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To investigate the tilt and decentration of the crystalline lens and the intraocular lens (IOL) relative to the corneal topographic axis using anterior segment ocular coherence tomography (AS-OCT). A sample set of 100 eyes from 49 subjects (41 eyes with crystalline lenses and 59 eyes with IOLs) were imaged using second generation AS-OCT (CASIA2, TOMEY) in June and July 2016 at Okayama University. Both mydriatic and non-mydriatic images were obtained, and the tilt and decentration of the crystalline lens and the IOL were quantified. The effects of pupil dilation on measurements were also assessed. The crystalline lens showed an average tilt of 5.15° towards the inferotemporal direction relative to the corneal topographic axis under non-mydriatic conditions and 5.25° under mydriatic conditions. Additionally, an average decentration of 0.11 mm towards the temporal direction was observed under non-mydriatic conditions and 0.08 mm under mydriatic conditions. The average tilt for the IOL was 4.31° towards the inferotemporal direction relative to the corneal topographic axis under non-mydriatic conditions and 4.65° in the same direction under mydriatic conditions. The average decentration was 0.05 mm towards the temporal direction under non-mydriatic conditions and 0.08 mm in the same direction under mydriatic conditions. A strong correlation was found between the average tilt and decentration values of the crystalline lens and the IOL under both non-mydriatic and mydriatic conditions (all Spearman correlation coefficients, r ≥ 0.800; all P < 0.001). When measured using second generation AS-OCT, both the crystalline lens and the IOL showed an average tilt of 4-6° toward the inferotemporal direction relative to the corneal topographic axis and an average decentration of less than 0.12 mm towards the temporal direction. These results were not influenced by pupil dilation and they showed good repeatability.

To examine factors affecting foveal avascular zone (FAZ) area in healthy eyes using swept-source optical coherence tomography angiography (OCTA). This prospective, cross-sectional study included 144 eyes of 144 individuals (77 women, 67 men) with a best corrected visual acuity of at least 20/20 and no history of ocular disorders. The area of the superficial FAZ was assessed using OCTA. Age, gender, central retinal thickness (CRT), retinal vascular density, refractive error, and axial length were examined to determine associations with FAZ area. The mean age of the subjects was 42.1 ± 20.2 years (range: 10-79 years). The mean FAZ area was 0.32 ± 0.11 mm2, while the mean retinal vascular density was 35.53 ± 0.92%. Multivariate regression analysis was performed using FAZ area as the dependent variable and age, gender, CRT, retinal vascular density, refractive error, and axial length as independent variables. The results of this analysis demonstrate that CRT and retinal vascular density were significantly associated with FAZ area in our sample (P < 0.001, R2 = 0.425). Age, gender, refractive error, and axial length were not significantly correlated with FAZ area, while CRT and retinal vascular density were negatively correlated with FAZ area (CRT: P < 0.001, R2 = 0.356; retinal vascular density: P < 0.001, R2 = 0.189). OCTA results suggest that CRT and retinal vascular density negatively affect FAZ area in healthy eyes.

Purpose To evaluate the cytotoxic effects of alteplase, a recombinant tissue plasminogen activator, and its additives on human retinal pigment epithelial (hRPE) cells. Study design Laboratory study. Methods We evaluated the cytotoxic effects of alteplase on human fetal RPE (hfRPE) cells, human induced pluripotent stem cell-derived RPE (hiPS-RPE), and ARPE-19 cells, as well as the cytotoxic effects of L-arginine and polysorbate 80, two additives of alteplase, on hfRPE cells. The effects of alteplase on the production of vascular endothelial growth factor (VEGF) and pigment epithelium-derived factor (PEDF) from hfRPE cells and the transepithelial resistance (TER) of hiPS-RPE cells were also assessed. The type of cell death induced by alteplase was investigated using ethidium homodimer III and FITC-Annexin V staining and terminal transferase deoxyuridine triphosphatase nick-end labeling. Results Alteplase reduced the viability of hfRPE cells significantly in a dose- and time-dependent manner. The reaction of hiPS-RPE and ARPE19 cells to alteplase was similar to that of hfRPE cells. Out of L-arginine and polysorbate 80, only treatment with L-arginine significantly reduced the viability of hfRPE cells. Alteplase (83 μg/ml, 6 h) had no significant effect on the production of VEGF and PEDF from hfRPE cells. Alteplase decreased the TER of hiPS-RPE cells in a dose- and time-dependent manner and induced necrosis as the type of cell death. Conclusion Alteplase can be cytotoxic to human RPE cells in a concentration- and time-dependent manner, with L-arginine being a possible causative factor.

The epithelial-mesenchymal transition (EMT) in retinal pigment epithelial (RPE) cells plays a central role in the development of proliferative vitreoretinopathy (PVR). The purpose of this study was to investigate the effect of AMP-activated protein kinase (AMPK), a key regulator of energy homeostasis, on the EMT in RPE cells. In this study, EMT-associated formation of cellular aggregates was induced by co-stimulation of cultured ARPE-19 cells with tumor necrosis factor (TNF)-α (10 ng/ml) and transforming growth factor (TGF)-β2 (5 ng/ml). 5-Aminoimidazole-4-carboxamide-1-β-D-ribofuranoside (AICAR), a potent activator of AMPK, significantly suppressed TNF-α and TGF-β2-induced cellular aggregate formation (p < 0.01). Dipyridamole almost completely reversed the suppressive effect of AICAR, whereas 5'-amino-5'-deoxyadenosine restored aggregate formation by approximately 50%. AICAR suppressed the downregulation of E-cadherin and the upregulation of fibronectin and α-smooth muscle actin by TNF-α and TGF-β2. The levels of matrix metalloproteinase (MMP)-2, MMP-9, interleukin-6, and vascular endothelial growth factor were significantly decreased by AICAR. Activation of the mitogen-activated protein kinase and mammalian target of rapamycin pathways, but not the Smad pathway, was inhibited by AICAR. These findings indicate that AICAR suppresses the EMT in RPE cells at least partially via activation of AMPK. AMPK is a potential target molecule for the prevention and treatment of PVR, so AICAR may be a promising candidate for PVR therapy.

To investigate the influence of subretinal injection pressure on the microstructure of the retina in a monkey model. After vitrectomy, balanced salt solution was injected subretinally into one eye each of four cynomolgus monkeys while controlling the injection pressure. Initially, a pressure of 2 psi was used, and this was gradually increased to determine the minimum required pressure. Subsequent injections were performed at two pressures: minimum (n = 13) and high (n = 6). To compare the influence of these injection pressures on retinal structure, optical coherence tomography (OCT) was performed before surgery and every week afterwards. The monkeys were euthanized and their eyes were enucleated at 1 or 6 weeks after the injections. The eyes were processed for light microscopy and transmission electron microscopy (TEM) as well as for TdT-mediated dUTP nick end labeling. The minimum pressure required to perform subretinal injection was 6 psi. After injection at this pressure, both OCT and microscopy showed that the retinal structure was well-preserved throughout the experimental period at all injection sites. Conversely, after injection at high pressure (20 psi) OCT images at all injection sites showed disruption of the ellipsoid zone (EZ) after 1 week. Microscopy indicated damage to the photoreceptor outer segment (OS) and stratification of the retinal pigment epithelium (RPE). After 6 weeks, OCT demonstrated that the EZ had become continuous and TEM confirmed that the OS and RPE had recovered. Photoreceptor apoptosis was absent after subretinal injection at both pressures. The retinal damage caused by subretinal injection increases depending on pressure, indicating that clinicians should perform subretinal injection at pressures as low as possible to ensure safety.

Purpose To determine the presence or absence of interocular differences in the foveal avascular zone (FAZ) area in healthy eyes. Study design Cross-sectional study. Methods We examined 236 healthy eyes of 118 consecutive subjects (mean age, 39.1 ± 18.9 years). We used swept-source optical coherence tomography angiography (OCTA) images of the FAZ to measure its area from both the superficial capillary plexus (SCP-FAZ) and the whole retinal capillary plexus (WCP-FAZ). We also investigated the relationship between interocular differences in SCP-FAZ and other factors such as: axial length, spherical equivalent, central retinal thickness, and retinal vascular density. Results There was no significant difference in the FAZ area between the right and left eyes in either the SCP-FAZ ( P  = 0.61) or WCP-FAZ ( P  = 0.80), and the FAZ areas of both eyes showed significant positive correlations (SCP-FAZ; P  < 0.001, R 2  = 0.884, WCP-FAZ; P  < 0.001, R 2  = 0.856). Bland–Altman plots showed that the mean interocular difference in SCP-FAZ area was 0.002 ± 0.037 mm 2 (95% confidence interval, -0.072–0.075 mm 2 ), and in the WCP-FAZ area, 0.050 ± 0.044 mm 2 (95% confidence interval, -0.036–0.137 mm 2 ). Multivariate regression analysis showed that none of the investigated factors were significantly associated with interocular differences in SCP-FAZ ( P  = 0.61, R 2  = 0.138). Conclusions There was no significant interocular difference in SCP- and WCP-FAZ areas in healthy eyes. The normal range of values for interocular difference in SCP-FAZ area was 0.002 ± 0.037 mm 2 and in the WCP-FAZ area, 0.050 ± 0.044 mm 2 .

Purpose To evaluate the 1-year treatment outcomes of intravitreal aflibercept injections (IVA) using a treat-and-extend regimen for polypoidal choroidal vasculopathy (PCV). Methods Thirty-seven eyes with treatment-naive PCV treated with IVA using a treat-and-extend regimen for 1 year were reviewed retrospectively. The main outcome measures were changes in the best-corrected visual acuity (BCVA) and central retinal thickness (CRT), and the treatment interval at 1 year. The predictive factors for patients who could not continue to extend the treatment interval because of poor response to IVA or recurrence were analyzed. Results The mean logarithm of the minimum angle of resolution BCVA improved from 0.37 at baseline to 0.21 at 1 year ( P  < 0.001). The mean CRT decreased from 342.3 μm at baseline to 196.6 μm at 1 year ( P  < 0.001). The mean treatment interval was 9.7 weeks at 1 year (4 weeks in 11 eyes [29.7%], 6 weeks in 1 eye [2.7%], 8 weeks in 2 eyes [5.4%], 10 weeks in 1 eye [2.7%], and 12 weeks in 22 eyes [59.5%]). A larger number of polypoidal lesions at baseline was predictive for patients who could not continue to extend the treatment interval. Conclusions IVA using a treat-and-extend regimen is effective for improving BCVA and CRT in eyes with PCV.

To investigate the occurrence ratio, localization, and surgical outcomes of intraretinal cystic spaces in idiopathic epiretinal membranes (ERMs). We retrospectively reviewed the charts of 432 eyes of 398 consecutive patients with idiopathic ERM who underwent vitrectomy and ERM peeling from January 2012 to September 2015. We selected cases with intraretinal cystic space prior to surgery, detected by spectral-domain optical coherence tomography. We then evaluated the effects of ERM peeling on intraretinal cystic spaces, best corrected visual acuity, and central retinal thickness at 6 months after surgery. Twenty-four eyes (5.5%) showed intraretinal cystic spaces before surgery, present in the inner retinal layer (the inner group) in 9 eyes, in the outer retinal layer (the outer group) in 6 eyes, and in both the inner and the outer retinal layers (the combined group) in 9 eyes. Additionally, 30 eyes with ERM but without any presence of intraretinal cystic space were selected randomly and classified as the no cyst group. At 6 months after surgery, the disappearance rate of cystic spaces was significantly greater for the outer group than for the inner group (83.3% and 11.1%, respectively, P = 0.011). The mean best corrected visual acuity improved significantly after surgery in the inner group, the outer group, and the no cyst group (P < 0.05 for all three groups) but did not improve in the combined group (P = 0.58). The mean central retinal thickness decreased significantly after surgery in the inner group, the combined group, and the no cyst group (P < 0.05). Intraretinal cystic spaces were observed in 5.5% of preoperative idiopathic ERM cases. Following surgery, the cystic spaces in the outer retinal layer disappeared at higher rates than those in the inner retinal layer, suggesting that the pathophysiologies of these cystic spaces are different.

Background We recently reported that lamellar macular hole (LMH) with lamellar hole-associated epiretinal proliferation (LHEP) can be effectively treated by embedding the LHEP into the retinal cleavage to improve foveal contour and visual acuity. Here, we report a case of LMH with LHEP for which we performed embedding of the LHEP combined with internal limiting membrane (ILM) inversion. We then evaluated the effects of this surgery on macular morphology and visual functions. Case presentation A 62-year-old man presented with visual disturbance (20/29) and metamorphopsia in his right eye. B-scan optical coherence tomography (OCT) imaging revealed the presence of both partial-thickness defect of the macula with degenerative retinal cleavage and LHEP at the surface of the retina. En face OCT imaging showed the absence of retinal fold. We performed phacoemulsification with intraocular lens implantation, vitrectomy, embedding of LHEP into the retinal cleavage, and ILM inversion. Three months after the surgery, both foveal contour and visual acuity (20/20) were improved and metamorphopsia was reduced. Conclusion Embedding of the LHEP combined with ILM inversion may be an effective treatment for LMH with LHEP.

To examine the usefulness of room temperature vulcanizing (RTV) silicone rubber as a barrier material for cell exclusion zone assays. We created barriers using three types of RTV silicone rubber with differing viscosities. We then assessed the adherence of these barriers to culture dishes and their ease of removal from the dishes. We tested the effect of the newly created barriers on the extracellular matrix (ECM) protein fibronectin by attaching and then removing them from fibronectin-coated culture dishes. We also conducted cell exclusion zone assays with MIO-M1 cells using this new barrier in order to measure cell migration. We used real time reverse transcription polymerase chain reaction (RT-PCR) and immunohistochemical staining to measure the effect of fibronectin on MIO-M1 cell migration and the effect of migration (with fibronectin coating) on basic fibroblast growth factor (bFGF) expression in MIO-M1 cells. Of the three types of RTV silicon rubber tested, KE-3495-T was the best in terms of adherence to the dish and ease of removal from the dish. When barrier attachment and removal tests were performed, this rubber type did not have an effect on the fibronectin that coated the dish. In the cell exclusion assay, removal of the barrier revealed that a cell-free area with a distinct margin had been created, which allowed us to conduct a quantitative assessment of migration. Fibronectin significantly promoted the migration of MIO-M1 cells (P = 0.02). In addition, both real time RT-PCR and immunohistological staining indicated that bFGF expression in migrating MIO-M1 cells was significantly higher than that in non-migrating cells (P = 0.03). RTV silicone rubber can be used to create an effective barrier in cell exclusion zone assays and allows simple and low-cost multi-parametric analysis of cell migration.