A serum- and feeder-free technique of culturing human corneal epithelial stem cells on amniotic membrane.

PURPOSE: To describe a simple technique of cultivating human corneal epithelial stem cells using an Epilife culture medium under serum- and feeder-free conditions. METHODS: Cadaveric donor limbal corneal epithelial cells were cultured on denuded amniotic membranes using an explant technique that was free of serum and feeder cells in the Epilife medium containing a growth supplement of defined composition. These cells were assessed by phase contrast microscope. The expressions of the proposed corneal epithelial stem cell markers (p63, ATP-binding cassette member 2 (ABCG2), and cytokeratin 15 and 19) and differentiation markers (cytokeratin 3, 12, connexin 43, and p75) were analyzed using reverse transcription polymerase chain reaction (RT-PCR) and immunocytochemical staining. RESULTS: Successful cultures were obtained, resulting in a monolayer to double layer cell sheets with a cobblestone-like morphology. RT-PCR and immunocytochemistry disclosed an expression of both putative limbal stem cell (LSC) markers and differentiation-associated markers in the cultured cells. Most of the cultured corneal epithelial cells that were immunopositive for putative LSC markers were smaller, more uniform, and closer to the limbal explant than cells positively stained with differentiation-associated markers. CONCLUSIONS: A serum- and feeder-free culture system using Epilife medium may grow human corneal epithelial equivalents, minimizing the risk of contamination during culture. The technique may also be useful for the clinical application of limbal stem cell culture.

Boston keratoprosthesis in epithelial downgrowth.

INTRODUCTION: To report a case of histologically proven epithelial downgrowth after multiple failed penetrating keratoplasties and glaucoma filtering surgeries that was successfully treated with Boston keratoprosthesis implantation. MATERIALS AND METHODS: A 61-year-old monocular patient had severe congenital ocular syphilis with secondary glaucoma. He had undergone many intraocular surgeries with a history of epithelial downgrowth, and he presented with a failed graft after 7 penetrating keratoplasties. Implantation of a corneal graft with an aphakic type of Boston keratoprosthesis was performed, combined with anterior vitrectomy. The main outcome measures were visual acuity, ocular inflammation and media clarity. RESULTS: Media clarity was restored and revealed severe retinal scarring and a pale optic nerve. Best corrected visual acuity of 20/400 was maintained without any further surgical intervention during 6 years follow up. No retroprosthesis membrane or epithelial growth behind the keratoprosthesis was observed. DISCUSSION: This is, to our knowledge, the first case of long-term successful treatment of epithelial downgrowth with a Boston keratoprosthesis. This approach might be considered a suitable treatment of epithelial downgrowth.

Euphorbia lactea sap keratouveitis: case report and review of the literature.

PURPOSE: To describe a case of Euphorbia lactea sap keratouveitis and to review all reported cases of ocular toxicity caused by Euphorbia species. METHODS: Case report and review of literature. RESULTS: A 79-year-old woman presented 34 hours after she felt some sap of an E. lactea plant spray into her right eye. Visual acuity was counting fingers at 1 m. Examination revealed ciliary injection, 90% corneal epithelial defect, marked stromal edema with Descemet folds, and anterior-chamber flare with a 1-mm hypopyon. There was no vitreitis, and funduscopy was unremarkable. No foreign body was seen on B scan ultrasound or computed tomography scan of the orbits. Corneal scraping excluded bacterial and herpetic keratitis. Intensive topical antibiotic therapy was started with cephalothin 5% and gentamicin 0.9%, and the pupil was dilated with atropine. Topical steroids were started once the epithelial defect had healed. Examination 11 weeks after the injury revealed minimal subepithelial corneal haze and marked improvement in visual acuity. CONCLUSIONS: To the best of our knowledge, this is only the third reported case of E. lactea sap keratouveitis. The clinical course of E. lactea sap keratouveitis is compared with that reported for other Euphorbia species.

Corneal perforation associated with pellucid marginal degeneration and treatment with crescentic lamellar keratoplasty: two case reports.

PURPOSE: To describe 2 cases of corneal perforation associated with pellucid marginal degeneration (PMD) and subsequent successful management with crescentic lamellar keratoplasty. METHODS: Two cases are discussed along with the relevant literature. RESULTS: Both patients presented with spontaneous corneal perforations and associated features consistent with PMD. In both cases, there was fellow eye evidence of PMD, and 1 of the patients was already known to our unit with this diagnosis. Initial conservative measures were unsuccessful, and both patients underwent crescentic lamellar keratoplasty as the definitive surgical management with a good visual outcome. CONCLUSIONS: Crescentic lamellar keratoplasty is an effective technique for the management of this rare complication of PMD.