Intraocular Immune Response in Human Uveitis: Time to Look Beyond Animal Models.

PURPOSE: To review the current and future approaches to investigating the intraocular immune response in human uveitis. DESIGN: Perspective. METHODS: Review of currently available methods for investigating the immune response in ocular tissues and fluids in patients with intraocular inflammation/ uveitis. The advantages and disadvantages of human studies have been compared to those of animal models of uveitis. RESULTS: Animal models, while being excellent tools for mechanistic studies, do not replicate the clinical and immunologic heterogeneity of human uveitis. Opportunities for immunological studies in human uveitis are mostly limited to histological studies, or sampling of intraocular fluids and peripheral blood. Histopathological studies can be enhanced by revisiting published historical data, tissue repositories, or autopsy specimens. Intraocular fluids can be investigated by a variety of techniques. Among these, flow cytometry and single-cell RNA sequencing (scRNAseq) provide single-cell resolution. While the current technology is costly and labor-intensive, scRNAseq is less limited by the low cellular yield from intraocular fluids and allows unbiased immune profiling enabling discovery of new cellular subsets. Immunological phenotypes uncovered from human data can be further investigated in animal studies. CONCLUSION: The diversity of the intraocular immune response in uveitis patients remains challenging but can be studied by multiple techniques including histopathology, flow cytometry, and scRNAseq. Human data can be combined with animal studies for translating uveitis research into novel therapies.

Imaging Cell Signaling in Tissues Using the IBEX Method.

The study of cell signaling within tissues can be enhanced using highly multiplexed immunohistochemistry to localize the presence and spatial distribution of numerous pathways of interest simultaneously. Additional data can also be gained by placing the identified proteins into the context of adjacent structures, stroma, and interacting partners. Here, we outline a protocol for using the recently described IBEX method on tissues. This is an open and simple cyclic immunohistochemistry approach suited to this application. We describe a simplified protocol and provide guidance on the method, using a 12-marker panel on human retina to demonstrate the approach.

Periodontitis and Outer Retinal Thickness: a Cross-Sectional Analysis of the United Kingdom Biobank Cohort.

Purpose: Periodontitis, a ubiquitous severe gum disease affecting the teeth and surrounding alveolar bone, can heighten systemic inflammation. We investigated the association between very severe periodontitis and early biomarkers of age-related macular degeneration (AMD), in individuals with no eye disease. Design: Cross-sectional analysis of the prospective community-based cohort United Kingdom (UK) Biobank. Participants: Sixty-seven thousand three hundred eleven UK residents aged 40 to 70 years recruited between 2006 and 2010 underwent retinal imaging. Methods: Macular-centered OCT images acquired at the baseline visit were segmented for retinal sublayer thicknesses. Very severe periodontitis was ascertained through a touchscreen questionnaire. Linear mixed effects regression modeled the association between very severe periodontitis and retinal sublayer thicknesses, adjusting for age, sex, ethnicity, socioeconomic status, alcohol consumption, smoking status, diabetes mellitus, hypertension, refractive error, and previous cataract surgery. Main Outcome Measures: Photoreceptor layer (PRL) and retinal pigment epithelium-Bruch's membrane (RPE-BM) thicknesses. Results: Among 36 897 participants included in the analysis, 1571 (4.3%) reported very severe periodontitis. Affected individuals were older, lived in areas of greater socioeconomic deprivation, and were more likely to be hypertensive, diabetic, and current smokers (all P < 0.001). On average, those with very severe periodontitis were hyperopic (0.05 ± 2.27 diopters) while those unaffected were myopic (-0.29 ± 2.40 diopters, P < 0.001). Following adjusted analysis, very severe periodontitis was associated with thinner PRL (-0.55 µm, 95% confidence interval [CI], -0.97 to -0.12; P = 0.022) but there was no difference in RPE-BM thickness (0.00 µm, 95% CI, -0.12 to 0.13; P = 0.97). The association between PRL thickness and very severe periodontitis was modified by age (P < 0.001). Stratifying individuals by age, thinner PRL was seen among those aged 60 to 69 years with disease (-1.19 µm, 95% CI, -1.85 to -0.53; P < 0.001) but not among those aged < 60 years. Conclusions: Among those with no known eye disease, very severe periodontitis is statistically associated with a thinner PRL, consistent with incipient AMD. Optimizing oral hygiene may hold additional relevance for people at risk of degenerative retinal disease. Financial Disclosures: Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article.

Adeno-associated virus gene therapy prevents progression of kidney disease in genetic models of nephrotic syndrome.

Gene therapy for kidney diseases has proven challenging. Adeno-associated virus (AAV) is used as a vector for gene therapy targeting other organs, with particular success demonstrated in monogenic diseases. We aimed to establish gene therapy for the kidney by targeting a monogenic disease of the kidney podocyte. The most common cause of childhood genetic nephrotic syndrome is mutations in the podocyte gene NPHS2, encoding podocin. We used AAV-based gene therapy to rescue this genetic defect in human and mouse models of disease. In vitro transduction studies identified the AAV-LK03 serotype as a highly efficient transducer of human podocytes. AAV-LK03-mediated transduction of podocin in mutant human podocytes resulted in functional rescue in vitro, and AAV 2/9-mediated gene transfer in both the inducible podocin knockout and knock-in mouse models resulted in successful amelioration of kidney disease. A prophylactic approach of AAV 2/9 gene transfer before induction of disease in conditional knockout mice demonstrated improvements in albuminuria, plasma creatinine, plasma urea, plasma cholesterol, histological changes, and long-term survival. A therapeutic approach of AAV 2/9 gene transfer 2 weeks after disease induction in proteinuric conditional knock-in mice demonstrated improvement in urinary albuminuria at days 42 and 56 after disease induction, with corresponding improvements in plasma albumin. Therefore, we have demonstrated successful AAV-mediated gene rescue in a monogenic renal disease and established the podocyte as a tractable target for gene therapy approaches.

Triazole-derivatized near-infrared cyanine dyes enable local functional fluorescent imaging of ocular inflammation.

Near-infrared (NIR) chemical fluorophores are promising tools for in-vivo imaging in real time but often succumb to rapid photodegradation. Indocyanine green (ICG) is the only NIR dye with regulatory approval for ocular imaging in humans; however, ICG, when employed for applications such as labelling immune cells, has limited sensitivity and does not allow precise detection of specific inflammatory events, for example leukocyte recruitment during uveitic flare-ups. We investigated the potential use of photostable novel triazole NIR cyanine (TNC) dyes for detecting and characterising activated T-cell activity within the eye. Three TNC dyes were evaluated for ocular cytotoxicity in-vitro using a MTT assay and optimised concentrations for intraocular detection within ex-vivo porcine eyes after topical application or intracameral injections of the dyes. TNC labelled T-cell tracking experiments and mechanistic studies were also performed in-vitro. TNC-1 and TNC-2 dyes exhibited greater fluorescence intensity than ICG at 10 µM, whereas TNC-3 was only detectable at 100 µM within the porcine eye. TNC dyes did not demonstrate any ocular cell toxicity at working concentrations of 10 µM. CD4+T-cells labelled with TNC-1 or TNC-2 were detected within the porcine eye, with TNC-1 being brighter than TNC-2. Detection of TNC-1 and TNC-2 into CD4+T-cells was prevented by prior incubation with dynole 34-2 (50 µM), suggesting active uptake of these dyes via dynamin-dependent processes. The present study provides evidence that TNC dyes are suitable to detect activated CD4+T-cells within the eye with potential as a diagnostic marker for ocular inflammatory diseases.

IBEX: an iterative immunolabeling and chemical bleaching method for high-content imaging of diverse tissues.

High-content imaging is needed to catalog the variety of cellular phenotypes and multicellular ecosystems present in metazoan tissues. We recently developed iterative bleaching extends multiplexity (IBEX), an iterative immunolabeling and chemical bleaching method that enables multiplexed imaging (>65 parameters) in diverse tissues, including human organs relevant for international consortia efforts. IBEX is compatible with >250 commercially available antibodies and 16 unique fluorophores, and can be easily adopted to different imaging platforms using slides and nonproprietary imaging chambers. The overall protocol consists of iterative cycles of antibody labeling, imaging and chemical bleaching that can be completed at relatively low cost in 2-5 d by biologists with basic laboratory skills. To support widespread adoption, we provide extensive details on tissue processing, curated lists of validated antibodies and tissue-specific panels for multiplex imaging. Furthermore, instructions are included on how to automate the method using competitively priced instruments and reagents. Finally, we present a software solution for image alignment that can be executed by individuals without programming experience using open-source software and freeware. In summary, IBEX is a noncommercial method that can be readily implemented by academic laboratories and scaled to achieve high-content mapping of diverse tissues in support of a Human Reference Atlas or other such applications.

Optogenetic therapy restores retinal activity in primate for at least a year following photoreceptor ablation.

All retina-based vision restoration approaches rely on the assumption that photoreceptor loss does not preclude reactivation of the remaining retinal architecture. Whether extended periods of vision loss limit the efficacy of restorative therapies at the retinal level is unknown. We examined long-term changes in optogenetic responsivity of foveal retinal ganglion cells (RGCs) in non-human primates following localized photoreceptor ablation by high-intensity laser exposure. By performing fluorescence adaptive optics scanning light ophthalmoscopy (AOSLO) of RGCs expressing both the calcium indicator GCaMP6s and the optogenetic actuator ChrimsonR, it was possible to track optogenetic-mediated calcium responses in deafferented RGCs over time. Fluorescence fundus photography revealed a 40% reduction in ChrimsonR fluorescence from RGCs lacking photoreceptor input over the 3 weeks following photoreceptor ablation. Despite this, in vivo imaging revealed good cellular preservation of RGCs 3 months after the loss of photoreceptor input, and histology confirmed good structural preservation at 2 years. Optogenetic responses of RGCs in primate persisted for at least 1 year after the loss of photoreceptor input, with a sensitivity index similar to optogenetic responses recorded in intact retina. These results are promising for all potential therapeutic approaches to vision restoration that rely on preservation and reactivation of RGCs.

Understanding immunity in a tissue-centric context: Combining novel imaging methods and mathematics to extract new insights into function and dysfunction.

A central question in immunology is what features allow the immune system to respond in a timely manner to a variety of pathogens encountered at unanticipated times and diverse body sites. Two decades of advanced and static dynamic imaging methods have now revealed several major principles facilitating host defense. Suborgan spatial prepositioning of distinct cells promotes time-efficient interactions upon pathogen sensing. Such pre-organization also provides an effective barrier to movement of pathogens from parenchymal tissues into the blood circulation. Various molecular mechanisms maintain effective intercellular communication among otherwise rapidly moving cells. These and related discoveries have benefited from recent increases in the number of parameters that can be measured simultaneously in a single tissue section and the extension of such multiplex analyses to 3D tissue volumes. The application of new computational methods to such imaging data has provided a quantitative, in vivo context for cell trafficking and signaling pathways traditionally explored in vitro or with dissociated cell preparations. Here, we summarize our efforts to devise and employ diverse imaging tools to probe immune system organization and function, concluding with a commentary on future developments, which we believe will reveal even more about how the immune system operates in health and disease.

Engineering adeno-associated viral vectors to evade innate immune and inflammatory responses.

Nucleic acids are used in many therapeutic modalities, including gene therapy, but their ability to trigger host immune responses in vivo can lead to decreased safety and efficacy. In the case of adeno-associated viral (AAV) vectors, studies have shown that the genome of the vector activates Toll-like receptor 9 (TLR9), a pattern recognition receptor that senses foreign DNA. Here, we engineered AAV vectors to be intrinsically less immunogenic by incorporating short DNA oligonucleotides that antagonize TLR9 activation directly into the vector genome. The engineered vectors elicited markedly reduced innate immune and T cell responses and enhanced gene expression in clinically relevant mouse and pig models across different tissues, including liver, muscle, and retina. Subretinal administration of higher-dose AAV in pigs resulted in photoreceptor pathology with microglia and T cell infiltration. These adverse findings were avoided in the contralateral eyes of the same animals that were injected with the engineered vectors. However, intravitreal injection of higher-dose AAV in macaques, a more immunogenic route of administration, showed that the engineered vector delayed but did not prevent clinical uveitis, suggesting that other immune factors in addition to TLR9 may contribute to intraocular inflammation in this model. Our results demonstrate that linking specific immunomodulatory noncoding sequences to much longer therapeutic nucleic acids can "cloak" the vector from inducing unwanted immune responses in multiple, but not all, models. This "coupled immunomodulation" strategy may widen the therapeutic window for AAV therapies as well as other DNA-based gene transfer methods.

Intraoperative Complications and Visual Outcomes of Cataract Surgery in Diabetes Mellitus: A Multicenter Database Study.

PURPOSE: To compare the visual outcome and the rate of intraoperative complications in eyes of diabetic and nondiabetic patients undergoing phacoemulsification over 15 years. DESIGN: Retrospective clinical cohort study. METHODS: Data of 179,159 eyes that underwent phacoemulsification at 8 centers were classified based on the presence or absence of diabetes mellitus. Visual acuity (VA) was defined as the best value of uncorrected or corrected distance measure available. For the VA analysis, eyes with co-pathologies or combined surgical procedures were further excluded, leaving a subset of 90,729 eyes. Main outcome measures were logarithm of the minimum angle of resolution (logMAR) VA at 4-12 weeks postoperatively, and rate of intraoperative complications. RESULTS: Cataract surgery in eyes of diabetic patients was associated with an improvement in mean VA of 0.48 logMAR (5 Snellen lines). Mean postoperative VA was slightly worse in diabetic compared to nondiabetic group (logMAR 0.23 vs 0.13; Snellen 20/30 vs 20/25; P < .0001) and the proportions of eyes achieving a visual gain of ≥3 Snellen lines (≥0.3 logMAR) was lower in the diabetic group (56.6% vs 63.5%; P < .0001). There was a linear relationship between diabetic retinopathy severity and worse postoperative visual acuity (ß coefficient 0.098 to 0.288; P < .0001). We observed higher rates of posterior capsule rupture (2.3% vs 1.6%; P < .001) and dropped nuclear fragments (0.3% vs 0.2%; P < .001) in the diabetic group. CONCLUSIONS: Postoperative VA negatively correlated with diabetes and diabetic retinopathy severity. Eyes of diabetic subjects had higher risks of posterior capsule rupture.

Gene Therapy of Dominant CRX-Leber Congenital Amaurosis using Patient Stem Cell-Derived Retinal Organoids.

Mutations in the photoreceptor transcription factor gene cone-rod homeobox (CRX) lead to distinct retinopathy phenotypes, including early-onset vision impairment in dominant Leber congenital amaurosis (LCA). Using induced pluripotent stem cells (iPSCs) from a patient with CRX-I138fs48 mutation, we established an in vitro model of CRX-LCA in retinal organoids that showed defective photoreceptor maturation by histology and gene profiling, with diminished expression of visual opsins. Adeno-associated virus (AAV)-mediated CRX gene augmentation therapy partially restored photoreceptor phenotype and expression of phototransduction-related genes as determined by single-cell RNA-sequencing. Retinal organoids derived from iPSCs of a second dominant CRX-LCA patient carrying K88N mutation revealed the loss of opsin expression as a common phenotype, which was alleviated by AAV-mediated augmentation of CRX. Our studies provide a proof-of-concept for developing gene therapy of dominant CRX-LCA and other CRX retinopathies.

IBEX: A versatile multiplex optical imaging approach for deep phenotyping and spatial analysis of cells in complex tissues.

The diverse composition of mammalian tissues poses challenges for understanding the cell-cell interactions required for organ homeostasis and how spatial relationships are perturbed during disease. Existing methods such as single-cell genomics, lacking a spatial context, and traditional immunofluorescence, capturing only two to six molecular features, cannot resolve these issues. Imaging technologies have been developed to address these problems, but each possesses limitations that constrain widespread use. Here we report a method that overcomes major impediments to highly multiplex tissue imaging. "Iterative bleaching extends multiplexity" (IBEX) uses an iterative staining and chemical bleaching method to enable high-resolution imaging of >65 parameters in the same tissue section without physical degradation. IBEX can be employed with various types of conventional microscopes and permits use of both commercially available and user-generated antibodies in an "open" system to allow easy adjustment of staining panels based on ongoing marker discovery efforts. We show how IBEX can also be used with amplified staining methods for imaging strongly fixed tissues with limited epitope retention and with oligonucleotide-based staining, allowing potential cross-referencing between flow cytometry, cellular indexing of transcriptomes and epitopes by sequencing, and IBEX analysis of the same tissue. To facilitate data processing, we provide an open-source platform for automated registration of iterative images. IBEX thus represents a technology that can be rapidly integrated into most current laboratory workflows to achieve high-content imaging to reveal the complex cellular landscape of diverse organs and tissues.

Label-free imaging of immune cell dynamics in the living retina using adaptive optics.

Our recent work characterized the movement of single blood cells within the retinal vasculature (Joseph et al. 2019) using adaptive optics ophthalmoscopy. Here, we apply this technique to the context of acute inflammation and discover both infiltrating and tissue-resident immune cells to be visible without any labeling in the living mouse retina using near-infrared light alone. Intravital imaging of immune cells can be negatively impacted by surgical manipulation, exogenous dyes, transgenic manipulation and phototoxicity. These confounds are now overcome, using phase contrast and time-lapse videography to reveal the dynamic behavior of myeloid cells as they interact, extravasate and survey the mouse retina. Cellular motility and differential vascular responses were measured noninvasively and in vivo across hours to months at the same retinal location, from initiation to the resolution of inflammation. As comparable systems are already available for clinical research, this approach could be readily translated to human application.

Intraocular Pressure Reduction After Real-world Cataract Surgery.

PRéCIS:: A large cohort undergoing cataract extraction was retrospectively analyzed to ascertain the degree of real-world intraocular pressure (IOP) reduction in normal eyes and those with glaucoma, and a predictive formula was developed. PURPOSE: The purpose of this study was to define the real-world degree of IOP reduction after cataract extraction to guide its role as an isolated intervention for glaucoma. MATERIALS AND METHODS: A retrospective analysis was carried out of clinical data collected in 8 clinical sites in the United Kingdom from an electronic medical record system between January 2006 and May 2015. A total of 20,508 eyes without known pathology and 2251 eyes from patients with glaucoma undergoing phacoemulsification and intraocular lens insertion were included. Eyes with intraoperative complications, undergoing additional procedures, axial lengths outside 22 to 26.5 mm, preoperative IOP under 6 mm Hg or over 30 mm Hg, and copathology, except for amblyopia or glaucoma, were excluded. The main outcome measure was the change in preoperative IOP compared with the next recorded visit for up to 12 weeks. RESULTS: In eyes without pathology, the mean reduction in IOP was 1.40 mm Hg (±3.74) compared with 1.03 (±5.02), P-value <0.001, in eyes with a diagnosis of glaucoma. A multiple linear regression model identified preoperative IOP, a glaucoma diagnosis, preoperative corrected visual acuity, age, and axial length as determinants of IOP reduction. The model was validated against an independent cohort. CONCLUSIONS: We quantify mean IOP reduction achieved in a real-world setting from cataract surgery alone. In glaucomatous eyes where angle closure is not differentiated, phacoemulsification alone yields only a modest reduction of IOP.

Intravenous indocyanine green dye is insufficient for robust immune cell labelling in the human retina.

It is not currently possible to reliably visualise and track immune cells in the human central nervous system or eye. Previous work demonstrated that indocyanine green (ICG) dye could label immune cells and be imaged after a delay during disease in the mouse retina. We report a pilot study investigating if ICG can similarly label immune cells within the human retina. Twelve adult participants receiving ICG angiography as part of routine standard of care were recruited. Baseline retinal images were obtained prior to ICG administration then repeated over a period ranging from 2 hours to 9 days. Matched peripheral blood samples were obtained to examine systemic immune cell labelling and activation from ICG by flow cytometry with human macrophage cultures as positive controls. Differences between the delayed near infrared ICG imaging and 488 nm autofluorescence was observed across pathologies, likely arising from the retinal pigment epithelium (RPE). Only one subject demonstrated ICG signal on peripheral blood myeloid cells and only three distinct cell-sized signals appeared over time within the retina of three participants. No significant increase in immune cell activation markers were detected after ICG administration. ICG accumulated in the endosomes of macrophage cultures and was detectable above a minimum concentration, suggesting cell labelling is possible. ICG can label RPE and may be used as an additional biomarker for RPE health across a range of retinal disorders. Standard clinical doses of intravenous ICG do not lead to robust immune cell labelling in human blood or retina and further optimisation in dose and route are required.

Gene Therapy for Glaucoma by Ciliary Body Aquaporin 1 Disruption Using CRISPR-Cas9.

Glaucoma is a common cause of blindness, yet current therapeutic options are imperfect. Clinical trials have invariably shown that reduction in intraocular pressure (IOP) regardless of disease subtype prevents visual loss. Reducing ciliary body aqueous humor production can lower IOP, and the adeno-associated virus ShH10 serotype was identified as able to transduce mouse ciliary body epithelium following intravitreal injection. Using ShH10 to deliver a single vector CRISPR-Cas9 system disrupting Aquaporin 1 resulted in reduced IOP in treated eyes (10.4 ± 2.4 mmHg) compared with control (13.2 ± 2.0 mmHg) or non-injected eyes (13.1 ± 2.8 mmHg; p < 0.001; n = 12). Editing in the aquaporin 1 gene could be detected in ciliary body, and no off-target increases in corneal or retinal thickness were identified. In experimental mouse models of corticosteroid and microbead-induced ocular hypertension, IOP could be reduced to prevent ganglion cell loss (32 ± 4 /mm2) compared with untreated eyes (25 ± 5/mm2; p < 0.01). ShH10 could transduce human ciliary body from post-mortem donor eyes in ex vivo culture with indel formation detectable in the Aquaporin 1 locus. Clinical translation of this approach to patients with glaucoma may permit long-term reduction of IOP following a single injection.

Single Eye mRNA-Seq Reveals Normalisation of the Retinal Microglial Transcriptome Following Acute Inflammation.

Background: Whether retinal microglia can maintain or restore immune homeostasis during and after inflammation is unclear. We performed single-eye mRNA-sequencing on microglia at different timepoints following a single inflammatory stimulus to characterise their transcriptome during and after resolution of endotoxin-induced uveitis (EIU). Experimental Approach:Cx3cr1CreER:R26-tdTomato (C57BL/6) male heterozygotes were administered tamoxifen via different regimes at 4-5 weeks of age. Four weeks post-tamoxifen, mice were injected intravitreally with 10 ng lipopolysaccharide (endotoxin induced uveitis, EIU). Six-hundred retinal microglia were obtained by FACS from individual naïve retinas and at 4 h, 18 h, and 2 weeks following EIU induction. Samples were sequenced to a depth of up to 16.7 million reads using the SMART-Seq v4 Ultra Low Input RNA kit. The data was analysed using Partek software and Ingenuity Pathway Analysis. Genes were considered differentially-expressed (DEG) if the FDR step-up p-value was ≤0.05 and the fold-change was ≥±2. Results: Flow cytometric analysis indicates that the Cx3cr1CreER:R26-tdTomato strain is both sensitive (>95% tagging) and specific (>95% specificity) for microglia when tamoxifen is administered topically to the eye for 3 days. During "early" activation, 613 DEGs were identified. In contrast, 537 DEGs were observed during peak cellular infiltrate and none at 2 weeks, compared to baseline controls (1,069 total unique DEGs). Key marker changes were validated by qPCR, flow cytometry, and fluorescence microscopy. C5AR1 was identified and validated as a robust marker of differentiating microglial subsets during an LPS response. Conclusion: Using EIU to provide a single defined inflammatory stimulus, mRNA-Seq identified acute transcriptional changes in retinal microglia which returned to their original transcriptome after 2 weeks. Yolk-sac derived microglia are capable of restoring their homeostatic state after acute inflammation.

Reduced Macular Vessel Density and Capillary Perfusion in Glaucoma Detected Using OCT Angiography.

AIMS: To evaluate retinal vasculature changes in primary open-angle glaucoma (POAG) and whether the functional visual loss correlates with parameters obtained using optical coherence tomography angiography (OCTA). MATERIALS AND METHODS: OCT and OCTA images were collected from 116 POAG eyes and 40 normal eyes in a prospective, cross-sectional observational study. Glaucomatous eyes were further divided into three groups according to a Glaucoma Staging System. Measurements of macular vessel density, ganglion cell complex (GCC), and disk retinal nerve fiber layer (RNFL) thickness were compared among groups. RESULTS: The macular vessel density, GCC, and RNFL are significantly reduced in POAG compared to normal eyes that also corresponds to the severity of glaucoma (Kruskal-Wallis test with Dunnett's correction; p < 0.0001). Visual field mean deviation correlates significantly with macular vessel density (p = 0.0028, r = 0.3), GCC (p < 0.0001, r = 0.6), and RNFL (p = 0.008, r = 0.36) in POAG. There are significant correlations between GCC and RNFL (p < 0.0001, r = 0.76) as well as macular vessel density (p < 0.0001, r = 0.48). Increased age also correlates with reduced macular vessel density in both normal (p = 0.0002, r = 0.49) and glaucomatous eyes (p < 0.0001, r = 0.48), but a greater proportionate reduction of vessel density is seen in glaucomatous eyes. CONCLUSION: Reduced macular vessel density occurs in POAG despite of age-related changes, which also correlates with reductions in RNFL and GCC measurements. OCTA can detect microstructural defects and offers potential to facilitate diagnosis of glaucoma.

A Database Study of Visual Outcomes and Intraoperative Complications of Postvitrectomy Cataract Surgery.

PURPOSE: To analyze the visual outcomes and rate of intraoperative complications of phacoemulsification surgery after prior pars plana vitrectomy (PPV). DESIGN: Retrospective, multicenter database study. PARTICIPANTS: Eyes that underwent phacoemulsification between June 2005 and March 2015 at 8 sites in the United Kingdom. METHODS: Study eyes were classified as vitrectomized (prior PPV group) or nonvitrectomized (reference group) depending on the vitreous state at the time of cataract surgery. Eyes with multiple intraocular surgeries or history of ocular diseases known to cause cataract progression or increased risk of intraoperative complications during phacoemulsification were excluded. MAIN OUTCOME MEASURES: Logarithm of the minimum angle of resolution (logMAR) visual acuity (VA), rate of intraoperative complications, and time interval to cataract surgery. RESULTS: Eyes in the prior PPV group (n = 2221) had worse preoperative logMAR VA (0.96±0.60 vs. 0.62±0.52, P < 0.0001), were from younger patients, and had longer axial lengths than the nonvitrectomized group (n = 136 533). At all postoperative time points measured up to 24 weeks, mean vision was poorer in the prior PPV group (0.41±0.47 vs. 0.17±0.29 at 4-12 weeks, P < 0.0001) and a smaller proportion of eyes achieved postoperative VA ≤0.30 logMAR (Snellen, ≥20/40) (60.8% vs. 86.5% at 4-12 weeks, P < 0.0001). The rate of posterior capsular rupture was not different between the prior PPV (1.5%) and the nonvitrectomized (1.7%) groups, but the incidences of zonular dialysis (1.3% vs. 0.6%) and dropped nuclear fragments (0.6% vs. 0.2%) were higher in the prior PPV group (P < 0.0001). The mean time interval between PPV and cataract surgery was 399 days. CONCLUSIONS: We found a significant improvement in VA with postvitrectomy cataract surgery. However, compared with eyes without prior PPV, there was a worse mean postoperative vision of 0.2 logMAR units, a higher rate of zonular dialysis and dropped nuclear fragments, and a similar rate of posterior capsule rupture.

Cataract Surgery Outcomes in Eyes With Primary Epiretinal Membrane.

Importance: Primary epiretinal membrane (ERM) is a common retinal disorder with a prevalence of 4% to 18.5%. Although ERM and cataracts commonly occur together, to our knowledge, no studies have investigated the outcome of cataract surgery alone in this setting. Objective: To analyze the visual outcome and cystoid macular edema risk with cataract surgery in eyes with primary ERM. Design, Setting, and Participants: In this retrospective clinical database study, data were collected from July 2003 to March 2015 from 8 locations in the United Kingdom. Cataract surgery data of 217 557 eyes were extracted from the electronic medical record of the UK National Health Service. After exclusion of 57 561 eyes with combined surgery, prior vitrectomy, copathology, and complications, 812 eyes with primary ERM and 159 184 reference eyes were analyzed. Main Outcomes and Measures: We report on visual acuity (VA), the incidence of cystoid macular edema, and the need for ERM surgery. Results: The mean (SD) age of patients in the ERM group was 73.7 (9.23) years, and 395 of 812 were men (46.8%). The mean (SD) age of patients in the reference group was 74.4 (12.19) years, and 65 265 of 159 184 were men (41%). Epiretinal membrane eyes assessed at 4 to 12 weeks postoperatively gained 0.27 (0.32) logMAR (approximately 3 Snellen lines), with 200 of 448 (44.6%) improving by 0.30 logMAR or more (≥3 Snellen lines) and 32 of 448 (7.1%) worsening by 0.30 logMAR or more. Reference eyes gained a mean (SD) of 0.44 (0.26) logMAR (approximately 4 Snellen lines), with 48 583 of 77 408 (62.8%) improving by 0.30 logMAR or more and 2125 of 77 408 (2.7%) worsening by 0.30 logMAR or more. Although all eyes with preoperative VA of 20/40 or less improved, only reference eyes with preoperative VA of more than 20/40 showed improvement. Cystoid macular edema developed in 57 of 663 ERM eyes (8.6%) (95% CI, 6.69-10.98) and 1731 of 125 435 reference eyes (1.38%) (95% CI, 1.32-1.45) (P < .001). Epiretinal membrane surgery was performed in 43 of 663 (6.5%) ERM eyes. Conclusions and Relevance: On average, VA improved 0.27 logMAR (approximately 3 Snellen lines) in eyes with ERM. Eyes with ERM and VA of 20/40 or less showed more benefit after cataract surgery than those with better preoperative vision. However, compared with eyes without ERM, higher rates of cystoid macular edema and a lower postoperative VA gain were noted.