Generation of 3D lacrimal gland organoids from human pluripotent stem cells.

Lacrimal glands are the main exocrine glands of the eyes. Situated within the orbit, behind the upper eyelid and towards the temporal side of each eye, they secrete lacrimal fluid as a major component of the tear film. Here we identify cells with characteristics of lacrimal gland primordia that emerge in two-dimensional eye-like organoids cultured from human pluripotent stem cells1. When isolated by cell sorting and grown under defined conditions, the cells form a three-dimensional lacrimal-gland-like tissue organoid with ducts and acini, enabled by budding and branching. Clonal colony analyses indicate that the organoids originate from multipotent ocular surface epithelial stem cells. The organoids exhibit notable similarities to native lacrimal glands on the basis of their morphology, immunolabelling characteristics and gene expression patterns, and undergo functional maturation when transplanted adjacent to the eyes of recipient rats, developing lumina and producing tear-film proteins.

RhoJ integrates attractive and repulsive cues in directional migration of endothelial cells.

During angiogenesis, VEGF acts as an attractive cue for endothelial cells (ECs), while Sema3E mediates repulsive cues. Here, we show that the small GTPase RhoJ integrates these opposing signals in directional EC migration. In the GTP-bound state, RhoJ interacts with the cytoplasmic domain of PlexinD1. Upon Sema3E stimulation, RhoJ released from PlexinD1 induces cell contraction. PlexinD1-bound RhoJ further facilitates Sema3E-induced PlexinD1-VEGFR2 association, VEGFR2 transphosphorylation at Y1214, and p38 MAPK activation, leading to reverse EC migration. Upon VEGF stimulation, RhoJ is required for the formation of the holoreceptor complex comprising VEGFR2, PlexinD1, and neuropilin-1, thereby preventing degradation of internalized VEGFR2, prolonging downstream signal transductions via PLCγ, Erk, and Akt, and promoting forward EC migration. After conversion to the GDP-bound state, RhoJ shifts from PlexinD1 to VEGFR2, which then terminates the VEGFR2 signals. RhoJ deficiency in ECs efficiently suppressed aberrant angiogenesis in ischemic retina. These findings suggest that distinct Rho GTPases may act as context-dependent integrators of chemotactic cues in directional cell migration and may serve as candidate therapeutic targets to manipulate cell motility in disease or tissue regeneration.

Preoperative hemoglobin A1c is associated with postoperative bleeding after vitrectomy for vitreous hemorrhage in patients with diabetic retinopathy.

We previously reported that a high HbA1c level 3 months before vitrectomy for vitreous hemorrhage or a large preoperative decrease in the HbA1c level over 3 months tended to increase the risk of rebleeding in diabetic retinopathy patients evaluated between 2010 and 2014. Here, we aimed to confirm these results with an extended study period and an increased number of operated eyes. This study included 121 diabetic patients who were admitted to Osaka University Hospital between 2010 and 2019 and who underwent vitrectomy for vitreous hemorrhage. Binomial logistic regression analysis was performed with the presence of postoperative bleeding as the outcome. The present study showed that the duration of the operation was associated with rebleeding (odds ratio = 1.02, p = 0.0016). A high HbA1c level just before vitrectomy tended to be associated with the bleeding (odds ratio = 1.27, p = 0.05), while preoperative HbA1c changes were not associated with rebleeding. The results of this study suggest that a high preoperative HbA1c level just before vitrectomy, not a decrease in HbA1c levels, in addition to the duration of the operation may increase the risk of postoperative bleeding after vitrectomy in diabetic retinopathy patients.

Continuous Corneal Endothelial Damage by Chlorhexidine Alcohol Used for Disinfection in Nonophthalmic Surgery.

PURPOSE: To report a case of corneal endothelial damage caused by alcohol-containing chlorhexidine gluconate (CG-A) and its progression over time. METHODS: This was a case report. RESULTS: A 22-year-old man underwent neurosurgery under general anesthesia. CG-A (1%) was used for disinfection after the application of corneal protection tape. Postoperatively, the patient presented with hyperemia and swelling of the left conjunctiva and was referred to our department. Initial examination revealed left corneal epithelial erosion and corneal edema, which improved on postoperative day 14. The corneal endothelial cell density (ECD) was 3,345 cells/mm 2 on day 14, decreased rapidly to 2,090 cells/mm 2 on day 42, and slowly reduced to 1,122 cells/mm 2 on day 168. Thereafter, no decrease in ECD was observed. CONCLUSIONS: CG formulations can lead to a persistent decrease in ECD over several months, even after improvement of acute corneal edema.

What Happens 20 to 30 years After Radial Keratotomy? Case Series.

ABSTRACT: Radial keratotomy (RK) was commonly performed in the 1980s and 1990s. We aimed to clarify the current status of post-RK refractive correction and treatment. We retrospectively reviewed the charts of 70 patients with a history of RK. Of the 70 patients, 44 were identified for clinical outcomes. Refractive or therapeutic intervention (rigid gas-permeable contact lens fit, spectacle prescription, corneal surgery, and use of pilocarpine hydrochloride for photophobia) was possible in 59% of patients with postoperative visual deterioration after RK; in the remaining 41%, therapeutic intervention was not possible. Rigid gas-permeable contact lens fit for corneal irregular astigmatism was the most common refractive intervention and was effective in 36% of cases in the university hospital.

A Comprehensive Wavefront Assessment of Keratoconus Using an Integrated Scheimpflug Corneal Tomographer/Hartmann-Shack Wavefront Aberrometer.

OBJECTIVES: To characterize higher-order aberrations (HOAs) in different severities of keratoconus (KC) from the anterior and posterior corneal surfaces and whole eye using an integrated Scheimpflug corneal tomographer/Hartmann-Shack wavefront aberrometer. METHODS: This study included eyes with clinical KC, topographic KC (no clinical signs), fellow eyes with very asymmetric ectasia with normal topography and no clinical signs (VAE-NT), and control eyes. Corneal and ocular wavefront aberrations were obtained using an integrated Scheimpflug tomographer/Hartmann-Shack wavefront aberrometer. The diagnostic capability of distinguishing VAE-NT from the control was also tested. RESULTS: This study included 68 eyes with clinical KC, 44 with topographic KC, 26 with VAE-NT, and 45 controls. Clinical KC had significantly greater total HOAs and coma from the anterior and posterior corneal surfaces and whole eye than the other groups ( P <0.05). Although topographic KC had significantly greater values in all wavefront parameters than the control ( P <0.05), ocular and corneal HOAs did not differ between the VAE-NT and control groups. The coma from the anterior cornea in topographic KC was significantly greater than that in VAE-NT ( P <0.05); the coma from the posterior cornea and whole eye did not differ. Total HOAs from the anterior corneal surface exhibited the highest area under the receiver operating characteristic curve value of 0.774 (sensitivity, 73%; specificity, 78%). CONCLUSION: A comprehensive wavefront assessment can be used to quantitatively evaluate corneal and ocular HOAs across various severity of KC. Total HOAs from the anterior corneal surface exhibited the potential ability in distinguishing VAE-NT from the control eyes.

Clinical Trial of Autologous Cultivated Limbal Epithelial Cell Sheet Transplantation for Patients with Limbal Stem Cell Deficiency.

PURPOSE: To confirm the efficacy and safety of Good Manufacturing Practice (GMP)-compliant autologous cultivated limbal epithelial cell sheets in government-controlled clinical trials that adhered to Good Clinical Practice stipulations for patients with unilateral limbal stem cell deficiency (LSCD). DESIGN: A prospective, multicenter, open-label, uncontrolled, single-arm clinical trial. PARTICIPANTS: Ten consecutive eyes of 10 patients with unilateral LSCD were followed for 2 years after surgery. Preoperative LSCD stage was IIB in 4 eyes and III in 6 eyes. METHODS: A limbal tissue biopsy was obtained from the healthy eye, after which limbal stem cells were dissociated and cultivated on temperature-responsive culture surfaces. All cell sheets were fabricated in a GMP-grade facility under established standard operating procedures. Cell sheets were evaluated using defined shipment criteria before transplantation, and only those that met the criteria were used. The cell sheet was transplanted onto each of the patients' diseased eye after removing the conjunctival scar tissue that covered the corneal surface. The severity of LSCD was determined according to a staging method agreed on by global consensus, with eyes evaluated as being in stages IA-C representing successful corneal epithelial reconstruction. Diagnosis and staging of LSCD were determined by the trial's Eligibility Judgment Committee and Effect Assessment Committee using slit-lamp photographs including fluorescein staining. Both committees comprised 2 or 3 third-party cornea specialists, who were provided with information anonymously and randomly. MAIN OUTCOME MEASURE: Corneal epithelial reconstruction rate was the primary end point. RESULTS: Corneal epithelial reconstruction was successful in 6 of 10 eyes (60%) 1 year postoperatively and was significantly higher than the 15% clinically significant efficacy rate achieved by allogeneic limbal transplantation. The reconstruction rate was 70% of eyes 2 years postoperatively. Additionally, improvements in visual acuity were noted in 50% and 60% of eyes at 1 and 2 years, respectively. No clinically significant transplantation-related adverse events were observed. CONCLUSIONS: The efficacy and safety of cultivated limbal epithelial cell sheet transplantation were thus confirmed, and the cell sheet, named "Nepic," is now approved as a cellular and tissue-based product in Japan. FINANCIAL DISCLOSURE(S): Proprietary or commercial disclosure may be found after the references. FINANCIAL DISCLOSURE(S): Proprietary or commercial disclosure may be found after the references.

Simultaneous intravitreal aflibercept and gas injections for submacular hemorrhage secondary to polypoidal choroidal vasculopathy.

PURPOSE: To investigate the outcomes of intravitreal aflibercept and gas injections for submacular hemorrhage (SMH) associated with polypoidal choroidal vasculopathy (PCV). METHODS: We retrospectively reviewed the medical records of 22 eyes with SMH secondary to PCV that underwent intravitreal aflibercept and 100% perfluoropropane (0.3-0.5 mL) followed by 3-day prone positioning from August 2013 through November 2020. The primary outcome measure was best-corrected visual acuity (BCVA) at 12 months. RESULTS: The average SMH size was 13.0 ± 9.7 (range, 2.0-37.8) disc diameter. The complete, partial, and no displacement of the SMH was observed in 8 (36%) eyes, 9 (41%) eyes, and 5 (23%) eyes, respectively. The BCVA (logarithm of the minimum angle of resolution) continuously improved significantly from 0.81 ± 0.41 (Snellen equivalent, 20/125) at baseline to 0.48 ± 0.44 (20/60), 0.33 ± 0.39 (20/43), and 0.28 ± 0.45 (20/38), at 3, 6, and 12 months, respectively (P = 0.01 for 3 months; P < 0.001 for 6 and 12 months). The BCVA improved by 3 or more lines in 14 eyes (64%). Two eyes (9%) developed visually significant vitreous hemorrhage, and 1 (5%) eye developed rhegmatogenous retinal detachment; all were successfully treated with vitrectomy. The better BCVA at 12 months tended to be associated with lower height of the SMH at baseline (R2 = 0.171, P = 0.056) and a greater displacement of SMH (R2 = 0.244, P = 0.069). Worse BCVA at 12 months was associated with anticoagulant medication (P < 0.001). CONCLUSIONS: Intravitreal aflibercept and gas injections are effective and relatively safe for SMH associated with PCV, resulting in significant visual improvement.

Assessment of the corneal biomechanical features of granular corneal dystrophy type 2 using dynamic ultra-high-speed Scheimpflug imaging.

PURPOSE: To evaluate the corneal biomechanical features of eyes with granular corneal dystrophy type 2 (GCD2) by analyzing corneal biomechanical indices obtained using a Corvis ST (CST) dynamic ultra-high-speed Scheimpflug imaging device. METHODS: In this retrospective case-control study, 35 CST parameters were compared in normal eyes (control) and eyes of patients with GCD2 treated at Osaka University Hospital, Osaka, Japan. The parameters included the Corvis Biomechanical Index (CBI), which is important in differentiating eyes with keratoconus from normal eyes. We measured the deposition rates of lesions in the central 7-mm region of the eye and assessed the correlation between the deposition rate and the CBI. RESULTS: Twenty-one eyes with GCD2 and 23 control eyes were analyzed. Eyes with GCD2 showed significantly less corneal stiffness in 15 CST parameters than did control eyes. In particular, the CBI was remarkably higher in eyes with GCD2 than in control eyes (P = 0.000006). Additionally, the deposition rate and the CBI were positively correlated. CONCLUSIONS: GCD2 eyes had softer corneas than did control eyes in most biomechanical CST parameters, and one of the parameters (the CBI) was linked to the rate of deposited lesions. Since IOP may be underestimated in GCD2 eyes, management should be especially careful in GCD2 cases complicated by glaucoma.

Long-term course with iris changes after trabeculectomy for uveitic glaucoma associated with iris mammillation: a case report.

BACKGROUND: Iris mammillation is a rare disease characterized by the distribution of multiple nodules on the iris surface. The course of uveitic glaucoma with iris mammillation has never been reported. CASE PRESENTATION: A 56-year-old woman, who presented with unilateral decreased vision, visited our hospital for treatment of uveitic glaucoma in the right eye. Multiple nodules were scattered over the iris surface in that eye. This case was diagnosed as iris mammillation on clinical findings. After excluding malignant tumors such as melanoma, trabeculectomy was performed. The resected iris had no pathologically malignant findings. The iris nodules evolved to a sand-like appearance, and the intraocular pressure remained stable without recurrent inflammation 7 years after trabeculectomy. CONCLUSIONS: In a case of unilateral uveitic glaucoma with iris mammillation, filtration surgery was performed after excluding the presence of a malignancy, and the long-term postoperative course has been stable.

Comparison of Ocular Wavefront Aberration Measurements Obtained Using Two Hartmann-Shack Wavefront Aberrometers.

OBJECTIVES: To assess agreement between measurements of ocular wavefront aberrations obtained using the Pentacam AXL Wave (Oculus Optikgeräte GmbH) (Aberrometer A) and KR-1W (Topcon Corp) (Aberrometer B), both of which are based on the Hartmann-Shack principle. METHODS: In this prospective case-control study, ocular wavefront aberrations measurements were obtained using both aberrometers in patients with keratoconus (KC) and control participants. Ocular wavefront aberrations were measured through the natural pupil without dilation using both devices in a dark room. For both aberrometers, accommodation was inhibited by automatically adding fogging. The individual Zernike coefficients from the second to fourth order were compared between the two aberrometers for a 4-mm pupil diameter. RESULTS: Twenty-six KC and 29 control eyes were assessed. Statistically significant correlations ( P <0.05) were observed for all Zernike coefficients, except for Z 4-2 in the control group. Bland-Altman analysis indicated good agreement between aberrometers and no statistically significant differences in the control group. However, in the KC group, patterns of proportional error were observed in vertical coma Z 3-1 (r=0.338, P =0.008), trefoil Z 4-4 (r=0.701, P =0.003), secondary astigmatism Z 4-2 (r=0.348, P =0.025), and spherical aberrations Z 40 (r=0.407, P =0.012). CONCLUSIONS: The Zernike coefficient values measured by the two aberrometers were well correlated in the control and KC groups. However, in eyes with KC, Aberrometer B tended to present greater values in several Zernike coefficients than Aberrometer A, suggesting that wavefront measurements obtained using the two aberrometers are not interchangeable in patients with KC.

Long-Term Corneal Refractive Power Changes Two Decades After Radial Keratotomy With Microperforations.

ABSTRACT: We retrospectively examined corneal refractive power in three patients who had been followed up for more than 20 years after radial keratotomy (RK) with microperforations (MPs). All patients underwent RK in both eyes and were referred to our clinic because of postoperative decreased vision. MP was observed in five of the six eyes at the initial visit. The corneal refractive power of the anterior and posterior surfaces of the 6-mm-diameter cornea was examined using Fourier analysis based on corneal shape analysis using anterior segment optical coherence tomography. The spherical components decreased in all three cases. The asymmetry and higher-order irregularity components and fluctuations in corneal refractive power were markedly greater in the two cases with MP in both eyes. Fluctuations in corneal refractive power were observed at more than 20 years after RK with MP. Therefore, careful observation is necessary, even after a long-term postoperative follow-up period.

Optical Quality and Visual Performance With Different Toric Contact Lens Designs.

OBJECTIVES: To evaluate the optical quality of different toric contact lens (CL) designs and compare their on-eye visual correction performance. METHODS: Twenty soft CL wearers aged 20 to 39 years were enrolled. Two daily disposable silicone-hydrogel toric CLs were tested: the "Eyelid Stabilized Design" (ESD-CL) and prism-ballast design (PB-CL); a spherical daily disposable silicone hydrogel CL (spherical CL) was used as a control. On-eye performance was compared for corrected distance visual acuity (CDVA), astigmatism, and ocular higher-order aberrations (HOAs); astigmatism and ocular HOAs were measured with a wavefront sensor. The subjective quality of vision, rated for "blurred vision" and "double vision," lens rotation, and fitting were also compared. RESULTS: The ESD-CLs, PB-CL, and no-CL provided better CDVA than spherical CL ( P <0.05). Compared with spherical CL and no CL, PB-CL and ESD-CLs caused significantly lesser astigmatism ( P <0.05). Coma was significantly lesser with ESD-CLs than that with PB-CL ( P <0.05); total HOAs did not differ among the four conditions. The subjective ratings for blurred and double vision were significantly lower with ESD-CLs than those with spherical CL ( P <0.05). CONCLUSIONS: Toric CLs provide a better CDVA than spherical CLs. However, differences in coma and subjective symptoms may occur because of the design of toric CLs.

Co-ordinated ocular development from human iPS cells and recovery of corneal function.

The eye is a complex organ with highly specialized constituent tissues derived from different primordial cell lineages. The retina, for example, develops from neuroectoderm via the optic vesicle, the corneal epithelium is descended from surface ectoderm, while the iris and collagen-rich stroma of the cornea have a neural crest origin. Recent work with pluripotent stem cells in culture has revealed a previously under-appreciated level of intrinsic cellular self-organization, with a focus on the retina and retinal cells. Moreover, we and others have demonstrated the in vitro induction of a corneal epithelial cell phenotype from pluripotent stem cells. These studies, however, have a single, tissue-specific focus and fail to reflect the complexity of whole eye development. Here we demonstrate the generation from human induced pluripotent stem cells of a self-formed ectodermal autonomous multi-zone (SEAM) of ocular cells. In some respects the concentric SEAM mimics whole-eye development because cell location within different zones is indicative of lineage, spanning the ocular surface ectoderm, lens, neuro-retina, and retinal pigment epithelium. It thus represents a promising resource for new and ongoing studies of ocular morphogenesis. The approach also has translational potential and to illustrate this we show that cells isolated from the ocular surface ectodermal zone of the SEAM can be sorted and expanded ex vivo to form a corneal epithelium that recovers function in an experimentally induced animal model of corneal blindness.

Vascular tortuosity analysis in eyes with epiretinal membrane imaged by optical coherence tomography angiography.

BACKGROUND: This study aimed to evaluate macular vessel tortuosity using optical coherence tomography angiography (OCTA) and its association with visual outcomes in eyes undergoing surgery for epiretinal membrane (ERM). METHODS: The study included 22 consecutive patients who underwent vitrectomy for ERM between May 2019 and July 2020 and OCTA at Osaka University Hospital. All patients underwent ophthalmologic examinations, including swept-source OCTA. Standard vitrectomy was performed, and the patients were followed up for 6 months postoperatively. Distortion of retinal vessels was calculated using two parameters: the actual vessel length in the vessel section (VL) and the direct vessel branching point distance (BD) in the three quadrants (nasal, temporal, and superior-inferior) of the macula. We analyzed the correlation between these parameters and visual outcomes. RESULTS: Significantly longer VL was found at 1, 3, and 6 months postoperatively (p = 0.006, 0.008, and 0.022, respectively) in the temporal quadrant compared to baseline temporal VL. Significantly shorter VL was found in nasal quadrants at 1 and 3 months (p = 0.046 and p = 0.018) in the comparison of nasal baseline VL. VL/BDs were correlated with the same postoperative best-corrected visual acuity (BCVA) at 1, 3, and 6 months (p = 0.035, 0.035, and 0.042, respectively) in the superior-inferior quadrant. A significant association of changes in VL and BCVA was found at 3 and 6 months postoperatively in the nasal quadrant (p = 0.018 and 0.0455, respectively). CONCLUSIONS: Changes in vascular distortion after ERM surgery can be measured using OCTA. The change in vessels around the macula became more linear; this was associated with visual outcomes after surgery.

Characteristics of Higher-Order Aberrations in Different Stages of Keratoconus.

OBJECTIVES: To characterize higher-order aberrations (HOAs) in clinical and subclinical keratoconus (KC). METHODS: The study included 33, 36, and 26 patients with clinical, topographic (no clinical signs), and pretopographic (normal topography and no clinical signs) KC and 30 controls. Ocular and corneal HOAs for the 4-mm pupils were measured using a wavefront sensor and expanded up to the sixth order of Zernike polynomials. The magnitudes of trefoil, coma, tetrafoil, secondary astigmatism, and spherical aberration were calculated via Zernike vector analysis and used as HOA parameters along with total HOAs. Area under the receiver operating characteristic curve (AUROC) values for each wavefront parameter for pretopographic KC were compared. RESULTS: Control eyes and eyes with pretopographic KC had significantly lower ocular or corneal total HOAs and Zernike vector terms than those with clinical KC and topographic KC, except for ocular tetrafoil between topographic KC and pretopographic KC and spherical aberration among all groups. The AUROCs for corneal total HOAs and corneal coma for pretopographic KC and control eyes were 0.781 (100% sensitivity and 47% specificity) and 0.735 (73% sensitivity and 73% specificity), respectively. CONCLUSION: Corneal total HOAs and corneal coma exhibited a potential ability to discriminate pretopographic KC from normal control eyes.

Severe Ocular Complications Associated With Wearing of Contact Lens in Japan.

OBJECTIVES: To examine the severe ocular complications associated with contact lens wearing in Japan. METHOD: A questionnaire was sent to 964 ophthalmologist training facilities inquiring for cases of contact lens-associated complications from April 2016 to March 2018. The inclusion criteria were as follows: (1) corrected distance visual acuity ≤0.1 decimal after treatment for 3 months, (2) corneal perforation observed during follow-up, and (3) requiring surgery. A secondary analysis was conducted, inquiring for further information on the type of contact lens, clinical manifestations, and course of treatment. RESULTS: Forty-two patients with infectious keratitis met the inclusion criteria. Eight patients were users of rigid gas-permeable contact lens, and 34 were users of soft contact lens. Microbiological tests were positive in 73.0%. The organisms isolated in microbiological culture were bacteria in 11 patients (Pseudomonas aeruginosa in 9 patients), fungi in 2 patients, and Acanthamoeba in 14 patients. Ten patients were treated with local antibiotics, 11 with a combination of systemic antibiotics, and 21 with a combination of surgical approaches, including 13 with corneal transplantation. CONCLUSIONS: The major cause of serious contact lens-associated ocular complications was microbial keratitis, and P. aeruginosa and Acanthamoeba were the major pathogens in Japan.

Generation and validation of a PITX2-EGFP reporter line of human induced pluripotent stem cells enables isolation of periocular mesenchymal cells.

PITX2 (Paired-like homeodomain transcription factor 2) plays important roles in asymmetric development of the internal organs and symmetric development of eye tissues. During eye development, cranial neural crest cells migrate from the neural tube and form the periocular mesenchyme (POM). POM cells differentiate into several ocular cell types, such as corneal endothelial cells, keratocytes, and some ocular mesenchymal cells. In this study, we used transcription activator-like effector nuclease technology to establish a human induced pluripotent stem cell (hiPSC) line expressing a fluorescent reporter gene from the PITX2 promoter. Using homologous recombination, we heterozygously inserted a PITX2-IRES2-EGFP sequence downstream of the stop codon in exon 8 of PITX2 Cellular pluripotency was monitored with alkaline phosphatase and immunofluorescence staining of pluripotency markers, and the hiPSC line formed normal self-formed ectodermal autonomous multizones. Using a combination of previously reported methods, we induced PITX2 in the hiPSC line and observed simultaneous EGFP and PITX2 expression, as indicated by immunoblotting and immunofluorescence staining. PITX2 mRNA levels were increased in EGFP-positive cells, which were collected by cell sorting, and marker gene expression analysis of EGFP-positive cells induced in self-formed ectodermal autonomous multizones revealed that they were genuine POM cells. Moreover, after 2 days of culture, EGFP-positive cells expressed the PITX2 protein, which co-localized with forkhead box C1 (FOXC1) protein in the nucleus. We anticipate that the PITX2-EGFP hiPSC reporter cell line established and validated here can be utilized to isolate POM cells and to analyze PITX2 expression during POM cell induction.

Rubicon regulates A2E-induced autophagy impairment in the retinal pigment epithelium implicated in the pathology of age-related macular degeneration.

Waste product deposition and light stress in the retinal pigment epithelium (RPE) are crucial factors in the pathogenesis of various retinal degenerative diseases, including age-related macular degeneration (AMD), a leading cause of vision loss in elderly individuals worldwide. Given that autophagy in the RPE suppresses waste accumulation, determining the molecular mechanism by which autophagy is compromised in degeneration is necessary. Using polarized human RPE sheets, we found that bis-retinoid N-retinyl-N-retinylidene ethanolamine (A2E), a major toxic fluorophore of lipofuscin, causes significant impairment of autophagy and the simultaneous upregulation of Rubicon, a negative regulator of autophagy. Importantly, this impairment was reversed in Rubicon-specific siRNA-treated RPE sheets. In a retinal functional analysis using electroretinograms (ERGs), mice with the RPE-specific deletion of Rubicon showed no significant differences from control cre-expressing mice but presented partially but significantly enhanced amplitudes compared with Atg7 knockout mice. We also found that an inflammatory reaction in the retina in response to chronic blue light irradiation was alleviated in mice with the RPE-specific deletion of Rubicon. In summary, we propose that upregulating basal autophagy by targeting Rubicon is beneficial for protecting the RPE from functional damage with ageing and the inflammatory reaction caused by light-induced cellular stress.

Corneal Opacity Induced by Light in a Mouse Model of Gelatinous Drop-Like Corneal Dystrophy.

Gelatinous drop-like corneal dystrophy (GDLD) is a severe inherited corneal dystrophy characterized by subepithelial corneal amyloid deposition. We had previously succeeded in identifying the responsible gene, TACSTD2, and subsequently found that the epithelial barrier function is significantly decreased. As with GDLD patients, the knockout mice showed severe loss of tight junction, progressive opacity, and neovascularization in the cornea. We devised an easy method to confirm the loss of the corneal barrier function even before corneal opacity is observed. Furthermore, by using knockout mice, we were able to verify clinical findings, such as the wound healing delay and light-induced acceleration of the disease. This mouse model should prove to be a highly useful tool for investigating the pathology of GDLD and for developing new therapies.