微载波
间充质干细胞
细胞生物学
微泡
胞外囊泡
细胞外小泡
化学
细胞培养
CD63
间质细胞
背景(考古学)
生物
细胞
生物化学
癌症研究
小RNA
遗传学
基因
古生物学
作者
Hélder Bandarra-Tavares,Teresa Franchi-Mendes,Cristiana Ulpiano,Sara Morini,Navjot Kaur,Abigail Harris-Becker,Mohan C. Vemuri,Joaquim M. S. Cabral,Ana Fernandes‐Platzgummer,Cláudia L. da Silva
标识
DOI:10.1016/j.jcyt.2024.03.001
摘要
Cell therapies based on mesenchymal stromal cells (MSCs) have gained an increasing therapeutic interest in the context of multiple disorders. Nonetheless, this field still faces important challenges, particularly concerning suitable manufacturing platforms. Here, we aimed at establishing a scalable culture system to expand umbilical cord-derived Wharton's jelly MSC (MSC(WJ)) and their derived extracellular vesicles (EVs) by using dissolvable microcarriers combined with xeno(geneic)-free culture medium.MSC(WJ) isolated from three donors were cultured at a starting density of 1 × 106 cells per spinner flask, i.e., 2.8 × 103 cells per cm2 of dissolvable microcarrier surface area. After a 6-day expansion period of MSC(WJ), extracellular vesicles (EVs) were produced for 24 h.Taking advantage of an intermittent agitation regimen, we observed high adhesion rates to the microcarriers (over 90% at 24 h) and achieved 15.8 ± 0.7-fold expansion after 6 days of culture. Notably, dissolution of the microcarriers was achieved through a pectinase-based solution to recover the cell product, reducing the hurdles of downstream processing. MSC identity was validated by detecting the characteristic MSC immunophenotype and by multilineage differentiation assays. Considering the growing interest in MSC-derived EVs, which are known to be mediators of the therapeutic features of MSC, this platform also was evaluated for EV production. Upon a 24-h period of conditioning, secreted EVs were isolated by ultrafiltration followed by anion-exchange chromatography and exhibited the typical cup-shaped morphology, small size distribution (162.6 ± 30.2 nm) and expressed EV markers (CD63, CD9 and syntenin-1).Taken together, we established a time-effective and robust scalable platform that complies with clinical-grade standards for the dual production of MSC(WJ) and their derived EV.
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