Epithelial hyperproliferation and transglutaminase 1 gene expression in Stevens-Johnson syndrome conjunctiva.
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In Stevens-Johnson syndrome, pathological keratinization of the ordinarily nonkeratinized corneal and conjunctival mucosal epithelia results in severe visual loss. We examined conjunctiva covering cornea in five eyes in the chronic cicatricial phase of Stevens-Johnson syndrome. Normal conjunctiva from five age-matched individuals was studied also. The number of epithelial cells in Stevens-Johnson syndrome conjunctiva that were immunoreactive with a monoclonal antibody, Ki-67, to a nuclear antigen found only in proliferating cells was greater than normal (93.8+/-19.8 cells above 100 basal cells versus 12.8+/-0.5 cells above 100 basal cells; P = 0.009). In addition, although clinical inflammation was mild, massive lymphocytic infiltration was seen in the substantia propria of conjunctiva covering cornea. In situ hybridization documented transglutaminase 1 (keratinocyte transglutaminase) mRNA in suprabasal cells of the abnormally thickened conjunctival epithelium in all Stevens-Johnson syndrome patients. In contrast, no message was detected in normal conjunctival or corneal epithelia. Transglutaminase 1 is expressed during the terminal differentiation of keratinocytes where it helps synthesize cornified cell envelopes. We speculate that in Stevens-Johnson syndrome, epithelial hyperproliferation, and transglutaminase 1 gene expression lead to the pathological keratinization of ocular surface mucosal epithelia. (+info)
Ocular ochronosis in alkaptonuria patients carrying mutations in the homogentisate 1,2-dioxygenase gene.
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AIMS: To assess the involvement of the recently identified human homogentisate 1,2-dioxygenase gene (HGO) in alkaptonuria (AKU) in two unrelated patients with ochronosis of the conjunctiva, sclera, and cornea. METHODS: A mutation screen of the entire coding region of the HGO gene was performed using single stranded conformational analysis after polymerase chain reaction with oligonucleotide primers flanking all 14 exons of the HGO gene. Fragments showing aberrant mobility were directly sequenced. RESULTS: Two homozygous missense mutations, L25P and M368V, were identified, each of which leads to the replacement of a highly conserved amino acid in the HGO protein. CONCLUSIONS: The authors describe a novel mutation, L25P, in the German population and bring to 18 the total number of known HGO mutations. (+info)
Combined effect of Interceed and 5-fluorouracil on delayed adjustable strabismus surgery.
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AIMS/BACKGROUND: To discover a more reliable method of performing delayed suture adjustment as a basis to investigate whether delayed adjustment actually provides more stable results. In order to prevent the formation of postoperative adhesions and delay the time of adjustment, an animal study was performed to determine the combined effect of physical barriers, Viscoat and Interceed, and a pharmacological agent, 5-fluorouracil (5-FU). METHODS: 38 rabbit eyes were divided into three groups. After recession of the superior rectus muscle (SRM), 5-FU was applied beneath and over the SRM in group 5-FU. Group I-f had Interceed and 5-FU and group I-fv, Interceed, 5-FU, and Viscoat. Delayed adjustment was performed once on each SRM at 1, 2, and 3 weeks postoperatively. The possible length and the necessary force to adjust as well as the degree of adhesions were recorded. RESULTS: 5-FU delayed the adjustment for up to 1 week after surgery in three out of four eyes. Combined use of Interceed and 5-FU could delay the adjustment for up to 1 week after surgery in three out of five eyes. Addition of Viscoat could delay the adjustment for up to 1 week after surgery in four out of five eyes. Adjustment was possible on only one of four eyes thereafter. CONCLUSIONS: Combined use of Interceed, 5-FU, and Viscoat could delay the adjustment in rabbits until 1 week postoperatively. (+info)
Transforming growth factor-beta1, -beta2, and -beta3 in vivo: effects on normal and mitomycin C-modulated conjunctival scarring.
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PURPOSE: To compare the effects of the three human isoforms of transforming growth factor (TGF)-beta in vivo using a mouse model of conjunctival scarring, both in normal eyes and after treatment with MMC, with a view to delineating the role of this growth factor in glaucoma filtration surgery. METHODS: Application of recombinant human TGF-beta was assessed in a prospective, randomized study of mouse conjunctival scarring, in which subconjunctival TGF-beta1, -beta2, and -beta3 (all 10(-9) M) were compared with control (phosphate-buffered saline [PBS] carrier) and mitomycin C (MMC; 0.4 mg/ml) treatment at 6 hours, and 1, 3, and 7 days after surgery (six eyes/treatment/time point). Effects of TGF-beta2 on eyes previously treated with MMC were also assessed. Histologic studies of enucleated eyes were performed to analyze development of the scarring response, extracellular matrix deposition, and the inflammatory cell profile. RESULTS: All three isoforms of TGF-beta behaved in a similar manner in vivo, being associated with a rapid-onset and exaggerated scarring response compared with control and MMC treatment. TGF-beta-treated eyes showed evidence of an earlier peak in inflammatory cell activity (P < 0.05) and increased collagen type III deposition (P < 0.05). TGF-beta2 treatment significantly stimulated scarring after MMC application (P < 0.05). CONCLUSIONS: TGF-beta1, -beta2, and -beta3 appear to have similar actions in vivo and stimulate the conjunctival scarring response. Application of TGF-beta2 modified the effects of MMC. All TGF-beta isoforms may be potent modulators of the conjunctival scarring response. These studies indicate that TGF-beta2 may naturally modify the antiscarring effects of antimetabolites such as MMC in glaucoma filtration surgery. (+info)
Human anti-transforming growth factor-beta2 antibody: a new glaucoma anti-scarring agent.
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PURPOSE: Currently available anti-scarring regimens for glaucoma filtration surgery have potentially blinding complications and thus the need for alternative and safer agents. The effects of a new antibody to transforming growth factor (TGF)-beta2 on in vitro and in vivo conjunctival scarring and after glaucoma filtration surgery were investigated. METHODS: The activity of a novel recombinant monoclonal neutralizing antibody (mAb) to human TGF-2 (rhAnti-TGF-beta2 mAb) was studied in conjunctival fibroblast-mediated proliferation, migration, and collagen contraction. Its safety in subconjunctival administration was assessed in vivo, and, in a rabbit model of glaucoma filtration surgery, its effects on conjunctival scarring and filtration surgery outcome were investigated. RESULTS: The rhAnti-TGF-beta2 mAb effectively inhibited TGF-beta2-mediated conjunctival scarring activity in vitro, at 50% inhibitory concentrations (IC50) of less than 1 nM. It significantly improved glaucoma filtration surgery outcome in an animal model of aggressive conjunctival scarring compared with control (P = 0.0291) and was clinically safe, nontoxic, and well tolerated after subconjunctival administration. CONCLUSIONS: Subconjunctival rhAnti-TGF-beta2 mAb treatment significantly affects surgical outcome and effectively reduces conjunctival scarring both in vitro and in vivo. It appears safe for subconjunctival administration and when compared with mitomycin-C treatment histologically, much less destructive to local tissue. rhAnti-TGF-beta2 mAb may have potential as a new anti-scarring agent for use in glaucoma filtration surgery. (+info)
Tear fluid gelatinase B activity correlates with IL-1alpha concentration and fluorescein clearance in ocular rosacea.
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PURPOSE: To correlate tear fluorescein clearance with interleukin-1alpha (IL-1alpha) concentration and gelatinase B (matrix metalloproteinase [MMP]-9) activity in the tear fluid of patients with ocular rosacea and normal control subjects. METHODS: Gelatinase activity was evaluated by gelatin zymography in tear fluid obtained from 13 patients with ocular rosacea (including 1 patient with recurrent epithelial erosion, 2 with recurrent peripheral corneal infiltrates and vascularization, and 2 patients with epithelial basement membrane dystrophy) and 13 normal subjects with normal aqueous tear production and no irritation symptoms. Tear fluorescein clearance was evaluated by measuring fluorescence in tear fluid collected from the inferior meniscus 15 minutes after instillation of 5 microl of 2% Na-fluorescein with a CytoFluor II fluorometer. Pro-MMP-9 and IL-1alpha concentrations in the tear fluid were measured by enzyme-linked immunosorbent assay (ELISA). RESULTS: Compared with normal control subjects, patients with ocular rosacea had a greater delay of tear fluorescein clearance (P < 0.001), a higher tear IL-1alpha concentration (P < 0.001), and a greater pro-gelatinase B (92 kDa) activity (P < 0.001) in their tear fluid. The 84-kDa active form of gelatinase B was observed in 46% of the rosacea tear samples and none of the controls. The zymographic results were confirmed by ELISA that showed a significantly greater concentration of pro-MMP-9 (92 kDa) in the tear fluid of rosacea patients than controls. Delayed tear clearance was correlated with elevated tear IL-1alpha concentration (p=0.67, P < 0.001) and increased tear gelatinase B activity (p=0.84, P < 0.001). Tear IL-1alpha concentration was correlated with tear gelatinase B activity (p=0.58, P < 0.002). CONCLUSIONS: Gelatinase B (MMP-9) activity is greater in patients with ocular rosacea than in normal eyes. The majority of this activity is due to 92-kDa proform of this enzyme. This activity is correlated with delayed tear clearance and tear fluid concentration of interleukin-1alpha, a proinflammatory cytokine that has been reported to stimulate gelatinase B production. Elevated gelatinase B activity in ocular rosacea may be involved in the pathogenesis of the irritation symptoms, recurrent epithelial erosions, vascularization, and epithelial basement membrane dystrophy that develops in the corneas of patients with this condition. (+info)
Overexpression of MMP-1 and MMP-3 by cultured conjunctivochalasis fibroblasts.
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PURPOSE: To determine whether conjunctivochalasis, denoting redundant, loose, nonedematous inferior bulbar conjunctiva, is associated with increased expression and activity of matrix metalloproteinases (MMPS) over their tissue inhibitors (TIMPs). METHODS: Expression of transcripts and proteins of MMPs, TIMPs, and urokinase plasminogen activator (uPA) by cultured normal human conjunctival and conjunctivochalasis fibroblasts was determined by Northern hybridization, enzyme-linked immunosorbent assay (ELISA), and Western blot analysis, respectively. Gelatin and casein zymography and quantitative collagenase activity assay were performed in the serum-free conditioned media. RESULTS: Compared with normal conjunctival fibroblasts from six subjects, conjunctivochalasis fibroblasts from eight patients showed markedly increased transcript expression of MMP-1 (5- to 32-fold) and MMP-3 (4 to 30-fold), whereas that of MMP-2, TIMP-1, TIMP-2, and uPA was similar between the two groups. Protein levels were increased in the serum-free conditioned media of conjunctivochalasis fibroblasts for MMP-1 (3.5- to 7.6-fold) and MMP-3 (2.3- to 13-fold), determined by ELISA and Western blot analysis. There was increased caseinolytic activity of MMP-3 and collagenolytic activity of MMP-1 (2.2-fold) by conjunctivochalasis fibroblasts, whereas no difference was noted between these two types of fibroblasts in the protein and gelatinolytic activity of MMP-2 or expression of TIMP-1 and TIMP-2 proteins, although that of TIMP-1 transcript was slightly higher in some conjunctivochalasis fibroblasts. No expression of MMP-9 was detected. CONCLUSIONS: Overexpression of MMP-1 and MMP-3 mRNA by conjunctivochalasis fibroblasts is correlated with their increased protein levels and proteolytic activities. Collectively, these data help explain how conjunctivochalasis manifests excessive degradation of the conjunctival matrix and Tenon's capsule. (+info)
Key factors in the subjective and objective assessment of conjunctival erythema.
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PURPOSE: To establish objectively measurable characteristics of the conjunctival vasculature that correspond with the judgment of erythema by human observers. METHODS: Color images of bulbar conjunctiva from 21 subjects were digitally analyzed to extract the following variables characteristic of the scene: vessel width (W), number of vessels (V), proportion of area occupied by vessels (PA), relative redness both in vessels (RRV) and in the whole image (RRI), red-green difference both in vessels (RGV) and in the whole image (RGI), red-blue difference both in vessels (RBV) and in the whole image (RBI), and red hue value (RHV). These data were compared with subjective judgments by a panel of seven trained observers who independently rated erythema in the same images, using a 0 to 4 scale with decimal interpolation between grades. RESULTS: Correlation analysis indicated significant associations (P<0.05) between the mean response of the human observers and all the objective variables except RHV. Associations with the morphometric variables PA (R2 = 0.93) and V(R2 = 0.90) were markedly stronger than for the best colorimetric variable RBV (R2 = 0.62). CONCLUSIONS: Judgments of erythema made by human observers do not rely primarily on color but can be closely approximated by a univariate, linear model involving only the proportion of the scene occupied by vessels. Under the conditions of this study, grading of erythema by trained observers can be considered to constitute measurement to at least an interval level. (+info)