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医学免费论文:天花粉蛋白诱导宫颈癌HeLa细胞凋亡及APC基因去甲基化
【摘要】 目的:研究天花粉蛋白(TCS)抗宫颈癌作用机制。方法:MTT还原法检测TCS对宫颈癌HeLa细胞的抑制作用，流式细胞仪检测TCS对HeLa细胞周期及凋亡率的影响，MSP检测TCS作用前后APC基因启动子甲基化状态。结果:TCS对HeLa细胞的生长具有明显的抑制作用，呈剂量依赖性;TCS可以将HeLa细胞阻滞于S期;TCS诱导HeLa细胞发生凋亡,有剂量依赖性;80 mg/L TCS处理HeLa细胞48 h后， APC基因启动子出现去甲基化。结论:TCS对HeLa细胞有明显抑制作用，抑制机理与将HeLa细胞阻滞于S期、诱导细胞凋亡及APC基因启动子出现去甲基化有关，TCS有良好的抗宫颈癌临床应用前景。
【关键词】 天花粉蛋白;HeLa细胞;细胞凋亡;APC基因
Effects of Trichosanthin on Inducing Apoptosis of Hela Cells and Methylation of APC Gene in Cervical CancerCHEN Yong,ZHENG Rong,LIU Yong-zhen,CHEN Dong-li,CHEN Shuang-yun^ (Department of Traditional Chinese Medicine,Hubei College of Traditional Chinese Medicine,Wuhan,Hubei 430061;Department of Gynecology,Taihe Hospital,Yunyang Medical College,Shiyan,Hubei 442000,China)Abstract:Objective To investigate the mechanism of trichosanthin(TCS) against cervical carcinoma.Methods The inhibitory effect of TCS on Hela cells proliferation was detected by MTT assay.The cell cycle and apoptosis rate of Hela cells were determined by flow cytometry.The methylation status of APC gene was determined with MSP before and after treatment respectively.Results The growth of Hela cells was significantly inhibited with TCS and the inhibitory effect was dose-dependent;the differentiation of Hela cells was blocked at S-phase of the cell cycle;the apoptosis of Hela cells was induced with TCS in a dose-dependent manner;The demethylation of APC gene promoter was observed after treating with TCS at 80 mg/L for 48 h.Conclusion Trichosanthin could inhibit the proliferation of Hela cells,which may be related with the cell cycle block,apoptosis induction and demethylation of APC gene promoter.TCS may have a wide clinical respect in cervical cancer treatment医.学全.在.线网站www.med126.com.
Key words:Trichosanthin;HeLa cells;Apoptosis;APC gene
天花粉蛋白(Trichosanthin，TCS)为中药天花粉的有效成分, 具有多种药理学活性，已应用于临床。近年来通过对天花粉蛋白结构和功能的深入研究,发现其具有抗肿瘤的作用。 TCS可用于临床抗绒癌及葡萄胎的治疗,并对多种肿瘤细胞(如胃癌、肝癌、结直肠癌、黑素瘤和白血病细胞等)有显著的杀伤作用,是一种有前途的抗肿瘤药物和免疫毒素弹头[1]。本研究应用TCS作用于宫颈癌HeLa细胞系，观察其对细胞增殖及凋亡的影响，并研究其对HeLa细胞腺瘤性结肠息肉病(adenomatous polyposis coli,APC)基因甲基化状态的影响，探讨TCS抗宫颈癌作用及其机制。
1 材料和方法
1.1 药品和试剂
天花粉蛋白注射液(1.2 g/L)(上海金山制药有限公司)，新生牛血清(四季青)，培养基DMEM(GBICO公司)，细胞毒性检测试剂盒及PI染料试剂盒(凯基生物科技发展有限公司)，组织基因组DNA提取试剂盒及DNA Ladder(北京天根生化科技有限公司)，DNA甲基化试剂盒(北京天漠科技开发有限公司)，Taq DNA聚合酶(Promega公司)，dNTP Mix(Amresco公司)，引物合成(上海生工生物工程技术服务有限公司)。
