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.

Abstract

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™). 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.