Aqueous outflow pathway glycosaminoglycans

doi:10.1016/0014-4835(81)90032-4

Experimental Eye Research

Volume 32, Issue 3, March 1981, Pages 265-277

https://doi.org/10.1016/0014-4835(81)90032-4 Get rights and content

Abstract

The acidic glycosaminoglycan distribution patterns of the aqueous outflow pathway, the iris-ciliary body, and sclera of the New Zealand Red rabbit were identified by analysis of the glycosaminoglycan moieties and by the use of zone electorphoresis. Alcian blue positive bands on cellulose acetate membranes were characterized by a determination of their electrophoretic mobility in two electrolyte systems and by a comparison to the electrophoretic mobility of standard reference glycosaminoglycans. To verify the identity of each band, specific glycosaminoglycan degrading enzymes were used to remove the glycosaminoglycans. Glycosaminoglycan samples equivalent to 5 μg uronic acid were treated with hyaluronate lyase, testicular hyaluronidase, chondroitin AC lyase and chondroitin ABC lyase. The glycosaminoglycans resistant to chondroitin ABC lyase were treated with nitrous acid and tested for solubility in cetyl pyridinium chloride. The scleral distribution pattern was hyaluronic acid, chondroitin sulfate and hybrid dermatan sulfate-chondroitin sulfate. The aqueous outflow pathway and iris-ciliary body distribution patterns were hyaluronic acid, keratan sulfate, heparan sulfate and hybrid dermatan sulfate-chondroitin sulfate.

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The GAG composition in TM extracts was determined by sequential enzymatic degradation with various GAG-degrading enzymes. The sources included TM from a few hours postmortem human eyes, stationary human anterior segment organ cultures, perfused human anterior segment organ cultures and densely confluent early-passage human TM cell cultures (Grierson and Lee, 1975; Schachtschabel et al., 1977; Knepper et al., 1981, 1996b; Schachtschabel et al., 1982; Rohen et al., 1984; Acott et al., 1985, 1988; Yue and Elvart, 1987; Tschumper et al., 1990; Acott, 1994a). Relatively good agreement was found in the amount of each type of GAG in the TM using these models and all of the normal GAGs were identified.
The extracellular matrix (ECM) of the trabecular meshwork (TM) is thought to be important in regulating intraocular pressure (IOP) in both normal and glaucomatous eyes. IOP is regulated primarily by a fluid resistance to aqueous humor outflow. However, neither the exact site nor the identity of the normal resistance to aqueous humor outflow has been established. Whether the site and nature of the increased outflow resistance, which is associated with open-angle glaucoma, is the same or different from the normal resistance is also unclear. The ECMs of the TM beams, juxtacanalicular region (JCT) and Schlemm's canal (SC) inner wall are comprised of fibrillar and non-fibrillar collagens, elastin-containing microfibrils, matricellular and structural organizing proteins, glycosaminoglycans (GAGs) and proteoglycans. Both basement membranes and stromal ECM are present in the TM beams and JCT region. Cell adhesion proteins, cell surface ECM receptors and associated binding proteins are also present in the beams, JCT and SC inner wall region. The outflow pathway ECM is relatively dynamic, undergoing constant turnover and remodeling. Regulated changes in enzymes responsible for ECM degradation and biosynthetic replacement are observed. IOP homeostasis, triggered by pressure changes or mechanical stretching of the TM, appears to involve ECM turnover. Several cytokines, growth factors and drugs, which affect the outflow resistance, change ECM component expression, mRNA alternative splicing, cellular cytoskeletal organization or all of these. Changes in ECM associated with open-angle glaucoma have been identified.
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An experimental model of pressure-induced optic nerve damage would greatly facilitate the understanding of the cellular events leading to ganglion cell death, and how they are influenced by intraocular pressure and other risk factors associated to glaucoma. The aim of the present report was to study the effect of a long-term increase of intraocular pressure in rats induced by intracameral injections of hyaluronic acid with respect to electroretinographic activity and retinal and optic nerve histology. For this purpose, hyaluronic acid was injected weekly in the rat anterior chamber of one eye, whereas the contralateral eye was injected with saline solution. The results showed a significant decrease of oscillatory potentials and a- and b-wave amplitude of the scotopic electroretinogram after 3 or 6 weeks of hyaluronic acid administration, respectively. These parameters were further reduced after 10 weeks of treatment with hyaluronic acid. No significant changes in anterior chamber angle structures from hyaluronic acid- and vehicle-injected eyes were observed, whereas a significant loss of ganglion cell layer cells and of optic nerve axons were detected in animals that received hyaluronic acid for 10 weeks, as compared to eyes injected with saline solution. In summary, present results indicate that the chronic administration of hyaluronic acid induced a significant decrease in the electroretinographic activity and histological changes in the retina and optic nerve that seem consistent with some features of chronic open-angle glaucoma. Therefore, this could be an experimental model to study the cellular mechanisms by which elevated intraocular pressure damages the optic nerve and the retina.
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The pathogenesis of many anterior segment disorders and ocular complications following surgery are secondary to the wound healing response. The extent of clinical damage observed is closely related to the amount of scarring and tissue contraction. Matrix metalloproteinases (MMPs) are a family of enzymes that play a vital role in all stages of the wound healing process. They degrade all extracellular matrix components and also have the ability to synthesize collagen and extracellular matrix members, and are therefore important in the remodeling of a wound. Overexpression of MMPs results in excessive extracellular matrix degradation, leading to tissue destruction and loss of organ function. In the case of the anterior segment, this may mean the loss of visual function. This review focuses on the role MMPs have in the development of various anterior segment disorders. The importance of MMPs in the wound healing response and its potential modulation to manipulate the scarring response is being recognized, and current developments will be described.
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AL-3037A (Sodium ferri ethylenediaminetetraacetate), a novel compound shown to stimulate the degradation of glycosaminoglycans, was evaluated for its effects on aqueous humor outflow and intraocular pressure (IOP) in four experimental models. Its effect on outflow facility was assessed in bovine and human ocular perfusion organ cultures. Its IOP effect was tested in normotensive and dexamethasone-induced ocular hypertensive rabbits. In bovine eyes, perfusion with AL-3037A (0.1% w/v, 2.3 m $M$ ) significantly increased the outflow facility well above the normal ‘wash-out’ effect. At 30 min after perfusion, the outflow facility of drug-treated eyes increased by 26.0 ± 2.8% (mean ± $S.E.(M.)$ ,n = 8), significantly higher than the 12.1 ± 2.8% increase in vehicle-treated eyes. This difference sustained throughout the study period (2 hr). The compound also enhanced aqueous outflow in perfused human anterior segments. In non-glaucomatous eyes, it produced a small decrease in IOP (15.4 ± 4.6%, n = 17), but in tissues derived from glaucoma patients, bolus administration of 3 mg (7 μ mol) of AL-3037A lowered the IOP by 52–68% (n = 2) lasting for at least 3 hr. This outflow-enhancing effect of AL-3037A in ex vivo studies was confirmed by in vivo results. In normotensive rabbits, oral (50 mg kg⁻¹), intravenous (10 mg kg⁻¹), or topical (2 mg; 50 μ l of 4% w/v solution) administration of AL-3037A produced maximum reduction of IOP, when compared to vehicle-treated animals, by 34.7 ± 3.5% (n = 10), 22.0 ± 4.6% (n = 10), and 21.6 ± 4.5% (n = 10), respectively. In dexamethasone induced ocular hypertensive rabbits, topical application of the compound (0.5 mg; 25 μ l of 2% w/v solution) reduced IOP significantly by 19.2 ± 0.4% (n = 7) at 3 hr after dosing. Importantly, the IOP lowering effect of AL-3037A did not diminish even after repeated treatments in consecutive days. Thus, in the four study models across three animal species, AL-3037A was demonstrated to be an efficacious ocular hypotensive compound whose effect is most likely mediated by augmentation of the aqueous outflow. Its proposed action on the metabolism of glycosaminoglycans may provide a new and unique mechanism for the treatment of glaucoma.
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Aqueous outflow pathway glycosaminoglycans

Abstract

Biochim. Biophys. Acta.

Am. J. Ophthalmol.

Biochim. Biophys. Acta.

Analyt. Biochem.

Analyt. Biochem.

Develop. Biology

Arch. Biochem. Biophys.

Exp. Eye Res.

Exp. Eye Res.

Analyt. Biochem.

Biochim. Biophys. Acta.

J. Biol. Chem.

Biochim. Biophys. Acta.

J. Biol. Chem.

Am. J. Ophthalmol.