Monitoring the biology stability of human umbilical cord-derived mesenchymal stem cells during long-term culture in serum-free medium

Mesenchymal stem cells (MSCs) are multipotent adult stem cells that have an immunosuppressive effect. The biological stability of MSCs in serum-free medium during long-term culture in vitro has not been elucidated clearly. The morphology, immunophenotype and multi-lineage potential were analyzed at passages 3, 5, 10, 15, 20, and 25 (P-3, P-5, P-10, P-15, P-20, and P-25, respectively). The cell cycle distribution, apoptosis, and karyotype of human umbilical cord-derived (hUC)-MSCs were analyzed at P-3, P-5, P-10, P-15, P-20, and P-25. From P-3 to P-25, the three defining biological properties of hUC-MSCs [adherence to plastic, specific surface antigen expression, multipotent differentiation potential] met the standards proposed by the International Society for Cellular Therapy for definition of MSCs. The cell cycle distribution analysis at the P-25 showed that the percentage of cells at G0/G1 was increased, compared with the cells at P-3 (P < 0.05). Cells at P-25 displayed an increase in the apoptosis rate (to 183 %), compared to those at P-3 (P < 0.01). Within subculture generations 3-20 (P-3-P-20), the differences between the cell apoptotic rates were not statistically significant (P > 0.05). There were no detectable chromosome eliminations, displacements, or chromosomal imbalances, as assessed by the karyotyping guidelines of the International System for Human Cytogenetic Nomenclature (ISCN, 2009). Long-term culture affects the biological stability of MSCs in serum-free MesenCult-XF medium. MSCs can be expanded up to the 25th passage without chromosomal changes by G-band. The best biological activity period and stability appeared between the third to 20th generations.