Experimental allergic aspermatogenic orchitis. III. Isolation of spermatozoal glycoproteins and their role in allergic aspermatogenic orchitis

Abstract

Four glycoproteins (GP1,2,3 and 4) rich in carbohydrate were isolated from guinea pig testes. GP1, 2, and 4 (one or more) were localized in the sperm acrosome by the indirect immunofluorescence technique. Purification consisted of delipidation with chloroform-methanol (2:1), acid extraction at pH 3.0, precipitation with 85% saturated ammonium sulfate, extraction with 5% trichloroacetic acid, and either gel filtration on agarose or ultrafiltration. The final purification steps were isoelectrofocusing or gel filtration on Sephadex G-75 followed by preparative slab gel electrophoresis at pH 8.6. Each glycoprotein appeared homogeneous by gel electrophoresis at pH 2.7 and 8.6, and by immunoelectrophoresis. The crude glycoprotein fraction from the agarose column was resolved into the three major components, GP1, 2, and 3, distinguished by their isoelectric points (pI 3.9, 4.4, and 5.0, respectively), electrophoretic mobilities at pH 8.6, and reactivities with antiserum in immunoelectrophoresis. GP4, isolated by ultrafiltration and Sephadex G-75 chromatography, was differentiated by the same criteria. Approximately 5 mg each of purified GP1, 3, and 4 and 2 to 3 mg of GP2 were isolated from 1000 g of wet guinea pig testes. GP1, 2, and 4 induced precipitating antibody in rabbits and goats. GP1 and GP4 induced allergic aspermatogenic orchitis in guinea pigs, an autoimmune disease characterized by infiltration of mononuclear cells around and within the seminiferous tubules, followed by extensive destruction of the germinal epithelium. The course of the disease induced by 2 mug of either GP1 or GP4 was essentially identical in time course and pathology to that induced by whole testicular homogenates or 1 mug of purified acrosomal protein (AP1).