Corneal endothelial cell inhibition of t cell activation

Purpose. Further study mechanisms of corneal endothelial cell inhibition of T cell activation. Methods. LEW rat corneal endothelial cells (CE cells) were grown to confluency in tissue culture plates. The murine Th2 clone E6 was used to measure effect of CE cells on IL-4, IL-5 and IL-6. The KZO mouse T cell hybridoma was used to measure IL-2. This hybridoma was also transfected with a plasmid containing bacterial lacZ reporter gene driven by three tandemly repeated NF-AT binding sites of IL-2 gene. T cells were stimulated with or without the prescence of a CE cell monolayer. Cell culture supernatant was harvested and cytokine production measured by ELISA. For β-gal activaty, KZO cells were loaded with fluorogenic β-galactosidase substrate FDG and analyzed by FACS. Viable cells were counted by trypan blue exclusion. Results. CE cells inhibit IL-4 production, but not IL-5 or IL-6, by E6 clone. CE cells inhibit IL-2 production of KZO cells stimulated by antigen and ARC, but not by PMA plus lonomycin. CE cells do not block NFAT driven β-gal activation induced by PMA plus lonomycin, but by antigen plus APC. Solid phase anti-CD3 mAb alone can stimulate KZO cells to make IL-2, which is also inhibited by CE cells. CTLA4-lg does not block IL -2 production by KZO cells stimulated by antigen plus APC. CE cells do not consume IL-4 or IL-2. CE cells also inhibit antigen induced hybridoma apoptosis. Conclusions. These results show that CE cells inhibit IL-2 production through early steps of TCR pathway.CE cell effect is independent of the presence of APC and does not on act costimulation. CE cells do not block IL5 and IL-6 production. CE cells also block apoptosis of T cell hybridoma.