The structure of the mite allergen Blo t 1 explains the limited antibody cross-reactivity to Der p 1.

The Blomia tropicalis (Blo t) mite species is considered a storage mite in temperate climate zones and an important source of indoor allergens causing allergic asthma and rhinitis in tropical and subtropical regions. Here, we report the crystal structure of one of the allergens from Blo t, recombinant proBlo t 1 (rproBlo t 1), determined at 2.1 Å resolution. Overall, the fold of rproBlo t 1 is characteristic for the pro-form of cysteine proteases from the C1A class. Structural comparison of experimentally mapped Der f 1/Der p1 IgG epitopes to the same surface patch on Blo t 1, as well as of sequence identity of surface-exposed residues, suggests limited cross-reactivity between these allergens and Blo t 1. This is in agreement with ELISA inhibition results showing that, although cross-reactive human IgE epitopes exist, there are unique IgE epitopes for both Blo t 1 and Der p 1.

Early onset wheeze associated with enhanced combined IL-1ß, IL-6, and IL-12/IL-23p40 in LPS-stimulated cord blood mononuclear cells.

BACKGROUND: Neonates with a family history of atopy are at higher risk for developing wheezing in early life. OBJECTIVE: From a birth cohort of at risk infants (first-degree family with atopic disease), we evaluated the influence of distinct intrinsic immunologic risk factors on wheezing disorders in the first 2 years of life. METHODS: Cord blood samples were collected from 195 eligible subjects of a birth cohort of 253 subjects. The subjects studied were those who developed wheezing (n=34) or eczema (n=29) in the first 2 years of life, and 65 healthy control infants. At the time of thawing the viability of the cells were median 70% (range 67.5%-72.5%). Cytokines from lipopolysaccharide (LPS)-stimulated mononuclear cells were analysed using fluorescent-activated cell sorting-array and their profiles were evaluated using factor analysis. RESULTS: Infants with wheeze were significantly associated with enhanced combined LPS stimulated IL-1ß, IL-6, and IL-12/IL-23p40 compared with healthy controls (P=0.003). This profile was also associated with the increased risk for wheeze at 2 years of age (OR=2.45; 95% CI=1.50-3.93, P=0.001). LPS-stimulated cytokine IL-8 was also significantly higher in the wheeze group compared with healthy controls and eczema (P=0.003). Intracellular staining showed that monocytes are main producers of IL-6 and IL-8 from cord blood mononuclear cells. Most of the subjects were non-atopic with 3/34 (9%) wheeze and 9/29 (31%) eczema subjects sensitized to the common dietary or inhalant allergens. CONCLUSION AND CLINICAL RELEVANCE: In infants at genetic risk of atopy, wheeze but not eczema in the first 2 years of life is associated with intrinsic hyperresponsive innate cytokine responses which might predispose infants to wheeze development. Distinct pre-symptomatic hyperresponsive innate immune responses risk factors were found to be associated with early onset wheeze disorders, but not eczema.

Cornulin, a marker of late epidermal differentiation, is down-regulated in eczema.

BACKGROUND: Eczema is a common chronic inflammatory skin disorder which shows strong genetic predisposition. To identify new potential molecular determinants of the disease pathogenesis, we performed a gene expression study in an eczema mouse model. This analysis identified a marked down regulation of the cornulin gene (CRNN), a member of the epidermal differentiation complex, in the eczema-like skin. We then investigated CRNN as an eczema candidate gene and studied its polymorphism and the expression in the skin of eczema patients. METHODS: An eczema-like phenotype was induced in mice by allergen (Der p2) patching. Gene expression analysis was performed with the subtractive suppression hybridization method and validated by real time PCR and the transmission disequilibrium test was used to test for genetic associations in 406 multiplex eczema families. RESULTS: Der p 2 patched mice developed a localized eczema and a Th 2 skewed systemic response. Real time PCR analysis confirmed a down regulation of CRNN mRNA in eczema-like skin in the mouse model and in human eczema. The CRNN polymorphism rs941934 was significantly associated with atopic eczema in the genetic analysis (P = 0.006), though only as part of an extended haplotype including a known associated variant (2282del4) in the filaggrin gene. CONCLUSIONS: CRNN mRNA expression is decreased in eczematous skin. Further studies are needed to verify whether the associated cornulin polymorphism contribute to the genetic susceptibility in eczema.

Corneal tattoo with tea infusion.

Except in animal models of cataractogenesis, the literature on the effects of tea infusion on ocular tissue is scant. In our patient, prolonged exposure to tea infusion may have led to a hypesthetic cornea with secondary limbal stem cell loss. In turn, the eye developed keratinization and corneal neovascularization.

Characterization of glutathione S-transferase from dust mite, Der p 8 and its immunoglobulin E cross-reactivity with cockroach glutathione S-transferase.

BACKGROUND: Sensitization to mite and cockroach allergens is common, and diagnosis and therapy of allergy can be further complicated by the presence of allergen isoforms and panallergens. Purified recombinant and native allergens are useful for studies to resolve such problems. OBJECTIVE: To assess the allergenicity of native and recombinant mite glutathione S-transferase (GST) (Der p 8) and study the IgE cross-reactivity between Der p 8 and cockroach GST. METHODS: Der p 8 cDNA encoding a new isoform was isolated and expressed in yeast. Native Der p 8 was affinity purified from mite extract. IgE reactivity to native and recombinant Der p 8 was assessed by ELISA using sera from allergic subjects from Taiwan, Singapore and Malaysia. IgE cross-reactivity between Der p 8 and cockroach GST was examined by IgE inhibition assays. RESULTS: Our Der p 8 cDNA encoded a basic isoform (pI=8.5) containing six polymorphic residues located at positions 46, 106, 149, 160, 167 and 184. At least 8 isoforms of native Der p 8 were detected by two-dimensionalgel and immunoblot analyses. Sera from Taiwanese asthmatics showed 96% and 84% IgE reactivity to native Der p 8 and recombinant Der p 8, respectively. Native Der p 8 showed 75% and 65% IgE reactivity with sera from Malaysia and Singapore, respectively. CONCLUSIONS: A high frequency of sensitization to mite GST among allergic subjects was observed but the titres of IgE reactivity were low. The IgE cross-reactivity between mite and cockroach GST suggests that GST is a panallergen.

Production of monoclonal antibodies for immunoaffinity purification and quantitation of Blo t 1 allergen in mite and dust extracts.

BACKGROUND: Blo t 1 is a cysteine protease-like allergen from Blomia tropicalis. Recombinant Blo t 1 binds up to 90% of IgE from allergic patients and shows limited cross-reactivity to Der p 1. The generation of monoclonal antibodies (mAbs) against Blo t 1 is important for the detection, isolation and characterization of the native form of the allergen. METHODS: Mice were immunized intramuscularly with naked plasmid DNA encoding Blo t 1 gene with in vivo electroporation and boosted intraperitoneally with recombinant Blo t 1. mAbs against Blo t 1 were generated using a methylcellulose-based hybridoma cloning kit. The native Blo t 1 was isolated by mAb affinity purification and its allergenicity was determined by ELISA. A two-site ELISA for Blo t 1 was developed using the mAbs generated. RESULTS: A DNA-based immunization protocol induced high titre Blo t 1-specific antibodies in mice. Six stable hybridoma clones secreting mAbs recognizing the native and recombinant Blo t 1 were generated. The native Blo t 1 was affinity-purified from a B. tropicalis extract and its allergenicity was determined at 63% using a panel of Singaporean and Malaysian mite allergic patients' sera. A two-site ELISA was developed, which showed a detection limit of 10 ng/mL of Blot t 1. CONCLUSION: Six Blo t 1 mAbs were successfully generated by DNA immunization. These mAbs are useful for nBlo t 1 immunoaffinity isolation and quantitative immunoassays for Blo t 1 in mite and environmental dust extracts.

Sensitization profiles of Malaysian and Singaporean subjects to allergens from Dermatophagoides pteronyssinus and Blomia tropicalis.

BACKGROUND: The house dust mites Dermatophagoides pteronyssinus (Der p) and Blomia tropicalis (Blo t) are the most common house dust mite species in Southeast Asia. To date, there have only been a few studies on the sensitization profile of the general populations in Southeast Asia to house dust mites. The aim of this study was to determine the profiles of Der p and Blo t sensitization among Singaporean and Malaysian subjects. METHODS: Enzyme-linked immunosorbent assay was used to detect specific IgE to Der p and Blo t mite crude extracts as well as purified Der p 1, Der p 2 and Blo t 5 allergens. Sera used were from 229 Singaporean subjects (124 with rhinitis, 105 without rhinitis) and 143 Malaysian subjects (94 adults and 49 children with asthma). RESULTS: The sensitization profile of rhinitis subjects to the dust mite allergens used in this study was as follows: Blo t extract positive: 91/124 (73%); Blo t 5 positive: 62/124 (50%); Der p extract positive: 61/124 (49%); Der p 1 positive: 53/124 (43%); Der p 2 positive: 45/124 (36%). The nonrhinitis subjects' sensitization profile was as follows: Blo t extract positive: 60/105 (57%); Blo t 5 positive: 24/105 (23%); Der p extract positive: 38/105 (36%); Der p 1 positive: 14/105 (13%); Der p 2 positive: 17/105 (16%). The study of Malaysian asthmatic adults showed that 39% of them were sensitized to Der p 1, 32% to Der p 2 and 37% to Blo t 5. Among the asthmatic children, sensitization to Blo t 5, Der p 1 and Der p 2 was 90, 57 and 39%, respectively. CONCLUSION: This study clearly revealed that dual sensitization to B. tropicalis and D. pteronyssinus is common in the general populations of Singapore and Malaysia. Sensitization to Blo t 5 is more prevalent than to Der p 1 and Der p 2.

Lack of human IgE cross-reactivity between mite allergens Blo t 1 and Der p 1.

BACKGROUND: The group 1 mite allergens are the most significant indoor allergens and they belong to the papain-like cysteine protease family. To date there is only one published report on the isolation and characterization of group 1 allergens from Blomia tropicalis mites. The aims of the study are to determine the cross-reactivity between group 1 allergens and to evaluate their clinical importance in allergic patients. METHODS: The full-length Blo t 1 gene was obtained by SMART RACE cDNA amplification method using gene-specific primers. The sequence alignment was performed using LOOK followed by three-dimensional homology modeling. The cDNA was expressed in Pichia pastoris as a secretory protein. Identification of native Blo t 1 in crude mite and spent mite medium extracts was done by Western immunoblot using monoclonal antibody. Allergenicity of recombinant Blo t 1 and native Der p 1 was examined by human IgE ELISA with 80 asthmatic sera. RESULTS: The cDNA sequence consists of 1105 base pairs, including 5'- and 3'-untranslating regions, encoding an open reading frame of 330 amino acid residues. The predicted molecular weight of the deduced protein was approximately 38 kDa. Blo t 1 shared 53 and 34% nucleotide and amino acid, respectively, sequence homology with Der p 1. Native Blo t 1 was detected in both crude mite and spent mite medium extracts, and its estimated molecular weight was about 26 kDa. The recombinant Blo t 1 reacted positively with IgE in 90 and 65% of sera from asthmatic children and adults, respectively, indicating that it is a major allergen. The correlation of human IgE reactivity between Blo t 1 and Der p 1 was low in these sera. CONCLUSION: The full-length cDNA encoding group 1 Blomia tropicalis mite allergen (designated as Blo t 1) has been characterized and expressed from local mites in Singapore. This fecal allergen showed high frequency of human IgE reactivity with asthmatic sera in the tropics and there was a low correlation of IgE reactivity between Blo t 1 and Der p 1.

Generation of monoclonal antibodies against Blo t 3 using DNA immunization with in vivo electroporation.

BACKGROUND: House dust mite allergy is closely associated with allergic diseases. Blomia tropicalis mite species is an important clinical species in the tropics. The cDNA clone encoding Blo t 3, a group 3 allergen from B. tropicalis, has been isolated in our laboratory. OBJECTIVE: This study was designed to generate Blo t 3-specific monoclonal antibodies (mAbs) for the detection, characterization and purification of this allergen. METHODS: Mice were immunized intramuscularly with naked plasmid DNA encoding Blo t 3 gene with in vivo electroporation. Hybridomas were generated by the fusion of the splenocytes to X63-Ag8.653 myeloma cells. Purified native Blo t 3 was obtained by mAb immuno-affinity purification and the allergenicity of native Blo t 3 was determined by human IgE enzyme-linked immunosorbent assay (ELISA). RESULTS: A panel of class-switched and high-affinity mAb recognizing a wide spectrum of Blo t 3 epitopes have been generated. These mAbs are useful for western immunoblot assay, sandwich ELISA and affinity purification of native Blo t 3. Allergenicity of native Blo t 3 protein was examined with 44 mite-allergic sera and approximately 57% of the tested sera had positive serum IgE reactivity to the native Blo t 3. CONCLUSIONS: These results demonstrated that intramuscular injection of naked DNA encoding Blo t 3 gene combined with in vivo electroporation is an effective and simple method to raise monoclonal antibodies that can be used for characterization and purification of Blo t 3.

Flap haze after epithelial debridement and flap hydration for treatment of post-laser in situ keratomileusis striae.

PURPOSE: To report a case of corneal haze after attempts to eliminate post-laser in situ keratomileusis (LASIK) lamellar striae. METHODS: Case report. RESULTS: A 24-year-old woman with visually significant flap striae 2 months after LASIK underwent manual epithelial debridement and flap hydration, refloating, and stretching to eliminate the striae. Three weeks after this intervention, she developed visually significant haze that was confined to the stroma of the flap. The haze slowly improved with use of a topical steroid. CONCLUSION: Stromal haze can develop after treatment of flap striae with epithelial debridement and hypo-osmolar irrigation. We speculate that these maneuvers may have induced cell death of anterior keratocytes and led to haze formation, as can occur after simple epithelial debridement and epithelial scrape-photorefractive keratectomy.

Aspergillus fumigatus keratitis after laser in situ keratomileusis.

PURPOSE: To report a case of Aspergillus fumigatus keratitis after a laser in situ keratomileusis (LASIK) enhancement procedure. METHOD: Case report. RESULTS: A 56-year-old woman developed an ulcer in the flap 13 days after LASIK enhancement. A 4-week course of fortified antibiotics for a presumed bacterial infection followed. The ulcer progressed, causing 60% thinning of the corneal stroma. A biopsy was performed 5 weeks after onset of symptoms, and antifungal agents were initiated. Cultures showed A. fumigatus. Her cornea perforated after the biopsy, requiring cyanoacrylate and lamellar overlay sutures, but the infiltrate resolved on antifungal agents. CONCLUSION: This report is the first description of Aspergillus keratitis after LASIK. We hypothesize that the infection became established on the stromal bed during surgery and led to melting, anteriorly through the flap and posteriorly through the stroma. Diagnosis was made by a corneal biopsy and inoculation of a wide array of media. This case demonstrates the need to consider atypical organisms, including fungi, in the differential diagnosis of post-LASIK infections when there is no response to therapy and highlights the role of corneal biopsy and flap lifting in the diagnosis of this condition.

Ophthalmic manifestations of neonatal onset multisystem inflammatory disease.

PURPOSE: To report the ophthalmic manifestations of neonatal onset multisystem inflammatory disease, a recently recognized, rare systemic disorder characterized by the triad of arthropathy, rash, and abnormal central nervous system development. METHOD: Case report. RESULTS: A 2-year-old female with neonatal onset multisystem inflammatory disease presented with visual acuity of fix and follow with each eye, bilateral optic nerve head pallor and gliosis, as well as marked sheathing of the peripapillary vessels. No vitreous inflammation or macular edema was found. Visual acuity was stable from the neonatal period through the 3 months of follow-up after the changes involving the optic nerve heads and peripapillary vessels were observed (a total of 33 months). CONCLUSIONS: This report provides a description of the ocular manifestations of neonatal onset multisystem inflammatory disease and supports earlier suggestions that swelling of the optic nerve heads can occur. In this case, optic nerve head pallor may have been a sequela of such swelling. The pathogenesis of neonatal onset multisystem inflammatory disease is unknown.

Subretinal fibrosis in patients with Vogt-Koyanagi-Harada disease.

OBJECTIVE: To describe subretinal fibrosis as a long-term complication of Vogt-Koyanagi-Harada (VKH) disease. DESIGN: Retrospective, clinic-based, cross-sectional study and clinical correlation. PARTICIPANTS: Ten patients with VKH disease and subretinal fibrosis were seen at two uveitis referral centers between 1977 and 1997. INTERVENTION: A review of the historical, clinical, and fluorescein angiographic features was performed. MAIN OUTCOME MEASURES: The prevalence, demographic and clinical features, and time to development of subretinal fibrosis were summarized. RESULTS: Subretinal fibrosis occurred in 20 eyes of 10 patients with VKH disease, an overall prevalence of 8% between the two institutions. Patient age ranged from 16 years to 48 years, with a median of 34.5 years. Five patients were Hispanic, one was mixed Hispanic and American Indian, three were Asian or mixed Asian and Caucasian, and one was African-American. Eight of the 10 patients were men. All patients were in the chronic, recurrent phase of their disease when they had subretinal fibrosis develop, and all patients had recurrent episodes of posterior uveitis. Presenting vision ranged from 20/20 to light perception, with a median acuity of 20/200. All patients were initially treated with oral and topical corticosteroids. Four patients required additional noncorticosteroid immunosuppressive therapy. Time from diagnosis of VKH disease to development of subretinal fibrosis ranged from zero (fibrosis present at time of diagnosis) to 27 years, with a median time of 10 months. The median time from diagnosis of VKH to development of subretinal fibrosis in Hispanic patients was 6.5 months, whereas in non-Hispanic patients it was 6.5 years. Final vision ranged from 20/25 to light perception, with a median acuity of 20/60. Seven of 20 eyes had a final visual acuity of 20/40 or better, and seven eyes saw 20/200 or worse. Five of the eyes with 20/200 or worse vision had fibrosis involving the fovea. CONCLUSIONS: Subretinal fibrosis occurs in a sizeable proportion of patients with VKH disease and may contribute to permanent loss of vision.

Ocular neovascularization in patients with uveitis.

Neovascularization is an infrequent but serious complication of uveitis. The retina and optic disk appear to be affected most often, although new blood vessels may arise from the iris, ciliary body, and choroid as well. Although neovascularization can usually be identified on careful clinical examination, some patients may require fluorescein angiography or UBM. Numerous neovascular growth and inhibitory factors have been identified experimentally. Clinically, however, uveitic neovascularization appears to be determined most directly by the severity of the inflammation and the presence of retinal nonperfusion. Virtually all patients with uveitic neovascularization deserve a trial of local or systemic corticosteroids. Laser photocoagulation can be considered in those patients who fail to respond to corticosteroid therapy, but only when retinal nonperfusion has been demonstrated on fluorescein angiography. Surgical excision of newly formed vessels is reserved for selected patients with CNV and uveitis, but should only be considered when corticosteroids and focal photocoagulation are ineffective or are otherwise contraindicated.

Protein sequence analysis and mapping of IgE and IgG epitopes of an allergenic 98-kDa Dermatophagoides farinae paramyosin, Der f 11.

BACKGROUND: A 98-kDa mite paramyosin (Der f 11) from Dermatophagoides farinae (Df) is highly allergenic, and its cDNA (Df642) has been cloned. This paper describes the sequence characteristics and the mapping of the immunodominant human IgE and IgG epitopes of Der f 11. METHODS: The protein sequence analysis was performed with a combination of FASTA, GCG, and CLUSTAL W computing packages. The whole cDNA insert and its PCR-derived DNA fragments were generated and expressed in E. coli. These overlapping recombinant peptides (F1 to F5) were used for B-cell epitope mapping with 18 mite-allergic sera by dot immunoassays. RESULTS: Df642 cDNA encodes a partial sequence that contains the 2nd to 26th 28-residue repeats and lacks the N-terminus and the C-terminus. The sequence identity of Der f 11 with other known paramyosins is 34-60%. The dominant IgE epitopes are located in peptides F1 and F4, whereas the dominant IgG epitopes are located in peptides F1 and F2. These peptides are more reactive than whole rDf642. CONCLUSIONS: Mite paramyosin is very similar to other known paramyosins. The human IgE and IgG epitopes are scattered throughout the entire molecule. Data also indicate the presence of unique IgE and IgG epitopes in Der f 11.

Sensitization to Blomia tropicalis and dermatophagoides pteronyssinus-a comparative study between Singapore and Taiwan.

Blomia tropicalis (Bt) and Dermatophagoides pteronyssinus (Dp) are the predominant domestic mites species in Singapore and Taiwan. This study aims to characterize and compare the mite sensitization profiles in both countries. Skin prick tests were performed on 203 Singaporeans with Dp and Bt crude extracts. In vitro IgE and IgG4 reactivity to extracts and specific allergens (Der p 1, Der p 2 Der p 5 and Blo t 5) were determined by immunoassays. Approximately 91% of the tested Singaporeans were skin test positive for both Bt and Dp. Both populations share similar frequencies of in vitro IgE reactivity to all the allergens tested, but they differ in the pattern and magnitude of allergen sensitization. Although Der p 1, Der p 2 and Blo t5 are major sensitizing allergens in both countries, Blo t 5 is a more potent one in Singapore, probably reflecting the high level of exposure to Bt. The unique major Bt and Dp allergens should be included for precise diagnosis and effective immuno-therapeutic treatment of mite allergy in both countries.

Uveal lymphoid infiltration. Report of four cases and clinicopathologic review.

PURPOSE: The purpose of the study was to report the clinicopathologic features of four patients with uveal lymphoid infiltration who were diagnosed by a conjunctival biopsy. DESIGN: A case series. PARTICIPANTS: Four patients at four institutions participated. MEASURES: The histopathologic findings of the conjunctival and episcleral biopsy specimens from four patients with the diagnosis of uveal lymphoid infiltration were recorded. The conjunctival specimens were placed in 10% neutral buffered formaldehyde solution or B5 solution and processed routinely for light microscopic examination and immunohistochemical analysis for B cells and T cells and kappa and lambda light chains. The clinical and histopathologic findings were compared with 47 previously reported cases of uveal lymphoid infiltration. RESULTS: Two men and two women (average age = 57 years) presented with diffuse punctate to coalescent yellow uveal infiltrates and epibulbar pink fleshy lesions. Ultrasonography of the lesions showed diffuse choroidal thickening and a lack of choroidal or scleral excavation. The conjunctival biopsy specimens showed diffuse infiltration of the substantia propria with B lymphocytes with variable kappa and lambda restriction. Scattered T lymphocytes also were present. Plasmacytoid cells and a prominent Grenz zone were present. CONCLUSIONS: Patients with solitary or multiple yellow uveal infiltrates should have careful conjunctival evaluation for the presence of pink conjunctival lesions. Biopsy specimens of the conjunctival lesion may establish the diagnosis of uveal lymphoid infiltration.

Vitritis as the primary manifestation of ocular syphilis in patients with HIV infection.

PURPOSE: To describe dense vitritis as the primary manifestation of ocular syphilis in three human immunodeficiency virus (HIV)-positive patients and to determine the response of these patients to the established regimen for neurosyphilis. METHODS: Anti-Toxoplasma gondii IgM and IgG antibody titers, tuberculin skin test, chest radiograph, and serum angiotensin-converting enzyme level were obtained because tuberculosis, sarcoidosis, and toxoplasmosis were in the differential diagnosis. Two of the three patients were not known to have HIV infection at the time of initial examination and consented to HIV testing. Treponemal and nontreponemal tests were performed on serum and cerebrospinal fluid to establish a definitive diagnosis. Treatment for neurosyphilis was initiated, and daily ophthalmic examinations were performed, with careful attention to signs commonly associated with syphilitic eye disease. RESULTS: All three patients exhibited improvement in visual acuity and resolution of vitreous haze. There was no evidence of other signs of posterior uveitis. The one patient for whom there has been a 6-month follow-up showed no sequelae of his eye disease. CONCLUSIONS: Human immunodeficiency virus-positive patients with syphilis may present atypically dense vitritis. In these patients, vitritis may be the first manifestation of syphilis. The regimen for neurosyphilis provides effective therapy. Moreover, in some patients, syphilitic vitritis may be the initial manifestation of HIV disease.

Crystallization and preliminary X-ray diffraction analysis of group 5 mite allergen from Dermatophagoides pteronyssinus.

Crystals of the 14-kDa group 5 allergen from Dermatophagoides pteronyssinus (Der p 5) have been obtained at low pH and diffract to 3-A resolution using a conventional x-ray source. The crystals belong to tetragonal space group P41212 or P43212, with unit cell parameters a = b = 114 A and c = 234 A. A self-rotation search revealed a 432 point symmetry and thus suggested 96 molecules in one unit cell, hence 12 monomers in each asymmetric unit.

Can zinc prevent apoptosis of anterior keratocytes after superficial keratectomy?

OBJECTIVE: To determine whether zinc, which blocks apoptosis in many systems, including in leukemic cells and possibly in retinal dystrophies, can prevent the unwanted loss of anterior stromal keratocytes after superficial keratectomy. METHODS: After mechanical central corneal epithelial debridement, the left eyes of nine New Zealand white rabbits were treated with 25 mM zinc chloride (ZnCl2) in Earle's salts minimal essential medium (MEM) either every 30 min (n = 3), every 2 h (n = 3), or every 4 h (n = 3). The left eyes of nine additional animals, divided into three equal groups, were deepithelialized, and each received pure culture medium at one of the same three frequencies. One eye of each of another six rabbits was deepithelialized but received no drops. After 24 h, all 24 animals were sacrificed and the globes were enucleated. The corneas were processed and sections were stained with hematoxylin and eosin. RESULTS: The cell count of MEM-treated corneas exceeded that of untreated corneas (p = 0.03, analysis of variance [ANOVA]), but there was no difference among eyes that received the different frequencies of MEM application (p = 0.36, ANOVA). Cell counts increased with frequency of zinc application, but the differences were not statistically significant (p = 0.09, ANOVA). Only in the group receiving the most frequent zinc applications were superficial keratocytes retained. This group also possessed a greater number of stromal keratocytes than untreated controls and MEM-treated corneas (p = 0.01). CONCLUSIONS: At a significantly high dosage, zinc can prevent loss of superficial keratocytes to a greater extent than culture medium alone can. Moreover, zinc has the advantage of preserving the most anterior layer of keratocytes. Retention of these cells may prevent the reactive overproliferation that constitutes haze after photorefractive surgery.