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Abstract Background Arginine deprivation was previously shown to inhibit retinoblastoma cell proliferation and induce cell death in vitro. However, the mechanisms by which retinoblastoma cells respond to arginine deprivation remain to be elucidated. Methods The human-derived retinoblastoma cell lines Y79 and WERI-Rb-1 were subjected to arginine depletion, and the effects on inhibiting cell growth and survival were evaluated. This study investigated potential mechanisms, including autophagy, cell cycle arrest and apoptosis. Moreover, the roles of the general control nonderepressible 2 (GCN2) and mechanistic target of rapamycin complex 1 (mTORC1) signaling pathways in these processes were examined. Results We demonstrated that arginine deprivation effectively inhibited the growth of retinoblastoma cells in vitro. This treatment caused an increase in the autophagic response. Additionally, prolonged arginine deprivation induced G2 cell cycle arrest and was accompanied by an increase in early apoptotic cells. Importantly, arginine depletion also induced the activation of GCN2 and the inhibition of mTOR signaling. We also discovered that the activation of SLC7A11 was regulated by GCN2 upon arginine deprivation. Knockdown of SLC7A11 rendered retinoblastoma cells partially resistant to arginine deprivation. Furthermore, we found that knockdown of GCN2 led to a decrease in the autophagic response in WERI-Rb-1 cells and arrested more cells in S phase, which was accompanied by fewer apoptotic cells. Moreover, knockdown of GCN2 induced the constant expression of ATF4 and the phosphorylation of 70S6K and 4E-BP1 regardless of arginine deprivation. Conclusions Collectively, our findings suggest that the GCN2‒SLC7A11 axis regulates cell growth and survival upon arginine deprivation through coordinating autophagy, cell cycle arrest, and apoptosis in retinoblastoma cells. This work paves the way for the development of a novel treatment for retinoblastoma.

Abstract The increasing prevalence of myopia worldwide presents a significant public health challenge. A key strategy to combat myopia is with early detection and prediction in children as such examination allows for effective intervention using readily accessible imaging technique. To this end, we introduced DeepMyopia, an artificial intelligence (AI)-enabled decision support system to detect and predict myopia onset and facilitate targeted interventions for children at risk using routine retinal fundus images. Based on deep learning architecture, DeepMyopia had been trained and internally validated on a large cohort of retinal fundus images (n = 1,638,315) and then externally tested on datasets from seven sites in China (n = 22,060). Our results demonstrated robustness of DeepMyopia, with AUCs of 0.908, 0.813, and 0.810 for 1-, 2-, and 3-year myopia onset prediction with the internal test set, and AUCs of 0.796, 0.808, and 0.767 with the external test set. DeepMyopia also effectively stratified children into low- and high-risk groups (p

Purpose: This study aims to report correlations between thyroid-stimulating immunoglobulin (TSI) and both clinical and radiological parameters in recent-onset symptomatic thyroid eye disease (TED) patients. Methods: A prospective cohort study of TED patients managed at the Chinese University of Hong Kong from January 2014 to May 2022. Serum TSI levels were determined with the functional assay. Outcomes included the Clinical Activity Score (CAS), marginal reflex distance1 (MRD1), extraocular muscle motility restriction (EOMy), exophthalmos, and diplopia. The radiological assessment included cross-sectional areas and signal of extraocular muscles on STIR-sequence MRI. Results: A total of 255 (197 female) treatment-naive patients, with an average onset age of 50 ± 14 years (mean ± s.d.), were included. Elevated pre-treatment TSI level was observed in 223 (88%) patients. There was a weak positive correlation between TSI and CAS (r = 0.28, P = 0.000031), MRD1 (r = 0.17, P = 0.0080), and the size of the levator palpebrae superioris/superior rectus complex (r = 0.25, P = 0.018). No significant correlation existed between TSI and STIR signals. The AUC and optimal cut-off value for clinical active TED were 0.67 (95% CI: 0.60–0.75) and 284% (specificity: 50%, sensitivity: 85%). In total, 64 patients received intravenous methylprednisolone (IVMP) during the study interval, and they had a higher baseline TSI level than those who did not have IVMP (P = 0.000044). Serial post-IVMP TSI among the 62 patients showed a significant reduction compared to the baseline level (P < 0.001). Both the baseline and post-IVMP TSI levels, and percentages of TSI changes were comparable between patients who responded and did not respond to the first course of IVMP. Conclusion: TSI can be a serum biomarker for the diagnosis, prognosis, and treatment response of TED. Further validation should be warranted.

This study explores the potential of Artificial Intelligence (AI) in early screening and prognosis of Dry Eye Disease (DED), aiming to enhance the accuracy of therapeutic approaches for eye-care practitioners. Despite the promising opportunities, challenges such as diverse diagnostic evidence, complex etiology, and interdisciplinary knowledge integration impede the interpretability, reliability, and applicability of AI-based DED detection methods. The research conducts a comprehensive review of datasets, diagnostic evidence, and standards, as well as advanced algorithms in AI-based DED detection over the past five years. The DED diagnostic methods are categorized into three groups based on their relationship with AI techniques: (1) those with ground truth and/or comparable standards, (2) potential AI-based methods with significant advantages, and (3) supplementary methods for AI-based DED detection. The study proposes suggested DED detection standards, the combination of multiple diagnostic evidence, and future research directions to guide further investigations. Ultimately, the research contributes to the advancement of ophthalmic disease detection by providing insights into knowledge foundations, advanced methods, challenges, and potential future perspectives, emphasizing the significant role of AI in both academic and practical aspects of ophthalmology.

Abstract Introduction The aim of this work is to determine the interocular differences in peripapillary retinal nerve fiber layer (p-RNFL) thickness and its associations among school children in Hong Kong. Methods We conducted a population-based study including 4034 children aged 6–8 years from the Hong Kong Children Eye Study (HKCES). All participants received comprehensive ocular examinations where p-RNFL thickness was measured using spectral-domain optical coherence tomography (SD-OCT). The degree of symmetry between both eyes was analyzed and represented by intraclass correlation coefficient (ICC) values. Multivariable linear regression analysis was used to investigate the associations between ocular and systemic factors with p-RNFL thickness difference. Results The study included 4034 children with a mean age of 7.61 ± 0.98 years. The mean global p-RNFL thickness was 106.60 ± 9.41 μm in right eyes and 105.99 ± 9.30 μm in left eyes. The ICC for global p-RNFL difference was 0.866 (95% CI 0.858–0.873, p

Abstract Failure to recognize samples from the classes unseen during training is a major limitation of artificial intelligence in the real-world implementation for recognition and classification of retinal anomalies. We establish an uncertainty-inspired open set (UIOS) model, which is trained with fundus images of 9 retinal conditions. Besides assessing the probability of each category, UIOS also calculates an uncertainty score to express its confidence. Our UIOS model with thresholding strategy achieves an F1 score of 99.55%, 97.01% and 91.91% for the internal testing set, external target categories (TC)-JSIEC dataset and TC-unseen testing set, respectively, compared to the F1 score of 92.20%, 80.69% and 64.74% by the standard AI model. Furthermore, UIOS correctly predicts high uncertainty scores, which would prompt the need for a manual check in the datasets of non-target categories retinal diseases, low-quality fundus images, and non-fundus images. UIOS provides a robust method for real-world screening of retinal anomalies.

Abstract Background A recent prospective demonstrated that cardiovascular risk factors in early childhood were associated with later cardiovascular events. However, the impact of secondhand smoke (SHS) on children is unclear. The aims of this study is to determine the effects of SHS exposure on the retinal vasculature of children. Methods This is a population-based cross-sectional study of children aged 6 to 8 years. All participants received comprehensive ophthalmic examinations and retinal photography. Data on SHS exposure was derived from a validated questionnaire. A validated deep-learning system was used to automatically estimate retinal arteriolar and venular calibers from retinal photographs. Associations of quantitative retinal vessel caliber values with SHS exposure, number of smokers in the household, and total number of cigarettes smoked were determined by analyses of covariance (ANCOVA) after adjusting for potential confounders. Test of trend was determined by treating categorical risk factors as continuous ordinal variables. Results Here we show children exposed to SHS have wider retinal arteriolar (CRAE 152.1 µm vs. 151.3 µm, p

Cyclic GMP-AMP (cGAMP) synthase (cGAS), a sensor of cytosolic DNA, recognizes cytoplasmic nucleic acids to activate the innate immune responses via generation of the second messenger cGAMP and subsequent activation of the stimulator of interferon genes (STINGs). The cGAS-STING signaling has multiple immunologic and physiological functions in all human vital organs. It mediates protective innate immune defense against DNA-containing pathogen infection, confers intrinsic antitumor immunity via detecting tumor-derived DNA, and gives rise to autoimmune and inflammatory diseases upon aberrant activation by cytosolic leakage of self-genomic and mitochondrial DNA. Disruptions in these functions are associated with the pathophysiology of various immunologic and neurodegenerative diseases. Recent evidence indicates important roles of the cGAS-STING signaling in mediating inflammatory responses in ocular inflammatory and inflammation-associated diseases, such as keratitis, diabetic retinopathy, age-related macular degeneration, and uveitis. In this review, we summarize the recently emerging evidence of cGAS-STING signaling in mediating ocular inflammatory responses and affecting pathogenesis of these complex eye diseases. We attempt to provide insightful perspectives on future directions of investigating cGAS-STING signaling in ocular inflammation. Understanding how cGAS-STING signaling is modulated to mediate ocular inflammatory responses would allow future development of novel therapeutic strategies to treat ocular inflammation and autoimmunity.

Abstract Neovascular age-related macular degeneration (nAMD), along with its clinical subtype known as polypoidal choroidal vasculopathy (PCV), are among the leading causes of vision loss in elderly Asians. In a genome-wide association study (GWAS) comprising 3,128 nAMD (1,555 PCV and 1,573 typical nAMD), and 5,493 controls of East Asian ancestry, we identify twelve loci, of which four are novel ( $$P \, < \, 1.19\times {10}^{-8}$$ P < 1.19 × 10 − 8 ). Substantial genetic sharing between PCV and typical nAMD is noted (r g = 0.666), whereas collagen extracellular matrix and fibrosis-related pathways are more pronounced for PCV. Whole-exome sequencing in 259 PCV patients revealed functional rare variants burden in collagen type I alpha 1 chain gene (COL1A1; $$P=1.05\times {10}^{-6}$$ P = 1.05 × 10 − 6 ) and potential enrichment of functional rare mutations at AMD-associated loci. At the GATA binding protein 5 (GATA5) locus, the most significant GWAS novel loci, the expressions of genes including laminin subunit alpha 5 (Lama5), mitochondrial ribosome associated GTPase 2 (Mtg2), and collagen type IX alpha 3 chain (Col9A3), are significantly induced during retinal angiogenesis and subretinal fibrosis in murine models. Furthermore, retinoic acid increased the expression of LAMA5 and MTG2 in vitro. Taken together, our data provide insights into the genetic basis of AMD pathogenesis in the Asian population.

Abstract Background Thyroid eye disease is an extrathyroidal manifestation of Graves’ disease and is associated with dry eye disease. This is the first systematic review and meta-analysis to evaluate the role of magnetic resonance imaging lacrimal gland parameters in thyroid eye disease diagnosis, activity grading, and therapeutic responses prediction. Methods Up to 23 August, 2022, 504 studies from PubMed and Cochrane Library were analyzed. After removing duplicates and imposing selection criteria, nine eligible studies were included. Risk of bias assessment was done. Meta-analyses were performed using random-effect model if heterogeneity was significant. Otherwise, fixed-effect model was used. Main outcome measures include seven structural magnetic resonance imaging parameters (lacrimal gland herniation, maximum axial area, maximum coronal area, maximum axial length, maximum coronal length, maximum axial width, maximum coronal width), and three functional magnetic resonance imaging parameters (diffusion tensor imaging-fractional anisotropy, diffusion tensor imaging-apparent diffusion coefficient or mean diffusivity, diffusion-weighted imaging-apparent diffusion coefficient). Results Thyroid eye disease showed larger maximum axial area, maximum coronal area, maximum axial length, maximum axial width, maximum coronal width, diffusion tensor imaging-apparent diffusion coefficient/ mean diffusivity, and lower diffusion tensor imaging-fractional anisotropy than controls. Active thyroid eye disease showed larger lacrimal gland herniation, maximum coronal area, diffusion-weighted imaging-apparent diffusion coefficient than inactive. Lacrimal gland dimensional (maximum axial area, maximum coronal area, maximum axial length, maximum axial width, maximum coronal width) and functional parameters (diffusion tensor imaging-apparent diffusion coefficient, diffusion tensor imaging-apparent diffusion coefficient) could be used for diagnosing thyroid eye disease; lacrimal gland herniation, maximum coronal area, and diffusion-weighted imaging-apparent diffusion coefficient for differentiating active from inactive thyroid eye disease; diffusion tensor imaging parameters (diffusion tensor imaging-fractional anisotropy, diffusion tensor imaging-mean diffusivity) and lacrimal gland herniation for helping grading and therapeutic responses prediction respectively. Conclusions Magnetic resonance imaging lacrimal gland parameters can detect active thyroid eye disease and differentiate thyroid eye disease from controls. Maximum coronal area is the most effective indicator for thyroid eye disease diagnosis and activity grading. There are inconclusive results showing whether structural or functional lacrimal gland parameters have diagnostic superiority. Future studies are warranted to determine the use of magnetic resonance imaging lacrimal gland parameters in thyroid eye disease.

Abstract Dysregulation of Th17 cell differentiation and pathogenicity contributes to multiple autoimmune and inflammatory diseases. Previously growth hormone releasing hormone receptor (GHRH-R) deficient mice have been reported to be less susceptible to the induction of experimental autoimmune encephalomyelitis. Here, we show GHRH-R is an important regulator of Th17 cell differentiation in Th17 cell-mediated ocular and neural inflammation. We find that GHRH-R is not expressed in naïve CD4+ T cells, while its expression is induced throughout Th17 cell differentiation in vitro. Mechanistically, GHRH-R activates the JAK-STAT3 pathway, increases the phosphorylation of STAT3, enhances both non-pathogenic and pathogenic Th17 cell differentiation and promotes the gene expression signatures of pathogenic Th17 cells. Enhancing this signaling by GHRH agonist promotes, while inhibiting this signaling by GHRH antagonist or GHRH-R deficiency reduces, Th17 cell differentiation in vitro and Th17 cell-mediated ocular and neural inflammation in vivo. Thus, GHRH-R signaling functions as a critical factor that regulates Th17 cell differentiation and Th17 cell-mediated autoimmune ocular and neural inflammation.

ABSTRACTTo evaluate the effectiveness of smartphone ophthalmoscope (SO) in teaching ophthalmoscopy, compared with direct ophthalmoscope (DO). In this cross-over study, 45 final-year medical students attending sessions at a single institution were randomly allocated to two groups (A and B). Both groups attended two training sessions. In the first session, Group A students were taught ophthalmoscopy using DO and Group B students using SO. In the second session, the training sessions were crossed over. A series of eye models with 10 letters placed on the inner surface were designed to assess the students’ skill on ophthalmoscopy. Students performed ophthalmoscopy on the eye models, recorded their findings, and completed a questionnaire of feedback on DO and SO. The main outcome measure was the score of ophthalmoscopy, assessed by the student correctly recording each letter (score 1 for each letter). For Group A, the mean score of ophthalmoscopy on the eye model using DO and SO was 3.9±2.4 and 8.2±2.2, respectively. For Group B, the mean score of ophthalmoscopy on the eye model using SO and DO 8.7±1.8 and 5.7±3.5 . Students scored significantly higher in ophthalmoscopy when using SO than DO (P

Abstract Introduction This study aimed to evaluate the habitual reading distance among non-myopic children and also myopic children with undercorrection and with full correction. Methods This was a population-based cross-sectional study with a total of 2363 children aged 6–8 years who were recruited from the Hong Kong Children Eye Study. Cycloplegic autorefraction, subjective refraction, habitual visual acuity, and best corrected visual acuity were measured. The entire reading process (9 min) was recorded using a hidden video camera placed 5 m away from the reading desk. Reading distances were taken at 6, 7, 8, and 9 min after the child began reading and were measured using a customized computer program developed in MATLAB. The main outcome was the association of habitual reading distances with refraction status. Habitual reading distances of children were documented via video camera footage. Results The habitual reading distances of undercorrected myopic children (23.37 ± 4.31 cm) were the shortest when compared to non-myopic children (24.20 ± 4.73 cm, P = 0.002) and fully corrected myopic children (24.81 ± 5.21 cm, P

Abstract Background To determine the association of the ANGPT2 gene with primary open-angle glaucoma (POAG) in Chinese. Methods Six single-nucleotide polymorphisms (SNPs) in ANGPT2 (rs2515487, rs2922869, rs13255574, rs4455855, rs13269021, and rs11775442) were genotyped in a total of 2601 study subjects from two cohorts. One is a Hong Kong Chinese cohort of 484 high tension glaucoma (HTG) and 537 normal tension glaucoma (NTG) patients, and 496 non-glaucoma control subjects. Another cohort is a Shantou Chinese cohort of 403 HTG and 135 NTG patients, and 543 non-glaucoma control subjects. Subgroup analysis by sex was conducted. Outcomes from different cohorts were combined for meta-analysis. Results The association of SNP rs11775442 with NTG in the Hong Kong cohort [P = 0.0498, OR = 1.24, 95% confidence interval (CI) 1.00–1.55] after adjusting for age and sex did not reach statistical significance after Bonferroni correction. Other SNPs were not significantly associated with NTG, HTG and POAG in individual cohort or in the combined analyses (P > 0.05). In the subgroup analysis by sex, SNP rs13269021 in the Shantou cohort, but not in the Hong Kong cohort, was significantly associated with NTG in males (P = 0.0081, OR = 1.67, 95% CI: 1.14–2.43) but not in females (P = 0.874). In the combined analyses by sex, no SNPs were significantly associated with NTG, HTG and POAG. Conclusions In the subgroup analysis by sex, a significant association was shown in SNP rs13269021 with NTG in Shantou males, but not in Hong Kong males. Further studies are needed to verify the association between ANGPT2 locus (rs13269021) and NTG in Chinese males.

Retinal detachment (RD) can result in the loss of photoreceptors that cause vision impairment and potential blindness. This study explores the protective effects of the oral administration of green tea extract (GTE) in a rat model of RD. Various doses of GTE or epigallocatechin gallate (EGCG), the most active ingredient in green tea catechins, were administered to Sprague Dawley (SD) rats with experimentally induced retinal detachment. The rats received sub-retinal injections of hyaluronic acid (0.1%) to induce RD and were given different doses of GTE and EGCG twice daily for three days. Notably, a low dose of GTE (142.9 mg/kg) caused significantly higher signal amplitudes in electroretinograms (ERGs) compared to higher GTE doses and any doses of EGCG. After administration of a low dose of GTE, the outer nuclear layer thickness, following normalization, of the detached retina reduced to 82.4 ± 8.2% (Mean ± SEM, p < 0.05) of the thickness by RD treatment. This thickness was similar to non-RD conditions, at 83.5 ± 4.7% (Mean ± SEM) of the thickness following RD treatment. In addition, the number of TUNEL-positive cells decreased from 76.7 ± 7.4 to 4.7 ± 1.02 (Mean ± SEM, p < 0.0001). This reduction was associated with the inhibition of apoptosis through decreased sphingomyelin levels and mitigation of oxidative stress shown by a lowered protein carbonyl level, which may involve suppression of HIF-1α pathways. Furthermore, GTE showed anti-inflammatory effects by reducing inflammatory cytokines and increasing resolving cytokines. In conclusion, low-dose GTE, but not EGCG, significantly alleviated RD-induced apoptosis, oxidative stress, inflammation, and energy insufficiency within a short period and without affecting energy metabolism. These findings suggest the potential of low-dose GTE as a protective agent for the retina in RD.

Abstract The retinoblastoma protein (pRb) functions as a cell cycle regulator controlling G1 to S phase transition and plays critical roles in tumour suppression. It is frequently inactivated in various tumours. The functions of pRb are tightly regulated, where post-translational modifications (PTMs) play crucial roles, including phosphorylation, ubiquitination, SUMOylation, acetylation and methylation. Most PTMs on pRb are reversible and can be detected in non-cancerous cells, playing an important role in cell cycle regulation, cell survival and differentiation. Conversely, altered PTMs on pRb can give rise to anomalies in cell proliferation and tumourigenesis. In this review, we first summarize recent findings pertinent to how individual PTMs impinge on pRb functions. As many of these PTMs on pRb were published as individual articles, we also provide insights on the coordination, either collaborations and/or competitions, of the same or different types of PTMs on pRb. Having a better understanding of how pRb is post-translationally modulated should pave the way for developing novel and specific therapeutic strategies to treat various human diseases.

PurposeThis study aims to investigate the effectiveness of interventions to control myopia progression. In this systematic review, the primary outcomes were mean differences (MD) between treatment and control groups in myopia progression (D) and axial length (AL) elongation (mm).ResultsThe following interventions were found to be effective (p

Both inflammation and anti-inflammation are involved in the protection of retinal cells. Antagonists of the hypothalamic growth hormone-releasing hormone receptor (GHRHR) have been shown to possess potent anti-inflammatory properties in experimental disease models of various organs, some with systemic complications. Such effects are also found in ocular inflammatory and neurologic injury studies. In experimental models of mice and rats, both growth hormone-releasing hormone receptor agonists and antagonists may alleviate death of ocular neural cells under certain experimental conditions. This review explores the properties of growth hormone-releasing hormone receptor agonists and antagonists that lead to its protection against inflammatory responses induced by extrinsic agents or neurologic injures in ocular animal models.

The rising prevalence of myopia is a major global public health concern. Economic evaluation of myopia interventions is critical for maximizing the benefits of treatment and the healthcare system. This systematic review aimed to evaluate the cost-effectiveness of interventions for treating myopia. Five databases were searched – Embase, Emcare, PubMed, Web of Science, and ProQuest – from inception to July 2022 and a total of 2,099 articles were identified. After careful assessments, 6 studies met the eligibility criteria. The primary outcomes of this systematic review were costs, quality-adjusted life years (QALYs), and incremental cost-effectiveness ratio (ICER). The secondary outcomes included utility values and net monetary benefits (NMB). One study determined the cost-effectiveness of photorefractive screening plus treatment with 0.01% atropine, 2 studies examined cost-effectiveness of corneal refractive surgery, and 3 studies evaluated cost-effectiveness of commonly used therapies for pathologic myopia. Corneal refractive surgeries included laser in situ keratomileusis (LASIK), femtosecond laser-assisted in situ keratomileusis (FS-LASIK), photorefractive keratectomy (PRK), and small-incision lenticule extraction (SMILE). Interventions for pathologic myopia included ranibizumab, conbercept, and photodynamic therapy (PDT). At an incremental cost of NZ$ 18 (95% CI 15, 20) (US$ 11) per person, photorefractive screening plus 0.01% atropine resulted in an ICER of NZ$ 1,590/QALY (US$ 1,001/QALY) (95% CI NZ$ 1,390, 1,791) for an incremental QALY of 0.0129 (95% CI 0.0127, 0.0131). The cost of refractive surgery in Europe ranged from €3,075 to €3,123 ([US$4,046 to $4,109 - adjusted to 2021 inflation). QALYs associated with these procedures were 23 (FS-LASIK) and 24 (SMILE and PRK) with utility values of 0.8 and ICERs ranging from approximately €14 (US$17)/QALY to €19 (US$23)/QALY. The ICER of LASIK was US$683/diopter gained (inflation-adjusted). The ICER of ranibizumab and PDT were £8,778 (US$12,032)/QALY and US$322,460/QALY respectively, with conbercept yielding a saving of 541,974 RMB (US$80,163)/QALY, respectively. The use of 0.01% atropine and corneal refractive surgery were cost-effective for treating myopia. Treating pathologic myopia with ranibizumab and conbercept were more cost-effective than PDT. Prevention of myopia progression is more cost-effective than treating pathologic myopia.

Myopic tilted disc is a common structural change of myopic eyes. With advancing ocular imaging technology, the associated structural changes of the eye, particularly the optic nerve head, have been extensively studied. These structural changes may increase patients’ susceptibility to axonal damage and the risk of developing serious optic neuropathies including glaucoma. They also lead to diagnostic difficulties of disease suspects and treatment dilemmas of patients, which implicate clinical practice and subsequently the health care system. In the context of the mounting prevalence of myopia worldwide and its implications to irreversible visual impairment and blindness, it is essential to gain a thorough understanding of the structural changes of myopia. Myopic tilted disc has been extensively investigated by different study groups. However, generalizing the knowledge could be difficult because of the variable definitions of myopic tilted disc utilized in these studies and the complexities of the changes. The current review aimed to clarify the concepts and discuss various aspects of myopic tilted disc, including the definitions, association with other myopia-related changes, mechanism of tilted disc development, structural and functional changes, and clinical implications.

Graphical Abstract

Glaucoma is the leading cause of irreversible blindness and visual impairment, affecting more than 80 million individuals worldwide. Oxidative stress and inflammation-induced neurodegenerative insults to retinal ganglion cells are the main pathogenesis of glaucoma. Retinal ganglion cells, the retinal neurons transmitting the visual signals to the visual cortex in the brain, have very limited regeneration or recovery capacity after damages. Apart from intraocular pressure-lowering treatments, there is still no clinically effective treatment to rescue the degeneration of retinal ganglion cells in glaucoma. Dietary antioxidants are easily accessible and can be applied as supplements assisting in the clinical treatments. Catechins, a chemical family of flavonoids, are the phenolic compounds found in many plants, especially in green tea. The anti-oxidative and anti-inflammatory properties of green tea catechins in vitro and in vivo have been well proven. They could be a potential treatment ameliorating retinal ganglion cell degeneration in glaucoma. In this review, the chemistry, pharmacokinetics, and therapeutic properties of green tea catechins were summarized. Research updates on the biological effects of green tea catechins in cellular and animal experimental glaucoma models were reviewed. In addition, clinical potentials of green tea catechins for glaucoma treatment were also highlighted.

Systems for automatic detection of a single disease may miss other important conditions. Here, the authors show a deep learning platform can detect 39 common retinal diseases and conditions.

Abstract The study aims to determine the prevalence of strabismus and its risk factors among school children in Hong Kong. This is a cross-sectional study involving 6–8 year old children from different districts in Hong Kong. 4273 children received comprehensive ophthalmological examination, cycloplegic auto-refraction, best corrected visual acuity (BCVA), anterior segment examination, cover/uncover test, ocular motility, and fundus examination. Demographic information, pre- and post- natal background, parental smoking status, and family history of strabismus were obtained through questionnaires. Strabismus was found among 133 children (3.11%, 95% CI 2.59–3.63%), including 117 (2.74%) exotropia and 12 (0.28%) esotropia cases (exotropia-esotropia ratio: 9.75:1). There was no significant difference in prevalence across age (6–8 years) and gender. Multivariate analysis revealed associations of strabismus with myopia (≤ − 1.00D; OR 1.61; 95% CI 1.03–2.52; P = 0.037) hyperopia (≥ + 2.00D; OR 2.49; 95% CI 1.42–4.39; P = 0.002), astigmatism (≥ + 2.00D; OR 2.32; 95% CI 1.36–3.94; P = 0.002), and anisometropia (≥ 2.00D; OR 3.21; 95% CI 1.36–7.55; P = 0.008). Other risk factors for strabismus included maternal smoking during pregnancy (OR 4.21; 95% CI 1.80–9.81; P = 0.001), family history of strabismus (OR 6.36; 95% CI 2.78–14.50, P 35 years; OR 1.65; CI 1.09–2.49, P = 0.018). The prevalence of strabismus among children aged 6—8 years in Hong Kong is 3.11%. Refractive errors, family history of strabismus and maternal smoking history during pregnancy are risk factors. Early correction of refractive errors and avoidance of maternal smoking during pregnancy are potentially helpful in preventing strabismus.

Abstract Retinitis Pigmentosa (RP) is a mostly incurable inherited retinal degeneration affecting approximately 1 in 4000 individuals globally. The goal of this work was to identify drugs that can help patients suffering from the disease. To accomplish this, we screened drugs on a zebrafish autosomal dominant RP model. This model expresses a truncated human rhodopsin transgene (Q344X) causing significant rod degeneration by 7 days post-fertilization (dpf). Consequently, the larvae displayed a deficit in visual motor response (VMR) under scotopic condition. The diminished VMR was leveraged to screen an ENZO SCREEN-WELL REDOX library since oxidative stress is postulated to play a role in RP progression. Our screening identified a beta-blocker, carvedilol, that ameliorated the deficient VMR of the RP larvae and increased their rod number. Carvedilol may directly on rods as it affected the adrenergic pathway in the photoreceptor-like human Y79 cell line. Since carvedilol is an FDA-approved drug, our findings suggest that carvedilol can potentially be repurposed to treat autosomal dominant RP patients.

Objective To develop and validate a real-world screening, guideline-based deep learning (DL) system for referable diabetic retinopathy (DR) detection.Design This is a multicentre platform development study based on retrospective, cross-sectional data sets. Images were labelled by two-level certificated graders as the ground truth. According to the UK DR screening guideline, a DL model based on colour retinal images with five-dimensional classifiers, namely image quality, retinopathy, maculopathy gradability, maculopathy and photocoagulation, was developed. Referable decisions were generated by integrating the output of all classifiers and reported at the image, eye and patient level. The performance of the DL was compared with DR experts.Setting DR screening programmes from three hospitals and the Lifeline Express Diabetic Retinopathy Screening Program in China.Participants 83 465 images of 39 836 eyes from 21 716 patients were annotated, of which 53 211 images were used as the development set and 30 254 images were used as the external validation set, split based on centre and period.Main outcomes Accuracy, F1 score, sensitivity, specificity, area under the receiver operating characteristic curve (AUROC), area under the precision-recall curve (AUPRC), Cohen’s unweighted κ and Gwet’s AC1 were calculated to evaluate the performance of the DL algorithm.Results In the external validation set, the five classifiers achieved an accuracy of 0.915–0.980, F1 score of 0.682–0.966, sensitivity of 0.917–0.978, specificity of 0.907–0.981, AUROC of 0.9639–0.9944 and AUPRC of 0.7504–0.9949. Referable DR at three levels was detected with an accuracy of 0.918–0.967, F1 score of 0.822–0.918, sensitivity of 0.970–0.971, specificity of 0.905–0.967, AUROC of 0.9848–0.9931 and AUPRC of 0.9527–0.9760. With reference to the ground truth, the DL system showed comparable performance (Cohen’s κ: 0.86–0.93; Gwet’s AC1: 0.89–0.94) with three DR experts (Cohen’s κ: 0.89–0.96; Gwet’s AC1: 0.91–0.97) in detecting referable lesions.Conclusions The automatic DL system for detection of referable DR based on the UK guideline could achieve high accuracy in multidimensional classifications. It is suitable for large-scale, real-world DR screening.

IntroductionGreen tea extract (GTE) alleviated ocular inflammations in endotoxin-induced uveitis (EIU) rat model induced by lipopolysaccharide (LPS) but the underlying mechanism is unclear.ObjectivesTo investigate the systematic and local mechanisms of the alleviation by untargeted metabolomics using liquid chromatography-tandem mass spectrometryMethodsSprague-Dawley rats were divided into control group, LPS treatment group, and LPS treatment group treated with GTE two hours after LPS injection. The eyes were monitored by slip lamp and electroretinography examination after 24 hours. The plasma and retina were collected for metabolomics analysisResultsIn LPS treated rats, the iris showed hyperemia. Plasma prostaglandins, arachidonic acids, corticosteroid metabolites, and bile acid metabolites increased. In the retina, histamine antagonists, corticosteroids, membrane phospholipids, free antioxidants, and sugars also increased but fatty acid metabolites, N-acetylglucosamine-6-sulphate, pyrocatechol, and adipic acid decreased. After GTE treatment, the a- and b- waves of electroretinography increased by 13%. Plasma phosphorylcholine lipids increased but plasma prostaglandin E1, cholanic metabolites, and glutarylglycine decreased. In the retina, tetranor-PGAM, pantothenic derivatives, 2-ethylacylcarinitine, and kynuramine levels decreased but anti-oxidative seleno-peptide level increased. Only phospholipids, fatty acids, and arachidonic acid metabolites in plasma and in the retina had significant correlation (p < 0.05, r > 0.4 or r < -0.4).ConclusionsThe results showed GTE indirectly induced systemic phosphorylcholine lipids to suppress inflammatory responses, hepatic damage, and respiratory mitochondrial stress in EIU rats induced by LPS. Phospholipids may be a therapeutic target of GTE for anterior chamber inflammation

Purpose: This study aims to compare dry eye parameters before and after COVID-19 infection in dry eye patients. Methods: We included 44 dry eye patients (88 eyes) from our existing dry eye cohort, with 22 belonging to the post-COVID-19 group due to a prior COVID-19 infection and the other 22 forming the non-COVID-19 group as they had no history of COVID-19. We examined and compared the dry eye parameters of the post-COVID-19 group, including the ocular surface disease index (OSDI), Schirmer’s test results (ST), non-invasive Keratography tear break-up time (NIKBUT), lipid layer thickness (LLT), Meibomian gland dysfunction (MGD), and the grading of papillae and follicles, both before and after the COVID-19 infection. We also compared the dry eye parameters difference of the post-COVID-19 group with the non-COVID-19 group. Results: The post-COVID-19 group was comprised of individuals with an average age of 38.36 ± 14.99 years, of which 82% were female. The time interval between the two tests was 16.92 ± 5.40 months, which did not differ significantly from the non-COVID-19 group. Compared to the pre-COVID-19 eyes, the post-COVID-19 eyes showed a significant decrease in the average LLT (52.86 ± 18.00 nm vs. 63.00 ± 22.40 nm, p < 0.001), as well as the maximum LLT (67.89 ± 20.81 nm vs. 78.48 ± 20.55 nm, p < 0.001). The MGD in both the upper (1.75 ± 0.84) and lower eyelids (1.43 ± 0.73) worsened after a COVID-19 infection. Additionally, the grading of papillae was worse following a COVID-19 infection (0.61 ± 0.69 vs. 0.16 ± 0.37, p < 0.001). The multivariate linear regression model revealed a negative association between COVID-19 infection and NIKBUT-average (β = −2.98, 95%CI: (−5.82, −0.15), p = 0.039), LLT-average (β = −14.12, 95%CI: (−22.66, −5.59), p = 0.001), and LLT max (β = −15.65, 95%CI: (−23.09, −8.20), p < 0.001). Conclusion: From preliminary results, we concluded that dry eye patients who have been infected with COVID-19 appear to have a more severe dry eye condition, as evidenced by lower LLT, worse papillae and MGD, and shorter NIKBUT. It is important to raise awareness of this potential long-term symptom of COVID-19, especially among existing dry eye patients.

Primary open-angle glaucoma (POAG) is highly heritable, yet not well understood from a genetic perspective. Here, the authors perform a meta-analysis of genome-wide association studies in 34,179 POAG cases, identifying 44 previously unreported risk loci and mapping effects across multiple ethnicities.

Inflammation is a critical pathophysiological process that modulates neuronal survival in the central nervous system after disease or injury. However, the effects and mechanisms of macrophage activation on neuronal survival remain unclear. In the present study, we co-cultured adult Fischer rat retinas with primary peritoneal macrophages or zymosan-treated peritoneal macrophages for 7 days. Immunofluorescence analysis revealed that peritoneal macrophages reduced retinal ganglion cell survival and neurite outgrowth in the retinal explant compared with the control group. The addition of zymosan to peritoneal macrophages attenuated the survival and neurite outgrowth of retinal ganglion cells. Conditioned media from peritoneal macrophages also reduced retinal ganglion cell survival and neurite outgrowth. This result suggests that secretions from peritoneal macrophages mediate the inhibitory effects of these macrophages. In addition, increased inflammation- and oxidation-related gene expression may be related to the enhanced retinal ganglion cell degeneration caused by zymosan activation. In summary, this study revealed that primary rat peritoneal macrophages attenuated retinal ganglion cell survival and neurite outgrowth, and that macrophage activation further aggravated retinal ganglion cell degeneration. This study was approved by the Animal Ethics Committee of the Joint Shantou International Eye Center of Shantou University and the Chinese University of Hong Kong, Shantou, Guangdong Province, China, on March 11, 2014 (approval no. EC20140311(2)-P01).

Ocular inflammation is a common complication of various eye diseases with wide consequences from irritations to potentially sight-threatening complications. Green tea is a popular beverage throughout the world. One of the proven health benefits of consuming green tea extract (GTE) is anti-inflammation. Catechins are the biologically active constituents of GTE. In in vitro and in vivo studies, GTE and catechins present inhibition of inflammatory responses in the development of ocular inflammation including infectious, non-infectious or autoimmune, and oxidative-induced complications. Research on the ocular inflammation in animal models has made significant progress in the past decades and several key disease mechanisms have been identified. Here we review the experimental investigations on the effects of GTE and catechins on various ocular inflammation related diseases including glaucoma, age-related macular degeneration, uveitis and ocular surface inflammation. We also review the pharmacokinetics of GTE constituents and safety of green tea consumption. We discuss the insights and perspectives of these experimental results, which would be useful for future development of novel therapeutics in human.

BackgroundAdvanced prediction of the daily incidence of COVID-19 can aid policy making on the prevention of disease spread, which can profoundly affect people's livelihood. In previous studies, predictions were investigated for single or several countries and territories. ObjectiveWe aimed to develop models that can be applied for real-time prediction of COVID-19 activity in all individual countries and territories worldwide. MethodsData of the previous daily incidence and infoveillance data (search volume data via Google Trends) from 215 individual countries and territories were collected. A random forest regression algorithm was used to train models to predict the daily new confirmed cases 7 days ahead. Several methods were used to optimize the models, including clustering the countries and territories, selecting features according to the importance scores, performing multiple-step forecasting, and upgrading the models at regular intervals. The performance of the models was assessed using the mean absolute error (MAE), root mean square error (RMSE), Pearson correlation coefficient, and Spearman correlation coefficient. ResultsOur models can accurately predict the daily new confirmed cases of COVID-19 in most countries and territories. Of the 215 countries and territories under study, 198 (92.1%) had MAEs 0.8. For the 215 countries and territories, the mean MAE was 5.42 (range 0.26-15.32), the mean RMSE was 9.27 (range 1.81-24.40), the mean Pearson correlation coefficient was 0.89 (range 0.08-0.99), and the mean Spearman correlation coefficient was 0.84 (range 0.2-1.00). ConclusionsBy integrating previous incidence and Google Trends data, our machine learning algorithm was able to predict the incidence of COVID-19 in most individual countries and territories accurately 7 days ahead.

Abstract Background Neovascular age-related macular degeneration (AMD) and polypoidal choroidal vasculopathy (PCV) are sight-threatening maculopathies with both environmental and genetic risk factors. We have previously shown relative risks posed by genes of the complement pathways to neovascular AMD and PCV. Methods In this study, we investigated the haplotype-tagging single nucleotide polymorphisms (SNPs) in the complement component 5 (C5) gene in 708 unrelated Chinese individuals: 200 neovascular AMD patients, 233 PCV patients and 275 controls. Six tagging SNPs in C5 were genotyped. Univariate single SNP association analysis, haplotype-based association analysis and gene-gene interaction analysis between C5 and other AMD-associated genes were performed. Results The results revealed none of the six tagging SNPs of the C5 gene had a significant association with neovascular AMD or PCV (P > 0.05). We also found insignificant haplotype-based association, and no significant SNP-SNP interaction between C5 and other genes (including C2-CFB-RDBP-SKIV2L, SERPING1, CETP, ABCG1, PGF, ANGPT2, CFH and HTRA1) for neovascular AMD and PCV. Conclusions This study showed no statistical significance in the genetic association of C5 with neovascular AMD or PCV in a Hong Kong Chinese population. Further studies in large samples from different populations are warranted to elucidate the role of C5 in the genetic susceptibility of AMD and PCV.

Background High blood pressure (BP) and obesity are becoming increasingly prevalent among children globally. Although prior studies have shown their adverse impacts on macrovascular health, less is known about their effects on microvascular heath. This study aims to evaluate the independent and synergistic effects of hypertensive BP and obesity on retinal vasculature in young children. Method and Results 1006 children aged 6 to 8 years were recruited from the Hong Kong Children Eye Study. Quantitative retinal vascular parameters, including central retinal arteriolar and venular equivalents and retinal arteriolar and venular fractal dimensions, were measured from retinal photographs following a standardized protocol. BP and body mass index were categorized according to reference values from American Academy of Pediatrics and International Obesity Task Force guidelines respectively. Children with hypertensive systolic BP had the narrowest central retinal arteriolar equivalents compared with children with either elevated or normotensive systolic BP (162.4, 164.6, and 167.1 µm; P‐trend

This review provides insights into the mechanism underlying the pathogenesis of myopia and potential targets for clinical intervention. Although the etiology of myopia involves both environmental and genetic factors, recent evidence has suggested that the prevalence and severity of myopia appears to be affected more by environmental factors. Current pharmacotherapeutics are aimed at inhibiting environmentally induced changes in visual input and subsequent changes in signaling pathways during myopia pathogenesis and progression. Recent studies on animal models of myopia have revealed specific molecules potentially involved in the regulation of eye development. Among them, the dopamine receptor plays a critical role in controlling myopia. Subsequent studies have reported pharmacotherapeutic treatments to control myopia progression. In particular, atropine treatment yielded favorable outcomes and has been extensively used; however, current studies are aimed at optimizing its efficacy and confirming its safety. Furthermore, future studies are required to assess the efficacy of combinatorial use of low-dose atropine and contact lenses or orthokeratology.

Uveitis is characterized by inflammatory lesions of intraocular structures. It is one of the important manifestations in patients with Reiter’s syndrome, an inflammatory arthritis, which is caused by enteric infection with bacteria, including Salmonella typhimurium. Corticosteroids remain the most frequently used therapies against uveitis associating with inflammatory arthritis. However, the long-term administration of steroids results in many side effects, and some uveitis patients do not respond to steroid treatment. Non-steroidal treatments are needed for uveitis patients. Our previous study found that Janus kinase (JAK) 1/2 inhibitor, ruxolitinib could suppress the expression of proinflammatory mediators in the ciliary body and iris. However, the impacts of ruxolitinib on ophthalmic features in uveitic eyes are still unknown. In this study, Salmonella typhimurium endotoxin-induced uveitis (EIU) was induced in Sprague Dawley rats by the injection of lipopolysaccharide (LPS). Compared with LPS-induced rats treated with water, ruxolitinib significantly attenuated the clinical manifestations, infiltrating cells and protein exudation in the aqueous humor, and retina–choroid thickening. Amplitudes of b-wave in both scotopic and photopic electroretinogram (ERG), and the amplitude of a-wave in scotopic ERG in EIU animals were alleviated by ruxolitinib. Collectively, we propose ruxolitinib could attenuate endotoxin-induced uveitis and rescue visual functions in rats by inhibiting the JAK2-STAT3 pathway.

Abstract Genetic associations for keratoconus could be useful for understanding disease pathogenesis and discovering biomarkers for early detection of the disease. We conducted a systematic review and meta-analysis to summarize all reported genetic associations for the disease. We searched in the MEDLINE, Embase, Web of Science, and HuGENET databases for genetic studies of keratoconus published from 1950 to June 2016. The summary odds ratio and 95% confidence intervals of all polymorphisms were estimated using the random-effect model. Among 639 reports that were retrieved, 24 fulfilled required criteria as eligible studies for meta-analysis, involving a total of 53 polymorphisms in 28 genes/loci. Results of our meta-analysis lead to the prioritization of 8 single-nucleotide polymorphisms (SNPs) in 6 genes/loci for keratoconus in Whites. Of them 5 genes/loci were originally detected in genome-wide association studies, including FOXO1 (rs2721051, P = 5.6 × 10−11), RXRA-COL5A1 (rs1536482, P = 2.5 × 10−9), FNDC3B (rs4894535, P = 1.4 × 10−8), IMMP2L (rs757219, P = 6.1 × 10−7; rs214884, P = 2.3 × 10−5), and BANP-ZNF469 (rs9938149, P = 1.3 × 10−5). The gene COL4A4 (rs2229813, P = 1.3 × 10−12; rs2228557, P = 4.5 × 10−7) was identified in previous candidate gene studies. We also found SNPs in 10 genes/loci that had a summary P value

Abstract Cigarette smoking is a major risk factor for age-related macular degeneration (AMD), in which progressive retinal pigment epithelial (RPE) cell degeneration is a major pathological change. Nicotine is a major biologically active component in cigarette smoke. It is continuously catabolized into cotinine, which has longer half-life and higher concentration in tissue cells and fluids. Here we hypothesized that continuous exposure of cotinine has more potent effects on human RPE cell properties than nicotine. Human RPE cell line (ARPE-19) was treated continuously with 1–2 µM of nicotine and/or cotinine for 7 days. RPE cells treated with 2 μM cotinine and nicotine-cotinine mixture has lower MTT signals without significant changes in cell apoptosis or integrity. Moreover, RPE cell migration was retarded under cotinine treatments, but not nicotine. Both nicotine and cotinine treatments attenuated the phagocytotic activity of RPE cells. In addition, cotinine and nicotine-cotinine mixture suppressed VEGF and IL-8 expression and upregulated TIMP-2 expression. Expressions of autophagy genes were upregulated by the cotinine treatment, whereas expressions of epithelial-to-mesenchymal transition markers were downregulated. In conclusion, our study, for the first time, demonstrated that cotinine, rather than nicotine, affects the properties of RPE cells in vitro, which could explain the smoking-induced RPE pathology.

Abstract Upon a drastic change in environmental illumination, zebrafish larvae display a rapid locomotor response. This response can be simultaneously tracked from larvae arranged in multi-well plates. The resulting data have provided new insights into neuro-behaviour. The features of these data, however, present a challenge to traditional statistical tests. For example, many larvae display little or no movement. Thus, the larval responses have many zero values and are imbalanced. These responses are also measured repeatedly from the same well, which results in correlated observations. These analytical issues were addressed in this study by the generalized linear mixed model (GLMM). This approach deals with binary responses and characterizes the correlation of observations in the same group. It was used to analyze a previously reported dataset. Before applying the GLMM, the activity values were transformed to binary responses (movement vs. no movement) to reduce data imbalance. Moreover, the GLMM estimated the variations among the effects of different well locations, which would eliminate the location effects when two biological groups or conditions were compared. By addressing the data-imbalance and location-correlation issues, the GLMM effectively quantified true biological effects on zebrafish locomotor response.

Summary: Background: Spectral-domain optical coherence tomography (SDOCT) can be used to detect glaucomatous optic neuropathy, but human expertise in interpretation of SDOCT is limited. We aimed to develop and validate a three-dimensional (3D) deep-learning system using SDOCT volumes to detect glaucomatous optic neuropathy. Methods: We retrospectively collected a dataset including 4877 SDOCT volumes of optic disc cube for training (60%), testing (20%), and primary validation (20%) from electronic medical and research records at the Chinese University of Hong Kong Eye Centre (Hong Kong, China) and the Hong Kong Eye Hospital (Hong Kong, China). Residual network was used to build the 3D deep-learning system. Three independent datasets (two from Hong Kong and one from Stanford, CA, USA), including 546, 267, and 1231 SDOCT volumes, respectively, were used for external validation of the deep-learning system. Volumes were labelled as having or not having glaucomatous optic neuropathy according to the criteria of retinal nerve fibre layer thinning on reliable SDOCT images with position-correlated visual field defect. Heatmaps were generated for qualitative assessments. Findings: 6921 SDOCT volumes from 1 384 200 two-dimensional cross-sectional scans were studied. The 3D deep-learning system had an area under the receiver operation characteristics curve (AUROC) of 0·969 (95% CI 0·960–0·976), sensitivity of 89% (95% CI 83–93), specificity of 96% (92–99), and accuracy of 91% (89–93) in the primary validation, outperforming a two-dimensional deep-learning system that was trained on en face fundus images (AUROC 0·921 [0·905–0·937]; p

Many contemporary neuroscience experiments utilize high-throughput approaches to simultaneously collect behavioural data from many animals. The resulting data are often complex in structure and are subjected to systematic biases, which require new approaches for analysis and normalization. This study addressed the normalization need by establishing an approach based on linear-regression modeling. The model was established using a dataset of visual motor response (VMR) obtained from several strains of wild-type (WT) zebrafish collected at multiple stages of development. The VMR is a locomotor response triggered by drastic light change, and is commonly measured repeatedly from multiple larvae arrayed in 96-well plates. This assay is subjected to several systematic variations. For example, the light emitted by the machine varies slightly from well to well. In addition to the light-intensity variation, biological replication also created batch-batch variation. These systematic variations may result in differences in the VMR and must be normalized. Our normalization approach explicitly modeled the effect of these systematic variations on VMR. It also normalized the activity profiles of different conditions to a common baseline. Our approach is versatile, as it can incorporate different normalization needs as separate factors. The versatility was demonstrated by an integrated normalization of three factors: light-intensity variation, batch-batch variation and baseline. After normalization, new biological insights were revealed from the data. For example, we found larvae of TL strain at 6 days post-fertilization (dpf) responded to light onset much stronger than the 9-dpf larvae, whereas previous analysis without normalization shows that their responses were relatively comparable. By removing systematic variations, our model-based normalization can facilitate downstream statistical comparisons and aid detecting true biological differences in high-throughput studies of neurobehaviour.

Inactivation of the retinoblastoma tumor suppressor gene (RB1) leads to genome instability, and can be detected in retinoblastoma and other cancers. One damaging effect is causing DNA double strand breaks (DSB), which, however, can be repaired by homologous recombination (HR), classical non-homologous end joining (C-NHEJ), and micro-homology mediated end joining (MMEJ). We aimed to study the mechanistic roles of RB in regulating multiple DSB repair pathways. Here we show that HR and C-NHEJ are decreased, but MMEJ is elevated in RB-depleted cells. After inducing DSB by camptothecin, RB co-localizes with CtIP, which regulates DSB end resection. RB depletion leads to less RPA and native BrdU foci, which implies less end resection. In RB-depleted cells, less CtIP foci, and a lack of phosphorylation on CtIP Thr847, are observed. According to the synthetic lethality principle, based on the altered DSB repair pathway choice, after inducing DSBs by camptothecin, RB depleted cells are more sensitive to co-treatment with camptothecin and MMEJ blocker poly-ADP ribose polymerase 1 (PARP1) inhibitor. We propose a model whereby RB can regulate DSB repair pathway choice by mediating the CtIP dependent DNA end resection. The use of PARP1 inhibitor could potentially improve treatment outcomes for RB-deficient cancers.

Previously, we identified RAD21R450C from a peripheral sclerocornea pedigree. Injection of this rad21 variant mRNA into Xenopus laevis embryos disrupted the organization of corneal stroma fibrils. To understand the mechanisms of RAD21-mediated corneal stroma defects, gene expression and chromosome conformation analysis were performed using cells from family members affected by peripheral sclerocornea. Both gene expression and chromosome conformation of cell adhesion genes were affected in cells carrying the heterozygous rad21 variant. Since cell migration is essential in early embryonic development and sclerocornea is a congenital disease, we studied neural crest migration during cornea development in X. laevis embryos. In X. laevis embryos injected with rad21 mutant mRNA, neural crest migration was disrupted, and the number of neural crest-derived periocular mesenchymes decreased significantly in the corneal stroma region. Our data indicate that the RAD21R450C variant contributes to peripheral sclerocornea by modifying chromosome conformation and gene expression, therefore disturbing neural crest cell migration, which suggests RAD21 plays a key role in corneal stroma development.

Retinal disease treatment by stem cell-based replacement relies on stem cell differentiation into retinal cells. We previously demonstrated that human periodontal ligament-derived stem cells can be directed into retinal lineage upon induction. Here, we report the transdifferentiation potential of human adipose-derived stem cells (ASCs) into retinal lineage and its enhancement by Notch signaling modulation. Human ASCs, isolated from abdominal fat, expressed mesenchymal but not hematopoietic stem cell markers, and they can differentiate into adipocytes, chondrocytes, and osteoblasts in vitro. Upon noggin/Dkk-1/IGF-1 induction, the treated ASCs showed elevated expression of retinal progenitor, retinal ganglion, and photoreceptor cell markers as well as the glutamate-evoked calcium response, which was not observed in the noninduced cells. Compared to the regular induction treatment, Notch signaling activation by JAG1 enhanced the expression of retinal progenitor and precursor markers without affecting the glutamate-evoked calcium response. In contrast, Notch signaling inhibition by DAPT showed more retinal ganglion cells, but delayed the response to glutamate stimulation. In summary, our results revealed that human ASCs possess a retinal transdifferentiation potential upon noggin/Dkk-1/IGF-1 induction, which can further be enhanced by Notch signaling activation.

[This corrects the article DOI: 10.1371/journal.pone.0149663.].

Cataract is the most common cause of visual loss in humans. A spontaneously occurred, autosomal dominant mouse mutant Secc, which displayed combined features of small eye, cataract and closed eyelid was discovered in our laboratory. In this study, we identified the mutation and characterized the cataract phenotype of this novel Secc mutant. The Secc mutant mice have eyelids that remain half-closed throughout their life. The mutant lens has a significant reduction in size and with opaque spots clustered in the centre. Histological analysis showed that in the core region of the mutant lens, the fiber cells were disorganized and clefts and vacuoles were observed. The cataract phenotype was evident from new born stage. We identified the Secc mutation by linkage analysis using whole genome microsatellite markers and SNP markers. The Secc locus was mapped at chromosome 1 flanked by SNPs rs3158129 and rs13475900. Based on the chromosomal position, the candidate cataract locus γ-crystallin gene cluster (Cryg) was investigated by sequencing. A single base deletion (299delG) in exon 3 of Cryga which led to a frame-shift of amino acid sequence from position 91 was identified. As a result of this mutation, the sequences of the 3rd and 4th Greek-key motifs of the γA-crystallin are replaced with an unrelated C-terminal peptide of 75 residues long. Coincidentally, the point mutation generated a HindIII restriction site, allowing the identification of the CrygaSecc mutant allele by RFLP. Western blot analysis of 3-week old lenses showed that the expression of γ-crystallins was reduced in the CrygaSecc mutant. Furthermore, in cell transfection assays using CrygaSecc mutant cDNA expression constructs in 293T, COS-7 and human lens epithelial B3 cell lines, the mutant γA-crystallins were enriched in the insoluble fractions and appeared as insoluble aggregates in the transfected cells. In conclusion, we have demonstrated that the Secc mutation leads to the generation of CrygaSecc proteins with reduced solubility and prone to form aggregates within lens cells. Accumulation of mutant proteins in the lens fibers would lead to cataract formation in the Secc mutant.

Retinal degeneration is often progressive. This feature has provided a therapeutic window for intervention that may extend functional vision in patients. Even though this approach is feasible, few promising drug candidates are available. The scarcity of new drugs has motivated research to discover novel compounds through different sources. One such example is Schisandrin B (SchB), an active component isolated from the five-flavor fruit (Fructus Schisandrae) that is postulated in traditional Chinese medicines to exert prophylactic visual benefit. This SchB benefit was investigated in this study in pde6cw59, a zebrafish retinal-degeneration model. In this model, the pde6c gene (phosphodiesterase 6C, cGMP-specific, cone, alpha prime) carried a mutation which caused cone degeneration. This altered the local environment and caused the bystander rods to degenerate too. To test SchB on the pde6cw59 mutants, a treatment concentration was first determined that would not cause morphological defects, and would initiate known physiological response. Then, the mutants were treated with the optimized SchB concentration before the appearance of retinal degeneration at 3 days postfertilization (dpf). The light sensation of animals was evaluated at 6 dpf by the visual motor response (VMR), a visual startle that could be initiated by drastic light onset and offset. The results show that the VMR of pde6cw59 mutants towards light onset was enhanced by the SchB treatment, and that the initial phase of the enhancement was primarily mediated through the mutants' eyes. Further immunostaining analysis indicates that the treatment specifically reduced the size of the abnormally large rods. These observations implicate an interesting hypothesis: that the morphologically-improved rods drive the observed VMR enhancement. Together, these investigations have identified a possible visual benefit of SchB on retinal degeneration, a benefit that can potentially be further developed to extend functional vision in patients.