Abstract
Purpose :
We have previously showed that ciliary neurotrophic factor (CNTF) suppresses expressions of rod and cone opsins as well as visual cycle enzymes (RPE65 isomerase and lecithin:retinol acyl transferase), thereby reducing the formation of the light sensitive visual pigments that mediate normal vision and photodamage-induced retinal degeneration. The purpose of this study was to test whether benzoic acid, which has been shown to promote CNTF expression in cell culture, and CNTF-deficiency can protect photoreceptors from light damage-induced necroptosis.
Methods :
Dark adapted wild-type (129S2/Sv strain) and Cntf-/- mice treated with different doses of sodium benzoate (NaB) were exposed to 15000 lux light to induce retinal photodamage. Expression of the necrosome proteins such as receptor interacting protein kinase 1 (RIPK1), RIPK3, and mixed lineage kinase domain-like protein (MLKL) in the retinas were analyzed by immunoblot and immunohistochemical analyses. Interaction of these necrosome proteins were determined by double-immunostaining and immunoprecipitation assays. Activation of the necrosomes was assessed by detecting phosphorylation of MLKL. Photoreceptor degeneration was evaluated by immunohistochemistry and immunoblot analysis of opsins and cone arrestin.
Results :
Intense light exposure induced a significant upregulation of both RIPK expression and MLKL phosphorylation in the WT mouse retinas. Interactions of the necrosome proteins were also enhanced in the WT retinas exposed intense light. Under the same light conditions, NaB treatment inhibited the RIPK upregulation and MLKL activation in a dose-dependent manner in WT retinas. Immunoblot and immunohistochemical analyses showed that intense light exposure caused degeneration of rods and cones in WT mice and NaB-treatment alleviated the light-induced degeneration of photoreceptors in WT. The same intense light exposure induced more greater activation of the necrosomes in the Cntf-/- retinas, as compared to WT mice. NaB-treatment, however, did not significantly inhibit RIPK3 upregulation, MLKL phosphorylation and retinal degeneration in the Cntf-/- mice exposed to the intense light.
Conclusions :
1) activation of the necrosomes contributes to light-induced retinal degeneration in both WT and Cntf-/- mice; 2) NaB inhibited photodamage-induced necrosome activation in the WT, but not in Cntf-/-, retinas.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.