Published papers

Purpose: To compare the safety of domestic and overseas donor corneas for keratoplasty. Objects and Method: This study was made on 490 eyes donated to us through eyebanks in the past 5 years through 2010. The series comprised 245 eyes from domestic donors and 245 eyes from USA. Results: Age of domestic donors averaged 68.8 years and that of overseas donors averaged 63.1 years. The difference was significant (p<0.01). Corneal endothelial density averaged 2713.3 and 2515.3/mm 2 respectively. The difference was significant (p<0.01). Primary graft failure occurred in 4 domestic eyes and none in overseas eyes. There was no difference in the positive rate of culture from corneal limbus or preservation medium between domestic and overseas donor eyes. Conclusion: The mean corneal endothelial density was higher in domestic than in overseas cornea. Primary graft failure was more frequent in domestic than overseas eyes. More strict quality control is desired for donated corneas.

Purpose: To investigate the relation between visual function, clinical findings, and visual symptoms in Stevens-Johnson syndrome (SJS) and to compare the results with Sjögren syndrome (SS) patients and normal subjects. Design: Cross-sectional comparative study. Methods: One hundred fifteen eyes of 59 consecutive patients with SJS and toxic epidermal necrolysis (TEN), 208 eyes of 104 healthy normal subjects, and 132 eyes of 66 SS patients were investigated in this multicenter study. All study subjects underwent tear function and ocular surface examinations, Landolt and functional visual acuity examinations, and the Japanese version of the NEI VFQ-25 (National Eye Institute Visual Function Questionnaire). Results: The mean ocular surface grading scores were significantly higher and the mean score of all 12 NEI VFQ subscales was significantly lower in the SJS patients compared to the SS patients and the normal subjects (P <.05). The conventional and functional logarithm of minimal angle of resolution (logMAR) visual acuities in SJS patients with minimal corneal complications were significantly higher and the mean total composite NEI VFQ scores were lower compared to SS patients. The conventional and functional logMAR visual acuities and the mean ocular surface grading scores in SJS with aqueous deficiency were significantly higher and the mean total composite NEI VFQ scores were lower compared to SS patients. Strong correlations between best-corrected logMAR functional visual acuities and either ocular surface grading scores or the composite NEI VFQ-25 scores were observed. Conclusions: The functional visual acuity examination reflects the severity of clinical ocular surface findings and vision-related quality of life more than the standard conventional visual acuity in SJS. © 2012 Elsevier Inc. All rights reserved.

Stem cell-based therapy has been proposed as a promising strategy for regenerating tissues lost through incurable diseases. Side population (SP) cells have been identified as putative stem cells in various organs. To examine therapeutic potential of SP cells in hypofunction of exocrine glands, SP cells isolated from mouse exocrine glands, namely, lacrimal and salivary glands, were transplanted into mice with irradiation-induced hypofunction of the respective glands. The secretions from both glands in the recipient mice were restored within 2 months of transplantation, although the transplanted cells were only sparsely distributed and produced no outgrowths. Consistent with this, most SP cells were shown to be CD31-positive endothelial-like cells. In addition, we clarified that endothelial cell-derived clusterin, a secretory protein, was an essential factor for SP cell-mediated recovery of the hypofunctioning glands because SP cells isolated from salivary glands of clusterin-deficient mice had no therapeutic potential, whereas lentiviral transduction of clusterin restored the hypofunction. In vitro and in vivo studies showed that clusterin had an ability to directly inhibit oxidative stress and oxidative stress-induced cell damage. Thus, endothelial cell-derived clusterin possibly inhibit oxidative stress-induced hypofunction of these glands. © AlphaMed Press.

Purpose: To evaluate visual function changes after punctal plug (PP) occlusion in patients with short breakup time of tear film (BUT) type of dry eyes. Methods: A prospective comparative study was performed in 43 eyes of 43 dry eye patients with short BUTbreakup time of tear film receiving PP occlusion. Patients were divided into 2 groups as those receiving only upper or only lower occlusion. Functional visual acuity measurements, tear function evaluations, and questionnaire on satisfaction with treatment in relation to presence of epiphora were performed before and 1 month after PP insertion. Results: Twenty eyes of 43 eyes (46.5%) with only lower occlusion and 17 eyes of 43 eyes (39.5%) with only upper occlusion showed satisfactory outcomes with treatment. One of 43 eyes (2.3 %) had epiphora. Vital staining scores and BUT values significantly improved in both groups after occlusion. The mean logarithm of the minimum angle of resolution functional visual acuity value significantly improved only in eyes receiving upper PP occlusion. Conclusion: Upper PP occlusion seems to be associated not only with improvement of tear stability, vital staining scores, and treatment-related satisfaction but also with better quality of visual function. Copyright © 2012 by Lippincott Williams & Wilkins.

The ocular surface is strongly affected by oxidative stress, and anti-oxidative systems are maintained in corneal epithelial cells and tear fluid. Dry eye is recognized as an oxidative stress-induced disease. Selenium compound eye drops are expected to be a candidate for the treatment of dry eye. We estimated the efficacy of several selenium compounds in the treatment of dry eye using a dry eye rat model. All of the studied selenium compounds were uptaken into corneal epithelial cells in vitro. However, when the selenium compounds were administered as eye drops in the dry eye rat model, most of the selenium compounds did not show effectiveness except for Se-lactoferrin. Se-lactoferrin is a lactoferrin that we prepared that binds selenium instead of iron. Se-lactoferrin eye drops suppressed the up-regulated expression of heme oxygenase-1, cyclooxygenase-2, matrix metallopeptidase-9, and interleukin-6 and also suppressed 8-OHdG production in the cornea induced by surgical removal of the lacrimal glands. Compared with Se-lactoferrin, apolactoferrin eye drops weakly improved dry eye in high dose. The effect of Se-lactoferrin eye drops on dry eye is possibly due to the effect of selenium and also the effect of apolactoferrin. Se-lactoferrin is a candidate for the treatment of dry eye via regulation of oxidative stress in the corneal epithelium. © 2012 Higuchi et al.

Previous observations in a rat model of a non-Sjögren's syndrome (non-SS) type of dry eye seen in users of visual display terminals (VDT) indicated that secretory vesicle (SV) accumulation in the lacrimal gland epithelia contributes to the condition. Here, to examine this possibility in humans, we compared the lacrimal gland histology and percent SV area in the cytoplasm of acinar epithelial cells using light microscopy and transmission electron microscopy, in patients with VDT work-related non-SS dry-eye (VDT group), SS-induced dry-eye, and autopsied normal controls. In addition, the VAMP8 (vesicle-associated membrane protein 8, an exocrine-pathway molecule) and Rab3D (mature vesicle marker) were histochemically examined in lacrimal gland tissue sections. The lacrimal gland acini were larger in the VDT group than in the SS group, and the percent SV area was significantly higher in the VDT group than in the normal controls (P = 0.021) or SS group (P = 0.004). Immunostaining revealed abnormal distributions of VAMP8 in the VDT and SS groups. Rab3D was more strongly expressed in the cytoplasm of acinar epithelial cells in the VDT group than in that of normal controls. The duration of VDT use was significantly longer in the VDT group than in the other groups. These findings suggest that excessive SV accumulation in the acinar epithelia may contribute to the reduced tear secretion in VDT users. © 2012 Kamoi et al.

PURPOSE. To investigate the expression, localization, and physiologic function of renin-angiotensin system (RAS) components in the mouse lacrimal gland. METHODS. Lacrimal glands and cultured lacrimal gland fibroblasts from wild-type (WT) BALB/c (H-2d) mice were used. Reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry were used to determine the expression and localization of the RAS components, prorenin/renin, angiotensin-converting enzyme (ACE), angiotensin II, angiotensin II type 1 receptor (AT1R), and angiotensin II type 2 receptor (AT2R) in the normal mouse lacrimal gland. To examine the change in tear secretion, mice received ARB (AT1R blocker) or AT2R antagonist. Tear secretion was assessed by cotton thread test before and after drug administration. RESULTS. The mRNAs coding for angiotensinogen, prorenin, ACE, and both AT1R and AT2R were found in normal lacrimal gland tissue and cultured lacrimal gland fibroblasts. Prorenin/renin and ACE were identified in myoepithelial cells around ducts and acini and in blood vessels. Angiotensin II, AT1R, and AT2R were observed in the ducts and interstitial fibroblasts. AT1R and AT2R were also localized in blood vessels. All the cultured lacrimal gland fibroblasts expressed angiotensin II, AT1R, and AT2R. Tear secretion increased in mice that received ARB. CONCLUSIONS. The results are consistent with the hypothesis that a tissue-specific RAS is present in the lacrimal gland, and suggest that fibroblasts are one of the cell types playing a role in the tissue RAS. Tissue RAS might be involved in tissue function of regulating tear secretion in the lacrimal gland. © 2012 The Association for Research in Vision and Ophthalmology, Inc.

Kabuki syndrome has long been clinically defined based mainly on its characteristic eye features. The recent discovery of MLL2 as a causative gene of Kabuki syndrome has enabled the extreme end of the phenotype to be explored. We herein report on two patients with striking visible congenital staphyloma at birth. A diagnosis of Kabuki syndrome was subsequently made in both patients based on a constellation of characteristic eye features, cardiac abnormalities and severe developmental delay, and finally by the confirmation of MLL2 mutations. In conclusion, congenital corneal staphyloma is a complication of Kabuki syndrome with MLL2 mutations. © 2012 Wiley Periodicals, Inc..

Purpose. The Tet-mev-1 mouse expressing a mitochondrial complex-II mutated SDHCV69E gene controlled by a tetracycline (Tet)-On/Off system can overproduce O2·- and is a versatile whole-animal model for studying mitochondrial oxidative stress. Here we report a series of age-dependent variations in corneal epithelium, endothelium, and parenchymal cells of the Tet-mev-1 mice relative to wild-type C57BL/6j mice. Methods. Measurements of (1) mitochondrial electron transport enzyme activities; (2) O2·- production; (3) carbonylated protein, and 8-hydroxydeoxyguanosine (8-OHdG) levels as markers of oxidative stress; (4) pathologic analyses under optical and electron microscopy; (5) hematoxylin-eosin or toluidine-blue staining; and (6) immunohistochemistry with an anti-β-catenin antibody were performed in the eye, especially the cornea. Results. Complex II-III activity was decreased by electron leakage between complex II and CoQ. This resulted in increased age-dependent intracellular oxidative stress in the eye of Tet-mev-1 mice. Corneal epithelialization was delayed in Tet-mev-1 mice after 20% ethanol treatment, as the number of cells and mitotic cells decreased in the corneal epithelium of Tet-mev-1 mice compared with that of wild type. The age-dependent decrease in cell number accelerated in the corneal endothelium cells. Moreover, it was suggested that the corneal thickness was decreased by thinning of parenchymal cells with age in Tet-mev-1 mice. Conclusions. These results suggest that mitochondrial oxidative stress with electron transport chain dysfunction can influence pathogenesis and progression of age-related corneal diseases, as well as generalized corneal aging acceleration. © 2012 The Association for Research in Vision and Ophthalmology, Inc.

Purpose: To investigate the incidence of intraocular pressure (IOP) elevation after Descemet's stripping endothelial keratoplasty (DSEK) and associated clinical findings. Methods: Forty-eight eyes of 46 patients who underwent DSEK were studied in this single-center retrospective study. IOP elevation was defined as the presence of an increase in IOP above 21 mmHg lasting at least two - months, the need to introduce topical/oral antiglaucoma medications for at least two months, and the need for surgical therapy for uncontrolled IOP. Patients with pre-existing glaucoma or patients who were using antiglaucoma drugs before DSEK were excluded according to the protocol of the study. The incidence of post-DSEK IOP elevation was investigated. Results: The incidence of IOP elevation after DSEK was 16.7%. Glaucoma therapy following DSEK was needed in eight eyes of eight patients. IOP elevations were diagnosed from two to ten months after DSEK. There was no statistical difference in the survival probabilities of patients with different preoperative diagnoses. Pre-op IOP was observed to be a risk factor of IOP elevation in this series. Conclusions: IOP elevation appears to be a common postoperative complication of DSEK. Patients without preexisting glaucoma but with higher pre-op IOP need to be followed up carefully after DSEK. © Japanese Ophthalmological Society 2012.

Aims: The objective of this study was to investigate the association between blood pressure (BP) and visual field defect progression in normal-tension glaucoma (NTG). Methods: The subjects were 35 consecutive patients with NTG who were admitted to Keio University Hospital for 24-hour intraocular pressure evaluation; all subjects underwent 6 or more visual field tests in either eye and were followed up for more than 5 years. Systolic and diastolic BPs were measured at 9.00 h. Subjects with negative regression lines and p values of less than 0.05 were classified as progressive; all others were defined as stable. Systolic BP, diastolic BP, and ocular perfusion pressure were compared between the 2 groups. Results: Twelve subjects showed progression in glaucomatous visual field defects (age 55.1 ± 15.1 years). Twenty-three subjects showed no progression (age 57.2 ± 10.9 years). Diastolic BP (stable group: 77.0 ± 12.1 mm Hg; progressive group: 67.7 ± 9.7 mm Hg; p = 0.03) was significantly lower in the progressive group than in the stable group. Multivariable analysis revealed that low diastolic BP is a significant risk factor for progression (OR 0.90; 95% CI 0.82-0.99). Conclusions: Low diastolic BP was associated with glaucomatous visual field progression in subjects with NTG. Copyright © 2012 S. Karger AG, Basel.

Purpose: To evaluate the efficacy of punctal plugs for patients with post-LASIK dry eye. Materials and methods: A prospective study was conducted on 25 eyes of 18 patients who underwent LASIK. All eyes fulfilled the Japanese dry-eye criteria and had not responded to conventional treatment with artificial tears by 1 month postsurgery. They were randomly divided into a plug and a non-plug group. Punctal plugs were inserted into the superior and inferior puncta in the plug group after the 1-month examination. Uncorrected and best corrected visual acuity (UCVA, BCVA), refraction, functional visual acuity (FVA) after 10 s eye opening (FVA 10), surface regularity index (SRI) after 10 s eye opening (SRI 10), corneal sensitivity, Schirmer test, fluorescein staining (FS), tear break-up time (TBUT), and symptoms were compared between the groups 1 and 3 months postsurgery. Changes in these data from 1 to 3 months were also compared between the groups. Results: The values in SRI 10, FS, and TBUT were significantly better in the plug group at 3 months. Changes in the values of UCVA, FVA 10, FS, TBUT, and symptoms between 1 and 3 months improved significantly in the plug group compared with the non-plug group. Conclusion: Our results suggest that punctal occlusion for both puncta is effective for patients with post-LASIK dry eye that cannot be controlled by artificial tears alone. Punctal plugs can improve not only the symptoms and tear function, but also the quality of vision, for example FVA. © Japanese Ophthalmological Society 2012.

Lutein, a xanthophyll of a carotenoid, is anticipated as a therapeutic product to prevent human eye diseases. However, its biological mechanism is still unclear. Here, we show the molecular mechanism of lutein's effect to reduce photodamage of the retina. We analyzed the light-exposed retinas of Balb/c mice given lutein-supplemented or normal diet. Visual function was measured by electroretinogram, and histological changes were observed. Immunohistochemical and immunoblot analyses were performed to analyze molecular mechanism. The reactive oxygen species induced in the retina was evaluated by fluorescent probes. In the mice after light exposure, reduction of a-wave and b-wave amplitudes in electroretinogram, indicating visual impairment, and thinning of the photoreceptor cell layer owing to apoptosis were both attenuated by lutein diet. Interestingly, γ-H2AX, a marker for double-strand breaks (DSBs) in DNA, was up-regulated in the photoreceptor cells after light exposure, but this increase was attenuated by lutein diet, suggesting that DSBs caused by photodamage contributed to the photoreceptor cell death and that this change was suppressed by lutein. Moreover, the expression of eyes absent (EYA), which promotes DNA repair and cell survival, was significantly up-regulated with lutein diet in the light-exposed retina. Therefore, lutein induced EYA for DNA repair, which could suppress DNA damage and photoreceptor cell apoptosis. Lutein reduced light-induced oxidative stress in the retina, which might contribute to promote DNA repair. The lutein-supplemented diet attenuated light-induced visual impairment by protecting the photoreceptor cells' DNA. © 2012 Elsevier Inc.

PURPOSE.: To evaluate the tear meniscus changes after punctal occlusion in dry eye patients using Visante optical coherence tomography (OCT) in a prospective controlled study. METHODS.: Thirty eyes of 15 symptomatic dry eye patients not responding to non-preserved artificial tears received additional upper and lower punctal occlusion with silicone plugs, and 30 eyes of 15 age-and sex-matched dry eye control patients received only non-preserved artificial tears treatment for 1 month. All subjects underwent tear meniscus height (TMH) measurements with Visante OCT. All study participants also underwent slitmicroscopy graticule scale TMH measurement, strip meniscometry testing, tear film break-up time measurement, ocular surface vital staining with fluorescein and Rose Bengal dyes, and the Schirmer-1 test. Both groups, dry eye and control group patients, were examined before and after 1-month treatment. Wilcoxon-matched pair test was performed. The study was conducted in compliance with the Tenets of the Declaration of Helsinki. RESULTS.: TMH measurements by OCT and slitlamp graticule scale significantly improved after punctal occlusion (p < 0.001) and remain unchanged in the dry eye control patients. Similarly, strip meniscometry scores, mean tear stability values, Rose Bengal, and fluorescein staining scores showed significant improvement after punctal occlusion (p < 0.05). Schirmer-1 test values tended to be higher after 1 month of treatment in both groups without any statistical significance (p > 0.05). CONCLUSIONS.: OCT TMH measurement appears to be effective in monitoring tear meniscus changes after punctal occlusion. OCT can be a valuable non-invasive and quick clinical tool for evaluation of treatment responses in dry eye patients. Copyright © 2012 American Academy of Optometry.

Objective: To determine the antioxidant supplementation effect on accommodation among VDT users. Design: a double blind randomized placebo controlled study. registered under clinical Trials.gov identifier no. ncT00877201. Participants and Controls: Fourty right eyes of 40 healthy VDT users (30 females, 10 males, mean age: 43.8±2.8 years, range: 40-49 years). 20 subjects (15 females, 5 males; mean age: 44.0±2.7 years, range: 40-49 years). Methods: subjects were required to take an antioxidant supplement, 20 age and sex matched subjects (15 females, 5 males; mean age: 43.6±3.1 years, range: 40-49 years) were required to take placebo medication for 4 weeks. Results: The mean of the change in accommodation power was significantly higher in the group receiving antioxidant supplements (0.20±0.50 Diopter (D)) compared to the placebo group (-0.12±0.48 (D)) (p<0.05). Conclusions: antioxidant supplementation was observed to improve accommodation in Japanese Visual Display Terminal (VDT) Users.

An imbalance between free radical generation and radical scavenging antioxidant systems results in oxidative stress, which has been associated with cell injury observed in many age-related diseases. The superoxide dismutase (SOD) family is a major antioxidant system, and deficiency of Cu,Zn-superoxide dismutase-1 (Sod1) in mice leads to many different phenotypes that resemble accelerated aging. In this study we examined the morphologic features and the secretory functions of the lacrimal glands in Sod1 -/- mice. Lacrimal glands showed atrophy of acinar units; fibrosis; infiltration with CD4 + T cells, monocytes, and neutrophils; increased staining with both 4-hydroxy-2-nonenal and 8-hydroxy-2′-deoxyguanosine; increases in apoptotic cells; and the presence of the epithelial-mesenchymal transition in senescent Sod1 -/- mice. Electron microscopy findings revealed evidence of epithelial-mesenchymal transition, presence of swollen and degenerated mitochondria, and the presence of apoptotic cell death in the lacrimal glands of senescent Sod1 -/- mice. These alterations were also associated with the accumulation of secretory vesicles in acinar epithelial cells, decreased production of both stimulated and nonstimulated tears, and a decline in total protein secretion from the lacrimal glands. Our results suggest that Sod1 -/- mice may be a good model system in which to study the mechanism of reactive oxygen species-mediated lacrimal gland alterations. © 2012 American Society for Investigative Pathology.

Purpose: To date, mouse lacrimal gland epithelial cells have been cultured successfully but only in cases involving newborn mouse lacrimal glands. In this work, we attempted to cultivate and characterize adult mouse lacrimal gland epithelial cells. Methods: Lacrimal glands were removed from newborn mice (C57B/6) and isolated lacrimal gland epithelial cells were seeded onto tissue culture treated or low adherent culture dishes in Cnt-07 culture medium with or without cholera toxin. Cultivated cells were characterized by immunostaining with pan-cytokeratin, α-smooth muscle actin, and lactoferrin antibodies. Lacrimal gland cells from 7-week-old green fluorescent protein (GFP) and non-GFP (C57B/6) mice were mixed and seeded onto uncoated dishes to assess sphere-forming efficiency. Cells were also seeded onto 3T3 cell feeder layers to assess colony forming efficiency. Results: Lacrimal gland epithelial cells were selectively cultured with cholera toxin, and cell type was verified by pancytokeratin and α-smooth muscle actin immunostaining. Sphere formation from single cells of adult mice was observed using specific medium and low adherent culture dishes. These cells could also undergo colony formation on 3T3 feeder cells. Conclusions: Adult mouse lacrimal gland epithelial cells were successfully cultivated in cholera toxin-containing medium, and were observed to form spheres from single cells. © 2012 Molecular Vision.

PURPOSE. To investigate the lipid and DNA oxidative stress as well as corneal and retinal effects after ultraviolet B (UV-B) exposure in mice, with or without silicon hydrogel soft contact lenses (SCL). METHODS. Twenty-eight C57BL6-strain male mice were divided into four groups: group I, control group with no SCL (SCL [-]) and no UV-B exposure (UV-B [-]); group II, senofilcon A SCL (senofilcon [+]) with UV-B exposure (UV-B [+]); group III, lotrafilcon A SCL (lotrafilcon [+]) with UV-B exposure (UV-B [+]); and group IV, no SCL (SCL [-]), but with UV-B exposure (UV-B [+]). All mice except group I received UV-B exposure for 5 days for a total dose of 2.73 J/cm 2. All mice underwent tear hexanoyl-lysine (HEL) and tear cytokine ELISA measurements, and fluorescein and rose bengal corneal staining before and after UV-B exposure. Corneal specimens underwent immunohistochemistry staining with CD45, HEL, 4-hydroxynonenal (4-HNE), and 8-hydroxy-20-deoxyguanosine (8-OHdG) antibodies and evaluation with electron microscopy. RESULTS. All mice without SCL but exposed to UV-B developed corneal edema, ulcers, or epithelial damage compared with mice with senofilcon A SCL and exposure to UV-B. Tear HEL and cytokine levels significantly increased in mice without SCL after UV-B exposure. Immunohistochemistry showed a significantly higher number of cells positively stained for CD45, 8-OHdG, HEL, and 4-HNE in the corneas of mice without SCLs compared with those with senofilcon A after UV-B exposure. CONCLUSION. Silicon hydrogel SCL showed corneal and retinal protective effects, owing to UV blocking properties, against oxidative stress-related membrane lipid and cellular DNA damage. © 2012 The Association for Research in Vision and Ophthalmology, Inc.

The ocular surface is a frequent target tissue of mucosal chronic GVHD (cGVHD). We investigated the histopathological features of the conjunctival microvilli in patients with cGVHD. Conjunctival tissue specimens from patients with cGVHD or Sjögren's syndrome (SS) or from healthy individuals were examined by light microscopy and EM, impression cytology, and immunohistochemistry. The cGVHD conjunctivae showed significantly more metaplasia and fewer goblet cells than the SS and normal conjunctivae. Abundant CD8 + T cells infiltrated the basal epithelia in the cGVHD conjunctiva. The microvilli per standard epithelial unit and the secretory vesicles were counted by analyzing electron micrographs. The mean number of mucosal microvilli was significantly lower in the cGVHD than that in the SS or normal specimens, and the microvilli were significantly shorter, with a smaller height-width ratio. The mean number of secretory vesicles was also significantly lower, and the membrane-spanning mucin thinner, in the cGVHD compared with the SS and normal specimens. Thus, the conjunctival mucosal microvilli of cGVHD patients were significantly different in number and morphology from those of SS and normal subjects. These may be important factors affecting the stability of the tear-film layer and its contribution to cGVHD-related dry eye. © 2012 Macmillan Publishers Limited All rights reserved.