Immunophenotypic Study of the Cord Blood CD34+ Progenitor Stem Cells-Derived Dendritic Cells

Abstract

Background: Dendritic cells (DCs) are bone marrow-derived cells of both lymphoid and myeloid stem cellorigin that populate all lymphoid organs including the thymus, spleen, and lymph nodes, as well asnearly all no lymphoid tissues and organs. Although DCs are a moderately diverse set of cells, theyall have potent antigen-presenting capacity for stimulating naive, memory, and effector T cells. DCsare receiving increasing scientific and clinical interest due to their key role in anti-cancer hostresponses and potential use as biological adjuvants in tumour vaccines, as well as their involvementin the immunobiology of tolerance and autoimmunity.Objectives: To generate the dendritic cells (DCs) in vitro from purified cord blood stems progenitor cells anddetects the growth curve of cells and analyze the DC in vitro differentiation pathway. Also toinvestigate the immunophenotypic CD markers morphology of the surface DC cells and to describethe effect of different growth factors on their expression.Materials & Methods: For isolation and establishment in tissue culture of human DC, the cord blood wasobtained from placenta of newly vaginaly delivered women, who were admitted in Al KadhmyiaTeaching Hospital. The cells were cultured in complete RPMI growth medium supplemented withgrowth factors GM-CSF+IL-4 for seven days then the key surface CD markers were detected by usingthe immunocytochemistry technique.Results: The use of the CD34 monoclonal antibody in combination with CD45 monoclonal antibody hasincreased the opportunity of obtaining a reasonable amount of purified stem cells. Furthermore, thegrowth factors (GM-CSF+IL-4) that were supplemented to the complete medium played an importantrole in the differentiation of the CD34+ stem cells toward the DC cells. The growth factors GM-CSFand IL-4 when used in combination had made the difference and the rhythm of differentiation ,thecount of cells, resolution of results, shape and size of cells, and the state of DC cells better than usingeach of them alone. The IL-4 when used in combination with GM-CSF act as inhibitory factor for thegranulocyte and cells other than DC and at the same time has the capability to keep the DC cells inimmature state. The kinetics of DC development in cord blood cultures was also determined. Theseresults indicated that, the optimal culture of the cord blood-derived DC was 7-9 days. By using theimmunocytochemistry technique, the key CD markers of DC cells has been revealed since the day 6 ofculture. The immature DC cells were characterised with the positive expression of CD1a, CD11b andCD11c while the CD14 was negative.Conclusion: Isolation, purification and mobilization of stem cells obtained were reliable and confirmatory, andindicate that the positive differentiation of the cord blood CD34+ stem cells toward the DC cells wasachieved.Key words; CD34 cord blood stem cells, immunophenotyping, dendritic cells