Ultrasound-targeted microbubble destruction (UTMD) assisted delivery of shRNA against PHD2 into H9C2 cells.
Ultrasound-targeted microbubble destruction (UTMD) assisted delivery of shRNA against PHD2 into H9C2 cells.
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超声靶向微泡破坏 (UTMD) 辅助将针对 PHD2 的 shRNA 递送至 H9C2 细胞
DOI：
10.1371/journal.pone.0134629
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发表时间：
2015
期刊：
影响因子：
3.7
通讯作者：
Xie M
中科院分区：
文献类型：
--
作者：
Zhang L;Sun Z;Ren P;Lee RJ;Xiang G;Lv Q;Han W;Wang J;Ge S;Xie M
基因治疗对人类疾病具有巨大的潜力。开发有效的递送系统对其临床转化至关重要。最近的研究表明，微泡结合超声（US）可用于促进基因递送。本研究的目的是调查是否美国靶向微泡破坏（UTMD）和聚乙烯亚胺（PEI）（UTMD/PEI）的组合可以介导甚至更大的基因转染效率比UTMD单独和优化超声辐照参数。本研究的另一个目的是研究PHD 2-shRNA转染H9 C2细胞后的生物学效应。将pEGFP-N1或真核shPHD 2-EGFP质粒与包被白蛋白的微泡和PEI混合形成复合物用于转染。将这些添加到H9 C2细胞中后，将细胞暴露于具有各种参数的US。然后收获细胞并分析基因表达。UTMD/PEI在基因转染中显示出高效。结果表明，超声强度1.5W/cm ~ 2、微泡浓度300μl/ml、照射时间45 s、质粒浓度15μg/ml是转染的最佳条件。UTMD/PEI介导的PHD 2-shRNA转染H9 C2细胞后，PHD 2表达明显下调，HIF-1α及下游血管生成因子VEGF、TGF-β和bFGF表达明显上调。UTMD/PEI与白蛋白包被的微泡相结合，保证了治疗性基因递送的进一步研究。
Gene therapy has great potential for human diseases. Development of efficient delivery systems is critical to its clinical translation. Recent studies have shown that microbubbles in combination with ultrasound (US) can be used to facilitate gene delivery. An aim of this study is to investigate whether the combination of US-targeted microbubble destruction (UTMD) and polyethylenimine (PEI) (UTMD/PEI) can mediate even greater gene transfection efficiency than UTMD alone and to optimize ultrasonic irradiation parameters. Another aim of this study is to investigate the biological effects of PHD2-shRNA after its transfection into H9C2 cells. pEGFP-N1 or eukaryotic shPHD2-EGFP plasmid was mixed with albumin-coated microbubbles and PEI to form complexes for transfection. After these were added into H9C2 cells, the cells were exposed to US with various sets of parameters. The cells were then harvested and analyzed for gene expression. UTMD/PEI was shown to be highly efficient in gene transfection. An US intensity of 1.5 W/cm2, a microbubble concentration of 300μl/ml, an exposure time of 45s, and a plasmid concentration of 15μg/ml were found to be optimal for transfection. UTMD/PEI-mediated PHD2-shRNA transfection in H9C2 cells significantly down regulated the expression of PHD2 and increased expression of HIF-1α and downstream angiogenesis factors VEGF, TGF-β and bFGF. UTMD/PEI, combined with albumin-coated microbubbles, warrants further investigation for therapeutic gene delivery.