Inhibition of HIF-1 Signaling Reduces Visual Impairment in a Mouse Model of Multiple Sclerosis-like Optic Neuritis

Abstract

Purpose : Optic neuritis (ON) is often an early sign of multiple sclerosis (MS) and recent studies show a link between HIF-1 pathway activation and inflammation. This study aimed to determine if inhibition of the HIF-1 pathway using the HIF-1α antagonist Acriflavine (ACF) can reduce clinical progression and rescue the ocular phenotype in an experimental autoimmune encephalomyelitis (EAE) ON model.

Methods : EAE-related ON was induced in 40 female C57BL/6J mice by immunization with MOG33-55, Complete Freund’s Adjuvant and pertussis toxin. Twenty EAE mice received daily systemic injections of ACF at 5 mg/kg, 20 EAE mice were placebo injected and another 20 naive mice served as controls. Clinical progression was monitored using a 5-point EAE scoring scheme. Visual acuity was assessed weekly and RGC complex thickness was measured using optical coherence tomography (OCT). Retinal Ganglion Cells (RGC) and optic nerve axons were counted using RBPMS and PPD staining. Grades of demyelination (0-3 grading) and numbers of CD3+ cells in optic nerves were evaluated by H&E/LFB and immunohistochemistry. Differences were analyzed by ANOVA and Tukey’s post-hoc test.

Results : ACF-treated EAE animals show significantly less motor-sensory impairment (Area under curve EAE:51±3 vs. EAE+ACF: 28±3; p=0.0001). ACF treatment also preserved visual acuity (EAE:0.24±0.06 c/d vs. EAE+ACF: 0.290.006 c/d; p=0.01) and maintained RGC complex thickness when compared to untreated EAE mice (EAE:61±3µm vs. EAE+ACF:64±3µm; p=0.003). Higher RGC density (EAE:248±19 RBPMS cells/field vs. EAE+ACF: 275±16 RBPMS cells/field; p=0.02) and axon numbers (EAE:1310±562 axons/field vs. EAE+ACF: 2096±527; p=0.008) were observed in ACF treated EAE mice when compared to EAE animals in the placebo group. Grade of optic nerve demyelination was lessened (EAE:2.1±0.5 vs. EAE+ACF:0.9±0.9; p=0.047) and less infiltrating CD3+ cells were observed (EAE:98±32 cell/mm vs. EAE+ACF: 31±20 cells/mm; p=0.0001) in ACF treated EAE mice.

Conclusions : In EAE mice, daily ACF injections reduce motor-sensory impairments as well as increase visual system recovery, including preservation of the RGC density and optic nerve axon number. In conclusion, these improvements seem to be attributed to the inhibition of the HIF-1 pathway by ACF. Further experiments are needed to determine ACF’s mechanisms of action in more detail but is a promising approach for MS therapies.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.