物和试剂 5-烯丙基-7-二氟亚甲基白杨素(ADFMChR)，由湖南师范大学药学系药物合成化学教研室合成，分子量：259；性状：黄色粉末状；纯度：98％。白杨素购于美国Sigma公司。PRMI－1640培养基为美国Gibco公司产品；新生牛血清购自杭州四季青生物工程材料有限公司；二甲基亚砜（DMSO）为美国Amresco公司产品。 1.2 细胞与细胞培养 人肝癌细胞系(SMMC-7721)购于中国科学院上海细胞生物研究所(中国上海市)。用含10%小牛血清的RPMI-1 640培养基于37 ℃，5%CO2混合气体，95%湿