Cytokine profiles and their relevance to human transplantation
The aim of this project was to develop an in vitro functional assay for the prediction of allograft rejection following renal transplantation. This assay was also used to study acute GVHD following identical sibling bone marrow transplantation. Lymphocyte cytokine profiles were measured by ELISA (protein secretion) and flow cytometry (cytokine expression) following mitogen stimulation and MLR. In normal individuals, considerable inter-individual variations were found in both protein secretion (IL-2, IL- 4, IL-10 and IFN-γ) and cytokine expression (IL-2 and IFN-γ). Strong relationships were found between IL-2 protein and expression, IL-2 and IL-10 protein, and IL-10 and IFN-γ protein secretion. Analysis of cytokine gene polymorphisms showed no correlation between IFN-y protein secretion, frequency or gene polymorphism. Pre-transplant MLRs were set up between renal transplant patient/donors pairs and cytokine protein secretion (IL-2, IL-4, IL-6, IL-10 and IFN-γ) measured by ELISA. Analysis was performed to ascertain predictive factors of allograft rejection. Inter-individual variations were found for all cytokine profiles. Significant correlations were found between individual cytokine protein profiles including IL-10 and IFN-γ. In addition, a correlation was found between HLA-DR mismatching and both IL-10 and IFN-γ protein secreted in the MLR. Primary univariate analysis revealed that HLA and HLA-DR mismatching, female donor sex, MLR-stimulated IL-10, MLR-stimulated IFN-γ and spontaneous IL-4 secretion were associated with an increased risk of rejection. Multivariate analysis showed the strongest correlations for predicting risk of rejection to be female donor sex, HLA mismatching and MLR-stimulated IL-l 0 secretion. A combination of high HLA mismatching and high IL-l 0 secretion in MLR gave the highest risk of rejection (RR=25.5). Finally, cytokine secretion decreased when measured post-transplant. Prediction of graft survival could not be analysed due to the low number (n=6) of patients that suffered graft failure in the group. Cytokine protein secretion (IL-2, IL-4, IL-10 and IFN-γ) in MLR was also studied for prediction of GVHD after bone marrow transplantation. There was a very low MLR response by all BMT pairs, therefore analysis could not be performed.