核心蛋白聚糖在HepG_2细胞中的表达及作用  

Expression and effect of recombinant secreting decorin in HepG_2 hepatoma cell

作　　者：张玉成[1] 王雅丽[1] 杜珍武[1] 高申 吕俊峰[1] 张桂珍[1] 

机构地区：[1]吉林大学中日联谊医院,吉林长春130033

出　　处：《山东医药》2008年第7期30-32,共3页Shandong Medical Journal

基　　金：吉林省发展与改革委员会计划资助项目(2006-1550)

摘　　要：目的构建分泌型核心蛋白聚糖(DCN)真核表达载体,并观察其在肝癌细胞HepG2中的表达和抗肿瘤作用。方法应用PCR技术扩增DCN全长基因cDNA片段,与pcDNA3.1载体连接,并转化到大肠杆菌中扩增获得重组载体,应用双酶切、PCR以及测序鉴定此重组载体,脂质体介导转染HepG2,经G418筛选建立稳定转染细胞株,采用RT-PCR、免疫组化检测其表达。MTT检测细胞增殖活力,流式细胞仪分析细胞周期。结果RT-PCR可见转染组细胞mRNA表达明显增多,免疫组化可见转染组细胞DCN蛋白表达明显增高。细胞生长曲线显示转染组细胞生长缓慢,G1期细胞显著增多。结论本方法可成功建立稳定转染DCN的HepG2细胞株,并证实DCN能够抑制HepG2细胞株的生长。Objective To construct a recombinant eukaryotic expressing vector pcDNA3.1-DCN and make it express in HepG2 hepatoma cell so as to investigate the effect of resisting tumor. Methods Using PCR,the full length of decorin cDNA was amplified. The recombinant plasmid-decorin was constructed and transformed into the bacteria. Recombinant eukaryotic expressing vector pcDNA3.1-DCN was transfected into HepG2 hepatoma cell mediated by liposome, established stable transfected cell line using G418 screening and detect recombinant secreting decorin expression using RT-PCR and immunochemistry methods. The cell activity was detected by MTT method,cell cycle was analyzed by flow cytometry . Results DCN mRNA expression was significantly increasing in transfected group using RT-PCR method. DCN protein expression also was high in transfected group using immunochemistry method. Cell growth curve displayed cell growth was slow in transfected group. The percentage of G1 phase in transfected cells was higher than in control calls. Conclusion We successfully construct the stable transfected HepG2 cell line with DCN and verify DCN can inhibit HepG2 cell line growth.

关 键 词：核心蛋白聚糖 聚合酶链反应 转染 HEPG2 

分 类 号：R735.7[医药卫生—肿瘤]