Tear fluid concentration of mmp-8 is elevated in non-allergic eosinophilic conjunctivitis and correlates with conjunctival inflammatory cell infiltration.

BACKGROUND: To investigate tear fluid concentration of matrix metalloproteinase 8 (MMP-8) and its relation to conjunctival inflammatory cell infiltration in persistent non-allergic eosinophilic conjunctivitis (NAEC). METHODS: Two groups were included: 26 consecutive adult patients with NAEC (conjunctival eosinophils at least 1+ [1-10 eosinophils/slide], skin prick test [SPT] to common allergens negative), and 26 asymptomatic adult persons (no conjunctival eosinophils, SPT negative). MMP-8 tear fluid concentrations were determined by immunofluorometric assay, and conjunctival brush cytology samples from NAEC patients were used for MMP-8 immunocytochemistry. Gelatin zymography was used to illustrate proteolytic activity within the tear fluid samples. RESULTS: The mean MMP-8 concentration was significantly higher among NAEC patients (214.3 +/- 327.7 microg/l) than among healthy persons (50.4 +/- 62.3 microg/l, P < 0.0001). In the NAEC patients, tear fluid MMP-8 correlated with the numbers of conjunctival neutrophils (r = 0.66, P = 0.0002) as well as with goblet cells and columnar epithelial cells (r = 0.54 for both, P = 0.045), but not with the lymphocyte numbers (r = -0.36, P = 0.0741). By immunocytology, MMP-8 protein could also be detected in vivo in the inflammatory cell population within the conjunctiva. Zymography revealed that proteolysis was significantly higher in the NAEC group, and activated enzymes were practically found only in the NAEC group. CONCLUSIONS: The results showed that NAEC is an inflammatory condition characterized by increased tear fluid MMP-8 levels, probably derived from both inflammatory and structural conjunctival cells. The increased proteolytic activity in NAEC patients may indicate risk of conjunctival structural changes (remodeling).

Elevated expression and activation of matrix metalloproteinase 8 in tear fluid in atopic blepharoconjunctivitis.

PURPOSE: Matrix metalloproteinase 8 (MMP-8) is an effective collagenolytic enzyme that is associated with many ocular inflammatory diseases, such as uveitis, keratitis, and ocular rosacea. We studied the tear fluid concentration and activation of MMP-8 in atopic blepharoconjunctivitis (ABC) and the presence of the enzyme in conjunctival inflammatory cells in vivo. METHODS: Tear fluid samples were collected from 26 patients with ABC and 26 healthy controls. MMP-8 concentrations were determined by immunofluorometric assay, and its molecular forms and degrees of activation were studied by Western blotting. Conjunctival brush cytology samples from patients with ABC were used for MMP-8 immunocytochemistry. RESULTS: : The mean MMP-8 concentration was statistically significantly higher among the patients with ABC (545.6 +/- 879.3 microg/L) than among the healthy controls (50.4 +/- 62.3 microg/L, P = 0.0001). There was a statistically significant correlation between neutrophils detected in brush cytology and tear fluid MMP-8 (P = 0.032, r = 0.47). Both the control and ABC tear fluid samples contained predominantly the larger (60-80 kDa), highly glycosylated polymorphonuclear leukocyte-type MMP-8 isoform, as identified by Western blotting, but neither was found to contain the mesenchymal-type isoform. The active enzyme was in practice present only in the ABC samples. Immunostainings show the MMP-8 protein to be present in all the main inflammatory cell types within the conjunctiva. CONCLUSIONS: : A higher mean concentration and activation of MMP-8 is present in tear fluid in ABC. This finding probably reflects persistent inflammatory and collagenolytic activity associated with the disease.

Cellular characteristics of non-allergic eosinophilic conjunctivitis.

PURPOSE: This study examines the histology of conjunctival biopsy samples from patients with persistent allergic eosinophilic conjunctivitis (AEC) or non-allergic eosinophilic conjunctivitis (NAEC). METHODS: Fourteen patients with conjunctivitis and eosinophilia in cytology samples were included in the study. Seven had positive skin-prick tests (the AEC group) and seven had negative skin-prick tests (the NAEC group). Eight asymptomatic subjects with negative skin-prick tests served as a control group. In conjunctival biopsies eosinophils were identified with monoclonal antibodies. Mast cells were identified by specific immunostaining and tryptase-positive granules were counted around them. The percentage of degranulated mast cells was used as a measure of cell activation. Eosinophil and goblet cell numbers were counted, epithelial thickness was measured, and the symptoms were characterized and graded. RESULTS: The numbers of eosinophils in biopsies were higher in patients with AEC than in healthy controls (p = 0.010). The proportion of activated mast cells tended to be higher in AEC patients (65%) than in NAEC patients (48%) or control subjects (40%). Patients with AEC had more goblet cells than control subjects (p = 0.049) and their epithelial layer was thicker (p = 0.054). Patients with AEC had more severe symptoms than control subjects (p = 0.0005), whereas the symptoms of NAEC patients did not differ statistically from those of controls (p = 0.065). CONCLUSIONS: Patients with NAEC were characterized by mild eosinophilic inflammation and only minor structural conjunctival changes. The condition seems to run a relatively mild but persistent clinical course.

Effect of 0.03% tacrolimus ointment on conjunctival cytology in patients with severe atopic blepharoconjunctivitis: a retrospective study.

PURPOSE: To evaluate the efficacy and effect of tacrolimus ointment on conjunctival cytology in patients with atopic blepharoconjunctivitis or keratoconjunctivitis. METHODS: Ten patients with severe atopic blepharoconjunctivitis treated with 0.03% tacrolimus ointment once daily as an intermittent treatment were analysed retrospectively. The main outcome measures were clinical response to topical tacrolimus, adverse events and changes in the inflammatory cells obtained from conjunctival brush samples. RESULTS: Marked clinical responses in blepharitis and conjunctivitis symptoms were seen after a mean follow-up time of 6 weeks. Clinical scores decreased by 67% in blepharitis and 74% in conjunctivitis symptoms. No severe adverse events or signs of immunosuppression such as herpes simplex infections occurred. No significant changes occurred in visual acuity, refraction, anterior chamber, retina or intraocular pressure. Median decreases were 85% (p =0.01) in conjunctival eosinophils, 50% (p = 0.01) in neutrophils and 58% (p = 0.02) in lymphocytes. CONCLUSIONS: Tacrolimus ointment is potentially a safe and effective treatment for atopic blepharoconjunctivitis. Regular treatment of the eyelids once daily may also lead to clinical and cytological improvement of the conjunctivitis.

Tear fluid levels of MMP-8 are elevated in ocular rosacea--treatment effect of oral doxycycline.

BACKGROUND: Ocular rosacea (OcR) is a chronic inflammatory disease especially affecting lid margins. Previous studies have shown that it is accompanied by increased levels and activation of tear fluid gelatinases. Matrix metalloproteinase 8 (MMP-8; collagenase 2) levels and activation are commonly elevated in many inflammatory conditions. Therefore we studied here whether MMP-8 concentration and activation in tear fluid are increased also in OcR, and if an oral doxycycline regimen could rectify the situation. METHODS: Tear fluid samples were collected from 22 OcR patients and 22 healthy controls. The OcR patients were then treated with an oral doxycycline regimen for 8 weeks and tear fluid samples collected again after 4 and 8 weeks. Conjunctival brush cytology and patients' subjective symptoms were scored. MMP-8 concentrations in the tear fluid were assessed by immunofluorometric assay and the molecular forms and isoenzyme expression of MMP-8 were studied by Western immunoblotting. RESULTS: The mean MMP-8 concentration was statistically significantly higher in OcR (156.8+/-207.4 mug/ml) than in the normal subjects (53.5+/-66.7 mug/ml) (P=0.036), but decreased to 79.2+/-141.6 mug/l and 53.6+/-75.2 mug/l after 4 and 8 weeks doxycycline treatment, respectively. There was a statistically significant difference between the untreated OcR and the MMP-8 results after 4 or 8 weeks of oral doxycycline (P=0.041 and 0.069, respectively) and the OcR patients experienced statistically significant relief of their subjective symptoms (P=0.0001) after the doxycycline regimen. Both the normal and OcR tear fluid contained the larger, 60-80 kDa highly- glycosylated polymorphonuclear leukocyte-type MMP-8 isoform in Western immunoblotting, but not the 45-55 kDa less glycosylated mesenchymal-type isoform. MMP-8 activation was in practice present only in the OcR samples, and was inhibited by oral doxycycline. CONCLUSIONS: MMP-8 concentration and activation degree in tear fluid are increased in OcR, probably reflecting increased inflammatory activity. Doxycycline effectively reduces these pathologically excessive levels and activation of MMP-8, and relieves patients' subjective symptoms.

Group IIA phospholipase A(2) concentration of tears in patients with ocular rosacea.

PURPOSE: To determine the concentration of group IIA phospholipase A(2) (GIIAPLA(2)) in tears of patients with ocular rosacea, and to compare it with GIIAPLA(2) concentration in tears of age-matched healthy controls. METHODS: The GIIAPLA(2) concentration in tears was measured with a time-resolved fluoroimmunoassay in 21 patients with ocular rosacea (mean age 55.6+/-9.2 years) and in 21 normal subjects (mean age 53.4+/-8.2 years). Conjunctival brush cytology was carried out and eosinophils, neutrophils, lymphocytes, squamous epithelial cells, columnar epithelial cells, metaplastic changes and goblet cells were calculated separately. RESULTS: The GIIAPLA (2) concentration in tears was statistically significantly lower in patients with ocular rosacea (31.0+/-18.4 microg/ml, p=0.0099) and, more specifically, in patients who had dry eye (25.8+/-15.1 microg/ml, p=0.0034), compared to that in normal controls. There was no correlation between the GIIAPLA (2) content of tears and the conjunctival cells collected by the brush cytology. CONCLUSION: The tears of patients with dry eye symptoms due to ocular rosacea have decreased GIIAPLA (2) content. The pathogenic importance of this finding is discussed.