成肌细胞表达异源心钠素基因的研究  

Ectopic expression of atrial natriuretic factor gene in the primary myoblast

作　　者：朱记法 卢永昕 李锋 田俊 王玉 

机构地区：[1]汉口协和医院心血管内科,武汉市430022

出　　处：《中国分子心脏病学杂志》2004年第5期284-287,共4页Molecular Cardiology of China

摘　　要：目的研究成肌细胞表达异源心钠素(atrialnatriureticfactor,ANF)基因的可行性,为其在心血管疾病中的应用提供依据。方法以逆转录聚合酶链反应(RT-PCR)扩增人的心钠素基因,将其与质粒pcDNA3.0连接,构建质粒pcDNA3.0/ANF。将pcDNA3.0/ANF及对照pcDNA3.0分别转染体外培养的大鼠骨骼肌成肌细胞,筛选以获得稳定表达异源心钠素基因的细胞。收集筛选后的细胞培养基以放射免疫分析法检测心钠素的表达水平,第14d以定量PCR的方法检测成肌细胞内心钠素mRNA的水平。结果pcDNA3.0/ANF组重组心钠素的表达水平为174.19±52.43pg/ml,成肌细胞内mRNA含量(灰度值)为254.62±10.86,与对照组比较均有显著性差异,P<0.01。结论成肌细胞可以表达异源心钠素基因,这一方法有可能应用于心血管疾病的基因治疗。Objective To study the possibility of ectopic expression of atrial natriuretic factor （ANF） gene in myoblast and pave the way for the use of this gene expression way in cardiovascular fields. Methods ANF gene of human was got by RT-PCR. ANF gene and plasmid pcDNADNA3.0 were both digested by enzymes EcoRI and XbaI, the digestion fragments were ligated to construct plasmid pcDNADNA3.0/ANF. The mock pcDNADNA3.0 and pcDNADNA3.0/ANF were transduced into skeletal myoblast, which derived from the hindlimb skeletal muscle of Wisdar newborn rat. After being selected, the concentration of recombinant ANF in cell culture medium was detected by radioimmunologic assay. At the 14th day after selection, ANF mRNA in myoblast was detected by quantitative PCR. Results the recombinant ANF concentration of pcDNADNA3.0/ANF group was 174.19±52.43pg/ml, mRNA level was 254.62±10.86, which both were higher than the control group, p 〈0.01. Conclusions myoblast could stable express ANF gene, and this way would be possibly used in the gene therapy of cardiovascular disease.

关 键 词：成肌细胞 心钠素 基因表达 

分 类 号：R541[医药卫生—心血管疾病]