Elevated α₂M in the aqueous humor is associated with Glaucoma but not with PXF alone.

Abstract

Purpose : Glaucoma is a chronic and progressive optic nerve neuropathy, characterized by apoptosis of retinal ganglion cells. The proteins α₂M and TNF-α are known mediators in the apoptotic processes of ganglion cells. The purpose of our work was to study their involvement in the pathogenesis of Primary Open-Angle Glaucoma (POAG) and Pseudo- Exfoliation syndrome (PXF) by measuring their levels in the aqueous humor.

Methods : Aqueous humor samples were collected during standard cataract surgery. α₂M and TNF-α levels were measured using standardized versus human immunoglobulin as loading control. Ten micrograms of total aqueous humor protein were combined with 2X SDS loading buffer and transferred to nitrocellulose membranes. The α₂M and TNF-α proteins were detected using rabbit polyclonal antibodies. Goat anti-rabbit antibodies conjugated with horseradish peroxidase were used as secondary reagents. For digital quantification the membranes were scanned and analyzed using ImageJ software. Data was subjected to ANOVA and Dunnett's Post Hoc Contrasts.

Results : Overall 27 samples were collected, of which 7 samples from POAG patients, 8 from PXF patients (only one with glaucoma) and 12 control samples. In patients with POAG, both α₂M levels and TNF-α levels were significantly elevated compared to control eyes (0.93±0.28 versus 0.089±0.087, and 0.77±0.32 versus 0.058±0.037, respectively, p < 0.001). In patients with PXF, the mean level of TNF-α was significantly elevated compared to control (0.42±0.33, p < 0.01) while mean α₂M level was similar to control level (0.13±0.19). A single patient with PXF glaucoma had considerably elevated levels of α₂M, comparable to the levels in POAG patients (0.6 versus a range of 0.04-0.13 in PXF patients without glaucoma).

Conclusions : This work further documents a potential novel mechanism of the apoptotic process in ganglion cells and a potential biomarker for glaucoma, in contrast to PXF alone. If verified in larger studies, these proteins may further serve as therapeutic targets for glaucoma.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.