الفهرس يوجد فقط 14 صفحة متاحة للعرض العام
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المستخلص
Accumulating evidence suggests that a population of professional regulatory T cells (enriched within CD4+ CD25+ T cells) limits immune responsiveness exist in rodents and healthy human subjects. However their existence and role in renal transplant graft acceptor recipients is still unclear. The aim of this study was to develop ex-vivo assays to evaluate the ability of human T cells to regulate responses to allo-antigens presented by donor cells.
Initially peripheral blood samples from healthy volunteers were analysed to establish the assays. The proliferative responses of CD4+ T cells in the presence and absence of CD4+ CD25+ T cells among enriched CD4+ T cells and from whole PBMCs were assessed. First, tritiated thymidine incorporation into dividing cells, which provides an estimate of the total number of dividing cells in mixed lymphocyte reaction (MLR), and second, labelling of the responding population with carboxy fluorescein diacetate succinimidyl ester (CFSE), a fluorescent dye, that stains intra-cellular proteins, and partitioned equally between daughter cells at each cell division, were examined. Removal of CD4+ CD25+ T cells resulted in an increase in the proliferative response detected by both methods. The CFSE assay offered advantages as it provided additional information about the behaviour of defined subpopulations of leukocytes.
Twenty living related, renal transplant recipients were randomly selected for the study. They were 11 males and 9 females. All the recipients enjoyed stable graft function 5-16 years from the date of transplantation to the time of the assay. There was no significant difference between the recipients and their healthy donors with regard to the phenotypic analysis of CD25 and CD45RO expression in CD4+ T cells. Functional analysis of T regulatory cells by our CFSE assay revealed further subdivision of the transplant recipeints into two groups. Group A (referred to as the potential regulatory group) and group B (referred to as the defective regulatory group).
However, there was no significant difference between the two groups with regard to the phenotypic analysis of CD25 and CD45RO expression by CD4+ T cells. Significant proliferation of the CD4+ T cells was achieved in response to challenge with donor antigens compared to that observed following third party stimulation after depletion of CD25+ T cells in group A (P=0.039) (Table 4.a). While there was no significant difference in group B samples in the response of CD4+ T cells to donor and third party control after depletion of CD25+ T cells (P=0.133) (Table 4.b).
CONCLUSIONS:
from our work, we have constructed an ex vivo assay (CFSE assay) that enabled us to study the function of long term renal transplant recipients T regulatory CD4+CD25+ cells in a clear way for the first time. Our results showed that CD4+ CD25+ T cells were able to suppress the already formed memory cells against donor allo-antigens.