Effects of ascorbic acid on levels of fibronectin, laminin and collagen type 1 in bovine trabecular meshwork in organ culture

Abstract

PURPOSE. Fibronectin, laminin and collagen type I are important extracellular matrix products of trabecular meshwork cells. This study was performed to examine the effects of ascorbic acid, a significant component in the aqueous humor, on the levels of these proteins in trabecular meshwork cells maintained in organ culture. METHODS. The anterior segment of freshly enucleated bovine eyes was perfused in a modified organ culture system. Three cultures were set up simultaneously. One received serum-free medium containing 100 µg/ml of ascorbic acid, one received 250 µg/ml of ascorbic acid and one served as a control. After 72 h, the tissues were processed for paraffin sections and immunostaining was conducted using an avidin-biotin-peroxidase complex method. Western blot and dot blot assays were performed on tissue extracts. RESULTS. Compared with the controls, the staining for fibronectin and laminin was markedly enhanced in trabecular meshwork tissues treated with both concentrations of ascorbic acid. Increased collagen type I production by trabecular meshwork cells was also demonstrated in the presence of ascorbic acid. Western blot and dot blot results confirmed the immunostaining findings. CONCLUSIONS. Ascorbic acid promotes production of fibronectin, laminin and collagen type I by trabecular meshwork cells. The organ culture results are consistent with those obtained previously from tissue culture studies.