Isolation and Culture of Mesenchymal Stem Cells from Human Umbilical Cord Blood in Vitro

ObjectiveTo investigate the feasibility and optimal condition of isolation,purification and expansion of mesenchymal stem cells(MSCs) derived from human umbilical cord blood in vtro.MethodsHuman umbilical cord blood(HUCB) was collected from full term deliveries scheduled,all samples were obtained sterilely with 20 U/ml preservative free heparin.The cord mononuclear cells were isolated with lymphocyte separation medium(density 1.077 g/ml),purified and expanded with Mesencult~(TM) medium and acidic environment to produce adherent layer.The surface antigen expression of MSCs was detected with flow cytometry.ResultsThe HUCB-derived mononuclear cells,when seeded in specific medium,gave rise to adherent cells,which exhibited either an osteoclast or mesenchymal-like phenotype.After passage 3,these cells were able to be purified and expanded.6.6×10~5 primary MSCs reached a number of 9.9×10~8 after 10 expanded passage.Flow cytometry showed that MSCs did not express antigens CD_(34),CD_(11a) and CD_(11b),but express strongly CD_(29) and weakly CD_(71),which was identical to human bone marrow-derived MSCs.ConclusionMSCs in HUCB can be cultured and expanded in vitro,and could be a source of stem cells for experimental and clinical application.