Alterations of peripheral CD4^(+)CD25^(+)Foxp3^(+)T regulatory cells in mice with STZ-induced diabetes

Complications arising from abnormal immune responses are the major causes of mortality and morbidity in diabetic patients.CD4^(+)CD25^(+)T regulatory cells(Tregs)play pivotal roles in controlling immune homeostasis,immunity and tolerance.The effect of hyperglycemia on CD4^(+)CD25^(+)Tregs has not yet been addressed.Here we used streptozotocin(STZ)-induced diabetic mice to study the effects of long-term hyperglycemia on CD4^(+)CD25^(+)Tregs in vivo.Four months after the onset of diabetes,the frequency of CD4^(+)CD25^(+)Foxp3^(+)T regulatory cells was significantly elevated in the spleen,peripheral blood lymphocytes(PBLs),peripheral lymph nodes(pLNs)and mesenteric LNs(mLNs).CD4^(+)CD25^(+)Tregs obtained from mice with diabetes displayed defective immunosuppressive functions and an activated/memory phenotype.Insulin administration rescued these changes in the CD4^(+)CD25^(+)Tregs of diabetic mice.The percentage of thymic CD4^(+)CD25^(+)naturally occurring Tregs(nTregs)and peripheral CD41Helios1Foxp31 nTregs were markedly enhanced in diabetic mice,indicating that thymic output contributed to the increased frequency of peripheral CD4^(+)CD25^(+)Tregs in diabetic mice.In an in vitro assay in which Tregs were induced fromCD4^(+)CD25^(+)T cells by transforming growth factor(TGF)-b,high glucose enhanced the efficiency of CD4^(+)CD25^(+)Foxp3^(+)T inducible Tregs(iTregs)induction.In addition,CD4^(+)CD25^(+)T cells from diabetic mice were more susceptible to CD4^(+)CD25^(+)Foxp3^(+)TiTreg differentiation than those cells from control mice.These data,together with the enhanced frequency of CD4^(+)CD25^(+)Foxp3^(+)T iTregs in the periphery of mice with diabetes,indicate that enhanced CD4^(+)CD25^(+)Foxp3^(+)T iTreg induction also contributes to a peripheral increase in CD4^(+)CD25^(+)Tregs in diabetic mice.Our data show that hyperglycemia may alter the frequency of CD4^(+)CD25^(+)Foxp3^(+)T Tregs in mice,which may result in late-state immune dysfunction in patients with diabetes.

Biological features of intrahepatic CD4^(+)CD25^(+)T cells in the naturally tolerance of rat liver transplantation

The biological features of intrahepatic CD4^(+)CD25^(+)T regulatory cells in the naturally tolerance of rat liver transplantation were explored.Orthotopic liver transplan-tation was performed in two allogeneic rat strain combina-tions,one with fatal immunosuppression despite a complete major histocompatibility complex mismatch.The subjects were divided into three groups according to different donors and recipients[Tolerance group:LEW-to-DA;Rejection group:DA-to-LEW;Syngegnic group(control group):DA-to-DA].The proportion of intrahepatic CD4^(+)CD25^(+)T cells from three groups was determined by flow cytometry(FCM)in different time.The intrahepaitc CD4^(+)CD25^(+)T cells were isolated by magnetic activated cell sorting(MACS)method and identified by FCM.The Foxp3 mRNA was detected by reverse transcriptase polymerase chain reaction(RT-PCR).And their suppression on the proliferation of CD4^(+)CD25^(-)T effector cells was analyzed by cell proliferation assay in vitro.Beginning immediately after transplantation,the proportion of Treg cells increased over time in both allogeneic groups but was significantly greater in the Rejection group.The pro-portion of Treg cells declined after day 5,and such reduction was more dramatic in the Rejection group than in the Tole-rance group.Animals in the Tolerance group showed a second increase in the proportion after day 14.Intrahepatic CD4^(+)CD25^(+)T cells isolated from spontaneous tolerance models inhibited the proliferation of mixed lymphocyte reaction.The purity of CD4^(+)CD25^(+)Tcells sorted by MACS was 86%–93%.The CD4^(+)CD25^(+)T cells could specifically express the Foxp3 gene compared with CD4^(+)CD25^(-)T cells.In vitro,the spleen cells from LEW rats can irritate the proliferation of CD4^(+)CD25^(+)T cells more obviously than the syngegnic spleen cells.CD4^(+)CD25^(+)Tr cells could suppress the proliferation of CD4^(+)CD25^(-)T cells,but the inhibition was reversed by exo-genous IL-2(200 U/mL).The CD4^(+)CD25^(+)T regulatory cells specifically express the Foxp3 gene,which may play animpor-tant role in the induction of liver transplantation tolerance by suppressing the reaction of effective T cells.