Harry A. Quigley’s research while affiliated with Johns Hopkins University and other places

Optic neuropathies cause irreversible vision loss as retinal ganglion cells (RGCs) die. Transplantation of pluripotent stem cell (PSC)-derived RGCs offers one potential therapeutic avenue to restore vision in patients suffering from optic neuropathy if the donor neurons survive long-term in the recipient eye and develop synaptic connections within the retinal inner plexiform layer (IPL) and subcortical visual centers (1). Thus far, attempts at intravitreal RGC transplantation have been hampered by sequestration on the epiretinal surface without engraftment into the retinal parenchyma. In mouse retinal explant cultures, enzymatic digestion of the retinal internal limiting membrane (ILM) promotes migration of transplanted RGCs into the recipient retina (2). Herein, we examined donor RGC survival and engraftment in living, immunosuppressed mice, rats, and rhesus macaques and in post-mortem human retinal explant cultures. Using 3 separate human PSC lines and 3 independent methods of ILM disruption, we demonstrate that the ILM is a barrier to the retinal engraftment of intravitreally delivered human PSC-derived RGCs. ILM disruption is associated with greater donor RGC survival over 2-8 weeks and enables migration of donor neuronal somas into the endogenous RGC layer where cells elaborate dendrites into the IPL and extend axons that follow the course of the endogenous retinal nerve fiber layer into the optic nerve head. Critically, ILM disruption enables donor RGCs to synaptically integrate into IPL circuits, conferring light responsivity. These findings have important implications for enabling neuronal replacement therapies to restore vision in patients with optic neuropathy. SIGNIFICANCE STATEMENT Retinal ganglion cell (RGC) replacement and optic nerve regeneration through transplantation of stem cell-derived RGCs holds potential for restoring vision lost to optic neuropathies. Here we demonstrate that intravitreally transplanted human RGCs laminate the epiretinal surface without projecting neurites into the retinal parenchyma. However, enzymatic, developmental and surgical disruption of the internal limiting membrane not only improves graft survival, but also enables structural and functional engraftment, with dendrites that stratify the inner plexiform layer, axons that grow into the optic nerve head, and acquired responsivity to light. These observations identify a translatable approach to enable transplantation-based RGC replacement for the treatment of optic neuropathy.

This study measured the strain response to intraocular pressure (IOP) change in the sclera, choroid, and retina of glaucoma patients whose optic nerve head region was imaged by optical coherence tomography (OCT) prior to and after IOP-lowering by laser suturelysis following trabeculectomy surgery. The strain response was calculated from digital volume correlation of the prior and after images. The strain response of the sclera, choroid, and retina were compared to those previously published for the anterior lamina cribrosa (ALC). Mean strains were lowest in the retina and highest in the prelaminar neural tissue (PLNT). Maximum principal and maximum shear strains were significantly increased with greater IOP decrease in all 5 eye regions. Maximum principal and maximum shear strains in the anterior lamina cribrosa (ALC) and the sclera were significantly related (p=0.0094). ALC and PLNT had a negative radial strain, while the sclera had a positive radial strain (p=0.017). In conclusion, the strain response of the sclera, choroid, and retina are related to the magnitude of IOP change and smaller than those of the ALC. The strain response of the ALC and sclera are significantly related to each other.

Purpose The purpose of this study was to measure biomechanical strains in the lamina cribrosa (LC) of living human eyes undergoing intraocular pressure (IOP) increase. Methods Healthy control subjects and patients with glaucoma underwent optical coherence tomographic (OCT) imaging of the LC before and after wearing of swim goggles that increased IOP (57 image pairs, 39 persons). Digital volume correlation was used to measure biomechanical strains in optic nerve head tissue and change in depth of the anterior border of the LC. Results The mean IOP increase in both glaucoma and control eyes was 7.1 millimeters of mercury (mm Hg) after application of the goggles. Among glaucoma eyes, strains that were significant were: contractile Ezz (average = −0.33%, P = 0.0005), contractile Eθθ (average = −0.23%, P = 0.03), Emax (average = 0.83%, P < 0.0001), and Γmax (average = 0.95%, P < 0.0001), whereas the average anterior LC depth (ALD) decreased by 2.39 µm (anterior; P = 0.0002). In glaucoma eyes, shear strain Ezθ was greater with worse mean deviation (MD) and visual function index (P = 0.044 and P = 0.006, respectively, multivariate models). Strain compliance for Erθ, Ezθ, and Eθθ all increased with greater MD worsening prior to imaging (P = 0.04, P = 0.007, and P = 0.03). Conclusions LC strains were measurable 20 minutes after IOP increase, producing axial compression and greater peripheral strain than centrally. Some strain compliances were greater with worse existing visual field loss or with more progressive past field loss. Translational Relevance Biomechanical strains are related to measures of glaucoma damage, supporting the hypothesis that optic nerve head biomechanical responses represent a noninvasive biomarker for glaucoma.

PURPOSE. The purpose of this study was to delineate the neuroprotective mechanisms of topical 2% ripasudil (Rip), a Rho kinase (ROCK) inhibitor. METHODS. In 340 mice, scheduled 2% Rip or balanced salt solution (BSS) saline drops were intermittently, unilaterally delivered. Intracameral microbead glaucoma (GL) injection increased intraocular pressure (IOP) from 1 day to 6 weeks (6W), whereas other mice underwent optic nerve (ON) crush. Retinal ganglion cell (RGC) loss was assessed using retinal wholemount anti-RNA Binding Protein with Multiple Splicing (RBPMS) labeling and ON axon counts. Axonal transport was quantified with β amyloid precursor protein (APP) immunolocalization. Micro-Western (Wes) analysis quantified protein expression. Immunofluorescent expression of ROCK pathway molecules, quantitative astrocyte structural changes, and ON biomechanical strains (explanted eyes) were evaluated. ROCK activity assays were conducted in separate ON regions. RESULTS. At 6W GL, mean RGC axon loss was 6.6 ± 13.3% in Rip and 36.3 ± 30.9% in BSS (P = 0.04, n = 10/group). RGC soma loss after crush was lower with Rip (68.6 ± 8.2%) than BSS (80.5 ± 5.7%, P = 0.006, n = 10/group). After 6W GL, RGC soma loss was lower with Rip (34 ± 5.0%) than BSS (51 ± 8.1%, P = 0.03, n = 10/group). Axonal transport of APP within the unmyelinated ON (UON) was unaffected by Rip. Maximum principal mechanical strains increased similarly in Rip and BSS-treated mice. Retinal ROCK 1 and 2 activity was reduced by Rip in GL eyes. The pROCK2/ROCK2 protein ratio rose in the retina of BSS GL eyes, but not in Rip GL eyes. CONCLUSIONS. Topical Rip reduced RGC loss in GL and ON crush, with suppression of ROCK signaling in the retina and ON. The neuroprotection mechanisms appear to involve effects on both RGC and astrocyte responses to IOP elevation.

Purpose To introduce a discussion of glaucoma screening Methods/Results Glaucoma screening faces challenges including not satisfying important ethical principles for medical screening. Conclusions Approaches that would better provide care for glaucoma patients include eliminating loss to follow-up and identification of glaucoma among family members of known glaucoma patients.

Objective To measure mechanical strain of the lamina cribrosa (LC) after intraocular pressure (IOP) change produced 1 week after a change in glaucoma medication. Design Cohort study. Participants Adult glaucoma patients (23 eyes, 15 patients) prescribed a change in IOP-lowering medication. Intervention Noninvasive OCT imaging of the eye. Main Outcome Measures Deformation calculated by digital volume correlation of OCT scans of the LC before and after IOP lowering by medication. Results Among 23 eyes, 17 eyes of 12 persons had IOP lowering ≥ 3 mmHg (reduced IOP group) with tensile anterior-posterior Ezz strain = 1.0% ± 1.1% (P = 0.003) and compressive radial strain (Err) = −0.3% ± 0.5% (P = 0.012; random effects models accounting inclusion of both eyes in some persons). Maximum in-plane principal (tensile) strain and maximum shear strain in the reduced-IOP group were as follows: Emax = 1.7% ± 1.0% and Γmax = 1.4% ± 0.7%, respectively (both P < 0.0001 vs. zero). Reduced-IOP group strains Emax and Γmax were significantly larger with greater % IOP decrease (P < 0.0001 and P < 0.0001, respectively). The compliances of the Ezz, Emax, and Γmax strain responses, defined as strain normalized by the IOP decrease, were larger with more abnormal perimetric mean deviation or visual field index values (all P ≤ 0.02). Strains were unrelated to age (all P ≥ 0.088). In reduced-IOP eyes, mean LC anterior border posterior movement was only 2.05 μm posteriorly (P = 0.052) and not related to % IOP change (P = 0.94, random effects models). Only Err was significantly related to anterior lamina depth change, becoming more negative with greater posterior LC border change (P = 0.015). Conclusions Lamina cribrosa mechanical strains can be effectively measured by changes in eye drop medication using OCT and are related to degree of visual function loss in glaucoma. Financial Disclosure(s) Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article.

Purpose: The strain response of the mouse astrocytic lamina (AL) to an ex vivo mechanical test was compared between two protocols: eyes that underwent sustained intraocular pressure (IOP) increase and eyes after optic nerve crush. Methods: Chronic IOP elevation was induced by microbead injection or the optic nerve was crushed in mice with widespread green fluorescence. After 3 days or 6 weeks, eyes were inflation tested by a published method of two-photon fluorescence to image the AL. Digital volume correlation was used to calculate strains. Optic nerve axon damage was also evaluated. Results: In the central AL but not the peripheral AL, four strains were greater in eyes at the 3-day glaucoma time point than control (P from 0.029 to 0.049, n = 8 eyes per group). Also, at this time point, five strains were greater in the central AL compared to the peripheral AL (P from 0.041 to 0.00003). At the 6-week glaucoma time point, the strains averaged across the specimen, in the central AL, and the peripheral AL were indistinguishable from the respective controls. Strains were not significantly different between controls and eyes 3 days or 6 weeks after crush (n = 8 and 16). Conclusions: We found alterations in the ex vivo mechanical behavior in eyes from mice with experimental glaucoma but not in those with crushed optic nerves. The results of this study demonstrate that significant axon injury does not directly affect mechanical behavior of the astrocytic lamina.