Cellular and cytokine profile of cord and adult blood mononuclear cells
Umbilical cord blood (CB) has been used as a source of haemopoietic stem cells in both related and unrelated bone marrow transplantation and in both settings there appears to be a reduced incidence of graft-versus-host disease (GVHD) when compared to bone marrow transplantation. The aim of this study was to perform a phenotypic and cellular analysis of both cord and adult mononuclear cells in vitro in order to identify any immunological characteristic that could account for the apparent reduced incidence of GVHD observed, in vivo. For this, the cell surface phenotype, allogeneic cellular responses, cytokine secretion and expression of co-stimulatory molecule CD40L was studied. The phenotypic analysis was carried out by determining the proportion of cells expressing the CD4, CD8, CD16, CD19, CD45RA and CD45RO markers using 3 colour flow cytometric analysis. The results showed that in cord blood mononuclear cells, there was an increased number of cells expressing CD19+ whereas the number of cells expressing CD8+ was decreased compared to adult blood mononuclear cells (ABMNCs). In contrast the number of cells expressing CD4+ and CD16+ was similar in both cord blood mononuclear cells (CBMNCs) and ABMNCs. Further phenotypic analysis confirmed that cord blood contained primarily 'unprimed' T cells expressing CD45RA, while adult peripheral blood lymphocytes express mainly the CD45R 'memory' phenotype. The primary and secondary allogeneic responses of cord and adult mononuclear cells were assessed using the standard mixed lymphocyte reaction (MLR) and the primed lymphocyte test (PLT), respectively. These results showed that there was no significant difference in primary MLR responses of CBMNCs and ABMNCs. In contrast, CBMNCs had a decreased ability to mount a secondary proliferative response compared to ABMNCs. However this impaired ability to respond could be overcome by the addition of exogenous IL-2 to the cultures. The secretion of cytokines such as IL-2, IL-4, γIFN and TNF-α and the expression of CD40L on PMA- and ionomycin-activated CBMNCs and ABMNCs was also analysed by 3 colour flow cytometry. These results showed that upon activation CBMNCs produced reduced levels of cytokines and had reduced expression of CD40L. It is likely that the reduced cytokine production and CD40L expression observed is due to the predominance of CD45RA+ cells in CB compared to AB. In summary, this study has shown that CB and AB have different phenotypic and functional characteristics in vitro which may well explain the reduced incidence of GVHD observed in cord blood transplantation.