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To investigate the pathogenic features of the polypoidal lesions from the specimens of polypoidal choroidal vasculopathy extracted from human subjects.Seven specimens of polypoidal lesions extracted from five eyes of six patients (mean age, 60.16 ± 10.41 years) of polypoidal choroidal vasculopathy were examined. The polypoidal lesions were obtained by surgical excision. Thereafter, a histopathological analysis of the specimens was performed.The polypoidal lesions were oval nodules located underneath the retinal pigment epithelium. A pathological study of the lesions revealed that Bruch's membrane schisis was observed in all specimens and they were all located in the Bruch's membrane. The Bruch's membrane schisis and serosanguineous materials constituted the main structure of the lesions in five of the seven specimens, with small vessels being observed in two specimens. One specimen was composed of two polypoidal lesions of different characteristics, and one specimen had a neovessel membrane complex with several polypoidal lesions. Inflammatory cells and blood vessels were observed in the polypoidal lesion of the specimen with neovessel membrane complex.Polypoidal lesions of polypoidal choroidal vasculopathy are abnormalities of the Bruch's membrane. The lesions are characterized by the Bruch's membrane schisis, which is filled with serosanguineous materials. The lesions are progressive and may contain inflammatory cells and blood vessels.

Background:To evaluate whether better prognosis can be obtained from the continuous surgical treatment (CST) before fibrosis /scar formation in open anterior segment (AS) injury.Methods:Prospective cohort study.19 eyes of 19 patients were selected from the database of Eye Injury Vitrectomy Study, which is a multi-center prospective cohort study and began in 1990s, from January 1, 2020 to July 31, 2021. Of these, 9 patients who received CST were assigned to group 1, and 10 patients without continuous treatment after initial wound repairment were included in group 2. Comparison between two groups was conducted in final best corrected visual acuity (BCVA), presumably significant complications with X² test. Corneal leucoma area ratio, astigmatism and score of AS abnormalities were analyzed by Students’ t-test.Results:Gross appearance displays remarkable differences between two groups with observation on AS. The individually paradigm, such as corneal-scleral wound and intraocular abnormalities, can be noted over various post-traumatic time intervals. The final BCVA is better in group 1(P=0.011). The complications directly caused by AS injury, adhesive corneal leucoma, uneven anterior chamber, block of light pass through pupil and fibrosis/scar are more frequent in group 2 (P=0.011, 0.022, 0.037 and 0.040, respectively). Secondary glaucoma (three cases) and severe AS structure destruction (two cases) occurred only in group 2 (P=0.037 and 0.474, respectively).Conclusions:Relying on comprehensive evaluations, advantages of CST of initiating before fibrosis/scar formation in open AS injury show dominance over following nature course of wound healing after wound repairment.

Severe mechanical ocular trauma with no light perception (NLP) predicts a poor prognosis of visual acuity and enucleation of the eyeball. Since the innovative treatment concept of exploratory vitreoretinal surgery has developed and treatment technology has advanced, the outcomes of severe ocular trauma treatment in NLP patients have greatly improved. However, there remains a lack of unified standards for the determination, surgical indication, and timing of vitrectomy in NLP eye treatment. To address these problems, we aimed to create evidence-based medical guidelines for the diagnosis, treatment, and prognosis of mechanical ocular trauma with NLP. Sixteen relevant recommendations for mechanical ocular trauma with NLP were obtained, and a consensus was reached. Each recommendation was explained in detail to guide the treatment of mechanical ocular trauma associated with NLP.

Background Loeys-Dietz syndrome (LDS) is an autosomal dominant disorder of the connective tissue that has phenotypic overlap with Marfan syndrome，but the aortic root dissections can be more aggressive and ocular findings in LDS may be very different than Marfan syndrome. Methods Review of one case of LDS with novel retinal findings. Results A 30-year-old female with LDS was found to have retinal arterial macroaneurism (RAM) in the left eye. Local laser photocoatulation and intravitreal anti-VEGF was applied but exudative retinal detachment developed soon after that. Transscleral diode photocoagulation was then performed and subretinal fluid was resolved. Conclusion RAM is a unique finding of LDS related to a novel mutation of TGFBR1.

Purpose To delineate anatomic and visual outcomes of injured eye globes with perforating, and to develop the prognostic indicators for perforating eyes. Methods The case series study, from a multicenter prospective cohort database. To the date of December 31st, 2018, of 63 perforating globes were selected. All cases underwent vitreoretinal surgeries or enucleations, and were followed up for at least 6 months. Demographic characteristics, basic examination for traumatized eyes, and intraocular tissue damages were recorded by surgery-in-chief. At the follow-up visit, best corrected VA, intraocular pressure, the intraocular tamponade material, retinal anatomic outcome of eye-globes, and phthisis or enucleation were evaluated. Results Fifty injured eyes (79%) were caused by sharp objects and 13 eyes (21%) were injured by a missiles. Twenty-two injured eyes can be anatomically restored with final vision of more than 4/200 through vitreoretinal surgery. The PVR-C (OR = 5.67, P = 0.01), area of retinectomy more than 2 times of optic disk (OR = 5.16, P = 0.04), and macular damage (OR = 6.38, P = 0.01) were correlated with unfavorable outcomes. Conclusion The injured eyes with perforation can be saved through vitreoretinal surgery, the PVR-C, retinectomy more than 2 times of optic disk, and macular damage were independent risk factors for poor long-term prognosis.

To observe the ultrastructural outcomes of autologous transplantation of retinal pigment epithelium-partial-thickness choroidal (RPE-PTC) sheets in rabbits after 6 months.Eighteen pigmented rabbits were used in this study. Among them, nine rabbits were used for autologous transplantation of RPE-PTC sheets. Tissue sections were observed under a transmission electron microscope for one, three, and six months after transplantation, respectively.One, three, and six months after the autologous transplantation of RPE-PTC sheets, the inner and outer segments of photoreceptor cells were arranged regularly, and the connection between the inner and outer segments was normal. The inner structure of the RPE cells and tight junctions among them remained normal. Phagocytosis of outer segment of photoreceptor cells could also be observed in RPE cells. The structure of the Bruch's membrane appeared loose, rather than being dense as normal, and it was undulated after one and three months, while it became dense after six months. The graft and the bed were healed well, the boundary was unclear, and the graft was vascularized after one, three, and six months, respectively.Our findings revealed that the RPE-PTC sheets could quickly rebuild blood vessels, thereby maintaining the normal physiological functions of RPE cells, as well as the survival and functional status of photoreceptor cells for a long-time.

Age-related macular degeneration (AMD) is a major reason of blindness in the elderly. MicroRNAs are implicated in various pathological processes, including inflammation and apoptosis. In this study, we aim to investigate the biological functions of miR-21-3p in inflammation and apoptosis caused by lipopolysaccharide (LPS) in human retinal pigment epithelial (ARPE-19) cells. The miR-21-3p inhibitor and mimic were transfected into ARPE-19 cells for 48 hours, followed by exposed to LPS (10 μg/mL) for 24 hours. The mRNA and protein expression of IL-6 and MCP-1 were measured using real-time PCR (RT-PCR) and enzyme-linked immunosorbent assays. Cell viability, apoptosis, caspase 3 activity, cleaved caspase-3 and cleaved-PARP protein levels were detected to evaluate the effects of miR-21-3p on apoptosis. Additionally, the target relationship between miR-21-3p and regulator of G-protein signalling 4 (RGS4) was verified by dual luciferase reporter assay. RT-PCR analysis demonstrated that LPS induced miR-21-3p expression. Inhibition of miR-21-3p reduced the mRNA and protein levels of IL-6 and MCP-1. Apoptosis, caspase-3 activity, and cleaved-caspase 3 and cleaved PARP protein levels were repressed by the miR-21-3p inhibitor. However, overexpression of miR-21-3p showed the opposite results. Furthermore, we identified that miR-21-3p directly targeted the 3' untranslated region of RGS4. MiR-21-3p negatively regulated the expression of RGS4 both in mRNA and protein levels. Silencing RGS4 reduced the anti-inflammatory and anti-apoptotic effects of miR-21-3p inhibitor. Our results revealed that miR-21-3p inhibition targeted RGS4 to attenuate inflammatory responses and apoptosis caused by LPS in ARPE-19 cells.

Background and Objective In smoking cessation clinical research and practice, objective validation of self-reported smoking status is crucial for ensuring the reliability of the primary outcome, that is, smoking abstinence. Speech signals convey important information about a speaker, such as age, gender, body size, emotional state, and health state. We investigated (1) if smoking could measurably alter voice features, (2) if smoking cessation could lead to changes in voice, and therefore (3) if the voice-based smoking status assessment has the potential to be used as an objective smoking cessation validation method. Methods A systematic review of the scientific literature was conducted to compile studies on smoking status assessment based on voice features. We searched nine scientific databases for original studies involving the effects of smoking on voice features, the effects of smoking cessation on voice features. Results A total of 34 studies were identified for review. We found that fundamental frequency, jitter, shimmer, harmonics to noise ratio, and other voice features are affected by smoking and could be used to assess smoking status. Conclusion Speech assessment of smoking status based on voice features has potential as a smoking status validation method, as it is simple, reliable, and less time-consuming. Furthermore, this study provides recommendations for future research on the objective speech assessment of smoking status based on voice features.

Background: Closure of traumatic macular hole (TMH) can be achieved spontaneously or by surgical intervention. Thus far, there exist no prospective comparative studies that have analyzed the difference between the two modalities. This study aimed to compare the anatomical and visual recovery of eyes with TMH following either an immediate vitrectomy or six-month observation. Methods: This was a multicenter prospective comparative study. Eight centers participated in the study. Patient data from 40 eyes with a recent history of blunt ocular trauma and newly formed full-thickness TMH were recruited in this study. The participating patients selected between an early vitrectomy or a six-month observation after a doctor explained the potential benefits and risks of both strategies in an unbiased manner. Twenty-five patients underwent an immediate vitrectomy, and 15 patients received six-month observation. Patients were assessed by spectral-domain optical coherence tomography (SD-OCT) and best-corrected visual acuity (BCVA). Results: Closure rates were 66.7% for the observational group, and 100% for the surgical group (P=0.002). There were no vision-threatening ocular complications in both groups. For the observational group, the mean closure time was 2.5±1.6 months, and 80% of the hole closure occurred within 3 months; cystic edema on the edge of the hole at baseline was significantly more frequent in the non-closed subgroup than in the closed subgroup (P=0.03). There were no significant differences in the foveal microstructure and in the final visual outcome between the spontaneously closed cases and the surgically closed cases. Conclusions: TMH had a moderately high incidence of spontaneous closure, but an immediate vitrectomy achieved an even higher closure rate. Vitrectomy was effective and safe to treat TMH, while a 3-month observation for spontaneous closure may be an alternative modality for TMH management. Cystic edema on the edge of the hole may be an unfavorable factor for the spontaneous closure of TMH.

Purpose To investigate the frequency of primary angle closure (PAC) and primary angle closure glaucoma (PACG) in patients with retinal vein occlusion (RVO) based on a hospital population. Methods A total of 375 consecutive cases newly diagnosed with RVO by fluorescein fundus angiography at a single eye centre in Peking were reviewed. Gonioscopy was performed in all patients. Glaucoma was diagnosed according to the criteria of the International Society of Geographical and Epidemiological Ophthalmology. Retinal vein occlusion was classified as central retinal vein occlusion (CRVO), hemicentral retinal vein occlusion (HRVO), or branch retinal vein occlusion (BRVO), and as arteriovenous crossing RVO (AV-RVO), optic cup RVO (OC-RVO), optic nerve RVO without optical nerve head swelling (NONHS-RVO), or RVO with optical nerve head swelling (ONHS-RVO) based on the site of venous occlusion. Percentage of PAC or PACG for each type of RVO were calculated. Results PACG had a frequency of 4.1% [95% confidence interval (CI) 2.2-6.9%] in 317 RVO patients [5.3% (95% CI 2.0-11.2%) in CRVO, 8.8% (95% CI 1.9-23.7%) in HRVO, and 1.9% (95% CI 0.4-5.4%) in BRVO]. Primary angle closure (PAC) had a frequency of 2.9% (95% CI 1.4-5.5%) in RVO. PAC/PACG had a frequency of 11.5% (95% CI 6.3-18.9%) in CRVO, 8.8% (95% CI 1.9-23.7%) in HRVO and 3.1% (95% CI 1.0-7.1%) in BRVO. PAC/PACG was significantly more prevalent in NONHS-RVO [18.9% (95% CI 9.4-32.0%)] than in ONHS-RVO [6.5% (95% CI 2.1-14.5%)], AV-RVO [3.1% (95% CI 0.9-7.8%)], and OC-RVO [2.3% (95% CI 0.1-12.3%)]. Conclusion The overall frequency of PAC/PACG was much higher in patients with RVO (especially CRVO) than that in the general population. Eyes with PAC/PACG may undergo mechanical changes in the lamina cribrosa of the optic disc, resulting in RVO. Angle-closure conditions should be borne in mind when investigating Chinese patients with RVO.

A novel class of styryl sulfones were designed and synthesized as CAPE derivatives by our work team, which showed a multi-target neuroprotective effect, including antioxidative and anti-neuroinflammatory properties. However, the underlying mechanisms remain unclear. In the present study, the anti-Parkinson’s disease (PD) activity of 10 novel styryl sulfone compounds was screened by the cell viability test and the NO inhibition test in vitro. It was found that 4d exhibited the highest activity against PD among them. In a MPTP-induced mouse model of PD, the biological activity of 4d was validated through suppressing dopamine neurotoxicity, microglial activation, and astrocytes activation. With compound 4d, we conducted the mechanistic studies about anti-inflammatory responses through inhibition of p38 phosphorylation to protect dopaminergic neurons, and antioxidant effects through promoting nuclear factor erythroid 2-related factor 2 (Nrf2). The results revealed that 4d could significantly inhibit 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine/1-methyl-4-phenylpyridinium (MPTP/MPP+)-induced p38 mitogen-activated protein kinase (MAPK) activation in both in vitro and in vivo PD models, thus inhibiting the NF-κB-mediated neuroinflammation-related apoptosis pathway. Simultaneously, it could promote Nrf2 nuclear transfer, and upregulate the expression of antioxidant phase II detoxification enzymes HO-1 and GCLC, and then reduce oxidative damage.

Purpose To report the results and complications of scleral buckling for the treatment of rhegmatogenous retinal detachment (RRD) using 25-gauge chandelier endoillumination. Methods A total of 61 patients (61 eyes) with RRD were treated with scleral buckling. For the sclera buckling procedure, a 25-gauge chandelier was inserted through the pars plana for intra-ocular illumination, and retinal tears were identified and treated with episcleral cryotherapy under surgical microscope. On postoperative days 1, 3 and 7, the intra-ocular pressure was measured by a non-contact tonometer. On postoperative months 1 and 3, ultrasound biomicroscopy was used to examine the pars plana incision. Results In the surgical procedure, there was no lenticular or retinal damage due to the chandelier insertion. There was no conjunctival bleb formation at pars plana incision and no incidence of endophthalmitis after surgery. The mean intra-ocular pressure was 15.74 ± 2.98, 15.83 ± 2.76 and 16.14 ± 2.52 mmHg on postoperative days 1, 3 and 7, respectively. The one-time retinal reattachment rate was 93.4%. No visible vitreous incarceration was found in the incision of the pars plana. Conclusion There was no complication found due to the chandelier insertion in early postoperative period. Chandelier endoillumination is a feasible method for retinal visualization under surgical microscope during scleral buckling.

Younger patients who underwent vitrectomy for proliferative diabetic retinopathy (PDR) display more aggressive nature distinguished from the older patients. Preoperative anti-VEGF therapy has been widely used as an adjunct for PDR surgery. However, the effect of anti-VEGF administration in young diabetics has rarely been evaluated in previous studies. The purpose of this study was to evaluate the effects of ranibizumab pretreatment on vitrectomy surgery in young patients with PDR.This was a prospective nonrandomized comparative study. Young patients (40 years old) undergoing diabetic vitrectomy with or without ranibizumab pretreatment (25 eyes in each group) were analyzed in this study. The use of the drug was determined by the patients' own preference. The two surgical groups were matched according to a complexity score. Intravitreal injection of ranibizumab (IVR) was performed 3-5 days prior to the vitrectomy surgery in the IVR group. Intraoperative records including total surgical time, intraoperative bleeding, the use of endodiathermy, the frequency of relaxing retinotomies, the incidence of iatrogenic retinal breaks, and the use of perfluorocarbon liquid (PFCL) and silicone oil tamponade, and postoperative indices regarding recurrent vitreous hemorrhage (VH), neovascular glaucoma (NVG), recurrent retinal detachment, and visual outcome were evaluated between the two groups. All patients were followed up for one year after surgery.In young PDR patients, the severity of intraoperative bleeding was significantly lower in the IVR group than in the control group (P=0.04). The total surgical time was shorter in the IVR group than in the control group. However, the rate of relaxing retinotomy, the incidence of iatrogenic retinal breaks and the use of PFCL and silicone oil tamponade were not affected by IVR pretreatment but affected by the complexity score of the case. Early postvitrectomy hemorrhage occurred less frequently in the IVR group than in the control group (P0.001), Early visual recovery was better in the IVR group than in the control group (P=0.03). However, there were no significant differences in the development of late recurrent VH, NVG, recurrent retinal detachment, and final visual outcome.IVR pretreatment is a safe and effective adjunct to vitrectomy in reducing intraoperative and early postvitrectomy bleeding and should be suggested in young PDR patients. However, IVR does not reduce the incidence of intraoperative and late postoperative complications in these patients. The risk of iatrogenic retinal breaks and silicone oil use are closely correlated with the complexity score of the surgical cases.

Purpose To characterize the classification, incidence, diagnosis and prognosis of traumatic choroidal injuries. Methods Subjects were selected from the database of the Eye Injury Vitrectomy Study (EIVS) and were examined for occurrences of different categories of choroidal injuries. Standard photographs were collected. Anatomical and visual outcomes were assessed in patients with greater than 1 year of follow-up. Eyes that had no light perception (NLP) and/or phthisis bulbi were defined as having had unfavourable outcomes. The percentage of eyes with an unfavourable outcome was analysed for different types of choroidal injuries. Results Nine categories of choroidal injuries with distinctive features were identified in the EIVS database. The incidence and the percentage of eyes with an unfavourable outcome in each injury category were as follows: suprachoroidal effusion, 21.2% (7.2%); suprachoroidal haemorrhage, 12.8% (11.2%); massive suprachoroidal haemorrhage, 4.0% (64.9%); choroidal avulsion, 4.2% (92.2%); traumatic chorioretinal rupture, 1.8% (13.3%); choroidal rupture, 4.8% (6.8%); choroidal loss, 1.6% (79.3%); choroidal hole, 1.1% (5.3%); and choroidal damage at the wound site, 39.2% (17.7%). Conclusions Ocular trauma can cause a variety of choroidal injuries that have distinctive features, some of which are associated with a high frequency of unfavourable prognoses.

"AIM: To describe the complex, overlapping phenotype of four Chinese patients with inherited retinal dystrophies (IRDs) who harbored two pathogenic genes simultaneously. METHODS: This retrospective study included 4 patients affected with IRDs. Medical and ophthalmic histories were obtained, and clinical examinations were performed. A specific Hereditary Eye Disease Enrichment Panel (HEDEP) based on exome capture technology was used for genetic screening. RESULTS: Four patients were identified to harbor disease-causing variants in two different genes. Patient retinitis pigmentosa (RP) 01-II:1 exhibited both classical ABCA4 -induced Stargardt disease (STGD) 1 and USH2A-associated RP, patient RP02-III:2 exhibited both classical ABCA4-induced STGD1 and CDH23-associated RP, patient RP03-II:1 exhibited both USH2A-induced autosomal recessive retinitis pigmentosa (arRP) syndrome and SNRNP200-induced autosomal dominant retinitis pigmentosa (adRP), and patient RP04- II:2 exhibited USH2A-induced arRP syndrome and EYS-induced arRP at the same time. CONCLUSION: Our study demonstrates that genotype–phenotype correlations and comprehensive genetic screening is crucial for diagnosing IRDs and helping family planning for patients suffering from the disease."

Retinitis pigmentosa (RP) is a clinically and genetically heterogeneous disorder characterized by night blindness, visual field constriction, and severely reduced visual acuity. Despite a number of genes being implicated in RP pathogenesis, the genetic etiology of the disease remains unknown in many patients. In this study, our aim was to identify the disease-causing mutation of a large Chinese family with autosomal dominant RP (adRP). Targeted exon capture sequencing was initially performed to screen mutations in known disease-causing genes, followed by exome sequencing. In doing so, a heterozygous mutation in ADIPOR1 (c.929A > G) that results in an amino acid substitution (p.Y310C) was identified to co-segregate with the disease phenotype in this family. Adipor1 is wildly expressed throughout the body, but appears to be enriched in the photoreceptor inner and outer segments. The p.Y310C mutation, predicted to affect the structure and function of the protein, was confirmed to affect protein folding and its subcellular localization in vitro. In addition, knockdown of adipor1 expression in a zebrafish model with morpholino (MO) preferentially reduced the number of rod photoreceptors, with no effect on the number of cones, a phenotype that is characteristic of RP. Furthermore, the knockdown phenotype was partially rescued by injecting wild-type, but not mutant, human ADIPOR1 mRNA. We conclude that ADIPOR1 is a novel adRP-causing gene and plays an important role in rod development and maintenance.

Purpose To compare the outcome after choroidal neovascular (CNV) membrane excision and retinal pigment epithelium (RPE) transplantation and make further evaluation of two types of RPE grafts on the visual function in patients with neovascular age-related macular degeneration (AMD), complicated by massive subretinal haemorrhage. Methods We retrospectively reviewed 80 patients who underwent CNV membrane excision with or without RPE transplantation. Two types of RPE grafts were adopted, RPE–Bruch's membrane complex graft (subgroup 1) and RPE monolayer sheet graft (subgroup 2). Outcome measures included pre- and postoperative visual acuity score (VAS), clinical findings, complications and recurrence rates. Results The mean VAS [Early Treatment of Diabetic Retinopathy Study (ETDRS)] in the CNV membrane excision group was 11.06 ± 18.28 at baseline and 14.41 ± 16.86 at follow-up (p = 0.12) in a mean follow-up period of 24.35 ± 9.31 months. While in subgroup 1, VAS increased from 22.62 ± 23.72 to 35.50 ± 24.46 (p = 0.02) in a mean period of 20.63 ± 6.25 months. The percentage of visual acuity (VA) improvement (at least two or more lines changed) in subgroup 1 is 62.5%, which is significantly higher than that in CNV excision group (23.5%), p = 0.02. In subgroup 2, VAS increased from 16.61 ± 27.98 to 29.16 ± 23.80 (p = 0.02) in a mean period of 21.72 ± 11.09 months. The percentage of VA improvement in subgroup 2 is 58.0%, which is also significantly higher than that in CNV excision group (23.5%), p = 0.02. Postoperative VA elevation was comparable between the two subgroups (p = 0.05). Complications including retinal detachment, proliferative vitreal retinopathy and CNV recurrence occurred in both techniques. Central fixation stability was observed in eight eyes in subgroup 1 and five eyes in subgroup 2. Conclusions The autologous RPE transplantation can increase the vision of patients with haemorrhagic AMD. Two types of autologous RPE grafts were both effective and comparable in restoring visual function and central fixation stability.

Axonal tracing is useful for detecting optic nerve injury and regeneration, but many commonly used methods cannot be used to observe axoplasmic flow and synaptic transmission in vivo. Manganese (Mn(2+))-enhanced magnetic resonance imaging (MEMRI) can be used for in vivo longitudinal tracing of the visual pathway. Here, we explored the dose response and time course of an intravitreal injection of MnCl2 for tracing the visual pathway in rabbits in vivo using MEMRI. We found that 2 mM MnCl2 enhanced images of the optic nerve but not the lateral geniculate body or superior colliculus, whereas at all other doses tested (5-40 mM), images of the visual pathway from the retina to the contralateral superior colliculus were significantly enhanced. The images were brightest at 24 hours, and then decreased in brightness until the end of the experiment (7 days). No signal enhancement was observed in the visual cortex at any concentration of MnCl2. These results suggest that MEMRI is a viable method for temporospatial tracing of the visual pathway in vivo. Signal enhancement in MEMRI depends on the dose of MnCl2, and the strongest signals appear 24 hours after intravitreal injection.

Directed differentiation of human induced pluripotent stem cells (iPSCs) into retinal pigmented epithelium (RPE) holds great promise in cell replacement therapy for patients suffering from degenerative eye diseases, including age-related macular degeneration (AMD). In this study, we generated iPSCs from human dermal fibroblasts (HDFs) by electroporation with episomal plasmid vectors encoding OCT4, SOX2, KLF4, L-MYC together with p53 suppression. Intriguingly, cell reprogramming resulted in a metastable transcriptional activation and selective demethylation of neural and retinal specification-associated genes, such as OTX2, RX1 and SIX3. In contrast, RPE progenitor genes were transcriptionally silent in HDFs and descendant iPSCs. Overexpression of OCT4 and SOX2 directly stimulated the expression of OTX2, RX1 and SIX3 in HDFs and iPSCs. Luciferase and chromatin immunoprecipitation (ChIP) assays further identified an OCT4- and two SOX2-binding sites located in the proximal promoter of OTX2. Histone acetylation and methylation on the local promoter also participated in the reactivation of OTX2. The transcriptional conversion of RX1 and SIX3 genes partially attributed to DNA demethylation. Subsequently, iPSCs were induced into the RPE cells displaying the characteristics of polygonal shapes and pigments, and expressing typical RPE cell markers. Taken together, our results establish readily efficient and safe protocols to produce iPSCs and iPSC-derived RPE cells, and underline that the reactivation of anterior neural transcription factor OTX2, eye field transcription factor RX1 and SIX3 in iPSCs is a feature of pluripotency acquisition and predetermines the potential of RPE differentiation.

Inherited retinal dystrophies (IRDs) are a group of clinically and genetically heterogeneous diseases involving more than 280 genes and no less than 20 different clinical phenotypes. In this study, our aims were to identify the disease-causing gene variants of 319 Chinese patients with IRD, and compare the pros and cons of targeted panel sequencing and whole exome sequencing (WES). Patients were assigned for analysis with a hereditary eye disease enrichment panel (HEDEP) or WES examination based on time of recruitment. This HEDEP was able to capture 441 hereditary eye disease genes, which included 291 genes related to IRD. As RPGR ORF15 was difficult to capture, all samples were subjected to Sanger sequencing for this region. Among the 163 disease-causing variants identified in this study, 73 had been previously reported, and the other 90 were novel. Genes most commonly implicated in different inheritances of IRDs in this cohort were presented. HEDEP and WES achieved diagnostic yield with 41.2% and 33.0%, respectively. In addition, nine patients were found to carry pathogenic mutations in the RPGR ORF15 region with Sanger sequencing. Our study demonstrates that HEDEP can be used as a first-tier test for patients with IRDs.