The Study Of Umbilical Cord Blood Mesenchymal Stem Cells Supporting Hematopoiesis

Objective: Mesenchymal stem cell (MSC) is a kind of stem cells with self-renewal and multilineagedifferentiation potential. MSCs are adhesion and can be expanded in vitro. It has not been found withspecific cell surface antigen. It is generally believed that MSCs express the antigen of interstitial cell, butdo not express hematopoietic stem cell related surface antigens, MHC class II antigen and costimulatorymolecules. Studies of animal experiments and clinical trials showed that, hematopoietic stem celltransplantation combined with MSCs can shorten the engraftment of hematopoietic stem cells andneutrophil, platelet recovery time, and reduce the incidence and severity of graft rejection and graft versushost disease (GVHD), therefore MSCs has the research value and good application prospect inhematopoietic stem cell transplant field. The purpose of this study was to isolate, culture and identificationof human umbilical cord blood derived mesenchymal stem cells (UCBMSCs), and to analyze its supportcapability and mechanism of ex vivo expansion of cord blood CD34+cells. It will provide experimental andtheoretical basis for umbilical cord blood mesenchymal stem cells used in allogeneic hematopoietic stemcell transplantation.Methods:1. Density gradient centrifugation was used to isolate and in vitro amplify UCBMSCs;2. Todetect surface antigen of UCBMSCs by using flow cytometry and identify the ability of multilineagedifferentiation;3. UCBMSCs were co-cultured with hematopoietic stem cells, and then the proliferation ofhematopoietic stem cell and formation of colonies were observed;4. To detect the expression of adhesionmolecules and cytokines mRNA of UCBMSCs by using RT-PCR;5. ELISA was used to detect the cytokineof UCBMSCs culture supernatant.Results:1. The fusiform,and fibroblast like MSCs were successfully isolated from umbilical cordblood.2. UCBMSCs positively express CD44, CD29, CD90, CD166, CD73, CD105, CD13, CD59andHLA-ABC; negatively express CD45, CD34, CD14, CD31and HLA-DR.3. UCBMSCs can differentiateinto osteoblasts and adipocytes.4. The number of hematopoietic stem cells (HSCs) and colonies wereamplified in the group of only mesenchymal stem cells feeder layer, only adding exogenous cytokine andmesenchymal stem cells feeder layer and adding exogenous cytokines(co-culture group), but co-culturegroup was the highest. The difference was statistically significant.5. The mRNA expression of CD54,CD166, CD56, CD44, CD49d, SDF-1, CXCR4, CD106, IL-1α, IL-1β, IL-6, IL-7, IL-8, IL-11, M-CSF,GM-CSF, FLT3LG and LIF can be detected in UCBMSCs.6. GM-CSF, IL-7, IL-8, IL-11, SCF and SDF-1were detected in the culture supernatant of UCBMSCs.Conclusion: UCBMSCs as feeder layer can support the expansion of CD34+cells in vitro. Theexpression and secretion of cytokines and adhesion molecules plays an important role in the amplificationof CD34+cells in vitro. This conclusion provides experimental basis for the clinical application ofUCBMSCs.