Minicircle microporation-based non-viral gene delivery improved the targeting of mesenchymal stem cells to an injury site
Minicircle microporation-based non-viral gene delivery improved the targeting of mesenchymal stem cells to an injury site
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DOI：
10.1016/j.biomaterials.2016.05.057
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发表时间：
2016-09-01
期刊：
影响因子：
14
通讯作者：
Oh, Doo-Byoung
中科院分区：
文献类型：
--
作者：
Mun, Ji-Young;Shin, Keun Koo;Oh, Doo-Byoung
通过病毒和非病毒基因递送方法，已经实现了提高间充质干细胞（MSC）的治疗潜力的基因工程方法。由于潜在的致癌性、致病性和免疫原性风险，病毒方法在临床应用中具有严重的局限性，而非病毒方法由于MSC是难以转染的细胞而遭受低转染效率和瞬时弱表达。在这项研究中，微环，这是一个最小的表达载体的细菌序列，用于MSC转染作为一种非病毒基因递送方法。传统的阳离子脂质体方法对于MSC转染无效，因为它导致转染效率非常低（低于5%）。微穿孔，一种新的电穿孔方法，大大提高了微环的转染效率高达66%，在MSC中没有任何显着的细胞活力的损失。此外，小环微穿孔比质粒微穿孔产生更强和更长的转基因表达。当将携带萤火虫荧光素酶基因的微环微穿孔的MSC皮下注射到小鼠时，生物发光持续超过一周，而质粒微穿孔诱导的MSC的生物发光在小鼠中迅速下降并在三天内消失。通过微环微孔作为非病毒基因递送，工程化以过表达CXCR4的MSC显示出朝向损伤部位的归巢能力大大增加，如通过小鼠体内生物发光成像所证实的。综上所述，通过微环微穿孔工程化MSC有望增强MSC在临床应用中的治疗潜力。(C)2016爱思唯尔有限公司版权所有
Genetic engineering approaches to improve the therapeutic potential of mesenchymal stem cells (MSCs) have been made by viral and non-viral gene delivery methods. Viral methods have severe limitations in clinical application because of potential oncogenic, pathogenic, and immunogenic risks, while non-viral methods have suffered from low transfection efficiency and transient weak expression as MSCs are hard-to-transfect cells. In this study, minicircle, which is a minimal expression vector free of bacterial sequences, was employed for MSC transfection as a non-viral gene delivery method. The conventional cationic liposome method was not effective for MSC transfection as it resulted in very low transfection efficiency (less than 5%). Microporation, a new electroporation method, greatly improved the transfection efficiency of minicircles by up to 66% in MSCs without any significant loss of cell viability. Furthermore, minicircle microporation generated much stronger and prolonged transgene expression compared with plasmid microporation. When MSCs microporated with minicircle harboring firefly luciferase gene were subcutaneously injected to mice, the bioluminescence continued for more than a week, whereas the bioluminescence of the MSCs induced by plasmid microporation rapidly decreased and disappeared in mice within three days. By minicircle microporation as a non-viral gene delivery, MSCs engineered to overexpress CXCR4 showed greatly increased homing ability toward an injury site as confirmed through in vivo bioluminescence imaging in mice. In summary, the engineering of MSCs through minicircle microporation is expected to enhance the therapeutic potential of MSCs in clinical applications. (C) 2016 Elsevier Ltd. All rights reserved.