The Change Of Peripheral Regulatory T Cells In Liver Transplant Recipients And Its Clinical Significance

ObjectiveThe ultimate goal of solid organ transplantation(Tx) is to achieve immune tolerance,which can be defined as the maintenance of normal graft function without features of graft rejection and without the need for immunosuppressive drugs.At present,this goal is almost not achievable for the vast majority of transplant recipients. Nonetheless,in some cases immunosuppression is withdrawn and patients do not reject their grafts,especially in liver transplantation(OLT).Immune regulatory T cell(Treg) play a crucial role in inducing and maintaining allograft tolerance.In this study,we examined the impact of tacrolimus(FK506) regimens on induction of CD4~+CD25~+FOXP3~+ Tregs in the peripheral blood of stable liver transplantat recipients.MethodWe divided 37 liver transplant recipients with stable course who receiving FK506 therapy into two groups according to the blood levels of FK506 at the time of the study: 30 patients with low concentrations of FK506(low FK506:FK506＜6 ng/mL) and another 7 patients with high levels(high FK506:FK506≥6ng/mL).Thirty age-matched healthy subjects(HS) were used as normal controls.Heparinized peripheral blood was obtained,and peripheral blood monouclear cells(PBMC) were isolated by density centrifugation over Ficoll gradient from the blood samples.Then,the expression CD4, CD25,and FoxP3 in peripheral blood T cell were examined by flow cytometry.Flow cytometry was performed with a FACSCalibur(BD Biosciences) instrument and the data were analysed by using Cell Quest software.The percentage of CD4~+CD25~+ and CD4~+CD25~+FOXP3~+ T cells was determined using three-color flow cytometry.The absolute numbers of CD4~+CD25~+ and CD4~+CD25~+FOXP3~+ T cells were calculated based on the peripheral blood lymphocyte count at time of sampling.Differences in the level of CD4~+CD25~+ and CD4~+CD25~+FOXP3~+ Tregs among the different groups of patients were analyzed.The correlation between Tregs level and the blood level of FK506 was analysed by Spearman's correlation.ResultBoth the absolute numbers and percentages of CD4~+CD25~+ and CD4~+CD25~+FOXP3~+ Tregs in high FK506 recipients were decreased markedly compared with healthy subjects(P＜0.01) or low FK506 recipients(P＜0.05). However,there was slight but no statistically significant decrease in Tregs level between low FK506 recipients and healthy subjects(P＞0.05).Furthermore,the percentages of CD4~+CD25~+ and CD4~+FOXP3~+ Treg cells in stable liver transplant recipients correlated inversely to the plasma level of FK506([r=-0.479,P=0.003]and[r=-0.648,P＜0.001]).However,the ratios of these Tregs to CD4~+ T cells were no significant difference.There was no significant difference of Tregs level as well as their ratios to CD4~+ T cell between hepatoma recipients and non-hepatoma recipients(P＞0.05).And there was no significant difference between the two immunosuppression protocol receiving either FK506 alone(n=30) or FK506 plus with MMF(n=7) else(P＞0.05).Conclusion1.The absolute numbers and the percentages of blood CD4~+CD25~+ and CD4~+CD25~+Foxp3~+ Tregs were decreased in stable liver transplant recipients who receive FK506 therapy when compared with healthy subjects.2.The absolute numbers and the percentages of blood CD4~+CD25~+ and CD4~+CD25~+Foxp3~+ Tregs were correlated with the plasma level of FK506.It showed a steady decrease in association with increasing blood levels of FK506.The percentage of blood Treg levels correlated inversely to the plasma level of FK506.3.The ratios of those Tregs to CD4~+ T cells decreased trend when compared with the healthy subjects or low FK506 recipients,but there was no statistical significance.