Large-scale expansion of pre-isolated bone marrow mesenchymal stromal cells in serum-free conditions

Abstract

The regenerative potential of mesenchymal stromal or stem cells (MSCs) has generated tremendous interest for treating various degenerative diseases. Regulatory preference is to use a culture medium that is devoid of bovine components for stem cell expansion intended for therapeutic applications. However, a clear choice an alternative to fetal bovine serum (FBS) has not yet emerged. We have screened five different commercially available serum-free media (SFM) for their ability to support the growth and expansion of pre-isolated undifferentiated bone marrow-derived MSCs (BM-MSCs) and compared the results with cells grown in standard FBS-containing medium as control. In addition, based on initial screening results, BD Mosaic™ Mesenchymal Stem Cell Serum-free (BD-SFM) medium was evaluated in large-scale cultures for the performance and culture characteristics of BM-MSCs. Of the five different serum-free media, BD-SFM enhanced BM-MSCs growth and expansion in Cell STACK (CS), but the cell yield per CS-10 was less when compared to the control medium. The characteristics of MSCs were measured in terms of population doubling time (PDT), cell yield and expression of MSC-specific markers. Significant differences were observed between BD-SFM and control medium in terms of population doublings (PDs), cell yield, CFU-F and morphological features, whereas surface phenotype and differentiation potentials were comparable. The BD-SFM-cultured MSCs were also found to retain the differentiation potential, immune-privileged status and immunosuppressive properties inherent to MSCs. Our results suggest that BD-SFM supports large-scale expansion of BM-MSCs for therapeutic use.

Serum‐free human MSC medium supports consistency in human but not in equine adipose‐derived multipotent mesenchymal stromal cell culture

“…In contrast, human MACS II MSCs appeared similar to equine DMEM MSCs in the current study. Interestingly, in other studies, alterations of morphology depending on the serum‐free culture medium used had also been observed in human MSCs, the cells being more slender and spindle‐shaped but also growing in bundles as observed for equine MACS MSCs in this study.…”

“…These non‐platelet‐based serum‐free media for human MSC expansion have been studied to a similar extent as platelet lysate‐based media. The reports demonstrate that there are media which promote the growth of MSCs, but they also reveal that serum‐free culture was not successful in all media tested . Moreover, primary serum‐free culture was not always attempted and the results obtained with different media are partly conflicting , indicating that at least, the media formulations are highly specific for particular cell types and culture procedures.…”

“…The reports demonstrate that there are media which promote the growth of MSCs, but they also reveal that serum‐free culture was not successful in all media tested . Moreover, primary serum‐free culture was not always attempted and the results obtained with different media are partly conflicting , indicating that at least, the media formulations are highly specific for particular cell types and culture procedures. Therefore, in accordance with the recommendations published by the ISCT , it is of utmost importance to evaluate the impact of the specific serum‐free culture conditions on the particular type of MSC intended for cellular therapy.…”

“…Published studies on serum‐free culture media mainly focus on human MSCs, which were obtained from bone marrow in the majority of studies , but also from adipose tissue or birth‐associated tissues . In contrast, serum‐free culture of large animal MSCs has only been addressed in very few studies so far .…”

Clinical Translation of Tissue Engineered Medicinal Products

“…The major manufacturing constraint involves scale-up, consistency, repeatability, reproducibility, immature production technologies, process variability, process validation and potency testing related to clinical outcome. Moreover, the critical process controls to be integrated and automated to enhance cGMP compliances for large scale commercial manufacturing [20,21] . On the other hand, the drug dose requirements of TEMPs and severity of the condition of recipient varies from patient to patient is critical variable factors that impact the clinical outcome.…”