Incorporation of Viral Glycoprotein VSV-G Improves the Delivery of DNA by Erythrocyte Ghost into Cells Refractory to Conventional Transfection
Incorporation of Viral Glycoprotein VSV-G Improves the Delivery of DNA by Erythrocyte Ghost into Cells Refractory to Conventional Transfection
复制标题
病毒糖蛋白 VSV-G 的掺入可改善红细胞血影将 DNA 递送至传统转染难治的细胞中
DOI：
10.1007/s12010-016-2246-x
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发表时间：
2017-02
期刊：
影响因子：
--
通讯作者：
Wei CJ
中科院分区：
文献类型：
--
作者：
Liu X;Li YP;Zhong ZM;Tan HQ;Lin HP;Chen SJ;Fu YC;Xu WC;Wei CJ
本研究的目的是建立一种基于红细胞血影（EG）整合融合病毒糖蛋白水泡性口炎病毒糖蛋白G（VSV-G）的新型基因传递系统。收获VSV-G蛋白作为携带VSV-G转基因的Ad 293细胞的条件培养基，然后掺入EG中。用不同的转染试剂浓缩质粒DNA。荧光素酶表达构建体（pGL 3-对照）和DsRed表达盒（pCMV-DsRed）用于评估DNA/EG/VSV-G复合物的递送效率。Western印迹分析表明，VSV-G蛋白可以在酸性条件下静态孵育掺入EG中。凝聚的质粒DNA主要结合在EG的外表面，这可以通过电镜检测和电泳测量。EG/VSV-G复合物能显著促进pGL 3-control转染Ad 293细胞，荧光素酶活性比对照提高约4倍。pCMV-DsRed的递送也随着DsRed阳性Ad 293细胞的百分比从55%增加到约80%而增强。此外，在3 T3、HeLa、INS-1和骨髓干细胞（BMSC）细胞中的转染效率增加约2-3倍。最后，共聚焦显微镜分析表明，VSV-G的掺入显着增强EG的内吞到靶细胞。在本研究中，配制了一种由EG和融合VSV-G蛋白组成的新型非病毒DNA递送载体，即使在对经典转染有抗性的细胞中也显示出上级转染效率。
The objective of this study was to formulate a novel gene delivery system based on the erythrocyte ghost (EG) integrated with fusogenic viral glycoprotein vesicular stomatitis virus glycoprotein G (VSV-G). VSV-G proteins were harvested as condition medium of Ad293 cells carrying a VSV-G transgene and then incorporated into EG. Plasmid DNA was condensed by various transfection reagents. A luciferase expression construct (pGL3-control) and a DsRed expression cassette (pCMV-DsRed) were used to evaluate the delivery efficiency of DNA/EG/VSV-G complexes. VSV-G proteins could be incorporated into EG in static incubation under acidic conditions as evidenced by the Western blot analysis. Condensed plasmid DNA was bound mostly to the outer surface of EG, which could be detected by electromicroscopy and measured by electrophoresis. EG/VSV-G complexes stimulated the delivery of pGL3-control into Ad293 cells significantly with the luciferase activity increased about 4-fold as compared to that of the control. The delivery of pCMV-DsRed was also enhanced with the percentage of DsRed-positive Ad293 cells increased from 55 % to about 80 %. Moreover, the transfection efficiency in 3T3, HeLa, INS-1, and bone marrow stem cell (BMSC) cells increased about 2–3-fold. Finally, confocal microscopy analysis showed that incorporation of VSV-G significantly enhanced the endocytosis of EG into target cells. In the present study, a novel type of non-viral DNA delivery vehicle consisting of EG and fusogenic VSV-G proteins was formulated, which showed superior transfection efficiency even in cells resistant to classical transfection.
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影响因子：
4.8
作者：
Matos, Pedro M.;Marin, Mariana;Melikyan, Gregory B.
通讯作者：
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影响因子：
16.1
作者：
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影响因子：
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作者：
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通讯作者：
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作者：
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通讯作者：
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影响因子：
5.4
作者：
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通讯作者：
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