Chia Nan University of Pharmacy & Science Institutional Repository:Item 310902800/31643
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    標題: The vector-related influences of autophagic microRNA delivery by Lipofectamine 2000 and polyethylenimine 25K on mouse embryonic fibroblast cells
    作者: Lin, Chia-Wei
    Jan, Ming-Shiou
    Kuo, Jung-Hua Steven
    貢獻者: Chung Shan Med Univ, Inst Biochem Microbiol & Immunol
    Chia Nan Univ Pharm & Sci, Dept Pharm
    關鍵字: Polyethylenimine 25K (PEI)
    microRNAs (miRNAs)
    Lipofectamine 2000 (Lipo)
    日期: 2017-04-01
    上傳時間: 2018-11-30 15:51:18 (UTC+8)
    出版者: Elsevier Science Bv
    摘要: Despite the greater potential for clinical applications of autophagic microRNA (miRNA) delivery, the vector-related effects of such delivery on cells have not been fully explored. In this study, autophagic mmu-miR-494-3p (miR-494) in mouse embryonic fibroblast (MEF) cells was selected as a cargo miRNA, and two commonly used non-viral carriers (Lipofectamine 2000 (Lipo) and polyethylenimine 25K (PEI)), were used as delivery vectors to mechanistically elucidate its vector-related effects. The cellular uptake, nuclear localization, and quantitative miR-494 levels of the complexes of miR-494 with Lipo (miR-494 lipoplexes) were lower than those of the complexes of miR-494 with PEI (miR-494 polyplexes) in MEF cells. The indicator of autophagic activity (LC3 (micro-tubule-associated protein 1 light chain 3)-II/LC3-I ratio) in cells treated with miR-494 lipoplexes was higher than that in cells treated with miR-494 polyplexes. Lipo alone and PEI alone induced slight increases in the quantitative levels of miR-494 in cells, but Lipo resulted in higher gene and protein expressions of target Igf1, higher LC3-II/LC3-I ratios, and higher autophagosome formation than PEI. We also demonstrated that the delivery of miR-494 by Lipo was more involved in apoptotic caspase-3 pathways than such delivery by PEI. By applying knock-out atg5 gene in MEF cells, we found that autophagy played a protective role in cell survival and also affected cellular uptake, the quantitative level of miR-494, and target gene Igf1 regulation of delivery systems. Taken together, these results indicate that there are different degrees of responses in MEF cells for autophagic miR-494 delivery through the use of Lipo or PEI vectors that also induce autophagy in cells. Therefore, Lipo and PEI vectors cannot be treated as inert molecules, and their effects must be known and evaluated when they are used in autophagic miRNA delivery systems. Most importantly, understanding these vector,related effects on cells will be helpful in achieving optimal delivery of autophagic miRNAs. (C) 2017 Elsevier B.V. All rights reserved.
    显示于类别:[藥學系(所)] 期刊論文


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