Published papers

Aims/hypothesis: Diabetic retinopathy is a progressive neurodegenerative disease, but the underlying mechanism is still obscure. Here, we focused on oxidative stress in the retina, and analysed its influence on retinal neurodegeneration, using an antioxidant, lutein. Methods: C57BL/6 mice with streptozotocin-induced diabetes were constantly fed either a lutein-supplemented diet or a control diet from the onset of diabetes, and their metabolic data were recorded. In 1-month-diabetic mice, reactive oxygen species (ROS) in the retina were measured using dihydroethidium and visual function was evaluated by electroretinograms. Levels of activated extracellular signal-regulated kinase (ERK), synaptophysin and brain-derived neurotrophic factor (BDNF) were also measured by immunoblotting in the retina of 1-month-diabetic mice. In the retinal sections of 4-month-diabetic mice, histological changes, cleaved caspase-3 and TUNEL staining were analysed. Results: Lutein did not affect the metabolic status of the diabetic mice, but it prevented ROS generation in the retina and the visual impairment induced by diabetes. ERK activation, the subsequent synaptophysin reduction, and the BDNF depletion in the diabetic retina were all prevented by lutein. Later, in 4-month-diabetic mice, a decrease in the thickness of the inner plexiform and nuclear layers, and ganglion cell number, together with increase in cleaved caspase-3- and TUNEL-positive cells, were avoided in the retina of lutein-fed mice. Conclusions/interpretation: The results indicated that local oxidative stress that has a neurodegenerative influence in the diabetic retina is prevented by constant intake of a lutein-supplemented diet. The antioxidant, lutein may be a potential therapeutic approach to protect visual function in diabetes. © 2010 Springer-Verlag.

PURPOSE.: To prospectively investigate the effects of passive cigarette smoke exposure on the ocular surface and tear film in soft contact lens (SCL) wearers. METHODS.: Twelve right eyes of 12 SCL wearers without any ocular or systemic diseases and 10 right eyes of 10 subjects who never wore CLs were examined before and 2 h after 5 min of passive cigarette smoke exposure in a controlled smoke chamber. Tear evaporation rate measurement, tear film break-up time (TBUT) examination, ocular surface fluorescein, rose bengal stainings, and Schirmer I test were performed at each visit. RESULTS.: The mean tear evaporation rates, TBUTs, and vital staining scores were significantly worse in CL wearers compared with healthy control subjects. TBUTs showed significant worsening after passive smoke exposure in both groups. The mean tear evaporation rate and vital staining scores showed a significant increase with brief passive smoke exposure in subjects not wearing CLs but not in CL wearers. CONCLUSION.: Even brief passive exposure to cigarette smoke is associated with adverse effects on the ocular surface as evidenced by an increase in tear instability and damage to the ocular surface epithelia in SCL wearers and non-CL wearers. Copyright © 2010 American Academy of Optometry.

PURPOSE: To investigate the influence of warming the periocular region for 10 minutes after visual display terminal (VDT) work on the near reflex, and the differences in influence between lid closure, dry and wet warming, and the amounts of water vapor. SUBJECTS AND METHODS: Subjects in the first trial were 10 healthy volunteers (10 eyes). Following VDT, subjects wore either a warming eye steamer (water vapor: 200 mg/10 min) or an eye mask with their eyelids closed for 10 minutes. Subjects in the second trial were 5 healthy volunteers (5 eyes). Following VDT, subjects applied either a dry-warming sheet (water vapor: 0 mg/10 min) or a wet-warming sheet (water vapor: 20, 35, 160 and 200 mg/10 min) to there eyelids and the periocular region for 10 minutes. In both experiments, subjective amplitude of accommodation, pupillary reaction, convergence reaction, near vision, and breakup time were measured before and immediately after VDT work and after 2 experimental settings. RESULTS: The warming eye steamer significantly improved amplitude of accommodation (p<0.05), and near vision (p<0.05). Warming the periocular region with water vapor improved subjective amplitude of accommodation, and near vision, the effect of which depended on the amount of water vapor (p<0.05). CONCLUSIONS: Warming the periocular region with water vapor improved subjective amplitude of accommodation, near vision and the effect depended on the amount of water vapor.

In early neonates, the fetal circulatory system undergoes dramatic transition to the adult circulatory system. Normally, embryonic connecting vessels, such as the ductus arteriosus and the foramen ovale, close and regress. In the neonatal retina, hyaloid vessels maintaining blood flow in the embryonic retina regress, and retinal vessels take over to form the adult-type circulatory system. This process is regulated by a programmed cell death switch mediated by macrophages via Wnt and angiopoietin 2 pathways. In this study, we seek other mechanisms that regulate this process, and focus on the dramatic change in oxygen environment at the point of birth. The von Hippel-Lindau tumor suppressor protein (pVHL) is a substrate recognition component of an E3-ubiquitin ligase that rapidly destabilizes hypoxia-inducible factor as (HIF-αs) under normoxic, but not hypoxic, conditions. To examine the role of oxygen-sensing mechanisms in retinal circulatory system transition, we generated retina-specific conditional-knockout mice for VHL (Vhl α-CreKO mice). These mice exhibit arrested transition from the fetal to the adult circulatory system, persistence of hyaloid vessels and poorly formed retinal vessels. These defects are suppressed by intraocular injection of FLT1-Fc protein [a vascular endothelial growth factor (VEGF) receptor-1 (FLT1)/Fc chimeric protein that can bind VEGF and inhibit its activity], or by inactivating the HIF-1a gene. Our results suggest that not only macrophages but also tissue oxygen-sensing mechanisms regulate the transition from the fetal to the adult circulatory system in the retina. © 2010. Published by The Company of Biologists Ltd.

PURPOSE. To investigate the status of oxidative stress and histopathologic alterations in patients with conjunctivochalasis and compare the findings with those in healthy control subjects. METHODS. Eleven patients (n = 20 eyes) with Yokoi grade 3 conjunctivochalasis and 11 health control subjects (n = 22 eyes) were prospectively recruited. ELISA for tear hexanoyllysine (HEL) and inflammatory cytokines, tear film break-up time tests, Schirmer test measurements, and fluorescein and rose bengal vital staining were performed. Conjunctival specimens obtained during surgery for conjunctivochalasis and cataract underwent immunohistochemical staining for HEL+8- OHdG, MMP-3, and MMP-9, and positively stained cells were counted. Transmission electron microscopy was also performed, with staining for elastic fibers in the conjunctival stroma. RESULTS. The mean tear stability and vital staining scores were significantly worse in the conjunctivochalasis patients than in the control subjects. The tear HEL and tear cytokine levels showed significantly higher values in eyes with conjunctivochalasis. IL-1β and IL-6 levels showed a significant correlation with corneal epithelial damage. IL-1β and TNFα showed a significant correlation with 8-OHdG-stained cell counts. Specimens from patients with conjunctivochalasis revealed a significantly higher number of cells positively stained for HEL, 8-OHdG, MMP-3, and MMP-9 than did specimens from age- and sex-matched control subjects. Transmission electron microscopy showed decreased intercellular cohesiveness, with the conjunctival stroma showing an accumulation of elastic fibers. CONCLUSIONS. Lipid and DNA oxidative stress were present in the conjunctiva. Increased tear inflammation seemed to coexist with loss of conjunctival epithelial cohesiveness and increased collagenolytic activity, which may explain the conjunctival laxity observed in patients with conjunctivochalasis. © Association for Research in Vision and Ophthalmology.

Purpose: We previously reported the successful transplantation of corneal epithelium-like cells derived from mouse embryonic stem (ES) cells onto injured mouse cornea. Here, we tested whether nonhuman primate ES cells have ability to differentiate into corneal epithelial cells and whether monkey ES cell-derived corneal epithelium-like cells were applicable for the experimental transplantation to damaged cornea. Methods: Monkey ES cells were cultivated on type IV collagen-coated dishes for various days to induce differentiation into corneal epithelium-like cells. The differentiation was evaluated by reverse transcription-polymerase chain reaction and immunostaining. The corneal epithelium-like cells were transplanted to the injured mouse cornea. Reconstitution of the corneal epithelium was evaluated by immunostaining. Results: The cells cultured on type IV collagen showed cobblestone like appearance resembling epithelial cells. They expressed messenger RNA of pax6, p63, E-cadherin, CD44, proliferating cell nuclear antigen, keratin 3, and keratin 12. Protein expressions of pax6, keratin 3/12, p63, proliferating cell nuclear antigen, E-cadherin, and CD44 were confirmed by immunostaining. When the corneal epithelium-like cells were transplanted, they adhered to the corneal stroma, leading to formation of multiple cell layers. The grafted cells were stained with anti-human nuclear protein antibody, which crossreacted with nuclei of monkey cells but not with those of mouse cells. They retained the expressions of keratin 3/12, E-cadherin, and CD44. Conclusions: We induced corneal epithelium-like cells from monkey ES cells with moderate efficiency. The cells were successfully transplanted onto the injured mouse cornea. This is the first demonstration that nonhuman primate ES cells were induced to differentiate into corneal epithelium-like cells, which were applicable for transplantation to an animal model of corneal injury. © 2010 Lippincott Williams & Wilkins.

Purpose : To report two cases who received replacement of multifocal intraocular lens (IOL) by monofocal IOL due to postoperative dissatisfaction. Cases : A 75-year-old male received bilateral cataract surgery with implantation of diffractive multifocal IOL. Asthenopia and photophobia became worse after surgery. In accordance to his wish, the multifocal IOL was replaced by monofocal IOL 4 months later. His subjective complaints persisted but improved after treatment for essential blepharospasm. Another 52-year-old female received similar bilateral cataract surgery with implantation of refractive multifocal IOL. The lens was replaced by monofocal one due to photophobia 4 months later. Her subjective complaints persisted. Conclusion : These cases illustrate that replacement of multifocal by monofocal IOL due to dissatisfaction of patients need careful considerations.

Purpose: To evaluate the efficacy, sensitivity and specificity of confocal microscopy (CM) parameters: meibomian gland (MG) acinar longest diameter (MGALD), MG acinar shortest diameter (MGASD), inflammatory cell density (ICD), and MG acinar unit density (MGAUD) in the diagnosis of MG dysfunction (MGD). Design: Prospective, controlled, single-center study. Participants: Twenty MGD patients (9 males, 11 females; mean age, 63.5±16.5 years) and 26 age- and gender-matched control subjects (13 males, 13 females; mean age, 53.2±15.7 years) were recruited. Methods: All subjects underwent slit-lamp examinations, tear film break-up time (BUT) measurements, assessment of tear evaporation rate from the ocular surface (TEROS), vital stainings, Schirmer test, meibography, MG expressibility, and CM of the MG. Data were compared between the 2 groups using the Mann-Whitney and chi-square tests. Main Outcome Measures: The correlation between the clinical findings of tear functions, vital staining scores, and the 4 CM parameters were tested by Spearman's correlation coefficient by rank test. Receiver operating characteristic curve technique was used to evaluate the sensitivity, specificity, and cutoff values of CM parameters. Results: The mean tear film BUT, vital staining scores, TEROS values, MG expressibility, and MG dropout grades by meibography were significantly worse in MGD patients compared with controls (P<0.001). The mean values of the MGALD, MGASD, ICD, and MGAUD in MGD patients were significantly worse than those observed in the controls with CM. All CM parameters showed a strong, significant correlation with tear functions, ocular surface vital stainings, MG expressibility, and MG dropout grades. The cutoff values for MGALD, MGASD, ICD, and MGAUD in the diagnosis of MGD were 65 μm, 25 μm, 300 cells/mm2, and 70 glands/mm2, respectively. The sensitivity and specificity values of these parameters under these cutoff values were 90% and 81% for MGALD, 86% and 96% for MGASD, 100% and 100% for ICD, 81% and 81% for MGAUD. Conclusions: Confocal microscopy has the potential to diagnose the simple MGD with high sensitivity and specificity. The CM-based diagnostic parameters correlated significantly and strongly with the status of the ocular surface disease. Financial Disclosure(s): The authors have no proprietary or commercial interest in any of the materials discussed in this article. © 2010 American Academy of Ophthalmology.

Purpose: To evaluate the long-term efficacy of a biconvex accommodating intraocular lens (IOL) to restore near visual performance. Setting: Department of Ophthalmology, Keio University Hospital, Tokyo, Japan. Methods: This 4-year study comprised eyes that had cataract surgery with implantation of a 1CU accommodating IOL. At all postoperative visits, the following were assessed: subjective spherical equivalent (SE) refraction; uncorrected (UDVA) and corrected (CDVA) distance visual acuities; uncorrected (UNVA), distance-corrected (DCNVA), and corrected (CNVA) near visual acuities; change in minimum additional (add) power (diopters) to attain CNVA; and accommodation amplitude. A questionnaire was administered at the last visit. Results: Twelve eyes of 8 cataract patients (mean age 59.0 years ± 18.4 [SD]) were evaluated. Postoperatively, the mean SE at 1 year and 4 years was significantly more hyperopic than at 1 month (P<.05). The mean UDVA increased significantly at 4 years compared with 1 month (P<.05). There were no significant changes in CDVA, UNVA, CNVA, and DCNVA throughout the follow-up period. The change in the minimum add power to attain CNVA and in the subjective and objective accommodation amplitudes also did not change significantly over time. Eighty-three percent of patients reported being satisfied with the results. Conclusion: Four years after implantation of an accommodating IOL, most patients had good vision and were satisfied, although the accommodation amplitude for near vision was not sufficient throughout the follow-up. Financial Disclosure: No author has a financial or proprietary interest in any material or method mentioned. © 2010 ASCRS and ESCRS.

Purpose: To investigate the visual sensations experienced by patients during vitrectomy under retrobulbar anesthesia. Methods: 30 men and 45 women with a mean age of 65.3 ± 10.6 years underwent vitrectomy under retrobulbar anesthesia for macular disease. 28 eyes had an idiopathic epiretinal membrane, 13 had an idiopathic macular hole, 32 had macular edema (17 diabetic retinopathy and 15 retinal vein occlusion), and 2 had submacular hemorrhage. 49 patients with nonmacular disease underwent similar vitrectomy procedures and were used for comparison. An interview was conducted with the patient about his/her visual sensations during and within 3 h of the vitrectomy. Results: 70 (93.3%) of the patients reported seeing lights, 53 (70.7%) reported seeing colors, and 48 (64.0%) reported seeing movements or moving objects. Of the patients who reported seeing movements or moving objects, 44 (58.7%) reported seeing surgical instruments, and 5 (6.7%) saw the surgeon's fingers or hands. Patients with macular diseases tended to report more visual sensations than patients with nonmacular diseases. The patients' description and drawings appeared to arise mainly from the shadows cast by the intravitreal objects, and some patients perceived highly accurate details including the movements and color of the objects. Conclusions: Visual sensations are experienced by approximately 90% of the patients, and there may be a common mechanism by which patients perceive the intravitreal objects that are not focused on by the retina through the eye's optical system. © 2009 S. Karger AG, Basel.

PURPOSE. To demonstrate the conjunctival alterations in patients with Sjögren's (SSDE) and non-Sjögren's syndrome dry eye (NSSDE) using a new generation confocal microscope (HRTII/ RCM; Heidelberg Engineering, Heidelberg, Germany), in a prospective controlled study. METHODS. Twenty-eight right eyes of 28 patients with SSDE (28 women; mean age, 58.2 ± 14.3 years), 7 right eyes of patients with NSSDE (7 women; mean age, 66.1 ± 14.4 years), and 14 right eyes of 14 age- and sex-matched control subjects were studied. All subjects underwent the Schirmer test, tear film breakup time (BUT), vital staining, and confocal microscopy of the temporal bulbar conjunctiva. The density of conjunctival epithelial cells, epithelial microcysts, and conjunctival and corneal inflammatory infiltrates were also assessed. RESULTS. The tear quantity, stability, and vital staining scores were significantly worse in patients with SSDE or NSSDE than in control subjects (P < 0.001 and P < 0.05, respectively). Eyes of the patients with SSDE or NSSDE had a significantly higher density of conjunctival and corneal inflammatory infiltrates than did the control eyes (P < 0.001). Conjunctival inflammatory cell densities showed a negative correlation with tear stability and tear quantity and a positive correlation with the vital staining scores. Conjunctival epithelial cell densities were significantly lower in SSDE and NSSDE compared with control subjects (P < 0.05). The density of epithelial cysts was significantly higher in SS than in healthy control eyes (P < 0.001). CONCLUSIONS. Confocal scanning laser microscopy was an efficient and a noninvasive tool for the quantitative assessment of the conjunctival inflammation and epithelial cell densities as well as evaluation of conjunctival morphologic alterations, such as microcysts in patients with SSDE and NSSDE dry eye disease. © Association for Research in Vision and Ophthalmology.

To evaluate the risk factors of severe peripheral corneal ulcers. Twenty three eyes of 15 patients (9 eyes of 7 men and 14 eyes of 8 women) with severe peripheral corneal ulcers, who were treated during the past 10 years at Keio University Hospital, were reviewed about the original disease, status of ulcer, treatment, and visual prognosis. Fifteen eyes of 10 patients with Mooren's ulcer, 4 eyes of 3 patients with rheumatoid arthritis, and 4 eyes of 2 patients with Terrien corneal degeneration were included. Mooren's ulcer tends to advance widely. Systemic administration of immunosuppressive and corticosteroidal agents was required in addition to surgical treatment. Visual prognosis was poor in the Mooren's ulcer and perforated cases. Mooren's ulcer and perforated cases are risk factors in peripheral corneal ulcer.

Purpose: To evaluate the changes in functional visual acuity before and after neodymium:YAG (Nd:YAG) laser capsulotomy. Setting: Keio University Hospital, Tokyo, Japan. Design: Case series. Methods: Eyes that had previous cataract surgery with a clinical diagnosis of central posterior capsule opacity requiring Nd:YAG laser capsulotomy were evaluated. All patients had refractive error and corrected distance visual acuity (CDVA) measurements; slitlamp microscopy examinations, including posterior capsule opacification evaluation; high-contrast visual acuity measured at 5 m using a Landolt chart; 10% low-contrast visual acuity (LCVA); functional visual acuity; wavefront examination; and a Schirmer test without anesthesia. Results: The study enrolled 9 patients (4 women, 5 men; 10 eyes) with a mean age of 57.3 years ± 7.0 (SD) (range 47 to 68 years). Although there was no significant difference in CDVA before and after Nd:YAG capsulotomy, there was a significant improvement in the mean functional visual acuity, from 0.18 ± 0.11 logMAR (range -0.03 to 0.31 logMAR) to 0.08 ± 0.07 logMAR (range -0.04 to 0.18 logMAR) (P<.05). The LCVA improved significantly after Nd:YAG laser capsulotomy (P=.003). Higher-order aberrations and the root mean square of the 4th-order coefficients decreased significantly after capsulotomy, and these aberrations were significantly correlated with functional visual acuity and LCVA. Conclusion: Results indicate that measurement of functional visual acuity is useful in assessing the quality of vision in patients who have Nd:YAG laser capsulotomy after cataract surgery. Financial Disclosure: No author has a financial or proprietary interest in any material or method mentioned. © 2011 ASCRS and ESCRS.

Introduction: Based on data from the largest studies of dry eye to date-the Women's Health Study (WHS) and the Physicians' Health Study (PHS)-and other studies, it has been estimated that about 3.23 million women and 1.68 million men, for a total of 4.91 million Americans aged ≥ 50 years, have dry eye. Tens of millions more have less severe symptoms and probably a more episodic manifestation of the disease that is notable only during contact with some adverse contributing factor(s), such as low humidity or contact lens wear. Dry eye disease is a common yet frequently under-recognized public health problem whose etiology and management challenge clinicians and researchers involved in this field. Areas covered: Advances in the understanding of the disease have been made over the past 10 years in areas of epidemiology, pathogenesis, clinical manifestation, and possible therapy. Historical aspects and recent information in relation to the use of artificial tear substitutes and anti-inflammatory agents in dry eye disease, including topical cyclosporin and corticosteroids, autologous serum, tetracyclines and systemic immunosuppressants, are covered in this review. The reader will gain insight into the recent views on the pharmacological menu of treatments for dry eyes following the recommendations of the 2007 International Dry Eye Workshop. Expert opinion: Dry eye is a visually disabling disease, the treatment of which needs tailoring according to the type and severity of dry eye disease. © 2011 Informa UK, Ltd.

PURPOSE. To investigate the role of reactive oxygen species (ROS) as the prime initiators of the angiogenic response after alkali injury of the cornea and observe the effects of antioxidants in preventing angiogenesis. METHODS. The corneal epithelia of SOD-1-deficient mice or wild-type (WT) mice were removed after application of 0.15 N NaOH to establish the animal model of alkali burn. ROS production was semiquantitatively measured by dihydroethidium (DHE) fluorescence. Angiogenesis was visualized by CD31 immunohistochemistry. The effects of the specific NF-κB inhibitor DHMEQ, the antioxidant N-acetyl-L-cysteine (NAC), and hydrogen (H2) solution were observed. RESULTS. ROS production in the cornea was enhanced immediately after alkali injury, as shown by increased DHE fluorescence (P< 0.01). NF-κB activation and the upregulation of vascular endothelial growth factor (VEGF) and monocyte chemoattractant protein-1 (MCP-1) were significantly enhanced (P< 0.01), leading to a significantly larger area of angiogenesis. Angiogenesis in SOD-1-/- mice corneas were significantly higher in WT mice (P<0.01), confirming the role of ROS. Pretreatment with the specific NF-κB inhibitor DHMEQ or the antioxidant NAC significantly reduced corneal angiogenesis by downregulating the NF-κB pathway (P<0.01) in both WT and SOD-1-/- mice. Furthermore, we showed that irrigation of the cornea with hydrogen (H2) solution significantly reduced angiogenesis after alkali-burn injury (P<0.01). CONCLUSIONS. Immediate antioxidant therapy with H2-enriched irrigation solution is a new potent treatment of angiogenesis in cornea to prevent blindness caused by alkali burn. © 2011 The Association for Research in Vision and Ophthalmology, Inc.

Purpose To compare tear film thickness between normal subjects and aqueous tear deficiency dry eye patients by tear interferometry. Design Prospective case-control study. Methods Central precorneal tear film thickness was measured noninvasively using an interference thin-film thickness measurement device (Quore MSPA1100; Mamiya-OP). Tear film thickness of 14 eyes from 14 normal subjects and of 28 eyes from 28 aqueous tear deficiency dry eye patients were compared along with noninvasively measured tear meniscus height, DR-1 (Kowa) dry eye severity grading, fluorescein and rose bengal staining scores, tear film break-up time, and Schirmer test results. Among dry eye patients, 13 eyes underwent punctal occlusion, and tear film thickness was compared before and after the surgery. Results Tear film was significantly thinner in dry eye patients (2.0 ± 1.5 μm) than normal subjects (6.0 ± 2.4 μm; P < .0001). Tear film thickness showed good correlation with other dry eye examinations. After punctal occlusion, tear film thickness increased significantly from 1.7 ± 1.5 μm to 4.9 ± 2.8 μm (P = .001) with the improvement of tear meniscus height, fluorescein and rose bengal staining scores, tear film break-up time, and Schirmer test values. Conclusions Interferometric tear film thickness measurement revealed impaired precorneal tear film formation in aqueous tear deficiency dry eyes and was useful for showing the reconstruction of tear film after punctal occlusion surgery. Interferometry of precorneal tear film may be helpful for the evaluation of aqueous tear deficiency in conjunction with other dry eye examinations.