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OBJECTIVE To determine whether preoperative and/or postoperative central endothelial cell density (ECD) and its rate of decline postoperatively are predictive of graft failure caused by endothelial decompensation following penetrating keratoplasty to treat a moderate-risk condition, principally, Fuchs dystrophy or pseudophakic corneal edema. METHODS In a subset of Cornea Donor Study participants, a central reading center determined preoperative and postoperative ECD from available specular images for 17 grafts that failed because of endothelial decompensation and 483 grafts that did not fail. RESULTS Preoperative ECD was not predictive of graft failure caused by endothelial decompensation (P = .91). However, the 6-month ECD was predictive of subsequent failure (P < .001). Among those that had not failed within the first 6 months, the 5-year cumulative incidence (±95% confidence interval) of failure was 13% (±12%) for the 33 participants with a 6-month ECD of less than 1700 cells/mm2 vs 2% (±3%) for the 137 participants with a 6-month ECD of 2500 cells/mm2 or higher. After 5 years' follow-up, 40 of 277 participants (14%) with a clear graft had an ECD below 500 cells/mm2. CONCLUSIONS Preoperative ECD is unrelated to graft failure from endothelial decompensation, whereas there is a strong correlation of ECD at 6 months with graft failure from endothelial decompensation. A graft can remain clear after 5 years even when the ECD is below 500 cells/mm2. CLINICAL TRIAL REGISTRY clinicaltrials.gov Identifier: NCT00006411Arch Ophthalmol. 2010;128(1):63-69--

ABSTRACTFungal keratitis is commonly caused by Fusariumspecies and less commonly by Candidaspecies. Recent outbreaks of Fusariumkeratitis were associated with contact lens wear and with ReNu with MoistureLoc contact lens care solution, and biofilm formation on contact lens/lens cases was proposed to play a role in this outbreak. However, no in vitro model for contact lens-associated fungal biofilm has been developed. In this study, we developed and characterized in vitro models of biofilm formation on various soft contact lenses using three species of Fusariumand Candida albicans. The contact lenses tested were etafilcon A, galyfilcon A, lotrafilcon A, balafilcon A, alphafilcon A, and polymacon. Our results showed that clinical isolates of Fusariumand C. albicansformed biofilms on all types of lenses tested and that the biofilm architecture varied with the lens type. Moreover, differences in hyphal content and architecture were found between the biofilms formed by these fungi. We also found that two recently isolated keratitis-associated fusaria formed robust biofilms, while the reference ATCC 36031 strain (recommended by the International Organization for Standardization guidelines for testing of disinfectants) failed to form biofilm. Furthermore, using the developed in vitro biofilm model, we showed that phylogenetically diverse planktonic fusaria and Candidawere susceptible to MoistureLoc and MultiPlus. However, Fusariumbiofilms exhibited reduced susceptibility against these solutions in a species- and time-dependent manner. This in vitro model should provide a better understanding of the biology and pathogenesis of lens-related fungal keratitis.

• To elucidate the pathogenic mechanisms involved in onchocercal sclerosing keratitis in humans, we developed a model of onchocercal interstitial keratitis in guinea pigs. Onchocerca volvulus antigens injected intrastromally into corneas of preimmunized Hartley guinea pigs induced an intense stromal keratitis with corneal edema, neovascularization, and infiltration with acute and chronic inflammatory cells. This reaction subsided after two weeks. Repeated intrastromal injection resulted in an exacerbation of the keratitis and ultimately in residual scarring. These findings are consistent clinically and histopathologically with the chronic interstitial keratitis observed in humans. To define which antigens induce the corneal reaction, O volvulus antigens were separated by molecular sieve chromatography and injected intrastromally. The highest activity was shown to reside in the fraction containing molecules of intermediate molecular weight. This model will be useful in defining O volvulus antigens and their role in sclerosing keratitis as well as in elucidating the immune mechanisms involved.

Eleven patients with leukemia refractory to conventional therapy were treated with high doses of cytarabine (3 g/m2intravenously every 12 hours for six days) followed by daunorubicin (30 mg/m2/day intravenously for three days). Each patient received, in a double-masked, randomized manner, either 1% prednisolone phosphate or placebo eyedrops in each eye 12 hours before and along with the cytarabine so that we could determine whether a topical corticosteroid prevents corneal toxicity from systemic cytarabine. Ten of the 11 patients had a significant reduction in the intensity and duration of the symptoms and signs of corneal toxicity in the corticosteroid-treated eye (P<.0005). In the placebo-treated eye, pain, photophobia, and tearing occurred at a median time of 6.5 days after initiation of therapy but resolved in approximately one week.

Corneal inflammation, produced by direct interlamellar injection of clove oil, is histologically characterized primarily by polymorphonuclear leukocytic infiltration of the stroma. The polymorphonuclear leukocytes can be radiolabelled with tritiated thymidine systemically prior to their invasion of the cornea. A measure of the number of labelled polymorphonuclear leukocytes in the cornea can then be obtained by scintillation counting techniques, which provides an objective method for quantitating the degree of inflammation in the cornea.

• Four patients had idoxuridine-induced conjunctival cicatrization similar to ocular cicatricial pemphigoid develop, but in the treated eye only. Three of the four patients had chronic, recurrent herpes simplex epithelial and stromal keratitis. The fourth patient had Sjögren's syndrome. All received idoxuridine (0.1% drops and/or 0.5% ointment) and topical corticosteroids from one to 3½ years. Substantial morbidity resulted that included visual loss, stromal ulceration, corneal scarring, and keratinization. Conjunctival biopsy specimens showed cicatrization with a mixed inflammatory cell reaction and absence of goblet cells. Results of direct immunofluorescent microscopy of the conjunctiva were either negative or nonspecific for autoantibody. No circulating autoantibody was detected in any of the four patients.

• We studied, in a rabbit, the ocular toxicity of topical 1% solutions or suspensions of amphotericin B, flucytosine, miconazole nitrate, and ketoconazole. Flucytosine, miconazole, and ketoconazole did not retard the closure of 8.5-mm corneal epithelial defects; amphotericin B greatly retarded the closure of such defects. Amphotericin B produced dramatic pathologic changes in this model; these changes worsened with each day of therapy. Ketoconazole produced modest biomicroscopically and histologically detectable pathologic changes in the regenerating corneal epithelium; flucytosine and miconazole did not produce such changes.

• The effect of medroxyprogesterone acetate treatment in preventing or retarding corneal ulceration after alkali burns was studied in rabbits in an attempt to correlate dose and response. The eyes were treated for 21 days with a 0.01% or 1% suspension administered topically four times a day or medroxyprogesterone acetate, 4 mg or 30 mg, given subconjunctivally on alternate days. None of these regimens was noticeably effective, although at times there was a trend toward retarding ulceration. Thus, it was not possible to confirm earlier reports of substantial efficacy of medroxyprogesterone. Corneal stroma with epithelial defects and/or ulcers persistently showed infiltration with neutrophils in the superficial stroma within the epithelial defect, implicating the tears as the source of the inflammatory cells.

• The chemotherapeutic efficacy of acyclovir was evaluated by observing the potential of topical acyclovir to reduce the severity of herpes viral lesions and to control the multiplication of this virus in the experimentally induced primary cutaneous infection of guinea pigs. Topical acyclovir showed a substantially high therapeutic efficacy when treatment was initiated after the development of clinically overt skin lesions. The therapeutic response was dose dependent and clearly evident even when the treatment was initiated on day 2 after inoculation. Our results indicate that topical acyclovir treatment improved the cutaneous herpes simplex virus type 1 (HSV-1) infectious process by inhibiting multiplication of HSV-1 in the skin of guinea pigs.(Arch Dermatol 116:672-675, 1980)

Sclerosing keratitis is the major cause of blindness due to onchocerciasis; its pathogenesis is poorly understood. We have previously reported an immune-mediated model of experimental interstitial keratitis in guinea pigs following intrastromal challenge with soluble antigens from Onchocerca volvulus. This model system is ideal for evaluation of pathogenicity of multiple purified antigen preparations; however, reagents necessary for detailed immunologic analysis of the inflammatory cellular infiltrate are not yet available for guinea pigs. Because of the ready availability of these reagents for mice, a mouse model has been developed. The inflammatory response observed in this model was analogous to that seen in human onchocercal sclerosing keratitis as well as in the guinea pig model of onchocercal sclerosing keratitis. Granulocytes were present in the acute inflammatory response, whereas the chronic response showed lymphocytes, plasma cells, and histiocytes. Neovascularization and scarring of the corneal stroma was also observed. This model will be helpful in examining the mechanisms of immuno-pathogenesis and the contribution of the host genetic background to the disease.

Forty-one human conjunctival papillomas were histopathologically examined for koilocytosis and papillomavirus common antigen with a peroxidase-antiperoxidase technique immunospecific for genus-specific papillomavirus antigen. Koilocytotic cells, typically found in other papillomavirus-induced tumors, were present in 24 of the 41 specimens. Two of the 41 specimens were positive for viral antigen. The positive reactions were intranuclear within the superficial layers of the epithelium.

In a double-masked, randomized fashion, 11 patients with hematologic malignancies received 13 courses of high-dose cytarabine therapy, intravenously (3 g/m2every 12 hours for five to six days). Each patient received topical prednisolone phosphate 1% in one eye and 2-deoxycytidine 100 μM in the other eye every six hours. Topical therapy was initiated 12 hours before the first cytarabine dose and continued for up to ten days (until four to five days after completion of cytarabine therapy). Slit-lamp biomicroscopy was performed before therapy and then weekly for one month. 2-Deoxycytidine was equally as effective as the topical corticosteroid therapy in reducing photophobia and pain, microcysts, and punctate epithelial erosions, and each treatment gave results significantly better when compared historically to placebo-treated eyes.

Two histologically proven conjunctival papillomas (one from a 33-month-old boy and the other from a 28-year-old woman) were examined for the presence of papillomavirus by DNA molecular hybridization. The first case, a recurrent tumor known to be positive for papillomavirus structural antigen, demonstrated human papillomavirus DNA sequences that cross-hybridized to a human papillomavirus type 11 DNA probe. The second case, an initial tumor, which was negative for papillomavirus structural antigen, demonstrated no viral DNA sequences by hybridization.

Seven patients with type I diabetes mellitus (Group 1), seven with normoglycemic cystic fibrosis (Group 2), seven with hyperglycemic cystic fibrosis (Group 3), and ten age-matched control subjects underwent corneal fluorophotometry and quantitative specular microscopy. Group 1 had background microangiopathic retinopathy but no evidence of proliferative disease by fluorescein angiography. Significant increases in mean corneal endothelial permeability and mean pump rate occurred in Group 1, indicating a defect in the endothelial barrier function early in type I diabetes mellitus. Similar significant increases in mean corneal endothelial permeability and mean pump rate occurred in both cystic fibrosis groups. The greatest increase was found in Group 3, suggesting a primary defect in the endothelial barrier function in cystic fibrosis, aggravated by the hyperglycemic state. No morphologic abnormalities were noted in Group 1, but both cystic fibrosis groups had smaller mean cell areas than did the control group. There were significant differences in the morphologic and functional correlations between Groups 1 and 3, suggesting different mechanisms for the increased endothelial permeability in these two disorders.

• Levobunolol hydrochloride (0.5% and 1%) and timolol maleate (0.5%) are being compared in an ongoing, double-masked, randomized study of 141 patients with ocular hypertension or chronic open-angle glaucoma. Baseline intraocular pressure (IOP) in the three treatment groups ranged from 26 to 27 mm Hg. During the first 15 months of the study, the two drugs have not proved to be significantly different in ocular hypotensive efficacy, with overall mean IOP decreases of 6.8 to 7.6 mm Hg. In addition, the two concentrations of levobunolol have been equally effective in controlling IOP. Neither drug has been associated with any significant ocular side effects. Both drugs have produced significant decreases (five to ten beats per minute) in mean heart rate. The effect on mean blood pressure has been less pronounced: overall decreases have been less than 4 mm Hg for both systolic and diastolic blood pressure. The results of this ongoing study suggest that levobunolol is as effective and as safe as timolol for the long-term control of IOP.

• Thirty-three epikeratoplasties were performed on 31 keratoconic contact lens intolerant patients. Thirty-two of these procedures were anatomically successful with clear lenticules and flattening of the postoperative keratometric measurements in all but one eye. One patient's lenticule developed a persistent epithelial defect that eventually required its removal at 42 days. This patient subsequently underwent a successful penetrating keratoplasty. Of the 32 eyes that were anatomically successful, 20 eyes in 19 patients were able to achieve satisfactory visual improvement requiring either no correction or spectacle lenses. Twelve eyes in 11 patients were successfully refit with contact lenses for anisometropic refractive errors (11 eyes) and residual irregular astigmatism (one eye). Three patients (two in the non-contact lens group and one in the contact lens group) have subsequently undergone penetrating keratoplasty for further visual improvement. Our data suggest that the majority of keratoconic epikeratoplasties achieve a satisfactory visual result either unaided or with spectacles; however, those patients who wish to resume contact lens wear may do so successfully.

• A model of herpes simplex interstitial keratitis was developed in rabbits sensitized with live herpes simplex virus (HSV) type 2 and challenged intrastromally with live virus. This model was used to evaluate the effects of subconjunctival medroxyprogesterone acetate on the course of the disease and on collagenase levels in the treated corneas. Whether treated prophylactically or therapeutically, the medroxyprogesterone-treated groups had substantially less stromal infiltration and neovascularization than the controls. The clinical effects corresponded with a marked reduction in the polymorphonuclear leukocyte infiltrate histologically, and the suppression of latent and active collagenase activity in the treated corneas cultured in vitro. However, epithelial disease was exacerbated in prophylactically treated animals. Medroxyprogesterone appears to be useful in the treatment of herpetic interstitial keratitis as an anti-inflammatory agent as well as an inhibitor of collagenase production but, like the corticosteroids, it can exacerbate epithelial disease.

Sclerosing keratitis is the major cause of blindness due to onchocerciasis caused by the parasite Onchocerca volvulus. Although the importance of T cells in the pathogenesis of onchocerciasis has been suggested, their precise role in onchocercal sclerosing keratitis has not yet been defined. Using immunohistological techniques and a murine model of onchocercal sclerosing keratitis, we have performed a temporal analysis of the inflammatory T cells infiltrating into the cornea at Days 4, 7, and 21 following intrastromal challenge with soluble O. volvulusantigens into presensitized mice. The maximum number of CD3+T cells were observed in the corneal stroma at Day 21 when sclerosing keratitis was most severe. The majority (>85%) of the CD3+T cells were CD4+at all time points. A few infiltrating cells bore IL-2 receptors indicating possible activation of a small fraction of the T cells. These results suggest that CD4+T cells play an important role in onchocercal sclerosing keratitis.