New approach to isolate mesenchymal stem cells (MSC) from human umbilical cord blood

Human umbilical cord blood (HUCB) has been frequently used in clinical allogeneic haematopoietic stem cells (HSCs) transplantation. However, HUCB is poorly recognised as a rich source for mesenchymal stem cells (MSCs). The aim of this study, therefore, was to establish a new method for isolating large number of MSCs from the HUCB and proving that it is a good source of MSCs. HUCB samples were collected from women following their elective caesarean section. The new Clot Spot method was carried out by explanting HUCB samples into mesencult complete medium and maintained in 5% CO2 incubator at 37ºC. Confirmation for the presence of MSCs was established by quantitative and qualitative immunophenotyping of cells and using fluorescein isothiocyanate (FITC) attached to MSC phenotypic markers (CD29, CD73, CD44 and CD105). Haematopoietic antibodies (CD34, CD45) were used as negative control. MSCs differentiation was examined in neurogenic and adipogenic media. Immunocytochemistry for the embryogenic markers; Sex determining region Y-box 2 (SOX2), Oligodendrocyte (OLIG-4) and Faty acid binding protein-4 (FABP-4) was carried out. The conventional Rosset Sep method was carried out for comparison.MSC cultures using the Clot Spot method showed three-fold increase in proliferation, compared with conventional method. Cells expressed MSC markers CD29, CD73 and CD44 and CD105, but lacked expression of CD34 and CD45. Cells also expressed the neural SOX2, Olig4 and adipogenic FABP-4 markers following differentiation. In conclusion HUCB is a good source for MSCs on using this new technique.