Fig. 2
From: NLRX1 limits inflammatory neurodegeneration in the anterior visual pathway
Clinical disease and anterior visual pathway degeneration in WT and Nlrx1−/− active EAE mice. (A) WT and Nlrx1−/− mice were immunized with subcutaneous injections of MOG35 − 55 in complete Freund’s adjuvant (CFA) and intraperitoneal injection of pertussis toxin (PTX). Control WT and Nlrx1−/− mice received CFA and PTX without MOG35 − 55 (CFA-only controls). Mice were clinically monitored for (B) EAE onset and (C) EAE paralysis severity overtime (n = 22 WT, n = 23 Nlrx1−/−). EAE paralysis score data represents mean ± standard error of the mean (SEM). On day 42 post-immunization (PID 42) mice were euthanized, and eyes and optic nerves were collected for immunofluorescence staining (IF). (D) Retinas were dissected and flat-mounted for IHC and Brn3a + retinal ganglion cells (RGCs) were quantified in 12 regions of interests (ROIs) using a MATLAB semi-automated counting algorithm. Each RGC density/count data point corresponds to an individual mouse, showing the averaged counts from 12 regions in the retina (4 central, 4 middle, and 4 peripheral). (E) Representative IF images of Brn3a+ RGCs in flat-mounted retinas from CFA-only and EAE WT and Nlrx1−/− mice. (F) Average RGC density for EAE (n = 20 WT, n = 22 Nlrx1−/−) and control mice, (CFA-controls = 10, 5 WT and 5 Nlrx1−/−). (G) Representative IF images of Brn3a + RGCs and phosphorylated (Ser73) c-Jun (pc-Jun) expression in flat-mounted retinas from WT and Nlrx1−/− EAE mice. (H) Density quantification of total Brn3a + RGCs and pc-Jun + Brna3 + RGCs in WT and Nlrx1−/− EAE mice. (I) Correlation between pc-Jun and Brna3 intensity in RGCs from WT and Nlrx1−/− EAE mice. The mean intensity of each marker in a single RGC was measured in retina using Zeiss ZEN software. (J) Representative IF staining of wholemount retina from an Nlrx1−/− EAE mouse, demonstrating pc-JunlowBrn3ahigh RGCs (red arrows) and pc-JunhighBrn3alow RGCs (white arrows). (K) Representative IF images of neurofilament light chain (NFL) and (L) average NFL percent area positive in the optic nerve of EAE and control mice. RGC density and NFL percent area positive data are represented as mean ± SEM with statistical analysis by Student’s t-test or one-way ANOVA with Dunnett’s T3 multiple comparison test. Linear associations were determined by Pearson correlation analysis. *p < 0.05, ***p < 0.001, ****p < 0.0001