Fig. 1.

Microglia depletion suppresses EAU. C57BL/6 mice were fed PLX5622 (PLX), a Csf1r antagonist, or control diet starting 7 d before IRBP immunization (day 0). (A) Retinas were whole-mounted and stained for P2ry12 in naive mice and mice fed PLX for 3 and 7 d. Z-stack orthogonal view images of the entire thickness (including the inner and the outer plexiform layers and the ganglion cell layer) of the midperipheral retina were taken by confocal microscopy. Representative images from three to four retinas per time point are shown. (Scale bars: 100 μm.) (B) Time course of EAU clinical scores (n = 7 mice per group) by fundus observation. Scores were graded in a blinded manner on a scale between 0 and 4 in half-point increments as described previously. Briefly, trace chorioretinal lesions and minimal vasculitis were scored as 0.5. Mild vasculitis with small focal chorioretinal lesions (≤5) were scored as 1. Severe vasculitis with multiple chorioretinal lesions (>5) were scored as 2. A pattern of a linear chorioretinal lesion, subretinal neovascularization, and hemorrhage were scored as 3. Inflammation with large retinal detachment and severe hemorrhages were scored as 4. (C) Representative retinal fundus images indicating a score of 1.5 in control immunized mice (vasculitis and chorioretinal lesions) but absent in PLX-treated mice on day 21. (D) Histopathological EAU score on day 21 (n = 5 mice per group) were graded in a blinded manner on a scale between 0 and 4 in half-point increments as described previously. Briefly, focal nongranulomatous, monocytic infiltration in the choroid, ciliary body, and retina were scored as 0.5. Retinal perivascular infiltration and monocytic infiltration in the vitreous were scored as 1. Granuloma formation in the uvea and retina together with the presence of occluded retinal vasculitis, photoreceptor folds, serous retinal detachment, and loss of photoreceptors, were scored as 2. In addition, the formation of granulomas at the level of the retinal pigment epithelium and the development of subretinal neovascularization were scored as 3 and 4 according to the number and the size of the lesions. (E) Representative histopathological (H&E staining) images indicating a score of 1.5 (photoreceptor folds, vitreous cells, and retinal granulomas) in control mice and 0 (no inflammation) in PLX-treated mice. The average of scores from both eyes was determined as the score of the animal. (Scale bars: 100 μm.) (B and D) Mann–Whitney U test. Data are expressed as mean ± SEM. *P < 0.05; **P < 0.01; ***P < 0.001.