Oxidative Protein Modifications in Glaucomatous Trabecular Meshwork

Abstract

Abstract: : Purpose: To determine whether oxidative protein modifications are elevated in the glaucomatous trabecular meshwork and possibly play a role in glaucoma pathogenesis Methods:Trabecular meshwork (TM) was dissected from normal human cadaver eyes from the Cleveland Eye Bank and from primary open angle glaucoma (POAG) patients undergoing trabeculectomy at the Cole Eye Institute. Protein was extracted by homogenization in 100 mM Tris–Cl buffer pH 7.8 containing 5 mM dithiotheritol, 1mM SnCl₂, 50 mM NaHPO₄, 1mM diethylenetriaminepentaacetic acid, 100 mM butylated hydroxy toluene and 0.5% SDS. Western analyses for oxidative protein modification were performed with antibodies to carboxyethyl–pyrrole (CEP), hydroxynonenal (HNE), isolevuglandin (Iso[4]LGE₂) and argpyrimidine (AGEs). Results:HNE immunoreactivity was found in 9/12 POAG TM but in no normal TM (0/12). AGE immunoreactivity was found in 3/4 POAG TM and no normal TM (0/4). Iso[4]LGE₂ immunoreactivity was found in 8/8 POAG TM and in 2/8 normal TM. CEP immunoreactivity was not observed in either POAG or normal TM. Conclusions: Protein modifications generated from the oxidation of lipids and sugar appear to be more prevalent in glaucomatous TM than in normal TM. Such oxidative modifications may contribute to blockage of aqueous humor circulation and increased intraocular pressure in POAG. CR: N. Supported in part by The American Health Assistance foundation, NIH grants EY06603, EY14239, EY15638, GM21249, HL53315, The Foundation Fighting Blindness and The Cleveland Clinic Foundation.

Keywords: oxidation/oxidative or free radical damage • protein modifications-post translational • proteomics