Extracellular vesicles (EVs) produced by human bone marrow-derived mesenchymal stromal cells (hBM-MSCs) are currently investigated for their clinical effectiveness towards immune-mediated diseases. The large amounts of stem cell-derived EVs required for clinical testing suggest that bioreactor production systems may be a more amenable alternative than conventional EV production methods for manufacturing products for therapeutic use in humans. To characterize the potential utility of these systems, EVs from four hBM-MSC donors were produced independently using a hollow-fiber bioreactor system under a cGMP-compliant procedure. EVs were harvested and characterized for size, concentration, immunophenotype, and glycan profile at three separate intervals throughout a 25-day period. Bioreactor-inoculated hBM-MSCs maintained high viability and retained their trilineage mesoderm differentiation capability while still expressing MSC-associated markers upon retrieval. EVs collected from the four hBM-MSC donors showed consistency in size and concentration in addition to presenting a consistent surface glycan profile. EV surface immunophenotypic analyses revealed a consistent low immunogenicity profile in addition to the presence of immuno-regulatory CD40 antigen. EV cargo analysis for biomarkers of immune regulation showed a high abundance of immuno-regulatory and angiogenic factors VEGF-A and IL-8. Significantly, EVs from hBM-MSCs with immuno-regulatory constituents were generated in a large-scale system over a long production period and could be frequently harvested with the same quality and quantity, which will circumvent the challenge for clinical application.

中文翻译：

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从人骨髓间充质基质细胞中空纤维生物反应器生产胞外囊泡产生纳米囊泡，反映了生产细胞的免疫调节性抗原特征

目前研究了人类骨髓源间充质基质细胞（hBM-MSC）产生的细胞外囊泡（EVs）对免疫介导疾病的临床有效性。临床测试所需的大量源自干细胞的电动汽车表明，与传统的电动汽车生产方法相比，生物反应器生产系统可能是一种更易于接受的替代方法，用于生产用于人类的治疗性产品。为了表征这些系统的潜在效用，在符合cGMP的程序下，使用中空纤维生物反应器系统独立生产了四个hBM-MSC供体的电动汽车。收集电动汽车，并在整个25天的三个独立时间间隔内对其大小，浓度，免疫表型和聚糖谱进行表征。接种生物反应器的hBM-MSC保持较高的生存力，并保留了其三系中胚层分化能力，同时在检索后仍表达MSC相关标记。从四个hBM-MSC供体中收集到的EV除具有一致的表面聚糖谱外，还显示出大小和浓度的一致性。EV表面免疫表型分析显示，除了存在免疫调节性CD40抗原外，还具有一致的低免疫原性特征。EV货物的免疫调节生物标志物分析表明，有大量的免疫调节因子和血管生成因子VEGF-A和IL-8。重要的是，来自hBM-MSC的具有免疫调节成分的EV是在较长的生产周期内大规模生产的，并且可能以相同的质量和数量被频繁收获，