Genetic Characteristics and Clinical Manifestations of Foveal Hypoplasia in Familial Exudative Vitreoretinopathy.

PURPOSE: This study aimed to ascertain the occurrence of foveal hypoplasia (FH) in individuals diagnosed with familial exudative vitreoretinopathy (FEVR). DESIGN: Retrospective cohort study. METHODS: In this study, FEVR families and sporadic cases were diagnosed at the Eye and ENT Hospital, Fudan University, between 2017 and 2023. All patients attended routine ophthalmologic examinations and genetic screenings. The classification of FH was determined using optical coherence tomography (OCT) scans. The FH condition was classified into 2 subgroups: group A (FH being limited to the inner layers) and group B (FH affecting the outer layers). A total of 102 eyes from 58 patients were suitable for analysis. RESULTS: Forty-nine mutations in LRP5, FZD4, NDP, TSPAN12, KIF11, CTNNB1, and ZNF408 were examined and detected, with 26 of them being novel. Forty-seven eyes (46.1%) revealed FH. The majority (53.2%) were due to the typical grade 1 FH. Patients with mutations in LRP5 and KIF11 were found to exhibit a higher prevalence of FH (P = .0088). Group B displayed the lowest visual acuity compared with group A (P = .048) and the group without FH (P < .001). The retinal arteriolar angle in group B was significantly smaller than in group A (P = .001) and those without FH (P < .001). CONCLUSIONS: This study offers a new diagnostic approach and expands the spectrum of FEVR mutations. LRP5 and KIF11 were found to be more susceptible to causing FH in patients with FEVR. FEVR eyes with FH exhibited both greater visual impairment and reduced retinal arteriolar angles. The assessment of foveal status in patients with FEVR should be valued.

R-(+)-WIN55212-2 protects pericytes from ischemic damage and restores retinal microcirculatory patency after ischemia/reperfusion injury.

BACKGROUND AND PURPOSE: Cannabinoids are vasoactive substances that act as key regulators of arterial tone in the blood vessels supplying peripheral tissues and the central nervous system. This study aimed to investigate the potential of R-(+)-WIN55212-2 (WIN), a cannabinoid receptor 1 agonist (CB1), as a treatment for retinal ischemia/reperfusion (I/R) injury. EXPERIMENTAL APPROACH: Male Wistar rats were subjected to retinal I/R injury by increasing intraocular pressure in the anterior chamber. The rats were randomly divided into four groups: normal control, I/R, vehicle (pre-treated with dimethyl sulfoxide [DMSO] via intraperitoneal injection), and experimental (pre-treated with WIN at a dose of 1 ml/kg via intraperitoneal injection). The rats were sacrificed at different time points of reperfusion (1 hour, 3 hours, 6 hours, and 1 day) after inducing retinal I/R injury, and their retinas were collected for analysis. Oxygen-glucose deprived/reperfusion (OGD/R) was performed by initially perfusing the retinas with oxygenated artificial cerebrospinal fluid (ACSF), then switching to an OGD solution to simulate ischemia, followed by another perfusion with ACSF. Pericyte contraction and the "no-reflow" phenomenon were observed using infrared differential interference contrast (IR-DIC) microscopy and immunohistochemistry. Western blot, enzyme-linked immunosorbent assay (ELISA), and nitric oxide (NO) detection were used to explore the potential mechanism. KEY RESULTS: In both the OGD/R and I/R models, retinal pericytes exhibited persistent contraction even after reperfusion. The ability of WIN to regulate the tone of retinal pericytes and capillaries was specifically blocked by the BKCa inhibitor iberiotoxin (100 nM). WIN demonstrated a protective effect against retinal I/R injury by preserving blood flow in vessels containing pericytes. Pretreatment with WIN alleviated the persistent contraction and apoptosis of retinal pericytes in I/R-induced rats, accompanied by a reduction in intracellular calcium ion (Ca2+) concentration. The expression of CB1 decreased in a time-dependent manner in the I/R group. After I/R injury, endothelium-derived nitric oxide (eNOS) levels were reduced at all time points, which was successfully reversed by WIN therapy except for the 1 day group. Additionally, the downregulation of cyclic guanosine monophosphate (cGMP) and BKCa expression at 3 hours, 6 hours, and 1 day after I/R injury was restored by pretreatment of WIN. CONCLUSIONS & IMPLICATIONS: WIN exerted its protective effects on retinal I/R injury by inhibiting the contraction and apoptosis of pericytes through the CB1-eNOS-cGMP-BKCa signaling pathway, thus ameliorated the occlusion of retinal capillaries.

Adipose-derived stem cells show hepatic differentiation potential and therapeutic effect in rats with acute liver failure.

Hepatocyte transplantation contributes to the repair of liver damage, but hepatocyte resources are limited, making it difficult for this to become a routine treatment. Previous studies have confirmed that mesenchymal stem cells (MSCs) can be induced to differentiate into hepatocyte-like cells (HLCs) by adding different cytokine combinations in vitro, and they then play some roles of hepatocytes. Our previous studies found that the differentiation ability of stem cells is closely related to the origin of the tissue. To identify the mesenchymal stem cells that are most suitable for hepatic differentiation and the treatment of liver failure, we use a three-phase induction process in which human adipose-derived stem cells (hADSCs) and umbilical cord mesenchymal stem cells (hUCMSCs) are induced to differentiate towards HLCs in vitro, and rats with acute liver failure (ALF) induced by D-gal are cured by MSCs and MSC-derived HLCs (MSCs-HLC), respectively. We find that hADSCs are stronger than hUCMSCs in hepatic differentiation ability, and they have a better curative effect when using hADSCs-HLC or jointly using hADSCs and hADSCs-HLC, which has positive significance for hepatocyte regeneration, recovery of liver function and reduction of systemic inflammatory reaction, finally improving the survival rate of rats with acute liver failure.

Case report: Sympathetic ophthalmia after vitrectomies in a patient with Von Hippel-Lindau syndrome.

Background: Sympathetic ophthalmia (SO) is a rare but sight-threatening uveitis, and most observations have been made after typical manifestations occur. This report focuses on the choroidal changes detected by multimodal imaging at the presymptomatic stage of SO, which is implicated in the early recognition of SO. Case presentation: A 21-year-old woman suffered from decreased vision in the right eye and was diagnosed with retinal capillary hemangioblastomas associated with Von Hippel-Lindau syndrome. The patient underwent two 23-G pars plana vitrectomies (PPVs), soon after which typical signs of SO manifested. SO resolved quickly after the oral administration of prednisone and remained stable during the follow-up of more than 1 year. The retrospective analysis revealed preexisting bilaterally increased choroidal thickness, dots of flow void on the choroid, and choriocapillaris en-face slabs in optical coherence tomography angiography (OCTA) after the first PPV, which were all reversed by corticosteroid treatment. Conclusion: The case report highlights the involvement of the choroid and choriocapillaris at the presymptomatic stage of SO after the first inciting event. Abnormally thickened choroid and flow void dots suggested that SO had started and an ensuing surgery would run the risk of exacerbating SO. OCTA scanning of both eyes should be ordered routinely for patients with a history of trauma or intraocular surgeries, especially before the next surgical intervention. The report also suggests that non-human leukocyte antigen gene variation may also regulate the progression of SO, which requires further laboratory investigations.

Correlation between nutritional status screening by MNA-SF and acute stroke-associated infections in older adults.

PURPOSE: This study aimed to explore the correlation between nutritional status screening using the MNA-SF and stroke-associated infections (SAI) in older adults. METHODS: A retrospective study of patients aged over 70 years with acute stroke was conducted. The patients were divided into normal nutritional status, malnutrition risk, and malnutrition groups depending on their baseline MNA-SF scores. The correlation between nutritional status and SAI was identified using multivariate logistic regression. The receiver operating characteristic (ROC) curve was used to demonstrate the predicted value of MNA-SF. RESULTS: 497 patients were included, 101 (20.32%) developed SAI. 32.29% of patients with malnutrition developed SAI, while 25.14% of those with malnutrition risk developed SAI. Malnutrition (aOR 4.58, 95% CI 2.34-8.96, p < 0.001) and risk of malnutrition (aOR 3.70, 95%CI 2.01-6.85, p < 0.001) were independent risk factors for SAI in older stroke patients. The area under the curve (AUC) value of MNA-SF was 0.713. CONCLUSION: MNA-SF is a simple and effective nutritional screening tool for predicting the occurrence of SAI in older patients with acute stroke.

Chlorogenic acid enhances alveolar macrophages phagocytosis in acute respiratory distress syndrome by activating G protein-coupled receptor 37 (GPR 37).

BACKGROUND: Impaired alveolar macrophages phagocytosis can contribute to pathogenesis of acute respiratory distress syndrome (ARDS) and negatively impacts clinical outcomes. Chlorogenic acid (CGA) is a naturally occurring polyphenolic compound with potential anti-inflammatory and antioxidant bioactivities. Studies have shown that CGA plays a protective role in ARDS, however, the precise protective mechanism of CGA against ARDS, is still unclear. PURPOSE: The aim of this study was to investigate whether CGA enhances alveolar macrophages phagocytosis to attenuate lung injury during ARDS. METHODS: RAW264.7 cells were stimulated with lipopolysaccharides (100 µg/ml for 24 h) and treated with CGA (100, 200, and 400 µM CGA for 1 h) to measure pro-inflammatory cytokine levels, GPR37 expression and macrophages phagocytosis. Mouse models of ARDS induced by cecal ligation and perforation (CLP) surgery were treated with CGA (100 or 200 mg/kg) to investigate lung inflammatory injury and alveolar macrophages phagocytosis. Computational modeling was performed to examine potential binding sites of G protein-coupled receptor 37 (GPR37) with CGA, and the results were validated by interfering with the binding sites. RESULT: In vitro, CGA notably ameliorated inflammatory response and increased phagocytosis in lipopolysaccharides-induced RAW264.7 cells. In vivo, CGA administration significantly alleviated lung inflammatory injury, decreased the bacteria load in the lung, promoted alveolar macrophages phagocytosis and improved the survival rate in mice with CLP-induced ARDS. Moreover, CGA markedly upregulated the expression of GPR37 in vivo and in vitro. However, the protective effect of CGA against ARDS were reversed after silencing the expression of GPR37. CONCLUSION: CGA has a protective effect against ARDS and may enhance alveolar macrophages phagocytosis and attenuate lung inflammatory injury by upregulating GPR37 expression.

Clinical and Genetic Analysis of Retinitis Pigmentosa with Primary Angle Closure Glaucoma in the Chinese Population.

PURPOSE: Retinitis pigmentosa (RP) constitutes a class of common inherited retinal dystrophies. Patients with RP and comorbid primary angle-closure glaucoma (PACG) have been described, but the relationship between the diseases remains unclear. This study investigated the clinical and genetic characteristics of Chinese patients with RP and comorbid PACG. METHODS: Of 1356 patients with RP, we analyzed the genetic features of 39 RP patients with PACG using next-generation sequencing and reviewed their clinical characteristics. RESULTS: In total, 18 patients with acute PACG and 21 patients with chronic PACG were included in this study; their age at examination was 50.54 ± 12.99 years (range, 25.0-71.0 years), and their age at PACG onset was 46.04 ± 14.50 years (range, 24.9-68.0 years). Additionally, the mean lens thickness (LT) was 4.49 ± 0.44 µm, and the mean axial length (AL) was 22.63 ± 1.17 mm. Notably, the prevalence of PACG in patients with RP was 2.88%; this was higher than the prevalence in the general population. This could be explained by nanophthalmos, thickened lentis, ectopia lentis, or zonular insufficiency. Furthermore, patients with a shorter AL, a greater LT, iridociliary cysts, or nanophthalmos exhibited earlier development of PACG. Overall, 30 disease-causing variants spanning 17 genes were identified in 56.41% of the patients, and PRPH2 was the most common mutation gene. CONCLUSIONS: Our findings revealed that there is a strong association between RP and PACG. Furthermore, intraocular pressure (IOP) should be measured in patients with RP to protect them from the aggravated damage of an elevated IOP.

Nobiletin protects retinal ganglion cells in models of ocular hypertension in vivo and hypoxia in vitro.

Glaucoma, a common cause of blindness, is characterized by the progressive loss of retinal ganglion cells (RGCs). Growing evidence suggests that nobiletin (NOB) is a promising neuroprotective drug; however, its effects on glaucomatous neurodegeneration remain unknown. Using rat models of microbead occlusion in vivo and primary RGCs model of hypoxia in vitro, we first demonstrate that NOB reduces RGC apoptosis by a TUNEL assay, Hoechst 33342 staining and FluoroGold (FG) retrograde labeling. This effect does not depend on intraocular pressure (IOP) lowering. Additionally, NOB partially restored the functional and structural damage of inner retinas, attenuated Müller glial activation and oxidative stress caused by ocular hypertension. At 2 weeks after IOP elevation, NOB further enhanced Nrf2/HO-1 pathway in RGCs to withstand the cumulative damage of ocular hypertension. With the administration of HO-1 inhibitor tin-protoporphyrin IX (SnPP), the protective effect of NOB was attenuated. Overall, these results indicate that NOB exerts an outstanding neuroprotective effect on RGCs of glaucomatous neurodegeneration. Besides, interventions to enhance activation of Nrf2/HO-1 pathway can slow the loss of RGCs and are viable therapies for glaucoma.

Ocular Manifestations in a Chinese Pedigree of Familial Amyloidotic Polyneuropathy Carrying the Transthyretin Mutation c.401A>G (p.Tyr134Cys)

Familial amyloid polyneuropathy (FAP) caused by a genetic mutation in transthyretin (TTR) is an autosomal dominant hereditary disease. The retrospective, observational case series study presents the ocular clinicopathological findings of five cases carrying the TTR mutation c.401A>G (p.Tyr134Cys). Multimodal retinal imaging and electrophysiological examination, Congo red staining and immunohistochemical analysis of specimens, and genetic analyses were performed. Cases 1 and 2 were symptomatic with vitreous and retinal amyloid deposition and poor visual recovery. Case 3 had a symptomatic vitreous haze in the left eye with good postoperative visual recovery. The right eye of case 3 and the eyes of cases 4 and 5 were asymptomatic. Thicker retinal nerve fiber layer, retinal venous tortuosity with prolonged arteriovenous passage time on fluorescein angiography and retinal dysfunction detected by multifocal electroretinogram occurred even in asymptomatic eyes. Moreover, the internal limiting membrane from patients with FAP was stained positive for Congo red and transforming growth factor-ß1. The results highlight the amyloid deposition of mutant TTR in the optic disc and retina, even in the asymptomatic stage. The deposited amyloid leads to increased resistance to venous return and retinal functional abnormalities. Therefore, careful follow-up of structural and functional changes in the retina is needed, even in asymptomatic patients with FAP.

Rebuilding hippocampus neural circuit with hADSC-derived neuron cells for treating ischemic stroke.

BACKGROUND: Human adipose-derived stem cells (hADSCs) have been demonstrated to be a promising autologous stem cell source for treating various neuronal diseases. Our study indicated that hADSCs could be induced into neuron-like cells in a stepwise manner that are characterized by the positive expression of MAP2, SYNAPSIN 1/2, NF-200, and vGLUT and electrophysiological activity. We first primed hADSCs into neuron-like cells (hADSC-NCs) and then intracerebrally transplanted them into MCAO reperfusion mice to further explore their in vivo survival, migration, integration, fate commitment and involvement in neural circuit rebuilding. RESULTS: The hADSC-NCs survived well and transformed into MAP2-positive, Iba1- or GFAP-negative cells in vivo while maintaining some proliferative ability, indicated by positive Ki67 staining after 4 weeks. hADSC-NCs could migrate to multiple brain regions, including the cortex, hippocampus, striatum, and hypothalamus, and further differentiate into mature neurons, as confirmed by action potential elicitation and postsynaptic currents. With the aid of a cell suicide system, hADSC-NCs were proven to have functionally integrated into the hippocampal memory circuit, where they contributed to spatial learning and memory rescue, as indicated by LTP improvement and subsequent GCV-induced relapse. In addition to infarction size shrinkage and movement improvement, MCAO-reperfused mice showed bidirectional immune modulation, including inhibition of the local proinflammatory factors IL-1α, IL-1ß, IL-2, MIP-1ß and promotion proinflammatory IP-10, MCP-1, and enhancement of the anti-inflammatory factors IL-15. CONCLUSION: Overall, hADSC-NCs used as an intermediate autologous cell source for treating stroke can rebuild hippocampus neuronal circuits through cell replacement.

Nobiletin Alleviates Astrocyte Activation and Oxidative Stress Induced by Hypoxia In Vitro.

Increasing evidence indicates that nobiletin (NOB) is a promising neuroprotective agent. Astrocyte activation plays a key role in neurodegenerative disorders. Thus, this study aims to investigate the effects of NOB on astrocyte activation and the potential mechanisms. In this study, astrocytes were exposed to hypoxia injury for 24 h to induce activation in vitro. Glial fibrillary acidic protein (GFAP) was chosen as a marker of astrocyte activation. To evaluate the effects of NOB on the migration of activated astrocytes, we used a scratch wound healing assay and Transwell migration assay. In addition, the levels of reactive oxygen species (ROS), malondialdehyde (MDA), mitochondrial membrane potential, Nrf2 and HO-1 were measured to investigate the mechanisms of NOB in the activation of astrocytes. We found that NOB alleviated astrocyte activation and decreased GFAP expression during hypoxia. Simultaneously, NOB alleviated the migration of astrocytes induced by hypoxia. With NOB treatment, hypoxia-induced oxidative stress was partially reversed, including reducing the production of ROS and MDA. Furthermore, NOB significantly improved the mitochondrial dysfunction in activated astrocytes. Finally, NOB promoted Nrf2 nuclear translocation and HO-1 expression in response to continuous oxidative damage. Our study indicates, for the first time, that NOB alleviates the activation of astrocytes induced by hypoxia in vitro, in part by ameliorating oxidative stress and mitochondrial dysfunction. This provides new insights into the neuroprotective effects of NOB.

Genotypic spectrum and phenotype correlations of EYS-associated disease in a Chinese cohort.

BACKGROUND: To date, certain efforts have been made to investigate the clinical and genetic characteristics of patients with EYS mutations. However, data for Chinese patients are limited. OBJECTIVES: To perform a detailed phenotyping and genetic characterization of 55 Chinese patients with EYS-RD, and to identify risk factors for these clinical data. METHODS: A total of 55 patients with EYS-RD were recruited. Best-corrected visual acuity (BCVA), patient age, age at symptom onset, disease duration, and genetic information were collected. RESULTS: Thirty-six novel variants, three hot mutations of EYS (30.3%, c.6416G>A, c.6557G>A, c.7492G>C) and one hot region (49.06%, Laminin G domains) were identified. In all, 36.84% of the mutations occurred at base G site, and majority of mutations (56.56%) were missense. Late-truncating mutations are significantly more prevalent (41.30%). The mean age of onset was 15.65 ± 14.67 years old; it had no significant correlation with genotype. The average BCVA was 0.73 ± 0.93 LogMAR, and 61.8% of eyes had a BCVA better than 0.52 logMAR. BCVA was positively correlated with disease duration time. The mean MD was 23.18 ± 7.34 dB, MD showed a significant correlation with genotype and age. Cataract was present in 56.45% of patients, and 42.59% of patients showed an absence of pigmentation in the retina. Cataract and hyperpigmentation both showed a significant correlation with age. CONCLUSIONS: EYS-RD is associated with a moderate phenotype with onset around adolescence, but great variability. Our study largely enhances the current knowledge of phenotypic and genotypic characteristics of EYS-RD, which could pave the way for better management of these patients.

Frequency and phenotypic characteristics of RPE65 mutations in the Chinese population.

BACKGROUND: The retinoid isomerohydrolase RPE65 has received considerable attention worldwide since a successful clinical gene therapy was approved in 2017 as the first treatment for vision loss associated with RPE65-mediated inherited retinal disease. Identifying patients with RPE65 mutations is a prerequisite to assessing the patients' eligibility to receive RPE65-targeted gene therapies, and it is necessary to identify individuals who are most likely to benefit from gene therapies. This study aimed to investigate the RPE65 mutations frequency in the Chinese population and to determine the genetic and clinical characteristics of these patients. RESULTS: Only 20 patients with RPE65 mutations were identified, and RPE65 mutations were determined to be the 14th most common among all patients with genetic diagnoses. Ten novel variants and two hotspots associated with FAP were identified. A literature review revealed that a total of 57 patients of Chinese origin were identified with pathogenic mutations in the RPE65 gene. The mean best Snellen corrected visual acuity was worse (mean 1.3 ± 1.3 LogMAR) in patients older than 20 years old than in those younger than 15 years old (0.68 ± 0.92 LogMAR). Bone spicule-like pigment deposits (BSLPs) were observed in six patients; they were older than those without BSLP and those with white-yellow dots. Genotype-phenotype analysis revealed that truncating variants seem to lead to a more severe clinical presentation, while best corrected visual acuity testing and fundus changes did not correlate with specific RPE65 variants or mutation types. CONCLUSIONS: This study provides a detailed clinical-genetic assessment of patients with RPE65 mutations of Chinese origin. These results may help to elucidate RPE65 mutations in the Chinese population and may facilitate genetic counseling and the implementation of gene therapy in China.

Prevalence and genetic-phenotypic characteristics of patients with USH2A mutations in a large cohort of Chinese patients with inherited retinal disease.

AIMS: To investigate the frequency of USH2A mutation and the clinical and genetic differences between Usher syndrome type II (USH2) and retinitis pigmentosa (RP) in a large cohort of Chinese patients. METHODS: A total of 1381 patients with inherited retinal disease (IRD) were recruited. The phenotypic and genotypic information of patients with USH2A mutations was evaluated. RESULTS: The prevalence of patients with USH2A mutations was 15.75%, which was the most frequently detected gene in this cohort of patients. Hotspot of USH2A mutations was c.8559-2A >G and c.2802T >G. Patients with USH2 had an earlier and more serious decline of visual function and damage to retina structure than did patients with RP in the first 10 years (p<0.05), but there was no difference in the visual prognosis between the two groups when the course of disease exceeded 10 years (p>0.05). Missense variants had less severe consequences and were found more commonly in RP, whereas more deleterious genotypes were associated with an earlier onset of disease and were found more commonly in USH2. CONCLUSIONS: This study provides detailed clinical-genetic assessment of patients with USH2A mutations of Chinese origin, enabling precise genetic diagnoses, better management of these patients and putative therapeutic approaches.

Comprehensive analysis of genetic and clinical characteristics of 30 patients with X-linked juvenile retinoschisis in China.

PURPOSE: To provides the clinical and genetic characteristics of a series of Chinese patients with X-linked juvenile retinoschisis (XLRS) through multimodal imaging and next-generation sequencing. METHODS: Thirty patients (60 eyes) from 29 unrelated families of Chinese origin with XLRS were screened using multigene panel testing, and underwent a complete clinical evaluation. All variants identified in this study and reported in the Human Gene Mutation Database were analysed. RESULTS: Twenty-five distinct variants in the retinoschisin gene were identified, of which eight were novel, and one was de novo. Missense mutations were the most prevalent type, and mutation hot spot was localized in the discoidin domain. The mean Snellen best-corrected visual acuity was 0.28 ± 0.17. Of all eyes presenting with schisis, 92.86% had lamellar schisis and 62.5% had peripheral schisis. Schisis changes mostly involved inner and outer nuclear layers. X-linked juvenile retinoschisis (XLRS) patients had a high incidence of complications, and peripheral schisis was a risk factor for it. No obvious genotype-phenotype association was observed. CONCLUSION: This study provides comprehensive analyses of the genetic and clinical characteristics of XLRS in a cohort of Chinese patients. The fourth de novo mutation in RS1 was identified. And we show that XLRS has a wide spectrum of clinical characteristics; hence, molecular diagnosis is crucial for its diagnosis, differential diagnosis and genetic counselling. Peripheral schisis is a risk factor for the high incidence of complications, and no clear genotype-phenotype correlations were found.

Novel variants of ABCA4 in Han Chinese families with Stargardt disease.

BACKGROUND: Stargardt disease (STGD1) is a common recessive hereditary macular dystrophy in early adulthood or childhood, with an estimated prevalence of 1:8000 to 1:10,000. ABCA4 is the causative gene for STGD1. The current study aims at identifying the novel disease-related ABCA4 variants in Han Chinese families with STGD1 using next-generation sequencing (NGS). METHODS: In the present study, 12 unrelated Han Chinese families (19 males and 17 females) with STGD1 were tested by panel-based NGS. In order to capture the coding exons and the untranslated regions (UTRs) plus 30 bp of intronic flanking sequences of 792 genes, which were closely associated with usual ophthalmic genetic disease, we designed a customized panel, namely, Target_Eye_792_V2 chip. STGD1 patients were clinically diagnosed by experienced ophthalmologists. All the detected variants were filtered and analyzed through the public databases and in silico programs to assess potential pathogenicity. RESULTS: Twenty-one ABCA4 mutant variants were detected in 12 unrelated Han Chinese families with STGD1, containing 14 missense, three splicing, two frameshift, one small deletion, and one nonsense variants. Base on the American College of Medical Genetics (ACMG) guidelines, 8 likely pathogenic and 13 pathogenic variants were determined. The functional consequences of these mutant variants were predicted through in silico programs. Of the 21 mutant variants in ABCA4, two novel coding variants c.3017G > A and c.5167 T > C and one novel null variant c.3051-1G > A were detected in three unrelated probands. CONCLUSIONS: By panel-based NGS, 21 ABCA4 variants were confirmed in 12 unrelated Han Chinese families. Among them, 3 novel mutant variants were found, which further expanded the ABCA4 mutation spectrum in STGD1 patients.

Cell Development Deficiency and Gene Expression Dysregulation of Trisomy 21 Retina Revealed by Single-Nucleus RNA Sequencing.

Retina is a crucial tissue for capturing and processing light stimulus. It is critical to describe the characteristics of retina at the single-cell level for understanding its biological functions. A variety of abnormalities in terms of morphology and function are present in the trisomy 21 (T21) retina. To evaluate the consequences of chromosome aneuploidy on retina development, we identified the single-cell transcriptional profiles of a T21 fetus and performed comprehensive bioinformatic analyses. Our data revealed the diversity and heterogeneity of cellular compositions in T21 retina, as well as the abnormal constitution of T21 retina compared to disomic retina. In total, we identified seven major cell types and several subtypes within each cell type, followed by the detection of corresponding molecular markers, including previously reported ones and a series of novel markers. Through the analysis of the retinal differentiation process, subtypes of retinal progenitor cells (RPCs) exhibiting the potential of different retinal cell-type commitments and certain Müller glial cells (MGs) with differentiating potency were identified. Moreover, the extensive communication networks between cellular types were confirmed, among which a few ligand-receptor interactions were related to the formation and function of retina and immunoregulatory interactions. Taken together, our data provides the first ever single-cell transcriptome profiles for human T21 retina, which facilitates the understanding on the dosage effects of chromosome 21 on the development of retina.

Mutation Screening of mtDNA Combined Targeted Exon Sequencing in a Cohort With Suspected Hereditary Optic Neuropathy.

Purpose: Leber hereditary optic neuropathy (LHON) and autosomal dominant optic atrophy (ADOA) are the two commonest forms of hereditary optic neuropathy. The aim of this study was to comprehensively investigate the incidence and spectrum of mutations in patients with suspected hereditary optic neuropathy by combining mitochondrial DNA (mtDNA) genome-wide and targeted exon sequencing. Methods: A cohort of 1101 subjects were recruited to participate in the study, comprising 177 families (177 probands and their family members, a total of 537 subjects, including 254 patients) and 164 sporadic cases with suspected hereditary optic neuropathy, and 400 unrelated control subjects for genetic analysis: all subjects (including control subjects) underwent a comprehensive ophthalmologic examination and were subjected to sequencing analysis of mtDNA genome-wide and targeted exon. Overall, targeted exon sequencing was used to screen 792 genes associated with common hereditary eye diseases, and the mtDNA genome-wide were screened by next-generation sequencing. Results: We found variants detected in 168 (40.2%, 168/418) of the 418 patients screened. Among these, 132 cases (78.6%, 132/168) were detected with known LHON disease-causing mtDNA variants; 40 cases (23.8%, 40/168) were detected with nuclear DNA (ntDNA) variants, which included 36 cases (21.4%, 36/168) with detected OPA1 mutations, 4 patients (2.4%, 4/168) with detected OPA3 mutations, and 2 patients (1.2%, 2/168) with detected TMEM126A homozygous mutation. Coexistence variation (mtDNA/mtDNA [n = 16], ntDNA/ntDNA [n = 4], mtDNA/ntDNA [n = 7]) was found in 27 patients (16.4%, 27/165), including mtDNA/ntDNA coexistence variation that was detected in seven patients. Among these ntDNA mutations, 38 distinct disease-causing variants, including autosomal recessive heterozygous mutations, were detected, which included 22 novel variants and two de novo variants. Total haplogroup distribution showed that 34.5% (29/84) and 28.6% (24/84) of the affected subjects with m.11778G>A belonged to haplogroup D and M, with a high frequency of subhaplogroups D4, D5, and M7. Conclusions: The LHON-mtDNA mutations are the commonest genetic defects in this Chinese cohort, followed by the OPA1 mutations. To our knowledge, this is the first comprehensive study of LHON, ADOA, and autosomal recessive optic atrophy combined with mtDNA genome-wide and targeted exon sequencing, as well as haplogroup analysis, in a large cohort of Chinese patients with suspected hereditary optic neuropathy. Our findings provide a powerful basis for genetic counseling in patients with suspected hereditary optic neuropathy. Translational Relevance: We applied mtDNA genome-wide sequencing combined with panel-based targeted exon sequencing to explore the pathogenic variation spectrum and genetic characteristics of patients with suspected hereditary optic neuropathy, providing a comprehensive research strategy for clinical assistant diagnosis, treatment, and genetic counseling.

Next-generation sequencing-based clinical diagnosis of choroideremia and comprehensive mutational and clinical analyses.

BACKGROUND: To report the clinical and genetic findings from seven Chinese patients with choroideremia. METHODS: Five hundred seventy-eight patients with a clinically suspected diagnosis of retinitis pigmentosa (RP) underwent comprehensive ophthalmic examinations. Next-generation sequencing (NGS) was performed on samples from all patients. Detailed clinical characteristics of the patients with choroideremia identified in this study were assessed using multimodal imaging. RESULTS: Seven patients with choroideremia were identified, and six novel variants in CHM (c.1960 T > C p.Ter654Gln, c.1257del p.Ile420*fs1, c.1103_1121delATGGCAACACTCCATTTTT p.Tyr368Cysfs35, c.1414-2A > T, and c.1213C > T p.Gln405Ter, c.117-1G > A) were revealed. All variants were deleterious mutations: two were frameshifts, two were nonsense mutations, two were splicing mutations, and one was a readthrough mutation. The clinical phenotypes of these patients were markedly heterogeneous, and they shared many common clinical features with RP, including night blindness, constriction of the visual field and gradually reduced visual acuity. However, patients with choroideremia showed pigment hypertrophy and clumping, and chorioretinal atrophy, and a majority of patients with choroideremia presented with retinal tubulations in the outer layer of the retina. CONCLUSIONS: We provide a detailed description of the genotypes and phenotypes of seven patients with choroideremia who were accurately diagnosed using NGS. These findings provide a better understanding of the genetics and phenotypes of choroideremia.

Novel variants associated with Stargardt disease in Chinese patients.

PURPOSE: Stargardt disease (STGD) is the most frequent cause of hereditary macular dystrophy in childhood. Variants in the ABCA4, ELOVL4, PROM1, BEST1, and PRPH2 genes have been detected in patients with autosomal recessive or dominant STGD. This study was aimed at identifying the novel disease-associated variants in Chinese patients with STGD. METHODS: Ten Chinese families and two sporadic cases with STGD (n = 32) were enrolled in the study. All subjects underwent genetic analysis with next-generation sequencing (NGS), which was based on a specially customized capture panel targeting exons and untranslated regions (UTRs) of 792 genes related to common hereditary ophthalmopathy. Variants were analyzed to assess possible pathogenicity. RESULTS: Fourteen disease-associated variants of ABCA4 were detected in 9 Chinese families with autosomal recessive STGD, including 11 pathogenic variants and 3 likely pathogenic variants. Variant c.4253 + 4C > T in ABCA4 was identified as one de novo variant. Of the 14 distinct variants in ABCA4, 7 novel variants were found. In addition, one known variant of PROM1, c.1117C > T (p.Arg373Cys), was detected in one family and one sporadic case with autosomal dominant STGD, respectively. One novel missense variant of ELOVL4, c.59A > G (p.Asn20Ser), was found in one sporadic case with autosomal dominant STGD. The potential deleterious effects of these novel variants were confirmed through intensive analysis. CONCLUSION: By panel-based NGS, 8 novel disease-associated variants are identified in two genes responsible for STGD, including ABCA4 and ELOVL4. Our results further extend the mutation spectrum of these two genes in Chinese patients with STGD. One ABCA4 c.4253 + 4C > T variant is identified as a de novo splicing variant.