供转染的截短型PDGF-α受体高滴度腺病毒的制备及作用分析  

Preparation and analysis of high-titer adenoviruses for transfecting truncated PDGF-α receptor

作　　者：王海燕[1] 于长青[1] 祝之明[1] 晋军[1] 王利娟[1] 胡文辉[1] 

机构地区：[1]第三军医大学附属大坪医院高血压内分泌科重庆市高血压研究所,重庆400042

出　　处：《免疫学杂志》2002年第4期308-310,314,共4页Immunological Journal

基　　金：国家自然科学基金资助项目 (39870 3493972 5 0 13)

摘　　要：目的　制备高滴度供转染的切除功能结构域的血小板衍生生长因子α受体 (truncatedPDGF αR)重组腺病毒 ,观察其在对平滑肌细胞增殖的影响。方法　通过基因重组技术 ,切除PDGF α受体功能结构域 ,将cDNA片段克隆入腺病毒载体 ,重组质粒导入病毒包装细胞 2 93扩增制备高滴度的腺病毒 ,感染大鼠主动脉平滑肌细胞 ,3 H TdR掺入法测定腺病毒对PDGF AA介导的增殖反应影响。结果　重组腺病毒滴度最高为 1.4× 10 5CFU/mL ;截短型PDGF α感染平滑肌细胞 ,PDGF AA介导的3 H TdR掺入减少。结论　制备高滴度供转染的截短型PDGF α受体腺病毒 ,感染平滑肌细胞后 ,PDGF AA介导的增殖反应减弱。Objective To prepare high titer recombinant adenoviruses for transducting truncated platelet derived growth factor α receptor (truncated PDGF α R)and study the role of truncated PDGF α R in vascular smooth muscle cells (VSMC) of rats . Methods Functional and structural domain of PDGF α R were truncated and the cDNA fragment of truncated PDGF α R was inserted into the polyclonal sites of adenoviruses vector which contain cytomegalovirus(CMV) promotor. Recombinant adenoviruses was transfected into 293 packing cell line. The supernatants of virus was used to transfect VSMCs. Immunocytochemical staining and PT PCR were performed to identify truncated PDGF α R. PDGF AA induced proliferation of VSMC was assayed by 3H TdR incorporation. Results The highest titer of the adenoviruses was 1.4× 10 5 CFU/mL. Transducted VSMCs expressed truncated PDGF α R. Truncated PDGF α R can inhibit PDGF AA induced proliferation of VSMC. Conclusions High titer recombinant adenoviruses was successfully prepared. PDGF AA induced the proliferation of VSMC can be inhibited after VSMCs were transfected by adenoviruses containing the truncated PDGF α R.

关 键 词：转染 截短型PDGF-α受体 腺病毒 血小板衍生生长因子 滴度 

分 类 号：R346[医药卫生—基础医学]