HIF1α-dependent hypoxia response in myeloid cells requires IRE1α.

Cellular adaptation to low oxygen tension triggers primitive pathways that ensure proper cell function. Conditions of hypoxia and low glucose are characteristic of injured tissues and hence successive waves of inflammatory cells must be suited to function under low oxygen tension and metabolic stress. While Hypoxia-Inducible Factor (HIF)-1α has been shown to be essential for the inflammatory response of myeloid cells by regulating the metabolic switch to glycolysis, less is known about how HIF1α is triggered in inflammation. Here, we demonstrate that cells of the innate immune system require activity of the inositol-requiring enzyme 1α (IRE1α/XBP1) axis in order to initiate HIF1α-dependent production of cytokines such as IL1ß, IL6 and VEGF-A. Knockout of either HIF1α or IRE1α in myeloid cells ameliorates vascular phenotypes in a model of retinal pathological angiogenesis driven by sterile inflammation. Thus, pathways associated with ER stress, in partnership with HIF1α, may co-regulate immune adaptation to low oxygen.

Bex1 is essential for ciliogenesis and harbours biomolecular condensate-forming capacity.

BACKGROUND: Primary cilia are sensory organelles crucial for organ development. The pivotal structure of the primary cilia is a microtubule that is generated via tubulin polymerization reaction that occurs in the basal body. It remains to be elucidated how molecules with distinct physicochemical properties contribute to the formation of the primary cilia. RESULTS: Here we show that brain expressed X-linked 1 (Bex1) plays an essential role in tubulin polymerization and primary cilia formation. The Bex1 protein shows the physicochemical property of being an intrinsically disordered protein (IDP). Bex1 shows cell density-dependent accumulation as a condensate either in nucleoli at a low cell density or at the apical cell surface at a high cell density. The apical Bex1 localizes to the basal body. Bex1 knockout mice present ciliopathy phenotypes and exhibit ciliary defects in the retina and striatum. Bex1 recombinant protein shows binding capacity to guanosine triphosphate (GTP) and forms the condensate that facilitates tubulin polymerization in the reconstituted system. CONCLUSIONS: Our data reveals that Bex1 plays an essential role for the primary cilia formation through providing the reaction field for the tubulin polymerization.

The systemic antiangiogenic effect of intravitreal aflibercept injection in a mouse model of retinopathy of prematurity.

Retinopathy of prematurity (ROP) is a leading cause of childhood blindness and intravitreal anti-vascular endothelial growth factor (VEGF) injection is becoming a first-line choice for treatment of ROP. However, there is a major concern that intravitreally injected anti-VEGF agents could escape from the eye into the systemic circulation and impair systemic development. Moreover, escaped anti-VEGF agents could have an effect on the retina of the fellow eye. In this study, we investigated the hematogenous effect of a single intravitreal anti-VEGF injection in a mouse model of ROP. Here, we showed that single intravitreal aflibercept injection to one eye can affect body weight gain, the fellow eye, and renal vessels, although no apparent effect was observed in brain vessels. Furthermore, this hematogenous effect was dose-dependent. Our results provide very important insights into the clinical use of anti-VEGF agents for ROP treatment.

Prediction of postoperative visual acuity after vitrectomy for macular hole using deep learning-based artificial intelligence.

PURPOSE: To create a model for prediction of postoperative visual acuity (VA) after vitrectomy for macular hole (MH) treatment using preoperative optical coherence tomography (OCT) images, using deep learning (DL)-based artificial intelligence. METHODS: This was a retrospective single-center study. We evaluated 259 eyes that underwent vitrectomy for MHs. We divided the eyes into four groups, based on their 6-month postoperative Snellen VA values: (A) ≥ 20/20; (B) 20/25-20/32; (C) 20/32-20/63; and (D) ≤ 20/100. Training data were randomly selected, comprising 20 eyes in each group. Test data were also randomly selected, comprising 52 total eyes in the same proportions as those of each group in the total database. Preoperative OCT images with corresponding postoperative VA values were used to train the original DL network. The final prediction of postoperative VA was subjected to regression analysis based on inferences made with DL network output. We created a model for predicting postoperative VA from preoperative VA, MH size, and age using multivariate linear regression. Precision values were determined, and correlation coefficients between predicted and actual postoperative VA values were calculated in two models. RESULTS: The DL and multivariate models had precision values of 46% and 40%, respectively. The predicted postoperative VA values on the basis of DL and on preoperative VA and MH size were correlated with actual postoperative VA at 6 months postoperatively (P < .0001 and P < .0001, r = .62 and r = .55, respectively). CONCLUSION: Postoperative VA after MH treatment could be predicted via DL using preoperative OCT images with greater accuracy than multivariate linear regression using preoperative VA, MH size, and age.

Macrophages fine-tune pupil shape during development.

Tissue macrophages, which are ubiquitously present innate immune cells, play versatile roles in development and organogenesis. During development, macrophages prune transient or unnecessary synapses in neuronal development, and prune blood vessels in vascular development, facilitating appropriate tissue remodeling. In the present study, we identified that macrophages contributed to the development of pupillary morphology. Csf1op/op mutant mice, in which ocular macrophages are nearly absent, exhibited abnormal pupillary edges, with abnormal protrusions of excess iris tissue into the pupillary space. Macrophages located near the pupillary edge engulfed pigmented debris, which likely consisted of unnecessary iris protrusions that emerge during smoothening of the pupillary edge. Indeed, pupillary edge macrophages phenotypically possessed some features of M2 macrophages, consistent with robust tissue engulfment and remodeling activities. Interestingly, protruding irises in Csf1op/op mice were only detected in gaps between regressing blood vessels. Taken together, our findings uncovered a new role for ocular macrophages, demonstrating that this cell population is important for iris pruning during development.

THE CORRELATION BETWEEN AQUEOUS VASCULAR ENDOTHELIAL GROWTH FACTOR LEVEL AND CLINICAL ACTIVITY IN NEOVASCULAR AGE-RELATED MACULAR DEGENERATION.

PURPOSE: The aim of the current study was to investigate the correlation between the pretreatment aqueous level of vascular endothelial growth factor (VEGF) and clinical activity in neovascular age-related macular degeneration. METHODS: Patients with neovascular age-related macular degeneration treated by intravitreal ranibizumab injections and followed for 12 months were included in the current study. The treatment regimen consisted of three consecutive monthly intravitreal ranibizumab injections (loading treatment) followed by a pro re nata (PRN) treatment regimen. The aqueous VEGF levels were measured by enzyme-linked immunosorbent assay using aqueous humor samples obtained just before the first intravitreal ranibizumab injections. RESULTS: Sixty-four eyes of 64 patients were included in the current study. The mean number of intravitreal ranibizumab injections during 12 months was 4.6 ± 1.4, and 17 eyes had no recurrence after loading treatment. The mean aqueous VEGF level was significantly higher in eyes with recurrence after loading treatment than in eyes without recurrence (107.6 vs. 83.8 pg/mL, respectively; P = 0.04) and significantly higher in eyes with recurrence within 3 months after loading treatment than in other eyes (114.9 vs. 86.7 pg/mL, respectively; P < 0.01). CONCLUSION: Pretreatment aqueous VEGF level was significantly correlated with the likelihood of recurrence in neovascular age-related macular degeneration. The measurement of pretreatment aqueous VEGF level may be useful to determine the best treatment options for patients with neovascular age-related macular degeneration.

Comparison between wide-angle OCT angiography and ultra-wide field fluorescein angiography for detecting non-perfusion areas and retinal neovascularization in eyes with diabetic retinopathy.

PURPOSE: To compare the ability of wide-angle optical coherence tomography angiography (OCTA) with that of ultra-wide field fluorescein angiography (UWFFA) to detect non-perfusion areas (NPAs) or retinal neovascularization (NV) in eyes with diabetic retinopathy (DR). METHODS: Patients with DR underwent UWFFA using the Optos® panoramic 200Tx imaging system and wide-angle OCTA with 12 × 12 mm fields of five visual fixations using the PLEX Elite 9000®. We compared the abilities of UWFFA and OCTA to detect NPAs and NV. RESULTS: Fifty-eight eyes of 33 patients (mean age, 60.0 years old; female/male, 16/17) with DR were evaluated. NPAs were detected in 47 out of 58 eyes using UWFFA and in 48 eyes using OCTA. NVs were detected in 25 out of the 58 eyes using UWFFA and in 26 eyes using OCTA. The sensitivity for detection of NPA using OCTA was 0.98, and the specificity was 0.82. The sensitivity for detection of NV was 1.0, and the specificity was 0.97. CONCLUSION: The wide-angle OCTA seems to be clinically useful for the detection of NPAs or NV.

OPTICAL COHERENCE TOMOGRAPHY ANGIOGRAPHY REVEALS BLOOD FLOW IN CHOROIDAL NEOVASCULAR MEMBRANE IN REMISSION PHASE OF NEOVASCULAR AGE-RELATED MACULAR DEGENERATION.

PURPOSE: The aim of the study was to investigate blood flow in choroidal neovascular membrane in remission phase of neovascular age-related macular degeneration using optical coherence tomography (OCT) angiography. METHODS: OCT angiography was obtained in eyes with remission phase of neovascular age-related macular degeneration after treatments, defined as no exudative change (such as macular edema, subretinal fluid, and subretinal hemorrhage) observed in eyes without any treatment for neovascular age-related macular degeneration within the previous 6 months. Irregular blood flows shown in the segmentation of outer retina detected by OCT angiography were considered as blood flows in choroidal neovascular membrane. The vascular area and vessel density were obtained from OCT angiography images. RESULTS: Twenty eyes of 20 patients were included in this analysis. The blood flows in choroidal neovascular membrane were observed in all eyes (100%) using OCT angiography. The mean vascular area was 3.81 ± 3.41 mm and the mean vessel density of lesion was 28.9 ± 8.2%. The vessel density was significantly correlated with best-corrected visual acuity and duration of remission (best-corrected visual acuity: P = 0.008, r = -0.576; duration of remission: P = 0.017, r = -0.525, respectively). CONCLUSION: Optical coherence tomography angiography revealed that blood flows in choroidal neovascular membrane remained in eyes with clinically inactive neovascular age-related macular degeneration.

The effectiveness of vitrectomy for diffuse diabetic macular edema may depend on its preoperative optical coherence tomography pattern.

BACKGROUND: To investigate the effectiveness of vitrectomy for diffuse diabetic macular edema (DDME) and its dependence on optical coherence tomography (OCT) findings. METHODS: The records of 65 patients and 81 eyes who received vitrectomy for DDME and followed up for at least 6 months were retrospectively reviewed. All eyes were classified according to their morphological characteristics on OCT including sponge-like diffuse retinal thickening (SDRT: n = 13), cystoid macular edema (CME: n = 42), serous retinal detachment (SRD: n = 13), and the combination of all morphological characteristics (FULL: n = 13). The best-corrected visual acuity (BCVA) and spectral domain OCT were investigated preoperatively and at 1, 3, and 6 months postoperatively. RESULTS: At six months after surgery, BCVA in logMAR units was significantly improved in all groups except the SDRT group. The improvement was -0.04 ± 0.20 in the SDRT group (P = 0.504), -0.16 ± 0.24 in the CME group (P < 0.01), -0.32 ± 0.32 in the SRD group (P < 0.01), and -0.26 ± 0.19 in the FULL group (P < 0.01), and significantly better in eyes with subretinal fluid (SRF; the SRD and FULL groups) than in eyes without SRF (the SDRT and CME groups, P = 0.003). CONCLUSIONS: Vitrectomy can be a useful treatment option for DDME, particularly for eyes with SRF.

PHOTORECEPTOR OUTER SEGMENT LENGTH AND OUTER FOVEAL THICKNESS AS FACTORS ASSOCIATED WITH VISUAL OUTCOME AFTER VITRECTOMY FOR VITREOMACULAR TRACTION SYNDROME.

PURPOSE: To investigate the predictive factors for postoperative best-corrected visual acuity (BCVA) in patients with vitreomacular traction syndrome treated with vitrectomy. METHODS: The records of 21 patients with 21 eyes that underwent vitrectomy for vitreomacular traction syndrome and followed for at least 12 months were retrospectively reviewed. The BCVA and spectral domain optical coherence tomography findings were investigated preoperatively and at 1, 3, 6, and 12 months postoperatively. Axial length was measured preoperatively. The correlations between 12-month postoperative BCVA and preoperative parameters, including BCVA, age, axial length, central foveal thickness, outer foveal thickness, and photoreceptor outer segment length, were evaluated. RESULTS: Twelve-month postoperative BCVA was significantly negatively correlated with preoperative outer foveal thickness and photoreceptor outer segment length (outer foveal thickness: P = 0.029, r = -0.501; photoreceptor outer segment length: P = 0.022, r = -0.523, respectively) but not correlated with age, axial length, preoperative BCVA, and preoperative central foveal thickness (age: P = 0.346, r = 0.216; axial length: P = 0.333, r = 0.242; BCVA: P = 0.202, r = 0.290; central foveal thickness: P = 0.065, r = -0.410, respectively). CONCLUSION: Preoperative outer foveal thickness and photoreceptor outer segment length could be good predictive factors of postoperative BCVA in patients with vitreomacular traction syndrome.

Optical Coherence Tomography Angiography in a Patient with Optic Atrophy After Non-arteritic Anterior Ischaemic Optic Neuropathy.

A 75-year-old female noticed a lower visual field (VF) defect in the right eye. A diagnosis of non-arteritic anterior ischaemic optic neuropathy (NAION) was made. The lower VF defect in the right eye did not change after onset. Optical coherence tomography (OCT) angiograms on the disc and the macula showed decreased retinal perfusion in the upper retina of the right eye. Retinal nerve fibre layer loss and ganglion cell complex loss in the upper retina were also seen in the right eye. OCT angiography could non-invasively detect the decrease of the retinal perfusion due to NAION.

Anterior chamber paracentesis might prevent sustained intraocular pressure elevation after intravitreal injections of ranibizumab for age-related macular degeneration.

BACKGROUND/AIMS: To evaluate the efficacy of anterior chamber paracentesis for preventing sustained intraocular pressure (IOP) elevation after intravitreal ranibizumab (IVR) injections for age-related macular degeneration (AMD). METHODS: The medical records for all cases of exudative AMD treated with IVR injections and followed monthly for 12 months or longer were reviewed retrospectively. Anterior chamber paracentesis was performed just before IVR injections. A sustained IOP elevation was defined as 22 mm Hg or higher during 2 consecutive visits with an increase exceeding 6 mm Hg from baseline. RESULTS: One hundred and eleven eyes met the inclusion criteria, and none of these eyes had a sustained IOP elevation. CONCLUSIONS: Anterior chamber paracentesis before IVR injections may prevent sustained IOP elevations.

Null mutation of exocyst complex component 3-like does not affect vascular development in mice.

Exocyst is an octameric protein complex implicated in exocytosis. The exocyst complex is highly conserved among mammalian species, but the physiological function of each subunit in exocyst remains unclear. Previously, we identified exocyst complex component 3-like (Exoc3l) as a gene abundantly expressed in embryonic endothelial cells and implicated in the process of angiogenesis in human umbilical cord endothelial cells. Here, to reveal the physiological roles of Exoc3l during development, we generated Exoc3l knockout (KO) mice by genome editing with CRISPR/Cas9. Exoc3l KO mice were viable and showed no significant phenotype in embryonic angiogenesis or postnatal retinal angiogenesis. Exoc3l KO mice also showed no significant alteration in cholesterol homeostasis or insulin secretion, although several reports suggest an association of Exoc3l with these processes. Despite the implied roles, Exoc3l KO mice exhibited no apparent phenotype in vascular development, cholesterol homeostasis, or insulin secretion.

Efficacy of scleral imbrication on all quadrants in enucleated pig eyes.

PURPOSE: To evaluate the changes in axial length (AL) and corneal astigmatism induced by scleral imbrication on all quadrants in pig eyes. STUDY DESIGN: Experimental study METHODS: We produced scleral imbrications either on all quadrants or on 2 consecutive quadrants of 5 enucleated pig eyes. Scleral imbrications 8 mm wide were made at 8 mm from the limbus on each quadrant. We determined the AL using an electronic caliper and the corneal astigmatism using a keratometer before and after the 2 types of scleral imbrications and compared the changes in ocular AL and corneal astigmatism induced by the 2 surgical procedures. RESULTS: The AL reduction after the scleral imbrication on all quadrants (3.96 ± 0.56 mm) was larger than that on 2 quadrants (2.39 ± 0.41 mm) (P = .001). The change in corneal astigmatism induced by imbrication on all quadrants (2.98 ± 1.96 D) was less than that on 2 quadrants (5.95 ± 2.04 D) (P < .029). CONCLUSIONS: Scleral imbrication on all quadrants induced a shorter AL and less corneal astigmatism than did a standard scleral imbrication on 2 quadrants. Therefore, the former could be a more effective operation for retinal disorders associated with high myopia, including macular hole retinal detachment and myopic foveoschisis.

Assessment of mouse VEGF neutralization by ranibizumab and aflibercept.

PURPOSE: To assess the interaction between ranibizumab, aflibercept, and mouse vascular endothelial growth factor (VEGF), both in vivo and in vitro. METHODS: In vivo, the effect of intravitreal injection of ranibizumab and aflibercept on oxygen induced retinopathy (OIR) and the effect of multiple intraperitoneal injections of ranibizumab and aflibercept on neonatal mice were assessed. In vitro, the interaction of mouse VEGF-A with aflibercept or ranibizumab as the primary antibody was analyzed by Western blot. RESULTS: In both experiments using intravitreal injections in OIR mice and multiple intraperitoneal injections in neonatal mice, anti-VEGF effects were observed with aflibercept, but not with ranibizumab. Western blot analysis showed immunoreactive bands for mouse VEGF-A in the aflibercept-probed blot, but not in the ranibizumab-probed blot. CONCLUSIONS: Aflibercept but not ranibizumab interacts with mouse VEGF, both in vivo and in vitro. When conducting experiments using anti-VEGF drugs in mice, aflibercept is suitable, but ranibizumab is not.

Essential Roles of Exocyst Complex Component 3-like 2 on Cardiovascular Development in Mice.

Angiogenesis is a process to generate new blood vessels from pre-existing vessels and to maintain vessels, and plays critical roles in normal development and disease. However, the molecular mechanisms underlying angiogenesis are not fully understood. This study examined the roles of exocyst complex component (Exoc) 3-like 2 (Exoc3l2) during development in mice. We found that Exoc3l1, Exoc3l2, Exoc3l3 and Exoc3l4 are expressed abundantly in endothelial cells at embryonic day 8.5. The generation of Exoc3l2 knock-out (KO) mice showed that disruption of Exoc3l2 resulted in lethal in utero. Substantial numbers of Exoc3l2 KO embryos exhibited hemorrhaging. Deletion of Exoc3l2 using Tie2-Cre transgenic mice demonstrated that Exoc3l2 in hematopoietic and endothelial lineages was responsible for the phenotype. Taken together, these findings reveal that Exoc3l2 is essential for cardiovascular and brain development in mice.

Effect of internal limiting membrane peeling on postoperative visual acuity in macula-off rhegmatogenous retinal detachment.

PURPOSE: To investigate the effects of internal limiting membrane (ILM) peeling on visual acuity (VA) after rhegmatogenous retinal detachment (RRD) surgery. METHODS: This retrospective analysis examined the medical records of patients with RRD who underwent vitrectomy at 26 institutions. To detect prognostic factors of VA at 6 months postoperatively (post-VA), multivariate linear regression was performed with post-VA as the objective variable; ILM peeling, sex, age, preoperative VA (pre-VA), intraocular pressure, axial length, duration of RRD, and cataract surgery served as explanatory variables. Recurrence of RRD and epiretinal membrane formation within 6 months postoperatively were compared between groups of patients with and without ILM peeling, among patients with macula-on and macula-off RRD. RESULTS: The inclusion criteria were met by 523 eyes with a macula-on RRD and 364 eyes with a macula-off RRD. ILM peeling was performed in 85 eyes with a macula-on RRD and 57 eyes with a macula-off RRD. In eyes with a macula-on RRD, ILM peeling did not affect post-VA (p = 0.72). Vitrectomy without cataract surgery and poor pre-VA were significantly associated with poor post-VA (p = 0.01 and p < 0.001, respectively). In eyes with a macula-off RRD, ILM peeling, long duration of RRD, and poor pre-VA were significantly associated with poor post-VA (p = 0.037, p = 0.007, and p < 0.001, respectively). Recurrence of RRD and epiretinal membrane formation were similar between groups of patients with and without ILM peeling, among patients with macula-on and macula-off RRD. Retina sensitivity was not evaluated by microperimetry. CONCLUSION: ILM peeling did not affect post-VA in eyes with a macula-on RRD, whereas post-VA was worse in eyes with ILM peeling than in eyes without peeling, among eyes with a macula-off RRD.

Effect of intravitreal bevacizumab for retinopathy of prematurity on weight gain.

PURPOSE: To evaluate the short-term effect on body weight (BW) gain after intravitreal bevacizumab (IVB) for retinopathy of prematurity (ROP). METHODS: This was a retrospective 1:1 matched case-control study. Infants with ROP treated by IVB or photocoagulation (PC) at Shiga University of Medical Science Hospital between April 2010 and December 2019 were included in the study. To match BWs at treatment between the IVB and PC groups, 1:1 matching for BWs at treatment within 100 g was performed. The BW gains for the 7 days before treatment (pre-treatment week), the 7 days after treatment (first post-treatment week), and the period from 7 to 14 days after treatment (second post-treatment week) were compared between the IVB and PC groups. RESULTS: Following 1:1 matching, 13 infants in both groups were enrolled in the analysis. The weekly BW gain for the first post-treatment week was significantly lower in the IVB group compared with the PC group (86 g vs. 145 g; P = 0.046), whereas the weekly BW gains for the pre-treatment week (173 g vs. 159 g; P = 0.71) and the second post-treatment week (154 g vs. 152 g; P = 0.73) were comparable between the two groups. The short-term inhibitive effect of IVB on BW gain was particularly observed in infants weighing less than 1500 g at treatment (<1500 g: 47 g vs. ≥1500 g: 132 g; P = 0.03). CONCLUSION: IVB could have a short-term inhibitive effect on BW gain in infants with ROP, and this effect is more likely to occur in infants with a lower BW at the time of treatment.

An Attachment Including a +20-D Lens to Gain Extended Field Images in Wide-Angle Optical Coherence Tomography Angiography.

Purpose: This article evaluates the clinical usefulness of an attachment involving a +20-D lens to gain extended field images on wide-angle optical coherence tomography angiography (OCTA). Methods: An attachment with a +20-D lens was developed to take OCTA images of anterior segments, and it was used to obtain extended field images of the posterior segment in this study. Ten eyes of 5 individuals who did not have a history of ocular or systemic disease underwent wide-angle OCTA with a 12 × 12-mm center field using the PLEX Elite 9000 with and without the attachment. The ratio of the area of the center field to the area of the extended field with the attachment was calculated. Results: The mean area of the center image was 125 disc areas and that of the field extended by the attachment was 210 disc areas. The mean ratio between the center field and the extended field was 1.67. Conclusions: The attachment involving the +20-D lens seems to be clinically useful to gain extended field images on wide-angle OCTA.