Immunohistochemical monitoring of the effect of a synthetic fibronectin-like peptide (Arg-Gly-Asp) on the age-related changes in the isolated human corneoscleral tissue of glaucomatous eyes.

Fibronectin, an adhesion glycoprotein has been detected and localized in samples of the trabecular meshwork from eight normotensive and 30 glaucomatous human eyes of various ages by means of the indirect immunoperoxidase staining technique. Fibronectin concentration in the trabecular meshwork tissue was evaluated by morphometric analysis. Deposits of the adhesion glycoprotein fibronectin were shown to be spread in the ocular drainage outflow system from patients along with progressive primary open-angle glaucoma (POAG). The fibronectin level quantitatively evaluated in serial cross-sections of trabecular meshwork, appeared to be increased during ageing and more rapidly in the event of POAG development. The active amino acid sequence in fibronectin is an arginine-glycine-aspartic acid tripeptide (Arg-Gly-Asp) and it was shown that the synthetic Arg-Gly-Asp peptide specifically inhibited the adhesive function of fibronectin in trabecular meshwork samples when incubated for 30 min at a concentration of 1-2 mg/ml. The peptide concentration necessary for a 50% decrease of the maximal fibronectin level in the trabecular meshwork specimen derived from patients with moderately advanced POAG stage, was about 1 mg/ml. Immunohistochemical staining exhibited a fainter fibronectin staining in trabecular tissues including the external trabecular layers and subendothelial region of Schlemm's canal, in samples incubated with the synthetic peptide compared with the same tissue explants before peptide treatment. It may be concluded that the adhesion control system is likely to play an important role in development and maintenance of tissue architecture and specialization of the normal human trabecular meshwork.

Changes in fibronectin staining in the human lens during ageing and cataractogenesis.

The content and localization of fibronectin, an extracellular glycoprotein, in the serial sections of lenses of normal human donors and cataractous patients of different ages were determined by the indirect immunoperoxidase staining technique. This was followed by the evaluation with quantitative morphometric analysis. It was shown that fibronectin was present in the area of cell contacts as single deposits of faint orange-brown stained material in the lens samples of young donors. The fibronectin level was raised in lens sections from aged donors. Its accumulation was detected mostly within the spaces of the lens fiber cells. At different stages of cataractogenesis a dramatic decrease of the fibronectin content was detected in the lens sections obtained from patients of different ages. A new linear spectrophotometric technique was developed for evaluation of the lens transparency, to correlate the lens opacity with corresponding histological data obtained from the immunostaining technique. Morphological studies performed further suggested that the lens fiber cell plasma membrane structures were deteriorated. This was observed as changes of fibronectin staining in the lens sections at different periods of human ageing and cataract development. It is concluded that a decrease of fibronectin staining in the human lens is an indication for the structural damage of the lens fiber cell plasma membranes during ageing and cataractogenesis.

Clinical, structural and molecular phototherapy effects of laser irradiation on the trabecular meshwork of human glaucomatous eyes.

The effects of argon laser trabeculoplasty (LTP) on intraocular pressure (IOP), outflow facility, the morphology of the trabecular meshwork (TM), and the pattern of extracellular glycoprotein fibronectin in trabeculum were studied in 46 eyes of patients with primary open-angle glaucoma (POAG). The LTP was done with informed consent, anticipating that trabeculectomy would be carried out at a scheduled time (2 h to several months following laser therapy). We found that the magnitude of IOP reduction and the improvement in the facility of outflow achieved are directly dependent on the time course after LTP and laser-induced structural changes in trabecular tissue. Light microscopic and immunohistochemical evaluations of the TM specimens at earlier intervals after LTP revealed evidence of heat effects, with disruption and shrinkage of the TM collagenous components and accumulation of fibronectin deposits in the aqueous drainage channels as compared with the TMs of matched patients with POAG who did not receive laser treatment. Within 24 h after LTP, proteins of glaucomatous TMs excised from patients incorporated increased amounts of [3H]-leucine radioactive label; however, the amount of [3H]-leucine-labeled material was significantly depressed in later periods of evaluation. The specimens obtained at longer intervals after LTP showed partial or total occlusion of the intertrabecular spaces by extracellular debris; however, the amount of trabecular fibronectin was not significantly different from that measured 24 h after LTP. At least two potential mechanisms are proposed for the TM tissue response to laser treatment, including heat-induced stretching of the collagen in lamellae and fibronectin-mediated attachment of beams supporting an adhesive tightening of the trabecular components caused by LTP. The changes in laser-induced tissue responses appear to be the result of morphological repair of irradiation-injured trabecular tissue.

Fibronectin detection in drainage outflow system of human eyes in ageing and progression of open-angle glaucoma.

The content of the fibronectin, an extracellular glycoprotein in the drainage outflow system of human eyes was determined by the indirect immunoperoxidase staining technique. The degree of fibronectin accumulation in ocular tissues was evaluated by quantitative morphometric analysis. It was shown that the fibronectin level was elevated in the ocular drainage outflow system of humans in ageing and was rapidly increased at different stages of primary open-angle glaucoma development. Increased deposit of fibronectin in trabecular tissues, mainly, in the inner wall of Schlemm's canal and juxtacanalicular, or cribriform part of trabecular meshwork, was demonstrated along with ageing and glaucoma disease progression. A hypothesis underlying the glaucomatous process as an adhesive impairment was proposed.