超声靶向微泡破裂联合聚乙烯亚胺介导大鼠心肌母细胞Bax基因沉默  

Ultrasound-targeted microbubble destruction combined with polyethylenimine mediated Bax gene silence in H9c2 cells

作　　者：金佳美[1] 张宇辉[1] 朱烨[1] 张良平[2] 宗明[3] 陈明[1] 

机构地区：[1]同济大学附属东方医院心脏医学部,上海200120 [2]同济大学附属东方医院转化平台,上海200120 [3]同济大学附属东方医院检验科,上海200120

出　　处：《中华超声影像学杂志》2013年第5期438-442,共5页Chinese Journal of Ultrasonography

基　　金：上海市卫生局青年基金项目（2010Y012）;上海市浦东新区卫生系统优秀青年医学人才项目（PW Rq201004）;上海市浦东新区心血管疾病学科群（PWZxkq2010-01）

摘　　要：目的探讨超声靶向微泡破裂（ultrasound—targeted microbubble destruction，UTMD）联合聚乙烯亚胺（polyethyleneimine，PEI）的方法转染大鼠心肌母细胞（H9c2细胞）以达到沉默Bax基因的效果。方法构建靶向Bax基因的shRNA表达质粒，将其与微泡及PEI共同处理后加入H9c2细胞并予以超声辐照，实验分组为：①单纯质粒组；②Lipofectamine2000＋质粒组；③UTMD＋质粒组；④PEI＋UTMD＋质粒组；⑤PEI＋SonoVue＋质粒组（无超声辐照）。采用荧光显微镜和FCM法评估基因转染效率，应用RT—PCR检测BaxmRNA水平的变化，Western Blot检测蛋白表达情况。结果超声辐照对所构建的BaxshRNA表达质粒无破坏。H9c2细胞采用UTMD＋PEI处理后，转染率显著高于其他各组（P〈0．05）。PTPCR及Western Blot检测结果显示，PEI＋UTMD＋质粒组BaxmRNA及Bax蛋白表达抑制率分别为（71．41±4．91）％及（64．09±2．38）％，显著高于其余各组（P〈0．05）。结论超声靶向微泡破裂联合聚乙烯亚胺的方法能有效地沉默H9c2细胞Bax基因的表达。Objective To silence Bax gene in H9c2 cells by ultrasound-targeted microbubble destruetion（UTMD） combined with polyethylenimine. Methods The shRNA expression plasmid targeting Bax gene was co treated with microbubbles and polyethylenimine. Then the mixture added to H9c2 cells were irradiated hy ultrasound. Grouping： （1）plasmid group;（2）Lipofectamine2000 ＋ plasmid group; （3） UTMD ＋ plasmid group; （4）PEI ＋ UTMD ＋ plasmid group; @ PEI ＋ SonoVue ＋ plasmid group（without ultrasound irradiation）. The transfection efficiency was assessed by fluorescence microscope and FCM. The expression of Bax mRNA and protein were detected by RT-PCR and Western Blot. Results Bax shRNA wash＇ t destroyed by ultrasound exposure. The transfection efficiency of PEI ＋ UTMD ＋ plasmid group was higher than any other groups obviously （ P 〈0.05）. The result of RT-PCR and Western Blot showed inhibition efficiency of Bax mRNA and protein expression in PEI ＋ UTMD ＋ plasmid group were （69.41 ± 4.91 ） and （64. 09 ± 2. 38） % separately, which were higher than any other groups significantly （ P〈0.05）. Conclusions UTMD combined with polyethylenimine can inhibit the expression of Bax gene in H9c2 cells effectively.

关 键 词：超声检查 微气泡 心肌母细胞 RNA干扰 基因沉默 

分 类 号：R54[医药卫生—心血管疾病]