Introduction: For expansion of human mesenchymal stem cells (MSCs), autologous human serum is safer than fetal bovine serum in clinical situations. One of the problems with the use of autologous human serum is that its proliferative effect on MSCs varies widely between donors. The threefold goals of this study were: (1) to demonstrate an improved method for preparing human serum; (2) to identify growth factors predictive of proliferative potential; and (3) to identify a cytokine to promote MSC proliferation in human serum. Methods: Fresh blood was collected using a closed bag system containing glass beads. The bag was shaken at 20 °C for 30 minutes for rapid preparation, or kept stationary at 4 °C for 24 hours for slow preparation. Passage 0 synovial MSCs derived from four donors were cultured with 10 % conventional rapid preparation serum or modified slow preparation serum from four different donors. To perform the colony-forming unit assay, synovial MSCs were cultured in these serums. The protein expression profile in serum was analyzed using cytokine array. The candidate proteins were speculated from the correlation between the colony-forming ability and protein expression. As an evaluation of the candidate proteins, proliferation ability, surface marker phenotype and differentiation capability of synovial MSCs were examined. Results: Compared with rapid preparation serum, slow preparation serum resulted in a significantly higher total colony number and twofold higher expression levels of nine proteins (angiopoietin-1, BDNF, EGF, ENA-78, IGFBP-2, platelet-derived growth factor (PDGF)-AA, PDGF-AB/BB, RANTES and TfR). Colony number was positively correlated with PDGF-AA/AB concentrations. Exogenous PDGF-AA significantly promoted proliferation of synovial MSCs, whereas PDGF receptor (PDGFR) inhibitor decreased it. Addition of PDGFs or PDGFR inhibitor did not affect surface epitopes of synovial MSCs. Pretreatment with PDGFs or PDGFR inhibitor did not affect chondrogenic, adipogenic, or calcification potentials of synovial MSCs. Conclusion: Slow preparation serum contained higher concentrations of PDGF-AA/AB and increased the colony formation number of synovial MSCs. PDGF-AA/AB were indicators of the proliferative potential of human serum. Exogenous PDGF-AA increased proliferation of synovial MSCs without alteration of surface epitopes and differentiation potentials.
All Science Journal Classification (ASJC) codes
- Medicine (miscellaneous)
- Molecular Medicine
- Biochemistry, Genetics and Molecular Biology (miscellaneous)
- Cell Biology