梅花鹿角脱盘多肽的分离纯化及对乳腺癌细胞端粒酶活性的影响  

Isolation and purification of polypeptide from Sika Antlerbase and its effect on telomerase activities in breast cancer cells

作　　者：胡薇[1] 田玉华[1] 孟星宇[1] 李沐[1] 李婷[1] 李子彬[1] 吴炎[1] 

机构地区：[1]吉林农业大学生命科学院,吉林长春130118

出　　处：《上海中医药杂志》2014年第1期85-89,共5页Shanghai Journal of Traditional Chinese Medicine

基　　金：吉林省产业技术研究与开发项目(JF2012C002-1);吉林省长春市科技局优质梅花鹿产业化关键技术与开发重大科技专项(2008214)

摘　　要：目的分离纯化梅花鹿鹿角脱盘多肽;以小鼠乳腺癌细胞系MA782为体外研究对象,初步研究鹿角脱盘多肽对肿瘤细胞端粒酶活性的影响。方法采用酸提醇沉法提取鹿角脱盘多肽,再经Sephadex G-25凝胶过滤层析除盐,BCA法测定的多肽蛋白质含量,TRAP-银染法分析MA782细胞端粒酶活性的变化,再利用相对荧光定量PCR技术检测MA782细胞端粒酶催化亚基TERT基因的表达水平。结果分离纯化出鹿角脱盘多肽粗提物,其蛋白质含量为91.800%;TRAP-银染法检测MA782细胞端粒酶的活性发现,与对照组细胞比较,多肽使细胞端粒酶活性有所下降,典型的重复序列条带数依次减少,多肽处理24 h、48 h、72 h、96 h、120 h后,MA782细胞中相对端粒酶活力分别为91.4、78.3、66.5、47.8、29.2、20.5,呈时间依赖性;相对荧光定量PCR法分析结果表明,与对照组比较,TERT基因的表达水平降低。结论鹿角脱盘多肽对乳腺癌细胞端粒酶活性有一定的抑制作用,可为深入研究鹿角脱盘多肽抗肿瘤活性提供依据。Objective The polypeptide from sika antler plate was purified, and the mice breast cancer cell line MA782 was used as research objects in vitro to study the effect of polypeptide on telomerase activity. Methods Sika antler polypeptide was extracted by the method of acid extraction and alcohol precipitation, rotary evaporation in vaeeum and freeze-drying. Salt was eliminated by Sephadex G-25 Gel filtering chromatography. The protein concentration of total peptide was detected by the method of BCA. The changes of telomerase activity were analyzed by TRAP argentation. Relative quantitative fluorescence PCR method was used to determinate the expression level of telomerase catalytic subunit TERT. Results Crude extraction of the antler plate polypeptide was got by separation and purification, and its protein＇ s concentration was 91. 800%. The telomerase activity was determined by the method of TRAP- argentation. Compared with control group, the telomerase activity of the group was reduced markedly. The straps of typical repeated sequence was decreased gradually. The relative telomerase activity in MA782 cell treated with polypeptide after 24 h, 48 h, 72 h, 96 h and 120 h was 91.4, 78.3, 66.5,47.8, 29.2, and 20.5 respectively in a time-dependence manner. The relative quantitative fluorescence PCR showed that the level of TERT gene expression was reduced obviously. Conclusion Antler plate polypeptidc has inhibitory effect on telomerase activity of breast cancer cells. All results have provided theoretical foundation for studying the mechanism of the sika antler plate antitumor activities.

关 键 词：鹿角脱盘 多肽 分离纯化 端粒酶活性 

分 类 号：R285.5[医药卫生—中药学]