Cryopreservation Of Immature Dendritic Cells Derived From Human Cord Blood

In the treatment of severe burns, early and effective coverage of the wound is essential. Allogeneic skin transplantation is a good method, but the rejection after transplantation limits its application and leads to strong rejection in short time. Immunosuppressive drugs can prolongate skin allografts survival and increase the potential risk for life-threatening infections. Therefore, it is a effective method to prolong the survival time of graft allogenic skin by application of imDC which contain allogenic antigen.Dendritic cells are the most important antigen-presenting cells in vivo and they play essential roles in both native immunity and adaptive immunity. There are a series of differentiate from immature dendritic cells(imDC) to maure dendritic cells(mDC) after pathogens or alloantigens'stimulation. Immature dendritic cells which are insufficient of MHC-II and costimulatory moleculars can induce allogenetic T cell anergy and antigen-specific immune tolerance. As we all know the allogenic skins for clinical use after cryopreservation can maintain their fresh states in vitro, however, the main problerm is how to keep the cells with long-term state of immaturation and activation for the clinical use as well as their homologized skins. This experiment investigated the biological properties of cryopreserved imDCs derived from human cord blood to explore a effective and safe preservative method of imDC for clinical application.In this study, immature dendritic cells were generated from human cord blood (CB) monocyte cultured with rhGM-CSF and rhIL-4 and we plus 10 % DMSO or 5% DMSO and 6% HES as protective solute respectly for cryopreservation. ImDCs were frozen in - 80℃refrigerator, then cryopreserved in - 196℃liquid nitrogen( for at lest one month),and thawed in 35~40℃at last, which were called as freeze-thawed immature DC. TBBR were observed by the fluorescence microscope and cell morphology was observed by scanning electronic microscope. DCs were analyzed by expression of maturate surface antigens(CD80 CD83 and CD86) by flow cytometry and allo-stimulating abilities in allgenetic mixed lymphocyte reaction.