Mono- and biallelic variant effects on disease at biobank scale.

Identifying causal factors for Mendelian and common diseases is an ongoing challenge in medical genetics1. Population bottleneck events, such as those that occurred in the history of the Finnish population, enrich some homozygous variants to higher frequencies, which facilitates the identification of variants that cause diseases with recessive inheritance2,3. Here we examine the homozygous and heterozygous effects of 44,370 coding variants on 2,444 disease phenotypes using data from the nationwide electronic health records of 176,899 Finnish individuals. We find associations for homozygous genotypes across a broad spectrum of phenotypes, including known associations with retinal dystrophy and novel associations with adult-onset cataract and female infertility. Of the recessive disease associations that we identify, 13 out of 20 would have been missed by the additive model that is typically used in genome-wide association studies. We use these results to find many known Mendelian variants whose inheritance cannot be adequately described by a conventional definition of dominant or recessive. In particular, we find variants that are known to cause diseases with recessive inheritance with significant heterozygous phenotypic effects. Similarly, we find presumed benign variants with disease effects. Our results show how biobanks, particularly in founder populations, can broaden our understanding of complex dosage effects of Mendelian variants on disease.

Integrated single-cell multiomics uncovers foundational regulatory mechanisms of lens development and pathology.

Ocular lens development entails epithelial to fiber cell differentiation, defects in which cause congenital cataracts. We report the first single-cell multiomic atlas of lens development, leveraging snRNA-seq, snATAC-seq and CUT&RUN-seq to discover previously unreported mechanisms of cell fate determination and cataract-linked regulatory networks. A comprehensive profile of cis- and trans-regulatory interactions, including for the cataract-linked transcription factor MAF, is established across a temporal trajectory of fiber cell differentiation. Furthermore, we identify an epigenetic paradigm of cellular differentiation, defined by progressive loss of the H3K27 methylation writer Polycomb repressive complex 2 (PRC2). PRC2 localizes to heterochromatin domains across master-regulator transcription factor gene bodies, suggesting it safeguards epithelial cell fate. Moreover, we demonstrate that FGF hyper-stimulation in vivo leads to MAF network activation and the emergence of novel lens cell states. Collectively, these data depict a comprehensive portrait of lens fiber cell differentiation, while defining regulatory effectors of cell identity and cataract formation.

Organelle degradation in the lens by PLAAT phospholipases.

The eye lens of vertebrates is composed of fibre cells in which all membrane-bound organelles undergo degradation during terminal differentiation to form an organelle-free zone1. The mechanism that underlies this large-scale organelle degradation remains largely unknown, although it has previously been shown to be independent of macroautophagy2,3. Here we report that phospholipases in the PLAAT (phospholipase A/acyltransferase, also known as HRASLS) family-Plaat1 (also known as Hrasls) in zebrafish and PLAAT3 (also known as HRASLS3, PLA2G16, H-rev107 or AdPLA) in mice4-6-are essential for the degradation of lens organelles such as mitochondria, the endoplasmic reticulum and lysosomes. Plaat1 and PLAAT3 translocate from the cytosol to various organelles immediately before organelle degradation, in a process that requires their C-terminal transmembrane domain. The translocation of Plaat1 to organelles depends on the differentiation of fibre cells and damage to organelle membranes, both of which are mediated by Hsf4. After the translocation of Plaat1 or PLAAT3 to membranes, the phospholipase induces extensive organelle rupture that is followed by complete degradation. Organelle degradation by PLAAT-family phospholipases is essential for achieving an optimal transparency and refractive function of the lens. These findings expand our understanding of intracellular organelle degradation and provide insights into the mechanism by which vertebrates acquired transparent lenses.

Genetic correlation between circulating cytokines and risk of three ophthalmic diseases: a bidirectional two-sample Mendelian randomization study.

PURPOSE: Pathogenesis and the associated risk factors of cataracts, glaucoma, and age-related macular degeneration (AMD) remain unclear. We aimed to investigate causal relationships between circulating cytokine levels and the development of these diseases. PATIENTS AND METHODS: Genetic instrumental variables for circulating cytokines were derived from a genome-wide association study of 8293 European participants. Summary-level data for AMD, glaucoma, and senile cataract were obtained from the FinnGen database. The inverse variance weighted (IVW) was the main Mendelian randomization (MR) analysis method. The Cochran's Q, MR-Egger regression, and MR pleiotropy residual sum and outlier test were used for sensitivity analysis. RESULTS: Based on the IVW method, MR analysis demonstrated five circulating cytokines suggestively associated with AMD (SCGF-ß, 1.099 [95%CI, 1.037-1.166], P = 0.002; SCF, 1.155 [95%CI, 1.015-1.315], P = 0.029; MCP-1, 1.103 [95%CI, 1.012-1.202], P = 0.026; IL-10, 1.102 [95%CI, 1.012-1.200], P = 0.025; eotaxin, 1.086 [95%CI, 1.002-1.176], P = 0.044), five suggestively linked with glaucoma (MCP-1, 0.945 [95%CI, 0.894-0.999], P = 0.047; IL1ra, 0.886 [95%CI, 0.809-0.969], P = 0.008; IL-1ß, 0.866 [95%CI, 0.762-0.983], P = 0.027; IL-9, 0.908 [95%CI, 0.841-0.980], P = 0.014; IL2ra, 1.065 [95%CI, 1.004-1.130], P = 0.035), and four suggestively associated with senile cataract (TRAIL, 1.043 [95%CI, 1.009-1.077], P = 0.011; IL-16, 1.032 [95%CI, 1.001-1.064], P = 0.046; IL1ra, 0.942 [95%CI, 0.887-0.999], P = 0.047; FGF-basic, 1.144 [95%CI, 1.052-1.244], P = 0.002). Furthermore, sensitivity analysis results supported the above associations. CONCLUSION: This study highlights the involvement of several circulating cytokines in the development ophthalmic diseases and holds potential as viable pharmacological targets for these diseases.

White matter plasticity following cataract surgery in congenitally blind patients.

The visual system develops abnormally when visual input is absent or degraded during a critical period early in life. Restoration of the visual input later in life is generally thought to have limited benefit because the visual system will lack sufficient plasticity to adapt to and utilize the information from the eyes. Recent evidence, however, shows that congenitally blind adolescents can recover both low-level and higher-level visual function following surgery. In this study, we assessed behavioral performance in both a visual acuity and a face perception task alongside longitudinal structural white matter changes in terms of fractional anisotropy (FA) and mean diffusivity (MD). We studied congenitally blind patients with dense bilateral cataracts, who received cataract surgery at different stages of adolescence. Our goal was to differentiate between age- and surgery-related changes in both behavioral performance and structural measures to identify neural correlates which might contribute to recovery of visual function. We observed surgery-related long-term increases of structural integrity of late-visual pathways connecting the occipital regions with ipsilateral fronto-parieto-temporal regions or homotopic contralateral areas. Comparison to a group of age-matched healthy participants indicated that these improvements went beyond the expected changes in FA and MD based on maturation alone. Finally, we found that the extent of behavioral improvement in face perception was mediated by changes in structural integrity in late visual pathways. Our results suggest that sufficient plasticity remains in adolescence to partially overcome abnormal visual development and help localize the sites of neural change underlying sight recovery.

HSP90ß prevents aging-related cataract formation through regulation of the charged multivesicular body protein (CHMP4B) and p53.

Cataract is a leading ocular disease causing global blindness. The mechanism of cataractogenesis has not been well defined. Here, we demonstrate that the heat shock protein 90ß (HSP90ß) plays a fundamental role in suppressing cataractogenesis. HSP90ß is the most dominant HSP in normal lens, and its constitutive high level of expression is largely derived from regulation by Sp1 family transcription factors. More importantly, HSP90ß is significantly down-regulated in human cataract patients and in aging mouse lenses, whereas HSP90ß silencing in zebrafish causes cataractogenesis, which can only be rescued by itself but not other HSP90 genes. Mechanistically, HSP90ß can directly interact with CHMP4B, a newly-found client protein involved in control of cytokinesis. HSP90ß silencing causes upregulation of CHMP4B and another client protein, the tumor suppressor p53. CHMP4B upregulation or overexpression induces excessive division of lens epithelial cells without proper differentiation. As a result, these cells were triggered to undergo apoptosis due to activation of the p53/Bak-Bim pathway, leading to cataractogenesis and microphthalmia. Silence of both HSP90ß and CHMP4B restored normal phenotype of zebrafish eye. Together, our results reveal that HSP90ß is a critical inhibitor of cataractogenesis through negative regulation of CHMP4B and the p53-Bak/Bim pathway.

Rat Model of Type 2 Diabetes Mellitus Recapitulates Human Disease in the Anterior Segment of the Eye.

Changes in the anterior segment of the eye due to type 2 diabetes mellitus (T2DM) are not well-characterized, in part due to the lack of a reliable animal model. This study evaluated changes in the anterior segment, including crystalline lens health, corneal endothelial cell density, aqueous humor metabolites, and ciliary body vasculature, in a rat model of T2DM compared with human eyes. Male Sprague-Dawley rats were fed a high-fat diet (45% fat) or normal diet, and rats fed the high-fat diet were injected with streptozotocin intraperitoneally to generate a model of T2DM. Cataract formation and corneal endothelial cell density were assessed using microscopic analysis. Diabetes-related rat aqueous humor alterations were assessed using metabolomics screening. Transmission electron microscopy was used to assess qualitative ultrastructural changes ciliary process microvessels at the site of aqueous formation in the eyes of diabetic rats and humans. Eyes from the diabetic rats demonstrated cataracts, lower corneal endothelial cell densities, altered aqueous metabolites, and ciliary body ultrastructural changes, including vascular endothelial cell activation, pericyte degeneration, perivascular edema, and basement membrane reduplication. These findings recapitulated diabetic changes in human eyes. These results support the use of this model for studying ocular manifestations of T2DM and support a hypothesis postulating blood-aqueous barrier breakdown and vascular leakage at the ciliary body as a mechanism for diabetic anterior segment pathology.

Epidemiologic association and shared genetic architecture between cataract and hearing difficulties among middle-aged and older adults.

Age-related cataract and hearing difficulties are major sensory disorders that often co-exist in the global-wide elderly and have a tangible influence on the quality of life. However, the epidemiologic association between cataract and hearing difficulties remains unexplored, while little is known about whether the two share their genetic etiology. We first investigated the clinical association between cataract and hearing difficulties using the UK Biobank covering 502,543 individuals. Both unmatched analysis (adjusted for confounders) and a matched analysis (one control matched for each patient with cataract according to confounding factors) were undertaken and confirmed that cataract was associated with hearing difficulties (OR, 2.12; 95% CI, 1.98-2.27; OR, 2.03; 95% CI, 1.86-2.23, respectively). Furthermore, we explored and quantified the shared genetic architecture of these two complex sensory disorders at the common variant level using the bivariate causal mixture model (MiXeR) and conditional/conjunctional false discovery rate method based on the largest available genome-wide association studies of cataract (N = 585,243) and hearing difficulties (N = 323,978). Despite detecting only a negligible genetic correlation, we observe polygenic overlap between cataract and hearing difficulties and identify 6 shared loci with mixed directions of effects. Follow-up analysis of the shared loci implicates candidate genes QKI, STK17A, TYR, NSF, and TCF4 likely contribute to the pathophysiology of cataracts and hearing difficulties. In conclusion, this study demonstrates the presence of epidemiologic association between cataract and hearing difficulties and provides new insights into the shared genetic architecture of these two disorders at the common variant level.

A role for YAP/FOXM1/Nrf2 axis in oxidative stress and apoptosis of cataract induced by UVB irradiation.

This study aims to investigate the hypothesis that Yes-associated protein (YAP) significantly regulates antioxidant potential and anti-apoptosis in UVB-induced cataract by exploring the underlying molecular mechanisms. To investigate the association between YAP and cataract, various experimental techniques were employed, including cell viability assessment, Annexin V FITC/PI assay, measurement of ROS production, RT-PCR, Western blot assay, and Immunoprecipitation. UVB exposure on human lens epithelium cells (HLECs) reduced total and nuclear YAP protein expression, increased cleaved/pro-caspase 3 ratios, decreased cell viability, and elevated ROS levels compared to controls. Similar Western blot results were observed in in vivo experiments involving UVB-treated mice. YAP knockdown in vitro demonstrated a decrease in the protein expression of FOXM1, Nrf2, and HO-1, which correlated with the mRNA expression, accompanied by an increase in cell apoptosis, caspase 3 activation, and the release of ROS. Conversely, YAP overexpression mitigated these effects induced by UVB irradiation. Immunoprecipitation revealed a FOXM1-YAP interaction. Notably, inhibiting FOXM1 decreased Nrf2 and HO-1, activating caspase 3. Additionally, administering the ROS inhibitor N-acetyl-L-cysteine (NAC) effectively mitigated the apoptotic effects induced by oxidative stress from UVB irradiation, rescuing the protein expression levels of YAP, FOXM1, Nrf2, and HO-1. The initial findings of our study demonstrate the existence of a feedback loop involving YAP, FOXM1, Nrf2, and ROS that significantly influences the cell apoptosis in HLECs under UVB-induced oxidative stress.

Sengers Syndrome-Associated Mitochondrial Acylglycerol Kinase Is a Subunit of the Human TIM22 Protein Import Complex.

Acylglycerol kinase (AGK) is a mitochondrial lipid kinase that catalyzes the phosphorylation of monoacylglycerol and diacylglycerol to lysophosphatidic acid and phosphatidic acid, respectively. Mutations in AGK cause Sengers syndrome, which is characterized by congenital cataracts, hypertrophic cardiomyopathy, skeletal myopathy, exercise intolerance, and lactic acidosis. Here we identified AGK as a subunit of the mitochondrial TIM22 protein import complex. We show that AGK functions in a kinase-independent manner to maintain the integrity of the TIM22 complex, where it facilitates the import and assembly of mitochondrial carrier proteins. Mitochondria isolated from Sengers syndrome patient cells and tissues show a destabilized TIM22 complex and defects in the biogenesis of carrier substrates. Consistent with this phenotype, we observe perturbations in the tricarboxylic acid (TCA) cycle in cells lacking AGK. Our identification of AGK as a bona fide subunit of TIM22 provides an exciting and unexpected link between mitochondrial protein import and Sengers syndrome.

Acylglycerol Kinase Mutated in Sengers Syndrome Is a Subunit of the TIM22 Protein Translocase in Mitochondria.

Mutations in mitochondrial acylglycerol kinase (AGK) cause Sengers syndrome, which is characterized by cataracts, hypertrophic cardiomyopathy, and skeletal myopathy. AGK generates phosphatidic acid and lysophosphatidic acid, bioactive phospholipids involved in lipid signaling and the regulation of tumor progression. However, the molecular mechanisms of the mitochondrial pathology remain enigmatic. Determining its mitochondrial interactome, we have identified AGK as a constituent of the TIM22 complex in the mitochondrial inner membrane. AGK assembles with TIMM22 and TIMM29 and supports the import of a subset of multi-spanning membrane proteins. The function of AGK as a subunit of the TIM22 complex does not depend on its kinase activity. However, enzymatically active AGK is required to maintain mitochondrial cristae morphogenesis and the apoptotic resistance of cells. The dual function of AGK as lipid kinase and constituent of the TIM22 complex reveals that disturbances in both phospholipid metabolism and mitochondrial protein biogenesis contribute to the pathogenesis of Sengers syndrome.

Proteome Remodeling of the Eye Lens at 50 Years Identified With Data-Independent Acquisition.

The eye lens is responsible for focusing and transmitting light to the retina. The lens does this in the absence of organelles, yet maintains transparency for at least 5 decades before onset of age-related nuclear cataract (ARNC). It is hypothesized that oxidative stress contributes significantly to ARNC formation. It is in addition hypothesized that transparency is maintained by a microcirculation system that delivers antioxidants to the lens nucleus and exports small molecule waste. Common data-dependent acquisition methods are hindered by dynamic range of lens protein expression and provide limited context to age-related changes in the lens. In this study, we utilized data-independent acquisition mass spectrometry to analyze the urea-insoluble membrane protein fractions of 16 human lenses subdivided into three spatially distinct lens regions to characterize age-related changes, particularly concerning the lens microcirculation system and oxidative stress response. In this pilot cohort, we measured 4788 distinct protein groups, 46,681 peptides, and 7592 deamidated sequences, more than in any previous human lens data-dependent acquisition approach. Principally, we demonstrate that a significant proteome remodeling event occurs at approximately 50 years of age, resulting in metabolic preference for anaerobic glycolysis established with organelle degradation, decreased abundance of protein networks involved in calcium-dependent cell-cell contacts while retaining networks related to oxidative stress response. Furthermore, we identified multiple antioxidant transporter proteins not previously detected in the human lens and describe their spatiotemporal and age-related abundance changes. Finally, we demonstrate that aquaporin-5, among other proteins, is modified with age by post-translational modifications including deamidation and truncation. We suggest that the continued accumulation of each of these age-related outcomes in proteome remodeling contribute to decreased fiber cell permeability and result in ARNC formation.

Comparative Proteomic Analysis of Aqueous Humor Reveals Biochemical Disparities in the Eyes of High Myopic Patients.

Myopia accounts for a significant proportion of visual lesions worldwide and has the potential to progress toward pathological myopia. This study aims to reveal the difference in protein content in aqueous humor between high myopic and nonhigh myopic patients, as well as better understand the dysregulation of proteins in myopic eyes. Aqueous humor was collected for liquid chromatograph mass spectrometer (LC/MS) analysis from 30 individual eyes that underwent phacoemulsification and intraocular lens (IOL) implantation. Results showed that a total of 190 differentially expressed proteins were identified, which revealed their involvement in cell metabolism, immune and inflammatory response, and system and anatomical structure. Further analysis focused on 15 intensively interacted hub proteins, encompassing functions related to complement cascades, lipoprotein metabolism, and fibrin biological function. Subsequent validations demonstrated elevated levels of APOE (apolipoprotein E), C3 (complement 3), and AHSG (α-2-HS-glycoprotein) in the high myopia group (31 eyes of cataracts and 45 eyes of high myopia with cataracts). AHSG had a significant positive correlation with axial length in high myopic patients, with good efficacy in distinguishing between myopic and nonmyopic groups. AHSG may be a potential indicator of the pathological severity and participator in the pathological progress of high myopia. This study depicted differential expression characteristics of aqueous humor in patients with high myopia and provided optional information for further experimental research on exploring the molecular mechanisms and potential therapeutic targets for high myopia. Data are available via ProteomeXchange with the identifier PXD047584.

Potential Immune-Inflammatory Proteome Biomarkers for Guiding the Treatment of Patients with Primary Acute Angle-Closure Glaucoma Caused by COVID-19.

Primary acute angle-closure glaucoma (PAACG) is a sight-threatening condition that can lead to blindness. With the increasing incidence of COVID-19, a multitude of people are experiencing acute vision loss and severe swelling of the eyes and head. These patients were then diagnosed with acute angle closure, with or without a history of PACG. However, the mechanism by which viral infection causes PACG has not been clarified. This is the first study to explore the specific inflammatory proteomic landscape in SARS-CoV-2-induced PAACG. The expression of 92 inflammation-related proteins in 19 aqueous humor samples from PAACGs or cataract patients was detected using the Olink Target 96 Inflammation Panel based on a highly sensitive and specific proximity extension assay technology. The results showed that 76 proteins were significantly more abundant in the PAACG group than in the cataract group. Notably, the top eight differentially expressed proteins were IL-8, MCP-1, TNFRSF9, DNER, CCL4, Flt3L, CXCL10, and CD40. Generally, immune markers are related to inflammation, macrophage activation, and viral infection, revealing the crucial role of macrophages in the occurrence of PAACGs caused by SARS-CoV-2.

Low expression of TGF-ß2 and matrilin2 in human aqueous humour with acute primary angle closure.

Primary angle-closure glaucoma (PACG) is the leading cause of irreversible blindness in the world. Angle closure induced by pupil block and secondary iris synechia is the fundamental pathology of the PACG. The molecular mechanisms of angle closure have not yet been clearly illustrated. This study was designed to investigate the protein difference in the aqueous humour and explore new biomarker of the PACG. Aqueous humour (AH) was collected from patients with acute primary angle closure (APAC) and cataract (n = 10 in APAC group) and patients with cataract only (n = 10 in control group). Samples were pooled and measured using label-free proteome technology. Then, the differentially expressed proteins (DEPs) were verified by ELISA using independent AH samples (n = 20 each group). More than 400 proteins were revealed in both groups through proteomics. Comparing the two groups, there were 91DEPs. These proteins participate in biological activities such as inflammation, fibrosis, nerve growth and degeneration and metabolism. We found that the expression of transforming growth factor-ß2 and matrilin2 was downregulated in the APAC group. The two proteins are related to inflammation and extracellular matrix formation, which might be involved in angle closure. This study characterized DEPs in AH of the APAC and found a downregulated protein matrilin2.

Cataract-causing Y204X mutation of crystallin protein CRYßB1 promotes its C-terminal degradation and higher-order oligomerization.

Crystallin proteins are a class of main structural proteins of the vertebrate eye lens, and their solubility and stability directly determine transparency and refractive power of the lens. Mutation in genes that encode these crystallin proteins is the most common cause for congenital cataracts. Despite extensive studies, the pathogenic and molecular mechanisms that effect congenital cataracts remain unclear. In this study, we identified a novel mutation in CRYBB1 from a congenital cataract family, and demonstrated that this mutation led to an early termination of mRNA translation, resulting in a 49-residue C-terminally truncated CRYßB1 protein. We show this mutant is susceptible to proteolysis, which allowed us to determine a 1.2-Å resolution crystal structure of CRYßB1 without the entire C-terminal domain. In this crystal lattice, we observed that two N-terminal domain monomers form a dimer that structurally resembles the WT monomer, but with different surface characteristics. Biochemical analyses and cell-based data also suggested that this mutant is significantly more liable to aggregate and degrade compared to WT CRYßB1. Taken together, our results provide an insight into the mechanism regarding how a mutant crystalin contributes to the development of congenital cataract possibly through alteration of inter-protein interactions that result in protein aggregation.

A crystallin mutant cataract with mineral deposits.

Connexin mutant mice develop cataracts containing calcium precipitates. To test whether pathologic mineralization is a general mechanism contributing to the disease, we characterized the lenses from a nonconnexin mutant mouse cataract model. By cosegregation of the phenotype with a satellite marker and genomic sequencing, we identified the mutant as a 5-bp duplication in the γC-crystallin gene (Crygcdup). Homozygous mice developed severe cataracts early, and heterozygous animals developed small cataracts later in life. Immunoblotting studies showed that the mutant lenses contained decreased levels of crystallins, connexin46, and connexin50 but increased levels of resident proteins of the nucleus, endoplasmic reticulum, and mitochondria. The reductions in fiber cell connexins were associated with a scarcity of gap junction punctae as detected by immunofluorescence and significant reductions in gap junction-mediated coupling between fiber cells in Crygcdup lenses. Particles that stained with the calcium deposit dye, Alizarin red, were abundant in the insoluble fraction from homozygous lenses but nearly absent in wild-type and heterozygous lens preparations. Whole-mount homozygous lenses were stained with Alizarin red in the cataract region. Mineralized material with a regional distribution similar to the cataract was detected in homozygous lenses (but not wild-type lenses) by micro-computed tomography. Attenuated total internal reflection Fourier-transform infrared microspectroscopy identified the mineral as apatite. These results are consistent with previous findings that loss of lens fiber cell gap junctional coupling leads to the formation of calcium precipitates. They also support the hypothesis that pathologic mineralization contributes to the formation of cataracts of different etiologies.

High glucose promotes osteogenic differentiation of human lens epithelial cells through hypoxia-inducible factor (HIF) activation.

Cataract, a leading cause of blindness, is characterised by lens opacification. Type 2 diabetes is associated with a two- to fivefold higher prevalence of cataracts. The risk of cataract formation increases with the duration of diabetes and the severity of hyperglycaemia. Hydroxyapatite deposition is present in cataractous lenses that could be the consequence of osteogenic differentiation and calcification of lens epithelial cells (LECs). We hypothesised that hyperglycaemia might promote the osteogenic differentiation of human LECs (HuLECs). Osteogenic medium (OM) containing excess phosphate and calcium with normal (1 g/L) or high (4.5 g/L) glucose was used to induce HuLEC calcification. High glucose accelerated and intensified OM-induced calcification of HuLECs, which was accompanied by hyperglycaemia-induced upregulation of the osteogenic markers Runx2, Sox9, alkaline phosphatase and osteocalcin, as well as nuclear translocation of Runx2. High glucose-induced calcification was abolished in Runx2-deficient HuLECs. Additionally, high glucose stabilised the regulatory alpha subunits of hypoxia-inducible factor 1 (HIF-1), triggered nuclear translocation of HIF-1α and increased the expression of HIF-1 target genes. Gene silencing of HIF-1α or HIF-2α attenuated hyperglycaemia-induced calcification of HuLECs, while hypoxia mimetics (desferrioxamine, CoCl2) enhanced calcification of HuLECs under normal glucose conditions. Overall, this study suggests that high glucose promotes HuLEC calcification via Runx2 and the activation of the HIF-1 signalling pathway. These findings may provide new insights into the pathogenesis of diabetic cataracts, shedding light on potential factors for intervention to treat this sight-threatening condition.

Novel and known variants in GJA3 and LIM2 in congenital cataract families from North India.

BACKGROUND: To identify the underlying genetic defects in autosomal dominant (ADCC) and autosomal recessive (ARCC) congenital cataract families from North India. METHODS: Detailed family histories were collected, pedigrees drawn followed by slit-lamp examination and lens photography. Mutation screening was performed using Sanger sequencing in the known candidate genes for crystallins, connexins, and membrane proteins. The pathogenicity of identified variants was assessed bioinformatically. RESULTS: In two ADCC families (CC-281 and CC-3015) with posterior lenticonus cataract, a novel change c.263C > T (p.P88L) in GJA3 in CC-281 family and a previously reported substitution c.388C > T (p.R130C) in LIM2 in CC-3015 family was observed. In an ARCC family (CC-3005) having central pulverulent cataract, a novel frameshift deletion (c.764delT;p.L255R46fs) in GJA3 was detected. The observed variants segregated completely with phenotypes in the affected members and were neither present in unaffected family members nor in the ethnically matched 150 controls (tested for two novel variants), hence excluding these as polymorphisms. CONCLUSIONS: Present study identified two novel mutations i.e., c.263C > T;p.P88L and c.764delT;p.L255R46fs in GJA3 in an ADCC and an ARCC family having posterior lenticonus and central pulverulent cataract, respectively. In another ADCC family with posterior lenticonus cataract, a previously reported mutation c.388C > T;p.R130C in LIM2 was observed. R130 may be a mutation hotspot as previously ADCC families from different ethnicities (UK/Czechia, China, Spain, Japan) also harbored the same substitution, however, with different phenotypes i.e., nuclear pulverulent, membranous, nuclear, lamellar, and sutural/lamellar. Findings in present study thus expand the mutation spectrum and phenotypic heterogeneity linked with GJA3 and LIM2.

Safety, effectiveness, and cost-effectiveness of immediate versus delayed sequential bilateral cataract surgery in the Netherlands (BICAT-NL study): a multicentre, non-inferiority, randomised controlled trial.

BACKGROUND: In an ageing population, efficiency improvements are required to assure future accessibility of cataract care. We aim to address remaining knowledge gaps by evaluating the safety, effectiveness, and cost-effectiveness of immediate sequential bilateral cataract surgery (ISBCS) versus delayed sequential bilateral cataract surgery (DSBCS). We hypothesised that ISBCS is non-inferior to DSBCS, regarding safety and effectiveness, and being superior in cost-effectiveness. METHODS: We did a multicentre, non-inferiority, randomised controlled trial, which included participants from ten Dutch hospitals. Eligible participants were 18 years or older, underwent expected uncomplicated surgery, and had no increased risk of endophthalmitis or refractive surprise. Participants were randomly assigned (1:1) to either the ISBCS (intervention) group or DSBCS (conventional procedure) group, using a web-based system stratified by centre and axial length. Participants and outcome assessors were not masked to the treatment groups because of the nature of the intervention. The primary outcome was the proportion of second eyes with a target refractive outcome of 1·0 dioptre (D) or less 4 weeks postoperatively, with a non-inferiority margin of -5% for ISBCS versus DSBCS. For the trial-based economic evaluation, the primary endpoint was the incremental societal costs per quality-adjusted life-year. All analyses were done by a modified intention-to-treat principle. Costs were calculated by multiplying volumes of resource use with unit cost prices and converted to 2020 Euros (€) and US$. This study was registered with ClinicalTrials.gov, number NCT03400124, and is now closed for recruitment. FINDINGS: Between Sept 4, 2018, and July 10, 2020, a total of 865 patients were randomly assigned to either the ISBCS group (427 [49%] patients; 854 eyes) or DSBCS group (438 [51%] patients; 876 eyes). In the modified intention-to-treat analysis, the proportion of second eyes with a target refraction of 1·0 D or less was 97% (404 of 417 patients) in the ISBCS group versus 98% (407 of 417) in the DSBCS group. The percentage difference was -1% (90% CI -3 to 1; p=0·526), thereby establishing non-inferiority for ISBCS compared with DSBCS. Endophthalmitis was not observed or reported in either group. Adverse events were comparable between groups, with only a significant difference in disturbing anisometropia (p=0·0001). Societal costs were €403 (US$507) lower with ISBCS than with DSBCS. The cost-effectiveness probability of ISBCS versus DSBCS was 100% across the willingness-to-pay range of €2500-80 000 (US$3145-100 629) per quality-adjusted life-year. INTERPRETATION: Our results showed non-inferiority of ISBCS versus DSBCS regarding effectiveness outcomes, comparable safety, and superior cost-effectiveness of ISBCS. National cost savings could amount to €27·4 million (US$34·5 million) annually, advocating for ISBCS if strict inclusion criteria are applied. FUNDING: Research grant from The Netherlands Organization for Health Research and Development (ZonMw) and Dutch Ophthalmological Society.