No abstract

Studies of the effects of glucocorticoid (GC) and oxidative stress stimuli in differentiated cultures of human trabecular meshwork (HTM) cells have provided the rationale for our studies of a major new gene termed TIGR (trabecular meshwork inducible GC response). The TIGR clone was isolated by differential library screening using selection criteria based on the induction pattern of a new protein/glycoprotein found in HTM cultures after prolonged but not brief exposure to GCs. This GC induction patter matched the time course and dose response required for intraocular pressure elevation in patients receiving corticosteroids. The very large, progressive induction of TIGR combined with specific structural features of its cDNA suggested that TIGR should be considered a candidate gene for outflow obstruction in glaucoma. Among the properties of TIGR cDNA were a signal sequence for secretion, several structural features for interactions with glycosaminoglycans and other glycoproteins and putative sites for cell surface interactions. In addition, the leucine zippers in the structure were related to TIGR-TIGR oligomerization that was shown to occur with native and recombinant TIGR protein. The verification that TIGR was a major stress response protein in HTM cells following hydrogen peroxide (or phorbol esters) exposure provided a potential link between GC and oxidative mechanisms thought to be involved in glaucoma pathogenesis. Pharmacological evaluation showed that basic fíbroblast growth factory and transforming growth factor β decreased the GC induction of TIGR, and certain nonsteroidal anti-inflammatory drugs protected against both GC- and oxidation-induced stress responses in HTM cells. Our recent studies of TIGR’s genomic structure have shown motifs in the promoter region that suggest a basis by which multiple hormonal/environmental stimuli can regulate TIGR production and by which putative genetic alterations could lead to an overexpression of the protein. Further application of cell biology/biochemistry, molecular biology, genetic and histological approaches will be helpful in understanding the role of TIGR in different glaucoma syndromes.

Cultured trabecular meshwork (TM) cells are a valuable model system to study the cellular mechanisms involved in the regulation of conventional outflow resistance and thus intraocular pressure; and their dysfunction resulting in ocular hypertension. In this review, we describe the standard procedures used for the isolation of TM cells from several animal species including humans, and the methods used to validate their identity. Having a set of standard practices for TM cells will increase the scientific rigor when used as a model, and enable other researchers to replicate and build upon previous findings.

TGF-beta2 reduces outflow facility when perfused into cultured human anterior segments. Furthermore, TGF-beta2 affects the extracellular matrix of the trabecular meshwork in a manner that is consistent with the observed reduction in outflow facility. Although the distribution of accumulated fibrillar material was different in these perfused eyes than that in POAG, the difference could be due to variation in biomechanical environment for TM cells in cultured anterior segments compared with the living eye. Overall, these results support the hypothesis that elevated TGF-beta2 levels in the aqueous humor play a role in the pathogenesis of the ocular hypertension in POAG.

PSX eyes differ from POAG and JG eyes not only by their clinical or biomicroscopic appearance, but also by their normal TGF-beta 2 levels in aqueous humor. The fact that most of the POAG eyes with favorable bleb development had normal TGF-beta 2 levels indicated that there might be some relationship between bleb formation and TGF-beta 2 levels. On the other hand, the fact that eyes with less favorable bleb development had both low and high TGF-beta 2 levels indicated that other factors are also involved in the scarring of the filtration bleb.