Fumiyuki Araki

Presynaptic Dystroglycan–Pikachurin Complex Regulates the Proper Synaptic Connection between Retinal Photoreceptor and Bipolar Cells

Dystroglycan (DG) is a key component of the dystrophin–glycoprotein complex (DGC) at the neuromuscular junction postsynapse. In the mouse retina, the DGC is localized at the presynapse of photoreceptor cells, however, the function of presynaptic DGC is poorly understood. Here, we developed and analyzed retinal photoreceptor-specific DG conditional knock-out (DG CKO) mice. We found that the DG CKO retina showed a reduced amplitude and a prolonged implicit time of the ERG b-wave. Electron microscopic analysis revealed that bipolar dendrite invagination into the photoreceptor terminus is perturbed in the DG CKO retina. In the DG CKO retina, pikachurin, a DG ligand in the retina, is markedly decreased at photoreceptor synapses. Interestingly, in the Pikachurin−/− retina, the DG signal at the ribbon synaptic terminus was severely reduced, suggesting that pikachurin is required for the presynaptic accumulation of DG at the photoreceptor synaptic terminus, and conversely DG is required for pikachurin accumulation. Furthermore, we found that overexpression of pikachurin induces formation and clustering of a DG–pikachurin complex on the cell surface. The Laminin G repeats of pikachurin, which are critical for its oligomerization and interaction with DG, were essential for the clustering of the DG–pikachurin complex as well. These results suggest that oligomerization of pikachurin and its interaction with DG causes DG assembly on the synapse surface of the photoreceptor synaptic terminals. Our results reveal that the presynaptic interaction of pikachurin with DG at photoreceptor terminals is essential for both the formation of proper photoreceptor ribbon synaptic structures and normal retinal electrophysiology.

In Vivo Confocal Microscopy of Hereditary Sensory and Autonomic Neuropathy

Purpose: To observe the morphology of the corneal cells and corneal nerve fibers in patients with type IV or V hereditary sensory and autonomic neuropathy (HSAN) by in vivo confocal microscopy and elucidate the mechanism leading to the loss of corneal sensation in this disease. Methods: In vivo confocal microscopy was performed on the central cornea of the right eye in 3 patients with HSAN (ages 17, 20, and 32 years), and their corneal morphology was compared with that of 3 healthy subjects (ages 28, 30, and 36 years). Corneal sensation was tested with a Cochet-Bonnet esthesiometer. Results: The superficial epithelial cell density was lower in the HSAN patients compared with the healthy subjects (1525, 1225, and 1250/mm2 vs. 2225, 1750, and 2500/mm2), but the basal epithelial cell density of the patients was similar to that of the healthy subjects. Nerve bundles were clearly observed in the sub-basal nerve plexus layer of the cornea in the healthy subjects, but were undetectable at the central cornea in the patients with HSAN. The corneal sensation of the patients with HSAN was much weaker than that of the healthy subjects (2.79, 40.30, and 132.50 g/mm2 vs. 1.47, 1.47, and 1.47 g/mm2). Conclusions: Superficial keratopathy accompanied with neurotrophic keratopathy and tear film instability observed clinically agrees with the large keratinized cells in the superficial corneal epithelium by in vivo confocal microscopy in these patients. Our findings suggest that the loss of corneal nerves contributes to impairment of corneal sensation in patients with type IV or V HSAN.

Contributions of retinal direction‐selective ganglion cells to optokinetic responses in mice

In the mouse retina, there are two distinct groups of direction‐selective ganglion cells, ON and ON–OFF, that detect movement of visual images. To understand the roles of these cells in controlling eye movements, we studied the optokinetic responses (OKRs) of mutant mice with dysfunctional ON‐bipolar cells that have a functional obstruction of transmission to ON direction‐selective ganglion cells. Experiments were carried out to examine the initial and late phases of OKRs. The initial phase was examined by measurement of eye velocity using stimuli of sinusoidal grating patterns of various spatiotemporal frequencies that moved for 0.5 s. The mutant mice showed significant initial OKRs, although the range of spatiotemporal frequencies that elicited these OKRs was limited and the response magnitude was weaker than that in wild‐type mice. To examine the late phase of the OKRs, the same visual patterns were moved for 30 s to induce alternating slow and quick eye movements (optokinetic nystagmus) and the slow‐phase eye velocity was measured. Wild‐type mice showed significant late OKRs with a stimulus in an appropriate range of spatiotemporal frequencies (0.0625–0.25 cycles/°, 0.75–3.0 Hz, 3–48°/s), but mutant mice did not show late OKRs in response to the same visual stimuli. The results suggest that two groups of direction‐selective ganglion cells play different roles in OKRs: ON direction‐selective ganglion cells contribute to both initial and late OKRs, whereas ON–OFF direction‐selective ganglion cells contribute to OKRs only transiently.

Endogenous Candida albicans infection causing subretinal abscess

Purpose We report a case of Candida albicans endophthalmitis with subretinal abscess formation in a patient who underwent liver transplantation. Methods Case report. Results A 51-year-old Japanese woman complained of deep pain and ciliary injection in her right eye. Three months prior, the patient had undergone liver transplantation for cirrhosis caused by hepatitis C. A slit-lamp examination revealed intense anterior chamber inflammation with hypopyon and fundoscopy showed a yellowish-white subretinal mass lesion in the inferior peripheral fundus. Systemic and topical antibiotics did not prevent further progression of the infection. The patient underwent pars plana vitrectomy treatment three times and a histopathological study of a vitreous specimen revealed C. albicans to be the causative organism. Conclusion A subretinal abscess, previously reported in Nocardia, Pseudomonas, Staphylococcus, and Aspergillus infection cases, can also occur in patients infected with Candida. Therefore, Candida infection should be considered as a potential cause of subretinal abscess in organ transplant recipients.

Aqueous cytokine levels are associated with reduced macular thickness after intravitreal ranibizumab for diabetic macular edema

Purpose It is controversial whether the administration of anti-vascular endothelial growth factor drugs for diabetic macular edema (DME) affects intraocular inflammatory cytokines. In this study, we measured cytokine concentration in aqueous humor before and after intravitreal injection of ranibizumab (IVR). The aim was to determine changes in cytokine concentration and their effects on DME reduction. Methods Twelve patients (13 eyes) with DME received two IVR (0.5 mg) with a 1 month interval, and a total of 26 aqueous humor samples were obtained. Macular thickness was measured with an optical coherence tomography (OCT) using thickness-map mode with an Early Treatment Diabetic Retinopathy Study (ETDRS) 9-zone grid that was divided into two zones: a central circle with a diameter of 1 mm (zone1); and an outer circle with a diameter of 6 mm (zone2). Results The concentration of eotaxin-1 in aqueous humor samples decreased significantly after IVR. Baseline cytokine concentration was associated with IVR-induced DME reduction. In zone1, higher baseline concentration of interferon-induced protein (IP)-10, and in zone 2, higher baseline concentration of granulocyte-macrophage colony-stimulating factor, IP-10, and tumor necrosis factor (TNF) α; and lower baseline concentration of eotaxin-1, interleukin (IL)-5, and IL-8 were associated with improved DME. Cytokine changes were associated with IVR-induced DME reduction. In zone1, lower concentration of IP-10 compared to baseline or higher concentration of macrophage inflammatory protein (MIP) -α, and in zone 2, lower concentration of IL-5 compared to baseline, IL-8, and IP-10 or higher concentration of eotaxin-1 and MIP-1β were associated with improved DME. Conclusions These findings suggest that ranibizumab affects the concentration of cytokines in aqueous humor. Various cytokines contribute to a decrease in retinal thickness, both in the center of the macula and in a larger area of the retina.

Training system using Bionic-eye for internal limiting membrane peeling

This paper reports on a training system for internal limiting membrane (ILM) peeling. ILM peeling task is difficult even for skilled surgeons, because ILM is a quite thin film on retina and there is a risk to damage the retina at peeling. In order to obtain the skill of ILM peeling and even evaluate newly developed medical devices, the surgical model that can be used for quantitative evaluation is highly demanded. Therefore, here we propose a surgical model for eye surgery named “Bionic-eye”. Fundus of Bionic-eye has artificial ILM and retina. In addition, we assembled Bionic-eye on a whole body surgical model of human named “Bionic-humanoid”.

In vivo confocal microscopy of human cornea covered with human amniotic membrane

PurposeAmniotic membrane transplantation has been widely performed to reconstruct the surface of the eye and treat chemical burns or epithelial defects. However, we have difficulty observing the cornea through the opaque transplanted amniotic membrane by slit-lamp biomicroscopy. We investigated the use of confocal microscopy for observation of human corneas covered with amniotic membrane.MethodsHuman amniotic membrane was placed onto the normal corneas of five volunteers aged 22–24 years. Then, all layers of the covered corneas were observed by in vivo confocal microscopy.ResultsConfocal microscopy displayed the epithelium, basement membrane, and stroma of the amniotic membrane. It also displayed the corneal epithelium. Furthermore, corneal stromal keratocytes and the corneal endothelium were clearly observed through the amniotic membrane by confocal microscopy.ConclusionsWe demonstrated that in vivo confocal microscopy enabled us to observe all layers of corneas covered with amniotic membrane in normal human eyes. Our findings suggest that confocal microscopy may have advantages for clinical examination of the ocular surface, including all layers of the cornea.

Role of the Mouse Retinal Photoreceptor Ribbon Synapse in Visual Motion Processing for Optokinetic Responses

The ribbon synapse is a specialized synaptic structure in the retinal outer plexiform layer where visual signals are transmitted from photoreceptors to the bipolar and horizontal cells. This structure is considered important in high-efficiency signal transmission; however, its role in visual signal processing is unclear. In order to understand its role in visual processing, the present study utilized Pikachurin-null mutant mice that show improper formation of the photoreceptor ribbon synapse. We examined the initial and late phases of the optokinetic responses (OKRs). The initial phase was examined by measuring the open-loop eye velocity of the OKRs to sinusoidal grating patterns of various spatial frequencies moving at various temporal frequencies for 0.5 s. The mutant mice showed significant initial OKRs with a spatiotemporal frequency tuning (spatial frequency, 0.09 ± 0.01 cycles/°; temporal frequency, 1.87 ± 0.12 Hz) that was slightly different from the wild-type mice (spatial frequency, 0.11 ± 0.01 cycles/°; temporal frequency, 1.66 ± 0.12 Hz). The late phase of the OKRs was examined by measuring the slow phase eye velocity of the optokinetic nystagmus induced by the sinusoidal gratings of various spatiotemporal frequencies moving for 30 s. We found that the optimal spatial and temporal frequencies of the mutant mice (spatial frequency, 0.11 ± 0.02 cycles/°; temporal frequency, 0.81 ± 0.24 Hz) were both lower than those in the wild-type mice (spatial frequency, 0.15 ± 0.02 cycles/°; temporal frequency, 1.93 ± 0.62 Hz). These results suggest that the ribbon synapse modulates the spatiotemporal frequency tuning of visual processing along the ON pathway by which the late phase of OKRs is mediated.

Effect of intravitreal ranibizumab injection on aqueous humour cytokine levels in patients with diabetic macular oedema.

Editor, T here is evidence to indicate that inflammatory cytokines, as well as vascular endothelial growth factor (VEGF), play an important role in the development of diabetic macular oedema (DME). Recently, intravitreally injected ranibizumab (IVR) (Lucentis; Novartis, Basel, Switzerland and Genentech Inc., South San Francisco, CA, USA) has been shown to suppress inflammatory cytokines and chemokines that promote adhesion to vascular endothelium or mobilize inflammatory cells by inhibiting multiple isoforms of VEGF-A (Ferrara et al. 2006). Campochiaro et al. (2009) reported that VEGF levels in the aqueous humour were significantly higher in patients with DME (528 112 pg/ml) than in patients with branch retinal vein occlusion (35 3 pg/ml). Muether et al. (2014) measured VEGF levels in the aqueous humour of DME patients receiving IVR, using Luminex, and reported that VEGF levels were completely suppressed in all patients receiving IVR treatment. In this study, we focused on the inflammatory cytokines thought to be involved in DME and analysed its levels that could measure for systematically to identify the changes after treated with ranibizumab. A total of 26 aqueous humour samples obtained from 13 eyes of 12 patients with DME were collected. (Patients’ mean age, 62.5 11.9 years, mean Hb-A1c, 7.0 1.3%, visual acuity, logMAR 0.47 0.25, mean central foveal thickness, 570.0 109.8 lm). A history of focal laser photocoagulations in two eyes was noted. A history of pan-retinal laser photocoagulations was noted for eight eyes. Intravitreal injections of ranibizumab (IVR) was performed twice in each patient. Undiluted aqueous humour samples were obtained during the IVR treatment as follows: a pre-IVR sample was taken during the initial IVR, and a second sample (post-IVR sample) was taken after completion of the second round of IVR, which was conducted 1 month after the initial IVR treatment. Individuals who had undergone previous vitreous surgery or who had already received intravitreal injections of bevacizumab were excluded from this study. Approval for this prospective study was obtained from the Institutional Review Board of our institution. Informed consent was obtained prior to study entry. A multiplex bead-based immunoassay, Luminex 100 multiplex array assay (Luminex Corporation, Austin, TX, USA), was used to measure the levels of 36 cytokines and chemokines in each aqueous humour sample. Then, nine cytokines levels were detected, which were examined between pre-IVR and post-IVR. Of note, eotaxin-1 levels in aqueous humour samples significantly decreased after IVR (p < 0.05, paired t-test, Table 1). In addition, interleukin (IL)6 levels were also measured and analysed in samples from 12 eyes and also showed a tendency to decrease after IVR treatment. Although eotaxin-1 that was shown in this study has been reported for the characteristics in a field of age-related macular degeneration (AMD), this is the first study to show that IVR treatment reduces the levels of eotaxin-1 in the aqueous humour of DME patients. According to a previous report in AMD patients, eotaxin-1 (also known as CCL11) binds to the CCR3 receptor, and the subsequent signalling between this receptor and VEGF is particularly important to express choroidal neovascular endothelial cells in humans with AMD. Furthermore, blockade of CCR3 was more effective than VEGF neutralization in reducing choroidal neovascularization (CNV) (Kim et al. 2015). Although the pathogenesis of AMD and DME may differ, reducing eotaxin-1 levels would be expected to be a more effective function of DME therapy, due to the previously reported molecular biological interaction between VEGF and CCR3.

Primary orbital lymphomatoid granulomatosis

Lymphomatoid granulomatosis (LYG) is a rare angiocentric and angiodestructive Epstein–Barr virus (EBV)-associated B cell lymphoproliferative disorder that involves multiple organs including the lung, skin, kidney and central nervous system. Ocular LYG is an extremely rare tumour, which appears as ocular granulomatous inflammation (conjunctival infiltration,1 posterior uveitis,2 choroidal vasculitis,3 optic neuropathy,4 and retinal detachment5), with only a few reports in the medical literature. Furthermore, there are no reports of primary orbital LYG without pulmonary LYG. We describe the first case of primary orbital LYG diagnosed by biopsy of an orbital tumour. A 54-year-old man presenting with a 6-week history of painless proptosis and ptosis of the right eye was referred to our outpatient clinic. Eye examination revealed marginal blepharitis on the right eye (fig 1A), and the right eye movement was restricted in all directions. Funduscopy showed severe disc swelling in the right eye (fig 1A). His best-corrected visual acuity (BCVA) …