The Anterior Chamber Of Foxp3 ~ + Treg Cells And Corneal Allograft Rejection Reaction Mechanism Of Experimental,

Purpose: To do researches on the relationship between Foxp3+Treg cells in anterior chamber and corneal allograft immune reaction,and the regulatory function of c-rel on Foxp3+Treg cell's differentiation after allograft.Methods: Building the PKP model.A total of 40 BALA/c mice received penetrating keratoplasty (PKP) were randomly divided into two groups—allogeneic PKP group(A) and syngeneic PKP(B). After surgery, the rejection time of allograft was observed by slit-lamp. The inflammatory cells infiltrated in allograft and iris-ciliary body were evaluated by HE staining on day 7 and 14 in two groups;CD4+T cells,IL-17+T cells and Foxp3+Treg cells in allgrafts and iris-ciliary body were seen by immunofluorescence staining on day 7 and 14 in two groups;The cytokines of IL-17 in CD4+T cells and their transcription factors of T-bet(Foxp3) as well as the mRNA representation level of c-rel in NF-kB were detected by Real-time RT-PCR on day 7 and 14 after surgery.The transcription factors of T-bet(Foxp3) in allgrafts and iris-ciliary body and the level of protein express of IL-17 were seen by Western Blot in two groups on day 7 and 14.CD4+T cells,CD25+T cells and CD4+CD25+Foxp3+Treg cells were seen by flow cytometry in two groups on day 14.Results: The rejection time of allograft in A group averaged 14.0±1.15 days. There was no rejection in group B. Inflammatory cells infiltrated corneal limbus on day 7 and some inflammatory cells adhered to corneal endothelium in these two groups. A great number of inflammatory cells accompanied by neovascularization were observed in group A.On day 7,IL-17 and Foxp3 representation increased significantly by immunofluorescence in group A&B.On day 14,IL-17 and Foxp3 express was high in group A.IL-17 and Foxp3 express stay the same as day 7 in gourp B.On day 7 and 14,Foxp3 mRNA,CD25mRNA,and c-rel mRNA increased greatly in corneal of allogeneic PKP group A presented by real-time PT-PCR,while Foxp3mRNA,CD25mRNA,and c-rel mRNA increased only on day 7 but decreased on day 14 in group B.On day 7 in group A,IL-17mRNA express increased significantly but decreased on day 14;IL-17mRNA remained low in group B.Analysing the rate of Foxp3mRNA/ IL-17mRNA we knew that on day 7 it was much higher in group B than in group A.The changing tendency of c-relmRNA was the same to the that of Foxp3mRNA in each group and each time spot. On day 14 after Western Blot operation,Foxp3 expressed significantly in both group;IL-17 protein express totally increased on day 14 in group A while it remained low in group B;and the rate of Foxp3/IL-17 was higher in group B than in group A.Flow cytometry showed that on day 14 in group A CD4+CD25+Foxp3+Treg cells took 2.6% percentage of CD4+T cells,Foxp immunofluorescence intensity averaged 58.557;CD4+CD25+Foxp3+Treg cells in group B took 1.117% percentage of CD4+T cells with Foxp3 immunofluorescence intensity averaging 114.444.Conclusions: After corneal allograft,immunity microenvironment in camera oculi anterior greatly changed.This leads to a great number of Fopx3+Treg cells and IL-17+Th17 cells' infiltration in some parts of ocular tissue. Treg cells in different immunity microenvironment have different Foxp3 express level.The Treg cells which express Foxp3 highly have more powerful immunologic suppression function;while Th17 cells play an important role in the proinflammatory function after surgery. Different expressing level of Foxp3 may decide the different function state of Treg cells. The relative amount between Treg cells and Th17 cells are more important in the process of corneal allograft rejection than the Treg cells themselves;The rate of Foxp3 protein/IL-17 decides whether the rejection would happen. c-rel plays an crucial role in the developmental regulation of Foxp3+Treg cells within anterior chamber.