Novel approach for effective isolation of mesenchymal stem cells from human umbilical cord blood

Introduction: Human umbilical cord blood (HUCB) has been successfully used in clinical allogenic graft transplantation of haematopoietic stem cells (HSCs). However, HUCB has been poorly characterized as a source of Mesenchymal stem cells (MSCs). The aim of this study was to establish HUCB as a source of MSCs using a novel clot spot method. Method:HUCB was collected (n=9) from candidates consenting women, who underwent elective caesarean section. HUCB clot was meticulously explanted in MesenCult basal medium and incubated in 5%CO2 incubator at 37°C. Qualitative and quantitative immunophenotyping of cells was achieved using fluorescein isothiocyanate (FITC) labelled antibodies reactive with CD34, CD45, CD29, CD44, CD73 and CD105 antigen markers. MSCs were plated in neurogenic and adipogenic differentiation media. Immunocytochemistry was used for embryogenic markers SOX2, Olig4 and FABP-4 identification. Results: MSC cultures using the clot spot method showed morphological changes inkeeping with cell lineage differentiation within two weeks of culture with a three fold increase in number, compared with other methods. Cells were negative forHSC markers (CD34, D45), but strongly positive for MSCmarkers CD29 (97%), CD44, CD73 (95%) and CD105 (97%). MSCs expressed the neural SOX2, Olig4 and adipogenic FABP-4 markers. Conclusion: HUCB is a good source of MSCs for transplantation, using this novel technique