Research On T Cell Synergistic Activation By Human B7-2 And Tumor Specitific TCR

Objective Research on B7-2 gene modified tumor cells on lymphocytes recruitment and activation, as well as B7-2 gene modified tumor cells combined with tumor specitific T cell receptor(TCR) modified lymphocytes synergistic reaction on anti-tumor . Methods To clone B7-2 gene , construct in eucaryotic expression vector, and then transfer to tumor cells Bel-7402 by liposome transfection reagent,which expression hereafter was detected by RT-PCR and flow cytometry(FCM). To label Bel-7402 cells transfected by B7-2 gene with Carboxyfluorescein Diacetate Succinimidyl Ester (CFSE), and then co-culture with PBMC, afterward detect the apoptosis of CFSE positive cells. Recombinant adenovirus Ad-TCRVα12-2/Vβ7.1 carrying tumor specitific T cell receptor gene transfected peripheral blood mononuclear cells(PBMC) with the pass of infection, co-cultured with Bel-7402 cells transfected by B7-2 gene, and follow detect the apoptosis of Bel-7402 cells by FCM.Results The expression of B7-2 gene was up-regulated after the B7-2 gene transfected Bel-7402 cells. B7-2 gene modified Bel-7402 co-cultured with PBMC for 4h, and the FCM approach apoptosis rate was 12.63%, which was significantly increased compared with the control group (7.53%)(p <0.01). Ad-TCRVα12-2/Vβ7.1 modified PBMC and Bel-7402 cells co-cultured for 48h, B7-2 and TCRVα12-2/Vβ7.1 co-transfected group, B7-2 single tansfected group, TCRVα12-2/Vβ7.1 single tansfected group,and non-transfected group, respectively the 4 groups positive rates(binding rates) of the Bel-7402 binding with PBMC were 24.26%, 15.53%, 10.13% and 5.43%, and the binding rates between the two adjacent groups were significantly different(p <0.01). Compared with the other groups, the binding rate of the B7-2 and TCRVα12-2/Vβ7.1 co-transfected group was the highest.The apoptosis rate of Bel-7402 binding with PBMC of B7-2 and TCRVα12-2/Vβ7.1 co-transfected group was 48.40% which were both sygnificantly increased compared with group of B7-2 single tansfected group(36.56%) and TCRVα12-2/Vβ7.1 single tansfected group(24.36%)(p<0.01). Furthermore, it could be observed and determined with Laser Confocal Microscopy that the B7-2 modified tumor cells enhanced T cells recruitment and activation.Conclutions B7-2 modified tumor cells can increase lymphocytes recruitment and enhance its activation, in addition, which combined with TCR modified lymphocytes can synergistically recruit T cells to focus on tumor cells and enhance T cells tumor-killing effect.