A novel instrument to determine hue discrimination.

PURPOSE: We report and verify a novel hue discrimination instrument. We also investigate its efficiency to determine hue discrimination in persons with normal color vision. STUDY DESIGN: Experimental and clinical investigation. METHOD AND STUDY PARTICIPANTS: The instrument setup comprises an optical unit and examination unit. The optical unit is composed of the same 2 spectrometers and their controllers, which enables the independent emission of different spectral lights. Two independent bundle fibers connect the optical unit and the examination unit. Two different wavelength lights are illuminated on the bipartite upper and lower circular objectives with a visual angle of 2 degrees in the examination unit. The examinee recognizes the difference in the spectral lights between the bipartite targets. Persons with normal color vision are examined and the findings are confirmed using the Ishihara Test for Colour Deficiency. RESULTS: The instrument could generate spectral light from 450 to 650 nm within 2-nm accuracy. The spectral light showed a different light intensity according to the spectral centroid, ranging from 450 to 650 nm, but the difference could be adjusted and was negligible in terms of determination of hue discrimination using the power meter. Three width slits, 0.2 mm, 0.5 mm, and 1.0 mm, to homogenize the light path were investigated. The half-width wavelength was accurate on each spectral centroid; however, the 0.5 mm slit was suitable to generate an efficient light path. The hue discrimination differed among the study participants. In general, at short and long wavelength lights, the hue discrimination range was large: about 15 nm at 450 nm and about 10 nm at 650 nm. Between 470 and 620 nm, the hue discrimination showed good sensitivity and specificity between 8 and 2 nm depending on the targeting wavelength lights. Intraindividual variation was small, ranging from 3 to 1 nm, thus indicating good repeatability. The time to examine the hue discrimination was about 20 min. CONCLUSION: This newly invented instrument using two independent spectrometer units enabled the determination of hue discrimination. The instrument's sensitivity and specificity including its repeatability were confirmed and indicated that the instrument could be a clinically applicable method.

Laser flare-cell photometer: principle and significance in clinical and basic ophthalmology.

A slit-lamp examination is an indispensable and essential clinical evaluation method in ophthalmology, but, it is qualitative subjective. To complement its weaknesses in making a quantitative evaluation of flare intensity and number of cells in the aqueous humor in the eye, we invented the laser flare-cell photometer in 1988. The instrument enables a non-invasive quantitative evaluation of flare intensity and number of cells in the aqueous with good accuracy and repeatability as well as maneuverability equal to slit-lamp microscopy. The instrument can elucidate the pathophysiology in the blood-aqueous barrier (BAB) function in a variety of ocular disorders. The accuracy of the instrument makes it possible to investigate not only the pathophysiology of intraocular disorders but also the effects of various drugs and surgical procedures in BAB. The instrument does not only lighten the burden on patients in clinical examinations and study but it also helps minimize the sacrifice of experimental animals and improves the reliability of the results by minimizing inter-individual variations through its good repeatability. Here I shall relate how the instrument has been applied to clinical and basic studies in ophthalmology and what novel knowledge its application contributed to pathophysiology in ophthalmology.

A diagnostic method for herpes simplex keratitis by simultaneous measurement of viral DNA and virus-specific secretory IgA in tears: an evaluation.

PURPOSE: We performed simultaneous measurement of herpes simplex virus (HSV) DNA by real-time polymerase chain reaction (real-time PCR) and of HSV-specific secretory IgA antibody (HSV-sIgA) by enzyme-linked immunosorbent assay (ELISA) in tears obtained using Schirmer strips in order to investigate its diagnostic efficacy for herpes simplex keratitis (HSK). METHODS: A total of 59 affected eyes from 59 patients with clinically suspected HSK (HSK group) and 23 eyes from 23 healthy volunteers (control group) were enrolled in this study. The HSK group was divided into five subgroups: dendritic/geographic keratitis, disciform keratitis, necrotizing keratitis, atypical keratitis, and others. The tear samples were taken using Schirmer strips to determine the HSV DNA and HSV-sIgA levels. RESULTS: The overall sensitivity and specificity were 55.8 and 100 % for HSV DNA and 49.2 and 82.6 % for HSV-sIgA. The HSV DNA levels in the disciform keratitis subgroup (median, 3.1 × 10(2) copies/sample) were significantly lower than those in the dendritic/geographic keratitis subgroup (median, 2.3 × 10(4) copies/sample) (P < 0.05, Mann-Whitney test). The HSV-sIgA levels in the disciform keratitis subgroup (median, 50.0 NU/ml) were significantly higher than those in the control group (median, 18.0 NU/ml) (P < 0.05, Steel test). The positive and negative predictive values obtained by simultaneous measurement of HSV DNA and sIgA were 90.9 and 61.3 %, respectively. CONCLUSION: The combination of laboratory detection of HSV DNA by real-time PCR and of HSV-sIgA by ELISA using tear samples enables higher reliability in diagnosing the subgroups of HSK, although the HSV DNA value is relatively lower in disciform HSK than in dendritic/geographic HSK.

[Evaluation of dectin-1 and BAFF expression in conjunctival epithelial cells].

PURPOSE: To investigate the expression of dectin-1 protein in conjunctival epithelial cells and the expression of dectin-1 and B-cell activating factor belonging to the tumor necrosis factor family (BAFF) mRNA in in vivo conjunctival epithelial cells (CECs) and in vitro cultured CECs, and its difference in topographical change and etiology of disorders. SUBJECTS AND METHODS: 1. Investigation of dectin-1 and BAFF expression by cytodiagnosis of CECs. The subjects were 12 eyes of 12 healthy volunteers (control group), 6 eyes of 6 patients with Sjögren syndrome (Sjögren group) and 10 eyes of 10 patients with vernal keratoconjuctivitis (VKC group). CECs were sampled by impression cytology using nitrocellulose membrane. The expression of dectin-1 in CECs was detected by immunofluorescence and the quantitative determination of dectin-1 mRNA and BAFF mRNA expression was performed by real-time polymerase chain reaction(real-time PCR). 2. Investigation of dectin-1 and BAFF expression using cultured CECs. Cultured CECs which were divided into an OK-432 addition group (addition concentrations: 0.02, 0.1, 0.5KU/mL), a lipopolysaccharide (LPS) addition group (addition concentrations : 80, 160, 320 microg/mL) and an additive-free group were cultured. Quantitative determination of dectin-1 mRNA and BAFF mRNA expression in cultured CECs was performed by real-time PCR. RESULTS: 1. Investigation of dectin-1 and BAFF expression by cytodiagnosis of CECs. In the control group, there was no significant topographical difference in the expression of dectin-1 and the amount of dectin-1 mRNA among superior, inferior tarsal conjunctiva and temporal bulbar conjunctiva. The levels of dectin-1 mRNA expression were 1.5 (0.1-4.0) [median value (range)] for the control group, 2.6 (1.1-4.8) for the Sjögren group and 3.6 (1.7-16.6) for the VKC group. The VKC group showed a significantly higher level of dectin-1 mRNA than the control group (p < 0.01, Kruskal-Walles H-test). The levels of BAFF mRNA expression were 2.8 (0.2-13.8) [median value (range)] for the control group, 6.3 (2.1-15.1) for the Sjögren group and 11.2 (3.5-70.8) for the VKC group. The VKC group showed a significantly higher level of dectin-1 mRNA than the control group (p < 0.01, Kruskal-Walles H-test). Moreover, regarding the relationship between expression level of dectin-1 mRNA and that of BAFF mRNA in all the subjects, there was a significant correlation between them (r = 0.75, p < 0.001, Spearman's rank coefficient). The levels of dectin-1 mRNA expression in the moderate and severe VKC group 9.2 (2.6-16.6) [median value (range)] were significantly higher than those in mild VKC group 2.8 (1.7-3.8) (p < 0.05, Mann-Whitney U-test). The levels of BAFF mRNA expression in the severe and moderate VKC groups 17.4 (9.1-70.8) [median value (range)] were significantly higher than those in the mild VKC group 4.3 (3.5-11.2) (p < 0.05, Mann-Whitney U-test). 2. Investigation of dectin-1 and BAFF expression by cultured CECs. In the OK-432 addition group, the expression levels of dectin-1 mRNA were increased dose-dependently due to the OK-432 stimulation (p < 0.05, Kruskal-Wallis H-test). Moreover, regarding the relationship between the expression level of dectin-1 mRNA and that of BAFF mRNA in all the cultured conjunctival epithelial cells stimulated by OK-432, there was a significant correlation between them (r = 0.85, p < 0.005, Spearman's rank coefficient). CONCLUSIONS: We concluded that dectin-1 expression in CECs was demonstrated, and expression of both dectin-1 and BAFF in CECs is thought to be involved in pathologic aggravation of allergic inflammatory in patients with VKC.

[Follow-up study on patients with vernal keratoconjunctivitis undergoing topical 0.1% tacrolimus treatment].

PURPOSE: A retrospective study for evaluating the clinical course of patients with vernal keratoconjunctivitis (VKC) treated with topical tacrolimus ophthalmic suspension 0.1% (Tacrolimus). SUBJECTS AND METHODS: Subjects were 30 patients (24 men and 6 women) with VKC who were treated with a combined therapy of Tacrolimus and antiallergic ophthalmic solution, and could be followed up for six months. The subjects were divided into two groups: 1. A conversion treatment group in which Tacrolimus was substituted for a steroid ophthalmic solution [21 patients; average age 14.7 +/- 9.44 years (mean +/- SD)] and 2. An additional treatment group receiving Tacrolimus and anti-allergic ophthalmic solution [9 patients; average age 28.2 +/- 7.31 years (mean +/- SD)]. The therapeutic effects of the patients were evaluated chronologically using the ocular clinical score according to the papillae-limbus-cornea grading score and eosinophil cationic protein (ECP) levels in tears. RESULTS: Papillae-limbus-cornea grading scores were significantly decreased from 8 (median) points at instillation initiation to 5 points at the first month after initiation of Tacrolimus treatment (p < 0.01, Steel test). Tear ECP levels were significantly decreased from 3493.6 (median) ng/ml at instillation initiation to 205.6 ng/ml at the first month after initiation of Tacrolimus treatment (p < 0.05, Steel test). During the course, four cases of exacerbation were found among the 30 cases, but no infections of the anterior segment were found. CONCLUSION: The therapeutic effect of Tacrolimus eye drops for vernal keratoconjunctivitis was remarkable at one month after instillation initiation. For evaluating the effect of treatment and diagnosing exacerbation in VKC treated with Tacrolimus, a follow-up examination using clinical indexes such as the papillae-limbus-cornea grading score and ECP levels in tears is beneficial.

[Evaluation of chemokines in tears of patients with infectious keratitis].

PURPOSE: To investigate the chemokine profile in tears of patients with infectious keratitis. SUBJECTS AND METHODS: Subjects were 32 eyes of 16 patients with infectious keratitis and 5 eyes of 5 healthy volunteers as a control. The patients with infectious keratitis were classified into two groups of eyes: 10 with bacterial keratitis and 6 with Acanthamoeba keratitis. Tear fluid was obtained from both eyes of the patients with infectious keratitis and from the right eyes of the control subjects using filter paper. Chemokine concentration (unit: Odu/mm2) and its profile in tears was analyzed using an antibody-array. RESULTS: In terms of chemokine profile in the bacterial keratitis group, the expression volume of interleukin-8 (IL-8) and monocyte chemoattractant protein-1 (MCP-1) in the diseased eyes was significantly higher than in the healthy eyes (p < 0.05). The expression volume of mucosae-associated epithelial chemokines (MECs) in the diseased eyes of the bacterial keratitis group was significantly lower than in the healthy eyes of that group (p < 0.05). In the Acanthamoeba keratitis group, chemokines were not significantly increased in the diseased eyes compared with those in the healthy eyes. However, MCP-1 was increased in tears of the Acanthamoeba keratitis group. Regarding the chemokine ratio, the IL-8/MEC ratio in the diseased eyes of the Pseudomonas keratitis group and the MCP-1/IL-8 in the diseased eyes of the Acanthamoeba keratitis group showed a significantly high level (p < 0.05). CONCLUSION: We concluded that the analyses of the chemokine profile and chemokine ratio in the tears of infectious keratitis patients is useful as a clinical tear laboratory test to interpret the pathologic condition of infectious keratitis

[Evaluation of anti-Pseudomonas aeruginosa specific secretory IgA antibodies in tears of patients with Pseudomonas aeruginosa keratitis].

PURPOSE: To investigate the role of specific secretary IgA antibodies on lipopolysaccharide (LPS-sIgA) and exotoxin A (ETA-sIgA) in tear fluid produced by immunoresponse to P. aeruginosa. SUBJECTS AND METHODS: Subjects were divided into 3 groups; 41 eyes of 41 normal volunteers without a history of using contact lenses (CL) as controls, 32 eyes of 32 CL users without adverse events as a healthy CL wearer group (CL group), and 12 eyes of 12 patients with CL-related corneal ulcers from which P. aeruginosa was isolated (ulcer group). Tear fluid was obtained using filter paper, and the concentration of LPS-sIgA and ETA-sIgA in samples was determined using enzyme-linked immunosorbent assay (ELISA). RESULTS: The lower limit of concentration for LPS-sIgA and ETA-sIgA was 1.24 and 1.24 (U/mL), respectively. The positive eyes which exceeded this lower limit in LPS-sIgA were 17 of 22 eyes in the control group (median concentration 2.00U/ml, range 1.28 to 7.20), 15 of 25 eyes in the CL group (median 1.76 U/ml, range 1.24 to 6.92) and 1 of 12 eyes in the ulcer group (1.80 U/ml). The number of eyes which exceeded the lower limit in ETA-sIgA was 36 of 41 eyes in the control group (median concentration 6.56 U/ml, range 1.36 to 182), 28 of 32 eyes in the CL group (median 5.40 U/ml, range 1.56 to 29.20) and 5 of 12 eyes in the ulcer group (median 1.72 U/ml, range 1.40 to 2.16). There was no significant difference in LPS-sIgA and ETA-sIgA between the control group and the CL group, but they were significantly lower in the ulcer group than in the control group (p < 0.01, Steel's test). CONCLUSION: Tear fluid in normal, healthy CL wearers contains LPS-sIgA and ETA-sIgA which acts as a barrier to P. aeruginosa infectious keratitis.

Metalloproteinases in corneal diseases: degradation and processing.

Matrix metalloproteinases (MMPs) are zinc-dependent endopeptidases with the potential to degrade all types of extracellular matrix. The ADAM (a disintegrin and metalloproteinase) family of peptidases was recently identified as cleaving the extracellular domain of transmembrane proteins. This was termed ectodomain shedding. We investigated the MMP expression in patients with corneal diseases and the potential role of ADAMs in corneal pathophysiology. We detected upregulation of the active form of MMP-2 and MMP-9 in the tear fluid from patients with corneal melting or recurrent corneal erosion. Using human corneal epithelial cells, we observed ADAM17-dependent ectodomain shedding of soluble tumor necrosis factor receptor 1 and soluble interleukin-6 (IL-6) receptor (sIL-6R). The production of sIL-6R was also induced by messenger RNA splicing in the human corneal epithelial cells. IL-6/sIL-6R-induced signal transducer and activator of transcription 3 phosphorylation was observed in cultured human corneal fibroblasts, suggesting that IL-6 trans-signaling induced inflammatory cellular signaling in the human corneal fibroblasts. We demonstrated that MMPs are significantly upregulated in collagen-destructive disorders of the cornea. Additionally, we observed that ectodomain shedding by ADAMs in corneal epithelial cells mediated the production of soluble cytokine receptors. Trans-signaling of IL-6 can induce an inflammatory response in corneal stroma, indicating the significance of IL-6 trans-signaling in ocular surface inflammation. Thus, MMPs and ADAMs play an important role in the pathophysiology of corneal diseases.

Anti-inflammatory effect of IL-6 receptor blockade in corneal alkali burn.

We investigated the effect of soluble IL-6R (sIL-6R) blockade on corneal inflammation. Topical instillation of either anti-IL-6R antibody (MR16-1) or phosphate buffered saline (PBS) was applied after wounding BALB/c mice corneas with alkali burn. The vascularized area was significantly reduced in the MR16-1 group. The immunoreactivity of phosphorylated STAT3, Gr-1, and F4/80 diminished significantly in the MR16-1 group. Laser capture microdissection resulted in a significant down-regulation of the mRNA expressions of ICAM-1, MCP-1, and VEGF-A in the corneal stroma of the MR16-1 group. Adding a combination of recombinant IL-6 and sIL-6R resulted in a significant increase in the release of VEGF from human corneal fibroblasts. As the infiltration of inflammatory cells, the expression of phosphorylated STAT3, and the expressions of inflammatory-related molecules in the experimental model of corneal inflammation were significantly inhibited by topical instillation of MR16-1, we deduce that IL-6 trans-signaling plays a significant role in ocular surface inflammation and that the blockade of IL-6R contributes to the reduction in corneal inflammation.

Evaluation of eosinophilic inflammation in a novel murine atopic keratoconjunctivitis model induced by crude Dermatophagoides farinae antigen.

BACKGROUND: The purpose of this study is to conduct a histopathological research of the conjunctival findings and eosinophilic inflammation of novel atopic keratoconjunctivitis in a NC/Nga mouse model using crude Dermatophagoides farina. METHODS: NC/Nga mice were sensitized by repeated topical applications of an ointment containing Dermatophagoides farinae body (Dfb). They were then divided into 4 groups depending on the following topical ophthalmic treatment: DFb group undergoing topical ophthalmic ointment containing Dfb; DFco group undergoing topical instillation of allergen extracts of Dermatophagoides farinae; Ba group undergoing topical ointment with substrate alone and NT group without after-topical ophthalmic treatment. At 24 hours after the last ophthalmic treatment, histopathological examination was performed. The density of the subepithelial infiltration of the eosinophils was determined. Serum total IgE and house-dust-mite (HDM)-specific IgE antibody concentrations were determined by enzyme-linked immunosorbent assay (ELISA). RESULTS: In the DFb group, the conjunctiva showed similar findings to those of atopic keratoconjunctivitis, i.e. intraepithelial pseudotubular formation, Torus-form infiltration due to massive lymphocytes in the palpebral conjunctiva and gelatinous hyperplasia in the limbus with subepithelial granuloma composed of lymphocytes and eosinophils. Subepithelial infiltration of eosinophil density in the DFb group [878.4 ± 399.7cells/mm2 (mean ± SD)] was significantly higher than in the other 2 groups (DFco 85.6 ± 40.1 Ba 49.2 ± 32.3) (P < 0.001). Total serum IgE concentration and HDM-specific serum IgE antibody concentration in the DFb group and the DFco group were significantly higher compared with those in the NT group. CONCLUSIONS: Topical application of an ointment containing DFb to both the skin and eyes of NC/Nga mice can induce an atopic keratoconjunctivitis (AKC) model in these mice.

Differential contributions of impaired corneal sensitivity and reduced tear secretion to corneal epithelial disorders.

BACKGROUND/AIMS: To determine the possible roles of impaired corneal sensitivity and reduced tear secretion in various types of corneal epithelial disorders. METHODS: A total of 99 patients (179 eyes) with corneal epithelial disorders classified as persistent epithelial defects (PED), corneal erosion, or superficial punctate keratopathy (SPK) and 115 individuals (230 eyes) without apparent ocular surface disorders (controls) were enrolled in a prospective study. Corneal sensitivity was measured with a Cochet-Bonnet esthesiometer, and tear secretion was measured by the Schirmer test in each subject. RESULTS: Corneal sensitivity of eyes in the PED and corneal erosion groups was significantly lower than that in the control group. Schirmer test values for eyes in the SPK group were significantly reduced compared with those in the control group. CONCLUSION: A loss of corneal sensitivity may contribute to the development of PED and corneal erosion, whereas reduced tear secretion may be a contributing factor for SPK. Both results indicate the importance of corneal sensory innervation to the maintenance of corneal integrity.

Transconjunctival immunotherapy using cholera toxin B to treat experimental allergic conjunctivitis in a mouse model.

PURPOSE: We evaluated the use of immunological biomarkers in transconjunctival immunotherapy by using cholera toxin B for treating experimental allergic conjunctivitis in a mouse model. METHODS: Balb/c mice were sensitized using intraperitoneal injections of ovalbumin (OVA) and were then divided into two groups. The first group was treated by topical instillation of OVA after the instillation of combined OVA and cholera toxin B (CTB) solution B group). The second group was treated by topical instillation of OVA alone (allergy group). The control group consisted of nonsensitized mice undergoing topical OVA instillation only. The numbers of eosinophils and CD4-positive lymphocytes in the conjunctiva were determined histologically, and the observation of immunoglobulin A (IgA)-positive cells in the conjunctiva was performed by immunohistochemistry. Cytokine concentration in the conjunctiva was determined by the protein-array methods. Messenger RNA expression of T-cell-specific markers, such as T-bet, GATA-3, and Foxp3, in the conjunctiva was detected by reversed transcriptase polymerase chain reaction. RESULTS: The number of eosinophils and CD4-positive lymphocytes increased significantly in the allergy group compared with the control group (P < 0.001) but showed no difference between the CTB group and control group. Concentrations of interleukin 4 (IL-4) (P < 0.05), B-lymphocyte chemoattractant (P < 0.01), and thymus-expressed chemokine (P < 0.05) in the conjunctiva were significantly higher in the CTB group than in the other two groups. GATA-3 messenger RNA (mRNA) in the conjunctiva was expressed in the three groups, but T-bet and Foxp3 were not detected. CONCLUSION: Transconjunctival immunotherapy using CTB can be evaluated by histological examination of eosinophils and CD4-positive T cells, and a mucosal immunity-associated chemokine and a helper T-cell-17-associated chemokine as biomarkers.

Regulation of soluble interleukin-6 (IL-6) receptor release from corneal epithelial cells and its role in the ocular surface.

PURPOSE: Interleukin (IL)-6 signaling through its soluble receptor (sIL-6R) (IL-6 trans-signaling) plays an important role in various inflammatory states. We investigated production of sIL-6R in the corneal epithelium and examined the role of IL-6 trans-signaling in the cornea. METHODS: In-vitro experiments were performed using SV40-transformed human corneal epithelial cells (HCEC) and primary human corneal fibroblasts (HCF, keratocytes). Ectodomain shedding in HCEC was stimulated by adding phorbol myristate acetate (PMA, 3 µM: ) both with and without ectodomain shedding inhibition using TNF-α processing inhibitor-1 (TAPI-1, 250 ng/mL), and the concentration of sIL-6R in the culture medium was determined by enzyme-linked immunosorbent assay (ELISA). Expression of differential sIL-6R mRNA splicing (DS-sIL-6R) in HCEC was examined by using reverse transcription (RT)-PCR. The recombinant IL-6 or combination of recombinant IL-6/sIL-6R was added to HCF culture medium and phosphorylation of STAT3 was analyzed by Luminex assay. Tear fluid from patients with Sjögren syndrome was collected and analyzed by ELISA for sIL-6R concentration. RESULTS: In HCEC culture medium, sIL-6R release was increased significantly (P < 0.01) by adding PMA and this increased release of sIL-6R was inhibited significantly by adding TAPI-1, indicating the participation of ectodomain shedding in sIL-6R production. In RT-PCR, DS-sIL-6R expression was noted in HCEC. IL-6/sIL-6R-induced STAT3 phosphorylation was recognized in cultured HCF, suggesting IL-6 trans-signaling induced inflammatory cellular signaling in HCF. In the tear fluid of the patients with Sjögren syndrome, sIL-6R expression was up-regulated (Sjögren syndrome; 2.38 ± 0.98 ng/mL, normal control; 0.16 ± 0.34 ng/mL). CONCLUSIONS: Production of sIL-6R was induced by both ectodomain shedding and mRNA splicing in the corneal epithelium. IL-6 trans-signaling can induce an inflammatory response in corneal fibroblasts. The up-regulation of sIL-6R in inflamed ocular surfaces suggests a pivotal role of sIL-6R at the ocular surface.

[Development of non-invasive clinical examination methods for the anterior segment of the eye and their clinical significance].

1. PURPOSE: Slit-lamp microscopy is a principal ophthalmic clinical method, because it provides microscopic findings of the anterior segment of the eye noninvasively. Its findings, however, are qualitative and there are large inter-observer variations in their evaluation. Furthermore, slit-lamp microscopy provides morphological findings, but a functional evaluation is difficult. We developed two novel methods that establish a qualitative methodology of the slit-lamp microscope and the pathophysiology of the anterior segment of the eye. One is the flare-cell photometer to evaluate flare and cells in the aqueous humor of the eye and the other is an immunohistochemical examination method using tear fluid to evaluate ocular surface disorders. The comprehensive evaluation of these studies is herein overviewed. 2. INNOVATION OF THE FLARE-CELL PHOTOMETER AND ITS CLINICAL SIGNIFICANCE: The breakdown of the blood-aqueous barrier (BAB) causes an increase in protein (flare) and leakage of blood cells (cell) into the aqueous humor of the eye and the severity of BAB breakdown has a positive correlation with the intensity of flare and cells. The flare and cells in the aqueous can be observed qualitatively by slit-lamp microscopy. These findings are primarily distinguished in optics by light scattering. Therefore, detection of the intensity of light scattering due to flare and cells can evaluate the BAB function. The flare-cell photometer comprises 3 novel components: a laser beam system as an incident light, a photomultiplier to detect scattered light intensity and a computer-assisted system to operate the whole system and analyze detected scattered light signals due to flare and cells. The instrument enables us to quantitatively analyze the flare and cells non-invasively and accurately with a wide dynamic measurement range, resulting in a repeated examination of each individual case. It also enables the evaluation of inflammation in the aqueous not only postoperatively but also in endogenous uveitis, evaluation of the effects of anti-inflammatory drugs on BAB and evaluation of aqueous humor dynamics. Furthermore, repeating the examination can minimize inter-individual variations and reduce the number of animals in animal experiments. 3. PATHOPHYSIOLOGICAL EVALUATION METHODS OF OCULAR SURFACE USING TEAR FLUID: Sampling of tears can be performed noninvasively, but the obtainable volume is limited. Therefore, a determination of targeting biomarkers and a development of their micro-volume analysis methods play a crucial role in pathophysiological studies of the ocular surface. Targeting biomarkers should be determined according to the various specified bioactive substances such as eosinophil cationic protein (ECP), cytokines and others. A number of microvolume analysis methods, such as chemiluminescent enzyme immunoassay, immunochromatography, micro-array system and polymerase chain reaction method are used. Objective disorders in the studies include allergic conjunctivitis and infectious diseases such as herpetic keratitis. Quantitative evaluation methods for ECP concentration, antigen-specific secretory IgA in allergic diseases and herpetic keratitis, herpes simplex virus-DNA and cytokine and chemokine profile in tear fluid sampled by filter paper method were investigated. We developed a clinically applicable quantitative immunochemical method for ECP concentration in tear fluid. The results revealed that tear fluid analysis using the above mentioned methods is a clinically useful to investigate the pathophysiology of the ocular surface. 4. CONCLUSION: Laser flare-cell photometer and tear fluid analysis are potent clinical quantitative methods to investigate the pathophysiology of the anterior segment of the eye.

Demographic study of expulsive hemorrhages in 3 patients with infectious keratitis.

PURPOSE: To investigate the demography, causative bacteria, and clinical findings in 3 patients with expulsive hemorrhage. METHODS: The clinical records of 3 patients (3 eyes), who developed expulsive hemorrhage because of infectious keratitis and were treated at our hospital between December 2006 and January 2008, were investigated retrospectively. RESULTS: Three women, older than 70 years, with physical and mental disabilities because of senile dementia were studied. Two were residents at a nursing home. Basic corneal disorders included bullous keratopathy, cicatricial syphilitic keratitis, and traumatic keratitis because of a foreign body. All patients developed expulsive hemorrhage. Two patients underwent enucleation, and 1 underwent bulbar exenteration with sclerocorneal patch for expulsive hemorrhage. Bacterial culture in these cases isolated either Capnocytophaga sp. and penicillin-intermediate resistant Streptococcus pneumoniae or methicillin-resistant Staphylococcus aureus. CONCLUSIONS: This is the first recorded case of Capnocytophaga keratitis in Japan. Patients with dementia may develop severe ocular complications after infectious keratitis because of their inability to communicate.

[Allergy related factors in tears of patients with vernal keratoconjunctivitis undergoing topical 0.1% tacrolimus treatment].

PURPOSE: To investigate, using tear fluid analysis, the effects of topical 0.1% tacrolims therapy on the pathophysiology of vernal keratoconjunctivitis (VKC). SUBJECTS AND METHODS: Subjects were 6 eyes of 6 patients with VKC who underwent topical 0.1% tacrolims treatment twice a day and 5 eyes of 5 healthy volunteers as a control. Using the filter paper method, the tear fluid of the patients was sampled both before and 4 +/- 2 weeks after the treatment and once from the control subjects. Eosinophil cationic protein (ECP) in the tears was examined by the chemiimmunoluminescent enzyme immunoassay method and the chemokine profile of the tears was analyzed using an antibody-array. RESULTS: In terms of the chemokine profile, growth related oncogene (GRO) -alpha, eotaxin-2 and thymus and activation-regulated chemokine (TARC) in the VKC were elevated compared with those in the controls, but they decreased significantly after the treatment (p<0.05). ECP concentrations in the tears were 3092 +/- 1658 ng/ml (average +/- S. D.) for the pretreatment base-line and 464 +/- 775 for the posttreatment. ECP values for the pre-treatment time were statistically significantly higher than those for the post-treatment in 5 patients (p<0.05). CONCLUSION: Topical tacrolims treatment of VKC can suppress allergic inflammation associated chemokines such as eotaxin-2 and TARC.