Neuroserpin, a Serine Protease Inhibitor Providing Neuroprotection in the Retina

In glaucoma, loss of retinal ganglion cells (RGCs) and axons leads to blindness, and preservation of RGCs is a major therapeutic goal. High intraocular pressure (IOP) has been implicated in neurodegeneration of RGCs in primary open angle glaucoma (POAG) but in some cases the disease progresses even in absence of high IOP. There is substantial evidence for the involvement of proteases in causing damage to the optic nerve head and RGCs in glaucoma. Plasmin is a serine protease enzyme, and its activity is regulated by neuroserpin in the retina. Our group has shown that in glaucoma pathology, there is loss of plasmin inhibitory activity of neuroserpin in the retina. The proteolytic activity of plasmin has been implicated in mediating excitotoxicity-induced damage to retina and RGCs. Therefore, the aims of this thesis were to determine the effects of neuroserpin neutralization and over-expression and to determine if the upregulation of neuroserpin in RGCs was neuroprotective in an experimental glaucoma model. This was achieved using neuroserpin overexpression (NS+/+Tg) and neuroserpin knockout mice (NS-/-) models and monitoring them. Knockout of neuroserpin resulted in inner retinal structural and functional deficits starting from three months of age. NS-/- mice retina also showed synaptic deficits as evident by reduced levels of synaptic markers. Enhanced autophagy markers were identified in NS-/- retinas that accompanied apoptotic changes. Furthermore, the neuroserpin antibody was intravitreally administered to C57BL/6J mice to neutralise the neuroserpin expression. Neuroserpin neutralization resulted in reduced inner retinal function and structural deficits in both healthy and glaucoma mice. Neuroserpin neutralization also resulted in perturbations in synaptic markers with upregulation of autophagy response and apoptosis. Further, experimental glaucoma was induced in both NS+/+Tg and NS-/- mice to determine the changes in retinal structural and functional integrity. NS-/- mice with glaucoma demonstrated exacerbation of the functional and structural damage of inner retina. However, NS+/+Tg mice with glaucoma were relatively protected against inner retinal structural and functional damage compared to the corresponding controls. Glaucoma induction in NS-/- retina affected the levels of synaptic markers and led to elevation in autophagy and apoptotic signalling markers. No such changes were observed in NS+/+Tg mice subjected to high IOP. Finally, to understand the effect of neuroserpin upregulation on the retina in glaucoma conditions, neuroserpin was overexpressed in C57BL/6J mice inner retina using AAV gene therapy. The overexpression of neuroserpin in RGCs resulted in protection against inner retinal structural and functional deficits under experimental glaucoma conditions. These structural and functional changes were accompanied by reduced autophagy and apoptotic activation. In summary, neuroserpin plays an important role in providing neuroprotection in the retina and can serve as a potential therapeutic candidate to protect retinal neurons in glaucoma pathology. Future studies on the roles of neuroserpin in other neurodegenerative diseases will help understand the biochemical mechanisms underlying neuroserpin mediated neuroprotection.