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Purpose: Strip meniscometry quantitatively measures the volume of tears in the tear meniscus and has been reported to diagnose dry eyes in clinical settings conveniently, easily, and rapidly. In this study, we used a modified strip meniscometry to assess the applicability of measuring the tear volume in mice in experimental settings., Methods: Dry eye was induced in 11 9-week-old C57BL/6J wild-type male mice (11 right eyes) by exposing them to an air fan inside a small compartment for 5 hours for 2 consecutive days. Tear function tests, including the SMTube for mice (SMTM) for tear volume evaluation, break-up time, fluorescein staining score, and lissamine green staining score, were performed. The correlation between SMTM and other tear function parameters was assessed., Results: The mean SMTM value was 3.89 ± 0.603 mm before and 3.09 ± 0.625 mm after dry environment exposure (P = 0.0078*). The Spearman's correlation by rank test showed a strong positive correlation between SMTM and tear film break-up time and a strong linear negative correlation with fluorescein and lissamine green values., Conclusions: The SMTM was capable of rapidly measuring the minimum tear volume in mice and correlated well with tear function parameters and appears to be a promising new modality in the evaluation of dry eyes in mice.

Purpose: To evaluate the morphologic changes in the corneal subbasal nerve (CSN) plexus in wild-type (WT) mice after exposure to scopolamine-induced dry eye stress (DES) by using in vivo confocal microscopy., Methods: Twenty right eyes of twenty (n = 20) 8-week-old WT BALB/c male mice were investigated. The mice were divided into two experimental groups; 10 eyes of 10 mice exposed to DES for 28 days and 10 eyes of 10 mice were used as a control group. All mice underwent examinations for aqueous tear secretion quantity, tear film breakup time (TBUT), corneal vital staining. and corneal sensitivity thrice (pre-experiment, 2nd week, and 4th week). CSN density, tortuosity, reflectivity, and dendritic cell (DC) densities were examined., Results: The mean aqueous tear secretion (P < 0.0001) and TBUTs (P < 0.0001) were significantly decreased after DES. The mean corneal vital staining scores were significantly higher (fluorescein, P < 0.0001; lissamine, P < 0.0001), the mean TBUTs were significantly shorter (P < 0.0001), and the corneal sensitivities (P < 0.0001) were significantly lower in the dry eye-induced mice than the control mice. The mean CSN fiber density (P < 0.0001) and the reflectivity (P < 0.001) were significantly lower; the mean tortuosity and the mean DC density were significantly higher (P < 0.0001) in the dry eye mice., Conclusions: Our data demonstrated that prolonged exposure to DES resulted in alterations of CSN density; DC intensity, reflectivity, and tortuosity as well as in tear volume; TBUT; fluorescein and lissamine green staining scores; and the corneal sensitivity in WT mice.

Reactive oxygen species (ROS) are produced as a by-product during the mitochondrial respiration of the oxygen and potentially able to damage the tissues. Oxidative stress occurs as a result of the disruption of the balance between the anti-oxidant system and the pro-oxidant system found in cells. It has been accepted that overexpression of ROS can be induced in the ocular surface as a result of many acute and chronic diseases and even in normal aging. Recent studies demonstrated that oxidative stress damages the ocular surface and plays an important role in the mechanism of dry eye disease. There is a need to investigate the therapeutic modalities employing topical/systemic use of antioxidants in dry eye disease. This review will summarize the recent studies showing the important relationship between oxidative stress and dry eye disease.

Purpose: To investigate the morphologic changes in the corneal subbasal nerve (CSN) plexus in wild-type mice after exposure to environmental dry eye stress (EDES) using in vivo confocal microscopy (IVCM)., Methods: We examined 22 eyes of 8-week-old wild-type male mice (Balb/c, n = 11). The mice were exposed to an air fan inside a small compartment 5 hours/day for 3 days (EDES). Aqueous tear secretion and corneal epithelial damage were assessed. The CSNs were investigated by laser-scanning IVCM. Density; tortuosity; and reflectivity of CSNs; and dendritic cell (DC) densities were evaluated using semi-automated NeuronJ software., Results: EDES significantly decreased the aqueous tear secretion quantity (P = 0.0019) and significantly increased the corneal fluorescein (P = 0.005) and lissamine green staining scores (P = 0003). The CSN density showed a significant decrease after EDES exposure (before, 2813 ± 762 pixels/frame; after, 1906 ± 896 pixels/frame, P = 0.0071). The tortuosity and the reflectivity grades did not show statistically significant differences after EDES exposure (tortuosity, P = 0.307; reflectivity, P = 0.758). However, the mean DC density showed a significant increase after EDES exposure (before, 12.62 ± 5.94 cells/mm2; after, 15.93 ± 5.30 cells/mm2, P = 0.026)., Conclusions: Even short-term exposure to EDES induced alterations in the CSN plexus morphology including decreased subbasal corneal nerve density and increased amount of DCs in mice. The EDES mouse model is a promising model to study the ocular surface and corneal nerve changes associated with dry eye disease.

Purpose: The superoxide dismutase-1 knockout (Sod1-/-) mouse is an age-related dry eye mouse model. We evaluated the role of 2% rebamipide ophthalmic solution on the conjunctiva and ocular surface alterations in Sod1-/- mice., Methods: Rebamipide eye drops (2%) were instilled in six 50-week-old male Sod1-/- mice and six C57BL/6 strain wild-type (WT) male mice four times a day for 2 weeks. Aqueous tear secretion quantity and tear film breakup time measurements as well as vital stainings were performed. Immunohistochemistry staining of the conjunctiva was performed using SAM pointed domain-containing ETS transcription factor (SPDEF), transglutaminase-1, and involucrin antibodies. Quantitative RT-PCR was carried out to study mRNA expression of the same markers., Results: The mean tear quantities showed no significant changes in both mice strains after treatment (P = 0.24). The mean tear film breakup time (P = 0.003) and vital staining scores significantly improved in the Sod1-/- mice after treatment. Treatment with 2% rebamipide eye drops significantly decreased the corneal fluorescein (P = 0.0093) and Rose Bengal (P = 0.002) staining scores in the Sod1-/- mice. We showed a notable increase in SPDEF and a marked decrease in transglutaminase-1 and involucrin immunohistochemistry stainings, together with a significant increase in SPDEF (P = 0.0003) and a significant decline in transglutaminase-1 (P = 0.0072) and involucrin (P = 0.009) mRNA expression after treatment in the Sod1-/- mice., Conclusions: Topical use of 2% rebamipide drops was observed to improve conjunctival epithelial differentiation and suppress keratinization in the Sod1-/- mice.

Purpose: To evaluate cytokine levels in the aqueous humor (AqH) of eyes with bullous keratopathy (BK) and low endothelial cell density (ECD)., Methods: A total of 145 AqH samples (60 BK, 16 low ECD, 35 corneal diseases with normal ECD, and 34 normal controls) were collected from consecutive patients who underwent corneal transplantation or cataract surgery. None of the patients had any clinically apparent inflammation at the time of AqH collection. The AqH levels of cytokines (IL-1α, IL-1β, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p70, IL-13, IL-17A, IFN-α, IFN-γ, monocyte chemotactic protein [MCP]-1, TNF-α, E-selectin, P-selectin, soluble intercellular adhesion molecule [sICAM]-1, granulocyte-macrophage colony-stimulating factor [GM-CSF], macrophage inflammatory protein [MIP] -1α and MIP-1β) were compared among the groups., Results: The levels of IL-1α, IL-8, IL-17A, TNF-α, GM-CSF, MIP-1α, IFN-γ, and E-selectin in the AqH were significantly elevated in BK and low ECD eyes, compared with healthy controls (all P < 0.03). The levels of IL-4, IL-6, IL-10, IL-12p70, IL-13, MCP-1, MIP-1β, P-selectin, and sICAM-1 were significantly elevated only in BK eyes, compared with healthy control (all P < 0.001). There were no significant differences in AqH cytokine levels between corneal diseases with normal ECD and normal control eyes. Among BK eyes, the IL-6 and IFN-γ levels were elevated in eyes with pseudophakic BK (PBK), Fuchs' endothelial corneal dystrophy (FECD), postkeratoplasty, posttrabeculectomy, and postlaser iridotomy (LI) (all, P < 0.04), whereas IL-1α, IL-10, IL-17A, MIP-1β, and sICAM-1 levels were elevated only in PBK, postkeratoplasty, posttrabeculectomy, and post-LI eyes (all, P < 0.05)., Conclusions: Subclinical elevation of AqH cytokine levels may cause endothelial cell loss.

Purpose: A healthy conjunctiva secreting mucins is essential for maintaining the integrity of the ocular surface epithelium. We used Cu, Zn-superoxide dismutase 1-deficient mice (Sod1-/- mice) and investigated the effect of oxidative stress on the tear function, conjunctival phenotype, and ocular surface mucin expression., Methods: Fifty-week-old C57/B6 wild-type (WT) and Sod1-/- mice were used for evaluations of the tear film breakup time and periodic acid Schiff staining of the conjunctival specimens to detect goblet cell densities in the conjunctiva. Immunohistochemistry stainings with anti-Muc5AC, anti-Muc1, anti-4-hydroxy-2-nonenal, and anti-8-hydroxy-2'-deoxyguanosine antibodies were also performed. The mRNA expression levels of Muc1, Muc5AC, Spdef, involcurin, and transglutaminase 1 were quantified with real-time RT-PCR., Results: The mean goblet cell density in the aged Sod1-/- mice was significantly lower than the aged WT mice. The mean number of Muc5ac-positive cells was significantly lower in the aged Sod1-/- mice compared with the aged WT mice. The conjunctival epithelium in the aged Sod1-/- mice displayed marked staining with lipid and DNA oxidative stress markers. The mRNA expression of transglutaminase 1 and involcurin in the aged Sod1-/- mice was significantly higher than the aged WT mice. The Spdef mRNA expression in the aged Sod1-/- mice was also significantly lower than the aged WT mice., Conclusions: Elevated oxidative stress status appears to affect the conjunctival differentiation and alter the conjunctival epithelial phenotype with aging in the Sod1-/- mice.

Purpose: We determined whether functional visual acuity (VA) parameters and a dry eyes (DEs) symptoms questionnaire could predict DEs in a population of visual terminal display (VDT) users., Methods: This prospective study included 491 VDT users from the Osaka Study. Subjects with definite DE, diagnosed with the presence of DE symptoms, tear abnormality (Schirmer test ≤ 5 mm or tear breakup time [TBUT] ≤ 5 seconds), and conjunctivocorneal epithelial damage (total staining score of ≥3 points), or probable DE, diagnosed with the presence of two of them, were assigned to a DE group, and the remainder to a non-DE group. Functional VA was assessed, and DE questionnaires were administered. We assessed whether univariate and discriminant analyses could determine to which group a subject belonged. Sensitivity and specificity were assessed., Results: Of 491 subjects, 320 and 171 were assigned to the DE and non-DE groups, respectively. No significant differences were observed between DE and non-DE groups in Schirmer test value and epithelial damage, but TBUT value (3.1 ± 1.5 vs. 5.9 ± 3.0 seconds). The sensitivity and specificity of single test using functional VA parameters were 59% and 49% in functional VA, 60% and 50% in visual maintenance ratio, and 83% and 30% in frequency of blinking, respectively. According to a discriminant analysis using a combination of functional VA parameters and a DE questionnaire, six variables were selected for the discriminant equation, of which area under the curve (AUC) was 0.735. Sensitivity and specificity of diagnoses predicted by the discriminant equation were 85.9% and 45.6%, respectively., Conclusions: The discriminant equation obtained using functional VA measurement combined with a symptoms questionnaire may suggest the possibility for the first step screening of DE with unstable tear film. Since the questionnaire has an overall poor sensitivity and specificity, further amelioration may be necessary for the actual utilization of this screening tool., (Copyright 2014 The Association for Research in Vision and Ophthalmology, Inc.)

Purpose: We evaluated the levels of lipid oxidative stress markers and inflammatory cells from tears and conjunctiva of patients with Sjögren syndrome (SS) and normal subjects., Methods: We examined 31 eyes of 16 patients (16 females) with SS and 15 eyes of 10 healthy controls (2 males and 8 females) in this prospective study. All subjects underwent a Schirmer test, measurement of tear film break-up time, vital stainings, confocal microscopy of the conjunctiva, tear collection for hexanoyl-lysine (HEL), ELISA, and conjunctival brush cytology. Brush cytology samples underwent immunohistochemistry (IHC) staining with HEL and 4-hydroxy-2-nonenal (4HNE). Hematoxylin-eosin and IHC staining with HEL and 4HNE also were performed on conjunctival samples of SS patients and controls., Results: The tear stability and vital staining scores were significantly worse in eyes of SS patients compared to the controls. Conjunctival inflammatory cell density was significantly higher in SS subjects compared to controls. The numbers of conjunctival cells stained positively for HEL and 4HNE were significantly higher in SS patients compared to controls. Tear HEL concentrations correlated significantly with staining scores and inflammatory cell density in confocal microscopy. Conjunctival specimens also revealed higher numbers of cells stained positively for inflammatory markers, as well as HEL and 4HNE in the IHC stainings., Conclusions: Increase of the oxidative stress status in the conjunctiva of SS patients appears to have a role in the pathogenesis of dry eye disease. A close relationship may exist between reactive oxygen species (ROS) production, lipid peroxidation related membrane damage, and inflammatory processes in dry eye.

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-2'-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.

Purpose: To investigate the changes in functional visual acuity (VA) and higher order aberrations in dry eye patients., Methods: In this prospective comparative case series, 22 right eyes were classified into those with or without superficial punctate keratopathy (SPK) in the central cornea of 22 patients with Sjögren syndrome; 10 right eyes of 10 normal subjects served as the control. Serial measurements of VAs using a functional VA measurement system and higher order aberrations using a wavefront sensor were performed under blink-free conditions without topical anesthesia over a 10-second period. The parameters for each measurement were compared among the SPK-positive and -negative and normal groups. The correlation between those parameters was also analyzed., Results: Dry eye with SPK showed significant deterioration of visual function and optical quality compared with dry eye without SPK and in normal eyes, as detected by both the visual maintenance ratio (VMR; P < 0.05) and the variation of VA (P < 0.05) and by comalike and total higher order aberrations (P < 0.05). Moreover, the severity of epithelial damage at the central cornea correlated significantly with VMR (P < 0.01) and variation of VA (P < 0.01) as well as comalike (P < 0.05) and total higher order aberrations (P < 0.05). The dry eye group without SPK showed minor visual deterioration compared with normal eyes, as detected only by VMR (P < 0.05)., Conclusions: Optical disturbances at the central optical zone of the cornea in dry eye disease may affect visual performance. Functional VA measurement may be an applicable method of evaluating visual performance in dry eyes that is as efficient as wavefront aberration measurements.

Purpose: To prospectively evaluate the effect of controlled adverse chamber environment (CACE) exposure on tear function, including tear osmolarity, in subjects wearing narafilcon A versus those wearing etafilcon A soft contact lens (SCL)., Methods: Thirty-one healthy subjects with no history of contact lens wear (13 women, 18 men; average age, 30.5 ± 6.5 years) were randomly divided into age- and sex-matched groups (15 subjects wearing narafilcon A SCL; 16 subjects wearing etafilcon A SCL) and entered a CACE for 20 minutes. All subjects underwent tear osmolarity, tear evaporation rate, strip meniscometry, tear film breakup time, fluorescein vital staining, and functional visual acuity measurement before and after exposure to the controlled adverse chamber., Results: The mean blink rate increased with significant deteriorations in the mean symptom VAS scores, mean tear osmolarity, tear evaporation rate, strip meniscometry score, and tear stability with CACE exposure along with a decrease in visual maintenance ratio in functional visual acuity testing in etafilcon A wearers. The mean symptom VAS scores, mean tear evaporation rate, tear stability, blink rates, and visual maintenance ratios did not change significantly in narafilcon A wearers after CACE exposure., Conclusions: This study suggested marked tear instability, higher tear osmolarity, and increased tear evaporation with marked dry eye and visual symptomatology in nonadapted hydrogel SCL wearers, suggesting that silicone hydrogel SCLs may be suitable for persons who live and work in cool, low-humidity, and windy environments, as tested in this study.

Purpose: To evaluate the extent of corneal inflammation and the response to treatment in patients with Mooren's ulcer, by in vivo confocal microscopy (IVCM)., Methods: Twenty-two eyes of 15 patients with Mooren's ulcer were enrolled in this prospective study. All subjects underwent routine ophthalmic examinations, IVCM, and conjunctival histopathologic examination of specimens in patients undergoing conjunctival excision. Eyes with active ulcer were treated with topical corticosteroids and additional therapy, depending on the signs and symptoms. Eyes in remission continued to receive the previous treatment protocols. The relation between the severity of ulcer and inflammation status as assessed by IVCM was also studied. The endpoints were inflammatory cell density (ICD), counted by IVCM, and the extent of the limbal arc involved with ulcers., Results: Ten eyes had active corneal ulcers, and 12 eyes had been in remission for the past year. The mean ICD of eyes with active ulcers before treatment was 2092.7 ± 1538.6 cells/mm(2) (range, 835.3-7832.7; 95% CI, 3232.5-952.9). Nine of the eyes improved at 8 weeks, with a decrease in ICD to 249.1 ± 109.0 cells/mm(2) (range, 100.3-595.3; 95% CI, 329.8-168.3). One eye had corneal perforation, and ICD immediately before perforation was 1677.6 ± 247.6 cells/mm(2). The mean ICD of 12 eyes in remission was 357.5 ± 266.8 cells/mm(2) (range, 12.7-1127.0; 95% CI, 555.2-159.8). The correlation of the ICD and the extent of limbal involvement with ulcers was strong (R(2) = 0.8119)., Conclusions: ICD evaluated by IVCM is a useful and promising parameter for evaluation of the degree of inflammation in eyes with Mooren's ulcer and for assessment of response to treatment.

Purpose: To evaluate the specificity and sensitivity of strip meniscometry (SM) testing in conjunction with tear function tests in the diagnosis of dry eye (DE) disease and to investigate the effect of SM on reflex tearing., Methods: One hundred seven left eyes of 107 patients with definite DE disease according to the Japanese DE diagnostic criteria and 68 left eyes of 68 age- and sex-matched control subjects were studied. Tear meniscus height (TMH) measurements, fluorescein tear film break-up time (FTBUT), fluorescein (F) and rose bengal (RB) staining, and Schirmer's test-1 (ST) were also performed. The assessment of reflex tearing before and after SM application was assessed with a graticule scale at the slit lamp and by optical coherence tomography. The sensitivity and specificity of SM alone and in combination with tear function tests were also sought., Results: The SM scores, TMH measurements, FTBUTs, and STs were significantly lower in dry eye patients than in the controls (P < 0.001). The RB and F staining scores were significantly higher in the dry eye group than in the control group (P < 0.001). The meniscometry strips did not induce significant changes in relation to reflex tearing. SM had an acceptable sensitivity and specificity., Conclusions: SM is a swift, noninvasive, promising method of assessing tear meniscus volume. The combined SM and FTBUT examination appears to be a sensitive approach to the assessment of dry eye disease.

Purpose: To investigate the applicability of in vivo laser scanning confocal microscopy in the diagnosis and follow-up of ocular demodicosis infestation in a prospective controlled study., Methods: Fifteen right eyes of 15 patients with blepharitis associated with cylindrical dandruff (10 males, 5 females; mean age: 62.9 ± 9 years) and eight right eyes of eight age- and sex-matched control subjects underwent HRTII/RCM, evaluation of ocular symptom scores, tear function tests including vital stainings, Schirmer test, and tear clearance test, and evaluation of mite numbers in the eyelids., Results: In vivo confocal microscopy effectively disclosed the mites in the terminal bulbs of the eyelashes, which were not observed after treatment. Eyelids with demodicosis infestation showed marked inflammatory infiltrates around the meibomian glands and conjunctiva, which cleared with tea tree oil treatment., Conclusions: Laser scanning confocal microscopy seems to be an efficient noninvasive tool in the diagnosis and follow-up of ocular demodicosis infestation.

Purpose: To investigate the cytologic findings of superior bulbar conjunctiva in superior limbic keratoconjunctivitis (SLK) using laser scanning confocal microscopy and impression cytology in a prospective controlled study., Methods: Twenty-one eyes of 11 SLK patients (9 women, 2 men; mean age, 49.3 +/- 17.9 years) and 18 eyes of 9 control subjects (6 women, 3 men; mean age, 46.4 +/- 8.7 years) underwent tear function tests including vital stainings, Schirmer test, tear clearance test, digital confocal laser scanning microscopy, and conjunctival impression cytology. After confocal microscopy and impression cytology images were obtained, the mean individual epithelial cell area (MIECA), nucleocytoplasmic (N/C) ratio, and inflammatory cell density were analyzed. The correlation between confocal microscopy and impression cytology parameters was investigated., Results: The MIECA of SLK patients and control subjects in confocal microscopy was 786.54 +/- 463.88 microm(2) and 311.50 +/- 78.30 microm(2), respectively. The mean N/C ratio was 0.356 +/- 0.090 and 0.490 +/- 0.038, respectively. The MIECA and N/C ratio in impression cytology showed significant correlation with the corresponding confocal microscopy parameters (MIECA, P = 0.0028; N/C, P = 0.0051). The inflammatory cell density in confocal microscopy significantly correlated with superior bulbar conjunctival Rose-Bengal scores (P = 0.0264)., Conclusions: Laser scanning confocal microscopy seems to be an efficient noninvasive tool in the evaluation of phenotypic alterations of the conjunctival epithelium in SLK and may serve as an alternative for impression cytology. N/C ratio and inflammatory cell density appear to be two new promising parameters of in vivo confocal microscopy in the assessment of ocular surface disease in SLK.

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 hexanoyl-lysine (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-1beta and IL-6 levels showed a significant correlation with corneal epithelial damage. IL-1beta and TNFalpha 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.

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.

Purpose: To investigate the applicability and efficacy of a new and simple method of quantification of the volume of tear meniscus, termed "strip meniscometry," in the diagnosis of the dry eye syndromes in a prospective controlled study., Methods: One hundred eyes of 50 patients with dry eye (19 males; 31 females) aged between 18 and 76 years (mean, 54.3 years), as well as 80 eyes of 40 normal subjects aged from 15 to 70 years (mean, 50.8 years; 12 males, 28 females) were recruited in this study. The patients and the control subjects underwent strip meniscometry for 5 seconds, tear film lipid layer interferometry, tear film break-up time measurement, and ocular surface vital staining with fluorescein and rose bengal dyes and the Schirmer-1 test., Results: Strip meniscometry scores correlated with tear quantity and stability, ocular surface staining scores, and lipid layer interferometry grades and improved after 2 weeks of punctal plug occlusion., Conclusions: Strip meniscometry is a swift, noninvasive, promising new method that is expected to find application in the diagnosis and evaluation of the outcome of treatment of dry eye syndromes.

Purpose: Tryptase that is released by mast cell degranulation has recently been thought to play a key role in wound healing in allergic bronchitis. Conjunctival fibroblasts secrete mediators and extracellular matrices that could exacerbate inflammation and papillary formation in allergic conjunctivitis. This study was conducted to investigate the effect of tryptase on the proliferation of conjunctival fibroblasts and studied whether this effect was mediated by protease-activated receptor (PAR)-2., Methods: Conjunctival fibroblasts were cultured with or without tryptase (0.1 ng/mL to 1.0 microg/mL), and the proliferation rate was assessed after 48 hours. The effects of tryptase inhibitors (leupeptin, benzamidine) and a PAR-2 agonist (SLIGKV) were examined. The existence of PAR-2 mRNA and protein in conjunctival fibroblasts was examined by RT-PCR and Western blot analysis, respectively. The existence of PAR-2 in cultured conjunctival fibroblasts and conjunctival papillae from patients with vernal keratoconjunctivitis, as well as conjunctival tissue from normal subjects was examined by immunohistochemistry., Results: Conjunctival fibroblast proliferation was upregulated by tryptase in a dose-dependent manner (P < 0.001). Leupeptin and benzamidine inhibited tryptase-induced fibroblast proliferation (P < 0.05), and SLIGKV mimicked tryptase's effect. PAR-2 mRNA and protein were detected in cultured conjunctival fibroblasts using RT-PCR and Western blot analysis. PAR-2 immunoreactivity in both the cultured conjunctival fibroblasts and in stromal cells in excised conjunctival tissues was observed., Conclusions: Tryptase increased conjunctival fibroblast proliferation and this response appeared to be mediated by PAR-2. Mast cells are the most likely source of tryptase in the conjunctiva and may play an important role in chronic exacerbations with conjunctival papillary formation in allergic conjunctivitis.

Objective: The present study investigated the expression of ICAM-1 and VCAM-1 on fibroblasts with interleukin (IL)-4 and/or tumor necrosis factor (TNF)-alpha stimulation and assessed the effect of eosinophil adhesion on fibroblast viability., Methods: Primary cultured human corneal fibroblasts were incubated with IL-4, TNF-alpha, or their combination for 24 hours. Expression of ICAM-1 and VCAM-1 was examined by real-time quantitative PCR and flow cytometric analysis. Purified eosinophils were cocultured with activated fibroblasts, and the number of eosinophils adhered to fibroblasts and the number of damaged fibroblasts were counted using microscopy. In a separate trial, conjunctival and corneal impression cytology was performed on patients with atopic keratoconjunctivitis and corneal ulcers (eight eyes) to assess the status of the ocular surface epithelium and the presence of inflammatory cell infiltrates., Results: Real-time quantitative PCR and flow cytometric analysis revealed that both mRNA and protein of VCAM-1 and ICAM-1 were upregulated by IL-4 and TNF-alpha. IL-5-primed eosinophils adhered to the corneal fibroblasts treated with IL-4 and TNF-alpha, and the fibroblasts were damaged by eosinophil adherence. Anti-ICAM-1 antibody and anti-VCAM-1 antibody inhibited the eosinophil adherence to fibroblasts and the fibroblast damage. Impression cytology revealed extensive infiltration of neutrophil and eosinophils among isolated ocular surface epithelial cells with advanced squamous metaplasia., Conclusions: Corneal fibroblasts expressed ICAM-1 and VCAM-1 when activated with IL-4 and TNF-alpha. Eosinophils can adhere to the activated fibroblasts and can induce subsequent fibroblast damage through these adhesion molecules. Eosinophil adhesion to fibroblasts may possibly contribute to the pathogenesis of severe persistent allergic corneal ulcers.

Purpose: This was a prospective case-control study conducted to evaluate the effectiveness of the Tear Stability Analysis System (TSAS) for the assessment of patients with dry eye., Methods: The TSAS can take 10 consecutive corneal topograms at one per second for 10 seconds. Examinations using the TSAS were conducted in 26 eyes of 26 healthy control subjects and in 27 eyes of 27 patients with dry eye. Examinations were also conducted in 14 eyes of 14 patients before and after the insertion of punctum plugs. Surface regularity and asymmetry indices (SRI, SAI), as well as new tear stability regularity and asymmetry indices (TSRI, TSAI), derived from SRI and SAI, were analyzed., Results: The mean SRI and SAI in dry eyes were significantly greater than in control eyes (P < 0.05). The time-wise change of SRI and SAI was significantly different between dry eyes and control eyes (P < 0.05). TSRI and TSAI in dry eyes were also significantly greater than in control eyes. Punctum plug insertion was associated with a significant decrease in SRI and SAI (P < 0.05)., Conclusions: TSAS was effective in objectively assessing the tear stability in patients with dry eye. This system may be useful in noninvasive diagnosis of dry eye and evaluation of treatment effects.

Purpose: To synthesize an interference color chart for the specific tear lipid layer interference camera, DR-1, for the conversion of the tear film lipid layer thickness into color graphic information--that is, for the quantification of the tear interference image--by a colorimetric approach., Methods: Because the color of the tear lipid layer interference image is visualized by a white light source interference phenomena, to produce gamma-corrected red, green, and blue (RGB) values of a specific interference color at a certain tear lipid film thickness, XYZ tristimulus values of the Commission Internationale de l'Eclairage (CIE) were obtained. XYZ tristimulus values were calculated from the light source spectrum of the DR-1 camera, the color-matching function of CIE, and the reflectance of the tear interference image in wavelengths ranging from 380 to 780 nm. These calculated interference colors were synthesized ranging from 0 nm to 1000 nm of lipid film thickness to produce a color chart. The applicability of the new color chart in the analysis of the lipid layer thickness was tested on a healthy control subject with normal tear function and a patient with dry eye who had aqueous tear deficiency and meibomian gland obstruction., Results: The specific tear interference color chart for the DR-1 camera was obtained with RGB and XYZ tristimulus values. The interference chart ranged from 0 to approximately the 5th interference order. The interference colors from clinical DR-1 images could be converted to lipid thickness data by using the color chart system., Conclusions: A new tear interference color chart was developed in this study, which may be of benefit in converting tear interference color information to data describing the thickness of the tear film lipid layer.