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Mesenchymal Stem Cells Engineering: Microcapsules-Assisted Gene Transfection and Magnetic Cell Separation
ACS Biomaterials Science & Engineering ( IF 5.4 ) Pub Date : 2017-09-22 00:00:00 , DOI: 10.1021/acsbiomaterials.7b00482 Albert R. Muslimov 1, 2, 3 , Alexander S. Timin 1, 4 , Aleksandra V. Petrova 2, 5 , Olga S. Epifanovskaya 1 , Alena I. Shakirova 1 , Kirill V. Lepik 1, 3 , Andrey Gorshkov 2 , Eugenia V. Il’inskaja 2 , Andrey V. Vasin 2, 5 , Boris V. Afanasyev 1 , Boris Fehse 6 , Gleb B. Sukhorukov 7
ACS Biomaterials Science & Engineering ( IF 5.4 ) Pub Date : 2017-09-22 00:00:00 , DOI: 10.1021/acsbiomaterials.7b00482 Albert R. Muslimov 1, 2, 3 , Alexander S. Timin 1, 4 , Aleksandra V. Petrova 2, 5 , Olga S. Epifanovskaya 1 , Alena I. Shakirova 1 , Kirill V. Lepik 1, 3 , Andrey Gorshkov 2 , Eugenia V. Il’inskaja 2 , Andrey V. Vasin 2, 5 , Boris V. Afanasyev 1 , Boris Fehse 6 , Gleb B. Sukhorukov 7
Affiliation
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Stem cell engineering—the manipulation and functionalization of stem cells involving genetic modification—can significantly expand their applicability for cell therapy in humans. Toward this aim, reliable, standardized, and cost-effective methods for cell manipulation are required. Here we explore the potential of magnetic multilayer capsules to serve as a universal platform for nonviral gene transfer, stem cell magnetization, and magnetic cell separation to improve gene transfer efficiency. In particular, the following experiments were performed: (i) a study of the process of internalization of magnetic capsules into stem cells, including capsule co-localization with established markers of endo-lysosomal pathway; (ii) characterization and quantification of capsule uptake with confocal microscopy, electron microscopy, and flow cytometry; (iii) intracellular delivery of messenger RNA and separation of gene-modified cells by magnetic cell sorting (MACS); and (iv) analysis of the influence of capsules on cell proliferation potential. Importantly, based on the internalization of magnetic capsules, transfected cells became susceptible to external magnetic fields, which made it easy to enrich gene-modified cells using MACS (purity ∼95%), and also to influence their migration behavior. In summary, our results underline the high potential of magnetic capsules in stem cell functionalization, namely (i) to increase gene-transfer efficiency and (ii) to facilitate enrichment and targeting of transfected cells. Finally, we did not observe a negative impact of the capsules used on the proliferative capacity of stem cells, proving their high biocompatibility.
中文翻译：
间充质干细胞工程：微胶囊辅助基因转染和磁性细胞分离。
干细胞工程（涉及基因修饰的干细胞的操纵和功能化）可以大大扩展其在人类细胞治疗中的适用性。为了实现这一目标，需要可靠，标准化且具有成本效益的细胞处理方法。在这里，我们探索了多层磁性胶囊作为非病毒基因转移，干细胞磁化和磁性细胞分离以提高基因转移效率的通用平台的潜力。具体而言，进行了以下实验：（i）研究磁性胶囊内在化为干细胞的过程，包括将胶囊与内溶酶体途径的既定标志物共定位；（ii）使用共聚焦显微镜，电子显微镜和流式细胞术对胶囊摄取进行表征和定量；（iii）通过磁性细胞分选术（MACS）在细胞内递送信使RNA并分离基因修饰的细胞；（iv）分析胶囊对细胞增殖潜能的影响。重要的是，基于磁性胶囊的内在化，转染的细胞变得易受外部磁场的影响，这使得使用MACS富集基因修饰的细胞变得容易（纯度约95％），并且还影响了它们的迁移行为。总之，我们的结果强调了磁胶囊在干细胞功能化方面的巨大潜力，即（i）提高基因转移效率和（ii）促进转染细胞的富集和靶向。最后，我们没有观察到使用的胶囊对干细胞的增殖能力有负面影响，证明了它们的高生物相容性。
更新日期：2017-09-23
中文翻译：
间充质干细胞工程：微胶囊辅助基因转染和磁性细胞分离。
干细胞工程（涉及基因修饰的干细胞的操纵和功能化）可以大大扩展其在人类细胞治疗中的适用性。为了实现这一目标，需要可靠，标准化且具有成本效益的细胞处理方法。在这里，我们探索了多层磁性胶囊作为非病毒基因转移，干细胞磁化和磁性细胞分离以提高基因转移效率的通用平台的潜力。具体而言，进行了以下实验：（i）研究磁性胶囊内在化为干细胞的过程，包括将胶囊与内溶酶体途径的既定标志物共定位；（ii）使用共聚焦显微镜，电子显微镜和流式细胞术对胶囊摄取进行表征和定量；（iii）通过磁性细胞分选术（MACS）在细胞内递送信使RNA并分离基因修饰的细胞；（iv）分析胶囊对细胞增殖潜能的影响。重要的是，基于磁性胶囊的内在化，转染的细胞变得易受外部磁场的影响，这使得使用MACS富集基因修饰的细胞变得容易（纯度约95％），并且还影响了它们的迁移行为。总之，我们的结果强调了磁胶囊在干细胞功能化方面的巨大潜力，即（i）提高基因转移效率和（ii）促进转染细胞的富集和靶向。最后，我们没有观察到使用的胶囊对干细胞的增殖能力有负面影响，证明了它们的高生物相容性。
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