Perfusion-cultured bovine anterior segments as a model for studying TGFB2-induced ocular hypertension and glaucoma

Purpose: Primary open angle glaucoma (POA) is a leading cause of blindness worldwide. The most important risk factor is elevated intraocular pressure (IOP), which is due to glaucomatous damage to the trabecular meshwork (TM). Damaged TM does not function properly and creates an obstruction to aqueous humor outflow, and therefore elevates IOP. Transforming growth factor beta 2 (TGFβ2) is elevated in about 50% POAG patients, and is known to increase IOP in several experimental models. The purpose of this study was to determine if TGFβ2 induces IOP elevation in perfusion cultured bovine eyes, which are an important glaucoma research model. Methods: Fresh bovine eyes were obtained, transferred to the lab and carefully dissected. Vitreous humor, uvea, retina, retinal pigment epithelium, and lens were removed. The remaining anterior segment tissue, which contained the sclera, cornea, and TM, was mounted and sealed on a custom-made plexiglass dish with an O-ring using four screws. Perfusion medium was infused by a syringe pump at a constant infusion rate of 5 ul/min. After baseline IOP was established, bovine eyes were perfused with or without 10ng/ml TGFB2 for up to 7 days. IOP was recorded by a pressure transducer and a computerized system. Changes in IOP were calculated by subtracting baseline IOP from IOP post treatment. Conditioned medium was collected for Western immunoblotting (WB). Results: TGFβ2 increased IOP in treated bovine eyes by about 4.5 mmHg while the fellow control eye did not show significant changes in IOP. WB showed that fibronectin, a TGFβ2-inducible protein, was increased in perfusate collected from the TGFβ2 treated eyes. Conclusion: Our study showed that TGFβ2 is able to induce ocular hypertension in perfusion cultured bovine eyes. This will provide researchers a useful model to study POAG.