Differential Development of Umbilical Cord-Derived Mesenchymal Stem Cells During Long-Term Maintenance in Fetal Bovine Serum-Supplemented Medium and Xeno- and Serum-Free Culture Medium

Umbilical cord-derived mesenchymal stem/stromal cells (UC-MSCs) are believed to have potential for the treatment of various diseases; thus, many scientists have investigated the molecular mechanisms underlying the function of UC-MSCs and, for example, the appropriate media for large-scale UC-MSC expansion to prepare cells for real-world application. In this study, we investigated the cellular morphology, proliferation capacity, surface markers, cellular senescence signals, clonogenic potential, trilineage differentiation capacity, and secreted factors of human primary UC-MSCs in long-term culture from passage 2 (P2) to passage 10 (P10) with either conventional fetal bovine serum (FBS)-supplemented medium or commercial xeno- and serum-free medium (StemMACS (TM)). We found that the cells cultured in both media had similar morphology and marker expression. However, the proliferation kinetics as measured by the cell population doubling time differed in a passage (P2-P10)-dependent manner between the cells cultured in the two media; sustainable growth was observed in cells maintained in xeno- and serum-free medium. Moreover, significant differences in cellular senescence signals were observed, with more aging cells in the cell population cultured in FBS-containing medium. Colony numbers and the day that the first colony appeared were similar; however, UC-MSC colony sizes were smaller when cultured in FBS-containing medium. In addition, the multidifferentiation potential of UC-MSCs cultured in xeno- and serum-free StemMACS medium was maintained during long-term culture, but this potential was lost for adipogenic differentiation at P9. Moreover, secreted epidermal growth factor and vascular endothelial growth factor (VEGF)-A were detected in the conditioned media from UC-MSCs, whereas platelet-derived growth factor was not. Similar expression of these factors was observed in conditioned media of UC-MSCs cultured in StemMACS, but the VEGF level was higher in young UC-MSCs (P6) than in aged UC-MSCs cultured in FBS-supplemented Dulbecco's modified Eagle's medium/F12. Thus, StemMACS is better for UC-MSC expansion than conventional FBS-supplemented culture medium, especially when culturing UC-MSCs for real-world applications.