In Vitro Expansion of Human Mesenchymal Stem Cells: Choice of Serum Is a Determinant of Cell Proliferation, Differentiation, Gene Expression, and Transcriptome Stability

Abstract

Human bone marrow mesenchymal stem cells (hMSCs) represent an appealing source of adult stem cells for cell therapy and tissue engineering, as they are easily obtained and expanded while maintaining their multilineage differentiation potential. All current protocols for in vitro culture of hMSCs include fetal bovine serum (FBS) as nutritional supplement. FBS is an undesirable additive to cells that are expanded for therapeutic purposes in humans because the use of FBS carries the risk of transmitting viral and prion diseases and proteins that may initiate xenogeneic immune responses. In the present study, we have therefore investigated if autologous serum (AS) or allogeneic human serum (alloHS) could replace FBS for the expansion of hMSCs in vitro. We discovered that the choice of serum affected hMSCs at several different levels.First, hMSCs in AS proliferated markedly faster than hMSCs in FBS, whereas use of alloHS resulted in hMSC growth arrest and death. Second, hMSCs in FBS differentiated more rapidly toward mesenchymal lineages compared with hMSCs in AS. Interestingly, genome-wide microarray analysis identified several transcripts involved in cell cycle and differentiation that were differentially regulated between hMSCs in FBS and AS. Finally, several transcripts, including some involved in cell cycle inhibition, were upregulated in hMSCs in FBS at a late passage, whereas the hMSC transcriptome in AS was remarkably stable. Thus, hMSCs may be expanded rapidly and with stable gene expression in AS in the absence of growth factors, whereas FBS induces a more differentiated and less stable transcriptional profile. Stem Cells 2005;23:1357-1366

Cranioplasty with Adipose-Derived Stem Cells, Beta-Tricalcium Phosphate Granules and Supporting Mesh: Six-Year Clinical Follow-Up Results

“…The in vitro, in vivo and clinical bone‐forming capacity of ASCs in combination with various scaffold materials have been reported by many authors 14, 20, 35, 36, 37, 38, 39, 40, 41. In this series, the flow cytometric characterization of the ASCs expanded in autologous serum corresponded to previously published results for ASCs 42, 43. According to flow cytometry, and alkaline phosphatase staining results, the cells were of mesenchymal origin and had a capacity to differentiate into the osteoblastic lineage.…”

Inactivated human platelet lysate with psoralen: a new perspective for mesenchymal stromal cell production in Good Manufacturing Practice conditions

“…Serum-free medium is unable to promote MSC expansion unless several recombinant human growth factors (GF) such as platelet-derived growth factor (PDGF), fibroblast growth factor (FGF), transforming growth factor (TGF)-β, and epidermal growth factor (EGF) are added to the medium. However, 1) defining the optimal amount of GF is difficult, 2) only a few GFs are licensed for therapeutic use, 3) recombinant Human-GFs are expensive, and 4) isolated GF cannot replace the different physiological functions of FBS (15)(16)(17).…”

Concise Review: Culture Mediated Changes in Fate and/or Potency of Stem Cells

“…Genome-wide transcriptome and promoter methylation studies have shown that the expressed gene profile changes on culturing MSC in vitro with increased expression of, among others, cytoskeleton proteins and downregulation of cell cycle inhibitory genes [36], which is accompanied by progressive epigenetic changes [37]. There is also evidence that relatively minor changes to the culture conditions used to expand MSC, including culture at different oxygen tensions, use of fetal calf serum versus autologous serum, use of platelet lysate, or addition of growth factors (e.g., FGF2 or platelet-derived growth factor [PDGF]), affect MSC proliferation ability as well as the rate and degree of differentiation to the osteogenic, adipogenic, and chondrogenic lineages [36,[38][39][40][41]. These functional changes are associated with differences in expressed gene profile and epigenomic state of the cells [36,[38][39][40][41].…”