PURPOSE: To determine whether
retinal pigment epithelial (RPE) cells can inhibit in vitro
T-cell activation during
inflammatory conditions. METHODS: Primary cultured RPE cells were established from normal
C57BL/6 mice. Target bystander
T cells were established from normal
splenic T cells with anti-CD3
antibodies.
T-cell activation was assessed for proliferation by both examining [(3)H]-
thymidine incorporation and the production of
interferon (
IFN)gamma or IL-17, as determined by
ELISA. Expression of programed
cell death 1 ligand 1 (PD-L1) on RPE or recombinant
mouse IFNgamma-pretreated RPE cells was evaluated using
oligonucleotide microarray,
RT-PCR,
immune staining, and
flow cytometry. Expression of programed
cell death 1 (
PD-1)(+) on target
T cells was evaluated by
flow cytometry. Anti-mouse PD-L1 or PD-L2
neutralizing antibodies or target
T cells from
PD-1 knockout donors were used for the
assay. RESULTS: IFNgamma-pretreated RPE greatly suppressed
activation of bystander
T cells, especially the IFNgamma production by the target
T cells (
Th1 cells, but not
Th17 cells) via direct cell contact. By examining cell surface candidate molecules, IFNgamma-pretreated RPE expressed much higher levels of PD-L1 compared with the control nontreated RPE. Although primary RPE did not express the costimulatory molecule, expression of the molecule was induced on the surface of IFNgamma-pretreated RPE. PD-L1(+) RPE in the presence of IFNgamma selectively suppressed
PD-1(+)
T-cell activation. IFNgamma-pretreated RPE in the presence of anti-PD-L1
neutralizing antibodies, but not anti-PD-L2, failed to suppress
T-cell production of IFNgamma. In addition, these RPE cells failed to suppress the production of IFNgamma by
CD4(+)
T cells from
PD-1 null donors. CONCLUSIONS: Suppression of
T-cell activation was obtained in cultures only when RPE expressed negative costimulators. Therefore, the authors propose that in vitro,
Th1 cytokine-exposed ocular resident cells can express this molecule and it is this expression that causes the suppression of the bystander Th1-type cells.
