|dc.description.abstract||Glaucoma is the leading cause of blindness worldwide after cataracts and it is estimated that the global prevalence of glaucoma will increase to 111.8 million by 2040 (1, 2). Primary open-angle glaucoma (POAG) is characterized by elevated intraocular pressure (IOP) owing to a build-up of aqueous humor (AH) outflow, which occurs predominantly in the conventional outflow pathway, the latter mainly comprised of the trabecular meshwork (TM) and Schlemm?s canal (SC) (3). Elevations in IOP cause damage to the retina, ganglion cell loss as well as damage to the optic nerve being characteristic features of disease pathology (4). Damage to ganglion cells results in irreversible vision loss and IOP-lowering drugs remain the main focus of therapy (5). Most topically applied therapeutics either reduce AH production by the ciliary body or enhance its clearance through the uveoscleral (unconventional) route and there is a current unmet clinical need for therapies directly targeting the conventional outflow pathway.
Chapter 1 provides an overall introduction to AH outflow dynamics and the pathologies associated with the various forms of glaucoma. Treatments currently available and those in development are also considered.
In Chapter 2, results from an in vitro study exploring changes occurring to glaucomatous human Schlemm?s canal endothelial cells (SCEC) are presented. Glaucomatous SCEC displayed increased expression of fibrotic alpha-smooth muscle actin (α-SMA) and adhesion vascular endothelial cadherin (VE-cadherin) markers, associated with an increase in cell size, cell migration, and proliferation, as well as evidence of mitochondrial dysfunction compared to healthy SCEC. All of these changes have the potential to negatively influence AH outflow facility and thus IOP.
In Chapter 3, a novel approach to enhancement of aqueous outflow facility by siRNA-mediated downregulation of SC endothelial tight junction (TJ) proteins, ZO-1, and tricellulin, resulting in an increase in paracellular permeability of SCEC, was tested in a clinically relevant setting. Systemic or localized use of glucocorticoids can result in dangerously high ocular hypertension (OHT), and there is currently a need for an effective means of rapid intervention. Data are presented in which siRNA-mediated suppression of SC endothelial TJs was used in a dexamethasone-induced murine model of steroid-induced OHT and resulted in significantly increased AH outflow facility and reduced IOP in these animals.
In Chapter 4, the use of the siRNA-mediated technique was also explored in a MYOCY437H transgenic mouse model of OHT, MYOC (myocilin) mutations having been encountered in rare hereditary forms of glaucoma and also in a proportion of adult-onset cases. In this study, a significant reduction in IOP was observed.
In the final chapter of this work (Chapter 5), an AAV-based therapeutic approach was assessed in the MYOCY437H mouse model. Chadderton and colleagues (6) have shown that AAV-mediated expression of NDI1, a yeast nuclear gene substituting for a number of mutations in the mitochondrial respiratory NADH-ubiquinone oxidoreductase complex (Complex 1) protects ganglion cells following intravitreal inoculation of the virus in a murine model of Leber hereditary optic neuropathy (LHON). Given evidence that has accumulated for mitochondrial dysfunction in glaucoma, an assessment was made of the possible therapeutic effects of NDI1 in MYOCY437H mice. NDI1 expression was obtained in both retina and in tissues of the anterior chamber (most obviously in corneal endothelia) in MYOCY437H mice following intravitreal and intracameral inoculation of virus. Data obtained from intracameral injections suggested a potentially beneficial effect in reducing IOP over a six-month period. However, further investigations are required in order to determine the mechanistic basis for such reduction.
In summary, the initial focus of this work lay in explorations of the physical, morphological and molecular differences between glaucomatous and healthy SCEC, which may bear relevance to future approaches to disease prevention. An siRNA-mediated strategy targeting steroid-induced glaucoma has been successfully validated in rodents, potentially representing a focused approach to clinical application in a sight-threatening acute scenario, the same approach having been assessed in a murine glaucoma model expressing a myocilin mutation. An AAV-based gene therapy approach has also been explored, providing encouraging initial results. Given that a significant number of POAG patients do not respond, or become resistant to current pressure-reducing medications, research in which new avenues of therapy are explored may hopefully eventually provide a meaningful benefit to such patients.||en