Loss of fenamate-activated K+ current from epithelial cells during corneal wound healing.
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PURPOSE: The corneal epithelium provides a barrier between the external environment and the cornea. It also serves as an ion transporting epithelium. Because of its proximity with the external environment, the corneal epithelium is frequently injured through physical or chemical insult. The purpose of this study was to determine whether corneal epithelial cell whole-cell currents change during corneal wound healing as the author of the present study has previously reported for corneal keratocytes and endothelial cells. METHODS: Rabbit corneal epithelial cells were injured by scraping, heptanol exposure, or freezing. The epithelium was allowed to heal for 12 to 74 hours. Cells were dissociated from corneas, and whole-cell currents were examined using the amphotericin-perforated-patch technique. RESULTS: Cells from the wounded corneal groups had significantly increased capacitance values, indicating increased surface area compared with that of control cells. As previously reported, the primary control whole-cell current was a fenamate-activated K+ current. An inwardly rectifying K+ current and a Cl- current were also observed. In epithelial cells from heptanol-wounded corneas, these conductances were generally unchanged. In cells from scrape- and freeze-wounded corneas, however, the fenamate-activated current was absent or significantly attenuated. CONCLUSIONS: As they do in corneal keratocytes and endothelial cells, K+ channels disappear during some models of corneal epithelial wound healing. In addition, cell capacitance, a measurement of cell surface area, increases. These results suggest that substantial K+ channel activity is not required for in vivo epithelial cell proliferation during corneal wound healing. (+info)
Amniotic membrane transplantation for ocular surface reconstruction.
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AIMS: To evaluate the efficacy of amniotic membrane transplantation (AMT) for ocular surface reconstruction. METHODS: 10 consecutive patients who underwent AMT were included. The indications were: group A, cases with persistent epithelial defect after corneal abscess (n = 1), radiation (n = 1), or chemical burn (n = 3); group B, cases with epithelial defect and severe stromal thinning and impending or recent perforation, due to chemical burn (two patients, three eyes) or corneal abscess (n = 2); group C, to promote corneal epithelium healing and prevent scarring after symblepharon surgery with extensive corneo-conjunctival adhesion (n = 1). Under sterile conditions amniotic membrane was prepared from a fresh placenta of a seronegative pregnant woman and stored at -70 degrees C. This technique involved the use of amniotic membrane to cover the entire cornea and perilimbal area in groups A and B, and the epithelial defect only in group C. RESULTS: The cornea healed satisfactorily in four of five patients in group A, but the epithelial defect recurred in one of these patients. After AMT three patients underwent limbal transplantation and one penetrating keratoplasty and cataract extraction. In group B amniotic membrane transplantation was not helpful, and all cases underwent an urgent tectonic corneal graft. Surgery successfully released the symblepharon, promoted epithelialisation and prevented adhesions in the case of group C. CONCLUSION: AMT was effective to promote corneal healing in patients with persistent epithelial defect, and appeared to be helpful after surgery to release corneo-conjunctival adhesion. Most cases required further surgery for visual and ocular surface rehabilitation. Amniotic membrane used as a patch was not effective to prevent tectonic corneal graft in cases with severe stromal thinning and impending or recent perforation. (+info)
Allo-limbal transplantation in patients with limbal stem cell deficiency.
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AIM: To report the outcome of a series of patients with stem cell deficiency who underwent allo-limbal transplantation and to describe a technique for this procedure. METHODS: Six consecutive patients underwent allo-limbal stem cell transplantation. The primary diagnosis included alkali burn (n = 2), trachoma (n = 1), chronic rosacea blepharitis and kerato-conjunctivitis (n = 1), aniridia (n = 1), and Stevens-Johnson syndrome (n = 1). The limbal rim consisted of peripheral cornea and perilimbal sclera. FK-506 was used postoperatively for immunosuppression. RESULTS: The length of follow up ranged from 3 to 24 months (mean follow up 11.8 (SD 9.3) months). The outcome was considered satisfactory in five of six cases. The corneal surface was completely epithelialised within 2 weeks, and there was a substantial improvement in vision and symptoms. One patient had recurrent epithelial defects related to eyelid abnormalities. No side effects associated with systemic immunosuppression were noted. CONCLUSION: Allo-limbal transplantation, with systemic immunosuppression with FK-506 is useful in reconstruction of the ocular surface with improvement in vision in patients with severe stem cell deficiency. (+info)
Effect of metalloproteinase inhibitor on corneal cytokine expression after alkali injury.
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PURPOSE: Interleukin (IL)-1alpha and IL-6 levels in the cornea are greatly elevated during the early stages after an alkali burn in mice. The authors investigated the effect of synthetic inhibitor of matrix metalloproteinase (SIMP) on the expression of inflammatory cytokines in alkali-burned murine corneas and evaluated the clinical appearance of the eyes. METHODS: After 0.5N NaOH-alkali burns to 400 corneas of ICR mice, 200 received 400 microg/ml of SIMP topically 4 times a day while 200 corneas were similarly treated with vehicle only. At days 4, 7 and 14 after injury, each cornea was assigned a clinical score for corneal opacity, corneal epithelial defect, hyphema and cataract. Extracts of injured corneas in each group were then assayed for cytokine production using ELISA systems for IL-1alpha, IL-1beta, IL-6 and tumor necrosis factor-alpha (TNF-alpha). RESULTS: The levels of IL-1alpha, IL-1beta and IL-6 were significantly lower in the SIMP-treated group than in the vehicle-treated group 7 days after the burn. However, levels of these cytokines were similar in the SIMP and non-SIMP groups at days 4 and 14. Levels of TNF-alpha did not differ between both groups at any postinjury time. In the SIMP-treated corneas, there was less opacification and hyphema formation and epithelial regeneration was faster. CONCLUSIONS: Topical application of SIMP in alkali-burned murine corneas reduced the expression of IL-1alpha, IL-1beta, and IL-6 and lessened the severity of the injury. (+info)
Nitric oxide synthase-II is expressed in severe corneal alkali burns and inhibits neovascularization.
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PURPOSE: Inducible nitric oxide synthase (NOS-II) is expressed in many inflammatory conditions. The implication of nitric oxide (NO) in angiogenesis remains controversial. The role of NOS-II and its influence on angiogenesis in corneal neovascularization is unknown and was investigated in this study. METHODS: A mouse model of corneal neovascularization induced by chemical cauterization was used. NOS-II mRNA expression was analyzed by reverse transcriptase-polymerase chain reaction, and NOS-II protein was studied in situ by immunohistochemical analysis of the cornea. The influence of NOS-II on neovascularization was determined by comparison of vessel development in "normal" wild-type mice and mice with a targeted disruption of the NOS-II gene. RESULTS: NOS-II mRNA was induced to very high levels after corneal cauterization and remained upregulated throughout the disease. Migratory cells in the center of the cauterization area expressed NOS-II protein. The neovascular response in mice lacking the NOS-II gene was significantly stronger than in wild-type mice, and the difference increased over time. CONCLUSIONS: These data are the first evidence that NOS-II is expressed in this model of sterile corneal inflammation. NOS-II expression inhibited angiogenesis in severe corneal alkali burns. (+info)
Long term results after autologous nasal mucosal transplantation in severe mucus deficiency syndromes.
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AIM: Severe mucus deficiency syndromes may require substitution of mucous membrane for re-establishment of the ocular surfaces. The long term results after autologous nasal mucosal transplantation were investigated. METHODS: 55 eyes of 50 patients with severe mucus deficiency syndromes were followed retrospectively after free autologous nasal mucosal transplantation-group A: patients after severe lye, acid, heat burns, or radiation (n=38 eyes), group B: patients with systemic mucosal disease (n=17 eyes). The results of routine clinical examination were recorded and patients were followed for a median of 37 months. 17 biopsies of transplanted nasal mucosa were studied by light microscopy and 22 patients by impression cytology before and at several intervals after mucosal transplantation. RESULTS: All nasal mucosal grafts healed well and no intraoperative complications occurred. During follow up 107 additional surgical procedures were performed including 16 lamellar and 21 penetrating keratoplasties. Subjective complaints improved in 44/47 patients with preoperative symptoms. Best corrected visual acuity at the end of follow up was increased in 23 eyes, 10 eyes (18. 2%) reached a final visual acuity equal to or greater than 20/200. Histopathologically, all (n=17) biopsies showed vital intraepithelial mucin producing goblet cells in the nasal mucosal graft (median 25 cells/field (400x magnification)). The mean density of goblet cells before transplantation was 48/mm(2) and after nasal mucosal grafting 432/mm(2) measured by impression cytology (p<0. 0001). CONCLUSIONS: Functional goblet cells persist in autologous nasal mucosa for up to 10 years after transplantation. In patients with severe mucus deficiency syndromes of different origin nasal mucosal transplantation can re-establish the ocular surface, substitute the mucus components of the tear film, improve symptoms of the patients, and facilitate a moderate increase in visual acuity. (+info)
Nerve growth factor promotes corneal healing: structural, biochemical, and molecular analyses of rat and human corneas.
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PURPOSE: A recent clinical report demonstrated that topical nerve growth factor (NGF) treatment in patients affected by corneal neurotrophic ulcers induced epithelial and stromal healing restoring corneal integrity. Mechanisms(s) undergoing these clinical NGF actions are still unclear. The aim of this study was to investigate the role of NGF in human and rat cornea physiopathology. METHODS: Expression of high-affinity NGF receptors, NGF-mRNA, and NGF protein was evaluated in human and rat normal corneas, in human and rat corneal epithelial cell cultures, in human corneal organ culture, and in the rat cornea after an experimental model of epithelial injury, by means of immunohistochemistry, in situ hybridization reverse transcription-polymerase chain reaction, and enzyme-linked immunosorbent assay. RESULTS: The resultant data demonstrated that NGF is a constitutive molecule present and produced in normal human and rat corneas. In vitro human and rat corneal epithelial cells produce, store, and release NGF and also express high-affinity NGF receptors (TrkA). In human organ culture, epithelium, keratocytes, and endothelium have been shown to bind exogenous radiolabeled NGF, and the epithelial cells' binding was increased after epithelium injury. In vivo, after rat corneal epithelial injury, a transient increase of corneal NGF levels was observed. Inhibition of endogenous NGF activity by neutralizing anti-NGF antibodies delayed the corneal epithelial healing rate, whereas exogenous administration of NGF accelerated healing. CONCLUSIONS: Taken together, the above findings show that NGF plays an important role in corneal physiopathology and suggest that this neurotrophin may exert therapeutic action in wide-spectrum corneal diseases. (+info)
Reconstruction of damaged corneas by transplantation of autologous limbal epithelial cells.
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BACKGROUND: Stevens-Johnson syndrome, ocular pemphigoid, and thermal or chemical burns can cause scarring and opacification of the cornea and loss of vision. Transplantation of epithelial cells from the limbus of the contralateral cornea can restore useful vision. However, this procedure requires a large limbal graft from the healthy eye and is not possible in patients who have bilateral lesions. METHODS: We took specimens of limbal epithelial cells from the healthy contralateral eyes of six patients with severe unilateral corneal disease. The epithelial cells were cultured and expanded on amniotic membrane. The amniotic membrane, together with the sheet of limbal epithelial cells, was transplanted to the denuded corneal surface of the damaged eye after superficial keratectomy to remove fibrovascular ingrowth. The mean (+/-SD) follow-up period was 15+/-2 months. RESULTS: Complete reepithelialization of the corneal surface occurred within two to four days of transplantation in all six eyes receiving transplants. By one month, the ocular surface was covered with corneal epithelium, and the clarity of the cornea was improved. In five of the six eyes receiving transplants (83 percent), the mean visual acuity improved from 20/112 to 20/45. In one patient with a chemical burn who had total opacification of the cornea, the acuity improved from the ability to count fingers at 40 cm to 20/200. No patient had recurrent neovascularization or inflammation in the transplanted area during the follow-up period. CONCLUSIONS: Transplantation of autologous limbal epithelial cells cultured on amniotic membrane is a simple and effective method of reconstructing the corneal surface and restoring useful vision in patients with unilateral deficiency of limbal epithelial cells. (+info)