Targeted Blocking Tim-3 Regulation Of CD8~+T Cells Of Glioma Experimental Research

Objective:In vitro experimental observation,target-blocking immune checkpoint Tim-3 which regulated CD8~+T cell kills glioma cell.Then preliminary discussing key molecules and immunologic mechanism.Method:Separated and purified the CD8~+T cell from the peripheral blood by Magnetic activated cell sorting.The addition of CD3 and CD28 antibodies in the CD8~+T cell,makes its proliferation.The addition of IL-12 up regulates Tim-3 expression.the CD8~+T cell is divided into groups,the closed group join Tim-3 antibody,the homotype Ig G join the control group.The CD8~+T cell and glioma cell lines U87/U251 establish the coculture system for 72 h in vitro.Collecting the glioma cells,lymphocytes,the supernatant from the coculture system.CCK-8 detects two groups glioma cell viability.CFSE detects the multipcation capacity of the CD8~+T cell.Elisa measures the standard of IFN-γ and TGF-β.Flow cytometry tests the apoptosis of glioma cell by Annexin V-FITC/PIResult:The proliferation capacity of two groups CD8~+T cell is lower than the normality cultural.The proliferation capacity in the homotype Ig G control group is also lower than the closed group(P<0.05).The cell viability of two glioma cell lines in the homotype Ig G control groups is higher than the closed groups.The glioma cell apoptosis capacity of the homotype Ig G control group is higher too(P<0.05).the standard of IFN-γ in the homotype Ig G control group is lower,but the standard of TGF-β is the other way around(P<0.05).Conclusion:Experimental results show Tim-3 gives play to influential role in the glioma immune escape process.Targeting against it can remarkable suppress two glioma cell line(U87and U251)proliferation,renews the kill ability and quantity of the CD8~+T cell,regains the immune ability which could against tumor.