Morphological alterations in corneal nerves of patients with dry eye and associated biomarkers.

The purposes of the present study were to (1) identify the relationship between dry eye symptoms and morphological changes in corneal subbasal nerves/ocular surfaces, and (2) discover tear film biomarkers indicating morphological changes in the subbasal nerves. This was a prospective cross-sectional study conducted between October and November 2017. Adults with dry eye disease (DED, n = 43) and healthy eyes (n = 16) were evaluated based on their subjective symptoms and ophthalmological findings. Corneal subbasal nerves were observed using confocal laser scanning microscopy. Nerve lengths, densities, branch numbers, and nerve fiber tortuosity were analyzed using ACCMetrics and CCMetrics image analysis systems; tear proteins were quantified by mass spectroscopy. Compared with the control group, the DED group had significantly lower tear breakup times (TBUT) and pain tolerance capacity, and significantly higher corneal nerve branch density (CNBD) and corneal nerve total branch density (CTBD). CNBD and CTBD showed significant negative correlations with TBUT. Six biomarkers (cystatin-S, immunoglobulin kappa constant, neutrophil gelatinase-associated lipocalin, profilin-1, protein S100-A8, and protein S100-A9) showed significant positive correlations with CNBD and CTBD. The significantly higher CNBD and CTBD in the DED group suggests that DED is associated with morphological alterations in corneal nerves. The correlation of TBUT with CNBD and CTBD further supports this inference. Six candidate biomarkers that correlate with morphological changes were identified. Thus, morphological changes in corneal nerves are a hallmark of DED, and confocal microscopy may help in the diagnosis and treatment of dry eyes.

A Novel Lid Hygiene Detergent Successfully Eliminated Demodex Mites, a Common Pathogen of Refractory Obstructive Meibomian Gland Dysfunction.

INTRODUCTION: Demodex mites are microscopic arthropods that have been shown to be responsible for chronic blepharitis. Although many reports have demonstrated positive effects of lid hygiene on demodicosis, some have produced conflicting results. We retrospectively evaluated the effect of lid margin cleansing with a novel lid hygiene detergent, Eye Shampoo, in patients with ocular demodicosis. METHODS: Outpatient clinic notes recorded over a 24-month period between June 30, 2016, and June 29, 2018, at Keio University Hospital in Japan were reviewed to identify patients diagnosed with ocular demodicosis who started cleansing their lid margins with Eye Shampoo at least once daily as a lid hygiene therapy. RESULTS: Five patients diagnosed with ocular demodicosis who started cleansing their lid margins using Eye Shampoo for lid hygiene were identified. In four cases, Demodex mites were successfully eliminated, and ocular conditions were ultimately improved with no recurrence. Active Demodex mites were still noted after 3 months of lid hygiene therapy in one case; further observation of this case was not available. There were no adverse effects of Eye Shampoo in any of these cases. CONCLUSION: Lid hygiene therapy using Eye Shampoo successfully eliminated Demodex mites, a common pathogen of refractory obstructive meibomian gland dysfunction.

Analysis of the Association between Galectin-3 Concentration in Tears and the Severity of Dry Eye Disease: A Case-Control Study.

This study aimed to investigate the relationship between the severity of dry eye disease (DED) and galectin-3 concentration (gal-3) and its cleavage (gal-3C) in tear fluid. Twenty-eight DED patients and 14 controls were recruited at Keio University Hospital. The lissamine green conjunctival staining (LG) score, fluorescein corneal staining (FL) score, tear film break-up time (TBUT), Schirmer's test, and ocular symptoms questionnaire score (dry eye questionnaire score, DEQS) were evaluated. Furthermore, the correlation between these parameters and the concentrations of gal-3 in tears (ng/µg) and the detection rate of gal-3C (%) were analyzed. Gal-3 concentration in tears was positively correlated with the LG score (R = 0.60, p < 0.01), FL score (R = 0.49, p < 0.01), and DEQS (R = 0.45, p < 0.01), and negatively correlated with the TBUT score (R = -0.40, p < 0.01) and Schirmer's I value (R = -0.36, p < 0.01). The detection rate of gal-3C in tears was significantly associated with the severity of DED, especially with the LG (p < 0.01) and FL (p < 0.01) scores. Therefore, the concentration of gal-3 and the detection rate of gal-3C in tears had a significant relationship with the severity of ocular surface barrier disruption.

Conjunctival Epithelial Ingrowth After Penetrating Keratoplasty.

PURPOSE: To report a case of conjunctival epithelial ingrowth after penetrating keratoplasty. METHODS: A 57-year-old woman with herpetic corneal keratitis, endotheliitis, and bullous keratopathy underwent penetrating keratoplasty (PKP) and secondary cataract surgery. One month after cataract surgery, an epithelial ingrowth was observed at the 5 o'clock donor host junction. Ingrowth extended into the anterior chamber and along the iris surface by 9 months. Another PKP was performed, and the excised graft was submitted for histopathology. RESULTS: The graft showed CK13-positive and CK3-negative cells lining the endothelial surface, indicating the conjunctival origin of ingrown epithelium. Ten months postoperatively, no recurrence of ingrowth was observed. CONCLUSIONS: We experienced a rare case of conjunctival epithelial ingrowth after penetrating keratoplasty. There was no recurrence of the ingrowth after surgical removal, and the conjunctival origin may explain the relatively benign course of the complication.

Demodex-Mite Infestation in Cilia and its Association With Ocular Surface Parameters in Japanese Volunteers.

OBJECTIVES: To investigate the presence of Demodex mites on the eyelashes of a Japanese population and to explore its associations with subjective ocular symptoms and clinical ocular surface signs, including lid margin findings and fluorescein breakup time (FBUT). METHODS: Sixty-three Japanese Tokyo-based volunteers were examined (24 men and 39 women; mean±SD of age, 50.6±15.8 years). Eyelash sampling was performed by epilating three lashes from the center of the right upper eyelid. Eyelashes were tested for Demodex mites with a light microscope. The subjects completed questionnaires to assess subjective symptoms and risk factors for dry eye disease (DED) and underwent general examinations for DED, including FBUT and cornea-conjunctival staining with fluorescein. Meibomian gland function was also investigated in lid margin findings, vascularity and mucocutaneous junction movement, lid irregularity, plugging, pouting, and meibum secretion grade scores. RESULTS: Demodex mites were found in 20.6% (13/63) of the volunteers. Lid margin vascularity and meibum grades in the upper eyelid margins were significantly associated with the presence of Demodex mites. However, there were no significant differences in the diagnosis of meibomian gland dysfunction, FBUT, or ocular surface discomfort between the subjects with or without Demodex mites. CONCLUSIONS: Demodex mites are not rare in the cilia of a Japanese population. Lid margin vascularity and lower meibum quality may be associated with the presence of Demodex mites.

The evaluation of dry eye mobile apps for screening of dry eye disease and educational tear event in Japan.

PURPOSE: To evaluate the usefulness of a dry eye mobile application (app) for screening dry eye disease (DED) at educational tear events in Japan. METHODS: In this cross-sectional study, Japanese subjects visiting a "Tears Day" event were selected randomly. They completed questionnaires and underwent ophthalmic evaluations for DED (using Japanese revised diagnostic criteria) including a functional visual acuity (FVA) test. The app calculated FVA using the average of the continuous VA over 30 s. RESULTS: Sixty-three general-population subjects were included: 25 men and 38 women (average age, 50.8 ±â€¯15.9 years). The prevalence of DED was 66.7% (42 subjects); age was significantly higher among subjects with DED (55.2 ±â€¯3.4 vs. 48.1 ±â€¯2.7 years, p = 0.04; men, 54.0 ±â€¯7.3 vs. 47.0 ±â€¯3.0 years, p = 0.36; women, 55.5 ±â€¯3.9 vs. 50.6 ±â€¯3.8 years, p = 0.4). The prevalence of DED was significantly higher in women (p = 0.04). Tear film breakup time was significantly shorter (3.8 ±â€¯2.4 vs. 8.7 ±â€¯2.0, p = 0.04) and the meibum score was significantly higher (p = 0.02) among subjects with DED. Regarding the tear film breakup pattern, line and random breaks were most prevalent among DED. FVA showed a significant negative correlation with DED (r = -0.25, p = 0.047). CONCLUSIONS: The app might motivate people to perform quick tests with the expectation of getting easy DED screening. The number of subjects diagnosed with DED was relatively high.

Functional expression of hexahistidine-tagged beta-subunit of yeast F1-ATPase and isolation of the enzyme by immobilized metal affinity chromatography.

Mitochondrial ATP synthase (F1Fo-ATPase) catalyzes the terminal step of oxidative phosphorylation. In this paper, we demonstrate the functional expression of the hexahistidine-tagged beta-subunit of yeast ATP synthase and the purification of the F1-ATPase from yeast cells. A gene encoding the beta-subunit from Saccharomyces cerevisiae was modified to encode a protein of which the original N-terminus import signal sequence was replaced by a sequence containing the import signal sequence of a mitochondrial ATPase inhibitor, its processing site, and six consecutive histidines. Expression of the modified gene generated a functional F1Fo complex in host yeast cells lacking a functional copy of the endogenous ATP2 gene, as judged by growth of rescued cells on lactate medium. F1 was extracted from the yeast mitochondria by chloroform treatment and purified by immobilized metal affinity chromatography and gel filtration chromatography. The specific activity of the purified F1 was comparable to that of the wild-type enzyme, and the F1 contained all of the 5 known subunits (alpha, beta, gamma, delta, and epsilon). Moreover, the activity of the F1 was completely inhibited by the specific ATPase inhibitor protein, IF1. These results indicate that F1 containing the tagged beta-subunit is fully assembled and active. The application of this novel procedure simplifies the number of steps required for the isolation of F1 used for studying the molecular mechanism of catalysis and regulation of the enzyme.