Bioreactor Parameters for Microcarrier-Based Human MSC Expansion under Xeno-Free Conditions in a Vertical-Wheel System

Abstract

<jats:p>Human mesenchymal stem/stromal cells (hMSCs) have been investigated and proven to be a well-tolerated, safe therapy for a variety of indications, as shown by over 900 registered hMSC-based clinical trials. To meet the commercial demand for clinical manufacturing of hMSCs, production requires a scale that can achieve a lot size of ~100B cells, which requires innovative manufacturing technologies such as 3D bioreactors. A robust suspension bioreactor process that can be scaled-up to the relevant scale is therefore crucial. In this study, we developed a fed-batch, microcarrier-based bioreactor process, which enhances media productivity and drives a cost-effective and less labor-intensive hMSC expansion process. We determined parameter settings for various stages of the culture: inoculation, bioreactor culture, and harvest. Addition of a bioreactor feed, using a fed-batch approach, was necessary to replenish the mitogenic factors that were depleted from the media within the first 3 days of culture. Our study resulted in an optimized hMSC culture protocol that consistently achieved hMSC densities between 2 × 105–6 × 105 cells/mL within 5 days with no media exchange, maintaining the final cell population doubling level (PDL) at 16–20. Using multiple hMSC donors, we showed that this process was robust and yielded hMSCs that maintained expansion, phenotypic characteristic, and functional properties. The developed process in a vertical-wheel suspension bioreactor can be scaled to the levels needed to meet commercial demand of hMSCs.</jats:p>

Extracellular Vesicles as a Therapeutic Tool for Kidney Disease: Current Advances and Perspectives

“…The microcarrier culture systems can promote a homogeneous suspension that improves cell culture robustness and reproducibility [105]. This approach has been used to improve the expansion process of various MSC cultures [106], often combined with dynamic systems for suspension culture such as spinner flasks, stirred-tank bioreactors or wave bioreactors, allowing high-density cultures with low cell damage [107][108][109][110][111].…”

Bioprocessing of Human Mesenchymal Stem Cells: From Planar Culture to Microcarrier-Based Bioreactors

“…This both reduces microcarrier loss during draining and also preserves hMSC-secreted cytokines which are known to enhance expansion. In some cases, if the off-line medium analysis infers that the depletion of a specific known nutrient component will soon occur (according to the cell growth curve), a higher concentration of this component may be provided as an additive to compensate for the shortage and maintain the cell proliferation rates [32,83].…”

Turning Nature’s own processes into design strategies for living bone implant biomanufacturing: a decade of Developmental Engineering

“…Bioreactor systems allowing efficient expansion of progenitor cells should be used and evidence of this has already been provided for example in multi-stack and hollow-fiber bioreactors [ [125] , [126] , [127] ]. There exist several types of suspension culture using microcarriers in stirred bioreactors [ 128 , 129 ] such as vertical wheel systems [ 130 , 131 ], wave bags [ 132 ] and mostly small scale stirred systems (typically spinner flasks of 100-200 ml) [ [133] , [134] , [135] ]. Studies showing that BM-MSCs can be expanded in 5L stirred tanks [ 136 ] and 50L Mobius® bioreactors demonstrate that large scale production of single cell progenitor cells is feasible and could provide adequate quantities of cells for further differentiation.…”