Culture conditions for growth of clinical grade human tissue derived mesenchymal stem cells: comparative study between commercial serum-free media and human product supplemented media

Abstract

Background: Large scale high quality and low cost production of clinical grade mesenchymal stem cells (MSCs) is critical for facilitating clinical research and potential stem cell therapies. Use of Fetal Bovine/Calf serum (FBS/FCS) for the clinical application carries a risk for transmission of zoonosis, therefore alternative xeno-free cell culture techniques have been evaluated. The commercial products for the culture of MSCs in xeno-free and/or serum free media such as STEMPRO® MSC SFM and MesenGro® are available but possibly not cost effective for large scale production of MSCs. The usage of human serum and pooled human platelet lysate (pHPL) present xeno-free alternatives as growth supplements. Methods: In this report, we analyzed the growth potentials of human tissue derived MSCs from bone marrow (BM), umbilical cord tissue (UCT) and umbilical cord blood (UCB) in presence of xeno-free, serum free commercial culture media (STEMPRO® MSC SFM and MesenGro®) and serum containing media supplemented with pHPL, FBS and human serum. We also evaluated the role of surface of culture flasks for the optimal production of MSCs from these tissues by comparing the expansion potential of MSCs in culture flasks from Costar®, CellBIND®, Greiner® and Tarsons® brands. Results: We compared the growth potentials of these MSCs derived from these human tissues and found that pHPL supplemented culture media serves as an efficient, cost effective xeno-free media to produce clinical grade human MSCs for clinical applications. Interestingly, out of two commercial media, MesenGro® demonstrated better isolation and expansion potential than STEMPRO® MSC SFM indicating that composition of commercial media affected the MSCs isolation and expansion although MSCs showed similar phenotypic characteristics and post cryopreservation viability. The MSCs production was at best in culture flasks with CellBIND® surface treatment. Conclusion:We conclude that pHPL supplemented media provide cost effective and efficacious way to produce clinical grade MSCs from various human tissues as opposed to commercial media. We showed that UCT can be an alternative option for large number of MSCs and unique surface treatment of cell culture flasks such as CellBIND® facilitates large scale production of MSCs from various human tissues.

Conditioned medium: a new alternative for cryopreservation of equine umbilical cord mesenchymal stem cells

“…The reduction in cell viability after cryopreservation (M1–M3), compared to fresh cells (P2), is possibly due to osmotic stress which occurs during the freezing and thawing process, which proved to be more pronounced in cells cryopreserved in the absence of FBS (M2). The loss of cell viability was also observed in other studies after cryopreservation of different cell (Dhot et al, ; Vrhovac et al, ; Ribeiro et al, ; Inamdar and Inamdar, ; Kim et al, ; Munévar et al, ). It is noteworthy that the loss of viability did not affect the adhesion and clonicity shown in CFU‐F assay of UCIM‐MSC cryopreserved with medium M1 and M3, since the CFU‐F efficiency did not differ ( P < 0.05) in these groups to fresh cells.…”

Mass Production of Mesenchymal Stem Cells — Impact of Bioreactor Design and Flow Conditions on Proliferation and Differentiation

“…Numerous clinical studies have confirmed the safety of allogenic and autologous MSCs for treatment of human diseases [12,13]. Currently, MSCs are being investigated in terms of their therapeutic potential for inflammatory, autoimmune and degenerative conditions in preclinical and clinical studies [14,15].…”

“…As things stand, bone marrow aspirate represents the most often used source [22] of MSCs. In addition, MSCs are obtained from adipose tissue [26], placenta as well as umbilical tissue [15] and blood [27,28], and peripheral blood [29] (Figure 1). Other sources, such as periosteum, trabecular bone, synovia, skeletal muscle, deciduous teeth, fetal pancreas, lung, liver and amniotic fluid, have also been reported [15].…”

“…In addition, MSCs are obtained from adipose tissue [26], placenta as well as umbilical tissue [15] and blood [27,28], and peripheral blood [29] (Figure 1). Other sources, such as periosteum, trabecular bone, synovia, skeletal muscle, deciduous teeth, fetal pancreas, lung, liver and amniotic fluid, have also been reported [15]. But independent of the source, the low frequency of MSCs makes their direct collection for the majority of MSC applications unfeasible.…”

“…In order to fulfill the potential of MSCs as therapeutic agents for a wide range of applications, culture conditions need to be optimized to obtain clinical grade MSCs with defined safety standards at large scale [14,15,31,[41][42][43][44][45][46]. To achieve this, strategies for preparation of the required quantities of cells, have to be implemented, which allow the undifferentiated state to be preserved or direct stem cell differentiation into the desired lineage.…”