Modulation of mesenchymal stromal cell characteristics by microcarrier culture in bioreactors

Abstract

Mesenchymal stromal cells (MSCs) are promising candidates for cell therapy. Their therapeutic use requires extensive expansion to obtain a sufficiently high number of cells for clinical applications. State-of-the-art expansion systems, that is, primarily culture flask-based systems, are limited regarding scale-up, automation, and reproducibility. To overcome this bottleneck, microcarrier (MC)-based expansion processes have been developed. For the first time, MSCs from the perinatal sources umbilical cord (UC) and amniotic membrane (AM) were expanded on MCs. This study focuses on the comparison of flask- and Cytodex 1 MC-expanded MSCs by evaluating the influence of the expansion process on biological MSC characteristics. Furthermore, we tested the hypothesis to obtain more homogeneous MSC preparations by expanding cells on MCs in controlled large-scale bioreactors. MSCs were extensively characterized determining morphology, cell growth, surface marker expression, and functional properties such as differentiation capacity, secretion of paracrine factors, and gene expression. Based on their gene expression profile MSCs from different donors and sources clearly clustered in distinct groups solely depending on the expansion process-MC or flask culture. MC- and flask-expanded MSCs significantly differed from each other regarding surface markers and both paracrine factors and gene expression profiles. Furthermore, based on gene expression analysis, MC cultivation of MSCs in controlled bioreactor systems resulted in less heterogeneity between cells from different donors. In conclusion, MC-based MSC expansion in controlled bioreactors has the potential to reliably produce MSCs with altered characteristics and functions as compared to flask-expanded MSCs. These findings may be useful for the generation of MSCs with tailored properties for clinical applications.

Modulation of the Mesenchymal Stem Cell Secretome Using Computer-Controlled Bioreactors: Impact on Neuronal Cell Proliferation, Survival and Differentiation

“…The use of bioreactors has been suggested as a promising alternative to conventional static culture flasks for hMSC expansion914. So far, different bioreactors have been used for hMSC expansion, including fixed bed bioreactors, perfusion bioreactors, and the cultivation of MSCs as aggregates or through the use of microcarriers in stirred suspension bioreactor systems15.…”

Mesenchymal Stem Cell-derived Extracellular Vesicles: Toward Cell-free Therapeutic Applications

“…Other approaches to enhancing the purity of MSC-EVs preparations could include sequential centrifugation, filtration and ultracentrifugation followed by sucrose density gradient to remove contaminating protein aggregates, cell debris and genetic material [118][119][120]. To scale up the amount of EVs isolated, bioreactors could be used to culture the MSCs [121]. In this regard, a small number of studies have documented significant increases in EV yield from cells cultured in bioreactor systems compared to conventional tissue culture flasks [122].…”

Mesenchymal stem cell derived secretome and extracellular vesicles for acute lung injury and other inflammatory lung diseases

“…Recently, Chen et al[112] proposed a robust scalable manufacturing process for therapeutic EV through oncogenic immortalization of human embryonic stem cell (ESC)-derived MSC by transfection with the MYC gene. Another approach to scale up EV production could be the use of bioreactors to culture MSC[113]. Several studies have documented significant increases in EV yield from cells cultured in bioreactor systems when compared with conventional tissue culture flasks[114].…”