Abstract
Purpose :
Abnormal turnover of the extracellular matrix protein elastin has been associated with wet AMD, the subtype of AMD that involves pathological angiogenesis; increased Bruch’s membrane elastin layer degradation, elevated serum elastin derived peptides and elastin antibody levels are reported in wet AMD patients. Based on these results we hypothesized that increased elastase enzyme activity, the enzymes that degrade elastin protein are associated with AMD pathogenesis and elastase inhibition can have therapeutic potential in AMD.
Methods :
Elastase activity was measured in ocular tissues of wet AMD mouse models (JR5558 - a spontaneous mouse model and laser-induced choroidal neovascularization (CNV) model), stable monolayers of primary mouse retinal pigment epithelial (RPE) cells and AMD patient-iPSC derived RPE cells. Elastase inhibitor Alpha1 antitrypsin (A1AT) was injected intraperitoneally (100 mg/kg) in mouse models weekly and lesions were assessed using Optical coherence tomography, fundus imaging and fluorescein angiography. VEGF protein levels were quantified using ELISA and western blotting. Efficacy of A1AT was also explored in a mouse model with RPE specific overexpression of HTRA1, a serine protease with elastase activity. For statistical analysis t test and one-way ANOVA with Tukey’s multiple comparison were used.
Results :
Elastase activity was increased in the retinas of JR5558 and laser-CNV mice by 50-30% compared to control retinas; increased extracellular elastase activity was detected in supernatants of AMD patient-iPSC derived RPE and primary RPE cells from JR5558 mice compared to controls (n=4, p<0.05). A1AT reduced CNV lesion area and pathological ONL thinning in JR5558 (n=4, p<0.05) as well as in laser-CNV (n=7, p<0.01) mice. A1AT mitigated VEGF increase in elastase-exposed RPE monolayers and in JR5558 mouse retinas (n=3, p<0.05). A1AT also reversed an early AMD phenotype such as RPE/BrM thickening present in HTRA1 overexpressing mice (n=4, p<0.01).
Conclusions :
Our results establish that increased elastase activity is associated with AMD pathogenesis. A1AT can reverse the outer retinal alterations occur during early AMD and can ameliorate the severe wet AMD retinal phenotypes via inhibiting the pathological VEGF release. Hence, repurposing A1AT could be a novel therapeutic strategy for AMD.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.