Enhanced extracellular vesicle production and ethanol-mediated vascularization bioactivity via a 3D-printed scaffold-perfusion bioreactor system.
Enhanced extracellular vesicle production and ethanol-mediated vascularization bioactivity via a 3D-printed scaffold-perfusion bioreactor system.
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DOI：
10.1016/j.actbio.2018.11.024
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发表时间：
2019-09
影响因子：
9.7
通讯作者：
D. Patel;Christopher R. Luthers;M. Lerman;J. Fisher;S. Jay
中科院分区：
文献类型：
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作者：
D. Patel;Christopher R. Luthers;M. Lerman;J. Fisher;S. Jay
细胞外囊泡（EV）由于其固有的治疗性质以及其充当生物活性货物的载体的能力而在生物技术领域中引起了极大的兴趣。然而，缺乏成熟的生物制造平台和EV在体内的有限效力仍然是临床转化的关键瓶颈。在这项研究中，我们利用3D打印支架灌注生物反应器系统来评估动态培养对内皮细胞（EC）产生细胞外囊泡的反应。我们还研究了乙醇调节是否可以在3D生产系统中成功地用于相同的目的，乙醇调节先前被证明可以增强在标准2D培养条件下产生的EC衍生EV的血管化生物活性。我们的研究结果表明，在灌注生物反应器中的动态培养显着提高EV生产从人类EC。此外，结合动态培养使用乙醇调节诱导EC衍生EV的促血管形成生物活性，其与促血管生成lncRNA HOTAIR和MALAT1的EV水平增加相关。因此，这项研究代表了合理设计的EV效力增强的第一份报告之一，该EV效力增强在静态2D和动态3D EV生产系统之间是保守的，增加了用于各种应用的治疗性EC衍生EVs的可扩展生物制造的潜力。然而，基于EV的疗法的转化可能会受到生物制造挑战的阻碍。迄今为止，EV生产主要涉及使用组织培养瓶。在这里，我们首次报告了使用具有集成3D打印生物材料支架的管状灌注生物反应器系统从人内皮细胞生产EV。与常规组织培养系统相比，该系统将EV产量提高了100倍以上。此外，我们表明，我们小组先前在2D培养中开发的用于增强EV效力的乙醇调节方法与这种新系统兼容。因此，EV用于血管化应用的效力增强是可能的，即使生产率显著增加。
Extracellular vesicles (EVs) have garnered significant interest in the biotechnology field due to their intrinsic therapeutic properties as well as their ability to serve as vehicles for bioactive cargo. However, the lack of an established biomanufacturing platform and limited potency of EVsin vivoremain critical bottlenecks for clinical translation. In this study, we utilized a 3D-printed scaffold-perfusion bioreactor system to assess the response of dynamic culture on extracellular vesicle production from endothelial cells (ECs). We also investigated whether ethanol conditioning, which was previously shown to enhance vascularization bioactivity of EC-derived EVs produced in standard 2D culture conditions, could be employed successfully for the same purpose in a 3D production system. Our results indicate that dynamic culture in a perfusion bioreactor significantly enhances EV production from human ECs. Moreover, the use of ethanol conditioning in conjunction with dynamic culture induces pro-vascularization bioactivity of EC-derived EVs that is correlated with increased EV levels of pro-angiogenic lncRNAs HOTAIR and MALAT1. Thus, this study represents one of the first reports of rationally-designed EV potency enhancement that is conserved between static 2D and dynamic 3D EV production systems, increasing the potential for scalable biomanufacturing of therapeutic EC-derived EVs for a variety of applications.Statement of significanceExtracellular vesicles (EVs) have substantial therapeutic potential in a variety of applications. However, translation of EV-based therapies may be hindered by biomanufacturing challenges. EV production to date has predominantly involved the use of tissue culture flasks. Here, we report, for the first time, the use of a tubular perfusion bioreactor system with an integrated 3D-printed biomaterial scaffold for EV production from human endothelial cells. This system increases EV yield by over 100-fold compared to conventional tissue culture systems. Further, we show that an ethanol-conditioning approach that our group previously developed in 2D culture for enhancing EV potency is compatible with this new system. Thus, potency enhancement of EVs for vascularization applications is possible even with significantly increased production rate.