A comprehensive characterisation of large-scale expanded human bone marrow and umbilical cord mesenchymal stem cells
A comprehensive characterisation of large-scale expanded human bone marrow and umbilical cord mesenchymal stem cells
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DOI：
10.1186/s13287-019-1202-4
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发表时间：
2019-03-18
影响因子：
7.5
通讯作者：
Wright, Karina
中科院分区：
文献类型：
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作者：
Mennan, Claire;Garcia, John;Wright, Karina
背景间充质干/基质细胞（mesenchymal stem/stromal cells，MSCs）的制备需要成本低、安全、规模化。目前在组织培养塑料上扩增的方法是劳动密集型的，并且涉及几个开放式程序。我们已经使用封闭的中空纤维生物反应器来扩增来自骨髓（BM）和脐带（UC）的MSC的四种培养物，并且与在组织培养塑料上生长的平行培养物相比，对每种培养物评估广泛的表征概况。将骨髓细胞重新接种到Quantum（R）中，收获并在P1时进一步表征。酶促分离UC-MSC并在组织培养塑料上培养一次，然后将细胞加载到Quantum（R）中，在P1收获并表征。Quantum（R）衍生培养物在免疫特征、三系分化、对炎症刺激的反应和端粒长度方面进行了表型分析，结果14 +/-2天（P0）从Quantum（R）获得的骨髓细胞为23.116.2x10（6），13 +/-13天（P0）获得的BM-MSC为131 +/-84 x10（6）。1天（P1），而从Quantum（R）收获的UC-MSC在7 +/-2天（P1）后为168 +/-52x10（6）UC-MSC。P1时的Quantum和组织培养塑料扩增培养物在细胞表面标志物、多能性和塑料粘附方面符合MSC的标准，而整合素CD 29、CD 49 c和CD 51/61在Quantum扩增的BM-MSC上被发现升高。与组织培养塑料上的培养物相比，Quantum中的快速培养扩增不会导致端粒缩短。免疫调节基因的表达在供体之间是可变的，但显示所有的MSC上调吲哚胺2，3-双加氧酶（IDO）。结论这里呈现的结果表明，Quantum（R）可用于从骨髓和脐带组织扩增大量的MSC，用于下一代大规模生产，而不影响MSC的许多特性或潜在的治疗。使用Quantum（R），我们可以从一次生产运行中获得多个MSC剂量来治疗许多患者。总之，我们的研究结果支持开发更便宜的基于细胞的治疗方法。
BackgroundThe manufacture of mesenchymal stem/stromal cells (MSCs) for clinical use needs to be cost effective, safe and scaled up. Current methods of expansion on tissue culture plastic are labour-intensive and involve several open' procedures. We have used the closed Quantum (R) hollow fibre bioreactor to expand four cultures each of MSCs derived from bone marrow (BM) and, for the first time, umbilical cords (UCs) and assessed extensive characterisation profiles for each, compared to parallel cultures grown on tissue culture plastic.MethodsBone marrow aspirate was directly loaded into the Quantum (R), and cells were harvested and characterised at passage (P) 0. Bone marrow cells were re-seeded into the Quantum (R), harvested and further characterised at P1. UC-MSCs were isolated enzymatically and cultured once on tissue culture plastic, before loading cells into the Quantum (R), harvesting and characterising at P1. Quantum (R)-derived cultures were phenotyped in terms of immunoprofile, tri-lineage differentiation, response to inflammatory stimulus and telomere length, as were parallel cultures expanded on tissue culture plastic.ResultsBone marrow cell harvests from the Quantum (R) were 23.116.2x10(6) in 14 +/- 2days (P0) and 131 +/- 84x10(6) BM-MSCs in 13 +/- 1days (P1), whereas UC-MSC harvests from the Quantum (R) were 168 +/- 52x10(6) UC-MSCs after 7 +/- 2days (P1). Quantum (R)- and tissue culture plastic-expanded cultures at P1 adhered to criteria for MSCs in terms of cell surface markers, multipotency and plastic adherence, whereas the integrins, CD29, CD49c and CD51/61, were found to be elevated on Quantum (R)-expanded BM-MSCs. Rapid culture expansion in the Quantum (R) did not cause shortened telomeres when compared to cultures on tissue culture plastic. Immunomodulatory gene expression was variable between donors but showed that all MSCs upregulated indoleamine 2, 3-dioxygenase (IDO).Conclusions The results presented here demonstrate that the Quantum (R) can be used to expand large numbers of MSCs from bone marrow and umbilical cord tissues for next-generation large-scale manufacturing, without impacting on many of the properties that are characteristic of MSCs or potentially therapeutic. Using the Quantum (R), we can obtain multiple MSC doses from a single manufacturing run to treat many patients. Together, our findings support the development of cheaper cell-based treatments.