Development and clinical assessment of an artificial cornea.

Keratoprosthesis research has been a gradual, rather fragmentary process with advances being made by isolated groups of researchers. This has arisen partly because of poor funding in the area; research groups which have achieved commercial support have often had constraints upon the full disclosure of their findings. Despite these difficulties there has been real progress over the last decade by several independent groups. This article concentrates upon our own development of a hydrogel core-and-skirt keratoprosthesis, the Chirila KPro, in order to illustrate the scientific and clinical problems common to keratoprosthesis research. Pilot data from a clinical trial is presented and the priorities for future research are discussed.

The effect of contact lens wear on the central and peripheral corneal endothelium.

PURPOSE: To compare central and peripheral corneal endothelial cell morphometry in normal subjects and long-term contact lens wearers. METHODS: Endothelial cell density (ECD), coefficient of variation of cell area (CV), and percentage of six-sided cells were measured by contact specular microscopy in the corneal center and temporal periphery of both eyes of 43 long-term contact lens wearers and in 84 normal subjects who had never worn contact lenses. The latter group included 43 age- and sex-matched controls for the contact lens wearers. ECDs were corrected for magnification changes due to corneal thickness. RESULTS: Central ECD (2,723+/-366 cells/mm2, mean +/- SD) was significantly higher than peripheral ECD (2,646+/-394 cells/mm2) for the normal group (p = 0.01) but not for the contact lens wear group (2,855+/-428 cells/mm2 central, 2,844+/-494 cells/mm2 peripheral, p = 0.84). Peripheral CV was significantly higher than central for normal subjects and contact lens wearers and was significantly higher in both center and periphery in contact lens wearers than in controls. Central percentage of six-sided cells was significantly higher than peripheral for normal subjects and contact lens wearers and was lower in both center and periphery in contact lens wearers than in controls. CONCLUSIONS: Central ECD was significantly higher by 3% than peripheral ECD in normal subjects, but not in contact lens wearers. The results suggest that contact lens wear causes a mild redistribution of endothelial cells from the central to the peripheral cornea. A reversal of this redistribution after contact lens wear is discontinued for refractive surgery could mask mild central endothelial damage from the refractive procedure.

The value of routine donor corneal rim cultures in penetrating keratoplasty.

OBJECTIVE: To investigate the value of donor corneal rim cultures performed routinely at the time of penetrating keratoplasty. DESIGN: Retrospective review of Mayo Clinic medical records for all corneal transplantations for which donor rim cultures have been performed. MAIN OUTCOME MEASURES: Frequency of positive cultures, occurrence of endophthalmitis within 2 months of undergoing surgery, action taken in response to the culture results, and costs of cultures. RESULTS: Donor rim culture results were available for 1078 of 1083 consecutive transplantations performed from 1981 to 1995. Three cases of endophthalmitis (0.28%) and 1 suture abscess occurred. Rim cultures were negative in all of these cases. Action was documented in response to positive cultures in 17 cases (8.1%). The estimated average cost of routine rim cultures in 1994 was $137 per donor cornea. Bacterial or fungal cultures were positive in 209 (19.4%) cases. Two microorganisms were cultured simultaneously in 17 cases (1.6%) and 3 in 2 cases (0.2%). Staphylococcus coagulase-negative (130 cases [12.1%]), and Streptococcus species, viridans group (23 cases [2.1%]), were the most common isolates. Fifty-two (62.7%) of 83 coagulase-negative Staphylococcus species isolates tested were resistant to gentamicin. There were more positive cultures from corneas stored in Optisol (37/183 [20%]) than in Optisol GS (16/144 [11%]) (P = .03). Fewer cultures were positive from live donors (9/93 [10%]) compared with cadaveric donors (181/ 909 [20%]) (P = .02). Positive cultures were more frequent for corneas excised in situ (39/125 [31.2%]) than for those enucleated (152/851 [17.9%]) (P < .001). CONCLUSIONS: Despite differences in rates of positive donor rim cultures with different harvesting and storage techniques, for our practice, routine donor corneal rim cultures had no predictive value for infective complications of penetrating keratoplasty and, therefore, added an unnecessary expense to the management of our patients.

Influence of topical human epidermal growth factor on postkeratoplasty re-epithelialisation.

AIM: To test the efficacy and safety of recombinant human epidermal growth factor (hEGF) on corneal re-epithelialisation following penetrating keratoplasty. METHODS: A prospective, randomised, placebo controlled study was carried out in which patients were matched for diagnosis and received either hEGF ophthalmic solution (30 micrograms/ml or 100 micrograms/ml) or placebo in a double masked fashion. Matched pairs of patients received donor corneas from the same donor and were operated by the same surgeon on the same day. At the end of surgery all donor epithelium was removed mechanically. Patients were examined twice daily and fluorescein stained photographs were taken until the epithelium had closed. The area of the defect was measured by planimetry of the fluorescein stained defect on the photographs. RESULTS: There were no significant differences in re-epithelialisation of the donor cornea between the placebo group and the group treated with 30 micrograms/ml hEGF. Time until complete closure was slightly longer with 100 micrograms/ml hEGF compared with 30 micrograms/ml hEGF and with placebo. Mean healing rate of the epithelial defect with 100 micrograms/ml hEGF was significantly slower than in the other groups. CONCLUSION: No significant acceleration of corneal re-epithelialisation was demonstrated with the use of recombinant hEGF after penetrating keratoplasty in humans.

Keratometric results of penetrating keratoplasty with the Hessburg-Barron and Hanna trephine systems using a standard double-running suture technique.

PURPOSE: To compare final, sutures-out, keratometric results of penetrating keratoplasty using two different suction trephine systems and a standard double-running suture technique. METHODS: Keratometric data after final suture removal were compared retrospectively for three groups of transplants: Group 1, 7.50-mm Hessburg-Barron recipient trephine and 7.70-mm modified Lieberman donor punch (n = 70); Group 2, 7.50-mm Hessburg-Barron recipient trephine and 8.00-mm modified Lieberman donor punch (n = 18); Group 3, 7.50-mm Hanna recipient trephine and 7.75-mm Hanna donor punch (n = 68). RESULTS: Final average keratometry was 46.7 +/- 2.4 D (mean +/- SD, diopters), and final median keratometric astigmatism was 4.6 D (2.6-7.4 D, interquartile range), with no significant difference between groups. Final average keratometry values increased from preoperative values by 3.3 +/- 2.7 D. The correlation between preoperative average keratometry values and final values was low (rs = 0.2; p = 0.07). Average keratometry values increased by 2.3 D (median) after 10/ 0 suture removal (p < 0.0001) and by 0.2 D (median) after 11/0 suture removal (p = 0.09), with no significant difference between groups. There was a negative correlation between donor age and overall change in average keratometry (rs = -0.3; p = 0.002). Visual acuity improved by a median of 5 lines, with no significant difference between trephine systems. CONCLUSIONS: There was no statistically significant difference in keratometric results using these two suction trephines with a standard suture technique. Final graft curvature was greater with both suction trephines compared with previously published results of transplants using hand-held trephines with the same suture technique.

Lysosomal enzymes in corneal storage media and corneal graft outcome.

PURPOSE: The purpose of this study was to relate lysosomal enzyme activities in corneal storage media to the outcome of the transplanted corneas. METHODS: Corneal storage media from 358 transplanted corneas were frozen at -70 degrees C and kept for enzyme analysis. Corneas were stored in K-Sol (28), CSM (35), Dexsol (80), Index medium (five), Optisol (158), and Optisol GS (52). Activities of alpha-D-mannosidase, beta-glucuronidase, alpha-glucosidase, and N-acetyl-beta-glucosaminidase were assayed fluorometrically. Mayo Clinic records were examined for donor information, including cause of death and 2-month graft follow-up data. RESULTS: For all corneas, there was a low but significant correlation between activities of each enzyme and storage time (rs = 0.13-0.35; p = 0.02-0.0001), and donor age (rs = -0.14 to -0.23; p = 0.009-0.0001). There was no significant correlation of enzyme activity with 2-month endothelial cell density, structure, cell loss, or corneal thickness. Enzyme activities for four primary donor failures and six grafts with > 65% 2-month endothelial cell loss were not significantly different from those for the rest of the transplanted corneas. Enzyme activities were higher for corneas from donors with renal failure but not from those with diabetes mellitus. There was no significant difference in graft outcome for different cause-of-death groups. CONCLUSIONS: The activities of lysosomal enzymes released into corneal storage media are not useful as predictors of graft outcome.

Specular microscopy before and after enucleation of live donor eyes.

PURPOSE: To compare the results of specular microscopic examination of corneal endothelium before and after enucleation of eyes from live donors. METHODS: Endothelial cell density (ECD), coefficient of variation of cell area (CV), and percent hexagonal cells were compared for 34 cornea donors before enucleation of their eyes and after excision of the corneoscleral rims and placement in preservative media. RESULTS: There was no statistically significant difference in ECD, CV, or percent six-sided cells after enucleation. The pre-enucleation and post-enucleation ECD measurements were significantly correlated (rs = .85, P < .0001). Mean percentage change in ECD was -0.7% +/- 6.0%. CONCLUSIONS: There was no significant difference in ECD, CV, or percent six-sided cells between measurements taken from the epithelial side in vivo and those taken of the same corneas from the endothelial side in vitro after enucleation and corneoscleral rim excision. These findings suggest that it is reasonable to compare postkeratoplasty clinical measurements with those of the donor corneas taken in the eye bank.

Posterior amorphous corneal dystrophy. A new pedigree with phenotypic variation.

BACKGROUND: Posterior amorphous corneal dystrophy is a rare condition characterized by bilateral sheet-like opacification of the posterior stroma in association with corneal flattening and thinning. It has been reported in only four families, all from the United States. The authors report on a fifth family, the first from Britain, with nine affected individuals. METHODS: Slit-lamp photography, refraction, keratometry, pachometry, corneal topography, and specular microscopy were used to assess the family members. RESULTS: Two distinct forms of the disease were identified. All patients with the centroperipheral form were hypermetropic and had keratometry readings below 41.00 diopters and a central corneal thicknesses less than 0.50 mm. Those with the less severe peripheral form were less hypermetropic, some slightly myopic, and had keratometry readings above 41.00 diopters, but the central corneal thicknesses was similar to those with the centroperipheral form. No abnormalities of the endothelium were detected, and visual acuity was only mildly affected. The condition appears to be nonprogressive. CONCLUSION: Though the centroperipheral form of posterior amorphous corneal dystrophy is more likely to lead to presentation, most patients are asymptomatic. This dystrophy can be very subtle in its appearance and easily overlooked. This led the authors to suspect that the prevalence of this condition is higher than the few reports in the ophthalmic literature suggest.

Morphologic assessment of corneal endothelium by specular microscopy in evaluation of donor corneas for transplantation.

Our purpose was to evaluate the role of specular microscopy in the assessment of donor corneas for transplantation. We conducted retrospective analysis of specular microscopic evaluations of 1,000 consecutive donor corneas processed at Mayo Clinic Eye Bank from 1986 to 1993. Thirty-four of the 1,000 corneas were excluded from transplantation use on the basis of specular microscopic examination. Twenty-four corneas were excluded because of the presence of dark spots on the endothelium that did not clear with time. Large endothelial cells were found in six corneas on inspection, with a mean cell density of 1,160 cell/mm2 (795-1,597 cells/mm2). The remaining four excluded corneas showed evidence of endothelial trauma. Of 966 corneas not excluded, 520 (mean cell density 2,632 cells/mm2, range 1,621-4,590 cells/mm2) were transplanted at the Mayo Clinic, and the rest were distributed for transplantation elsewhere, when possible. Six of the corneas transplanted at the Mayo Clinic (1.2%) failed primarily. There ware no significant differences in the preoperative characteristics of the donor corneas between the donor failures and the clear grafts. Specular microscopic examination excluded 3.4% of donor corneas on the basis of unsatisfactory endothelium. Despite examination of the endothelium, six of 520 transplanted corneas (1.2%) suffered primary graft failure. Morphologic assessment of donor corneal endothelium by specular microscopy probably lessens, but does not eliminate, the risk of primary donor failure.

A quantitative study of the lateral spread of Müller cell responses to retinal lesions in the rabbit.

A wide variety of retinal pathology is associated with an increase in Müller glial cell expression of glial fibrillary acidic protein (GFAP). In this study the time course and spatial spread of the Müller cell GFAP response following argon laser photocoagulation lesions was examined in wholemounted rabbit retina. At 24 hours single focal lesions were surrounded by GFAP positive Müller cell end feet which declined in density with distance but extended as far as 2-3 mm from the lesion. The Müller cell reaction reached a maximal spread of 4-5 mm at 14 to 21 days and had started to contract by 30 days, leaving a core of GFAP positive processes immediately around the lesion site at 60 days. This zone of spread was much larger than the area of disrupted pigment epithelium. Isodensity plots did not reveal any correlation with the trajectory of retinal ganglion cell axons. The spread of reaction was more confined for lesions within the visual streak than in the dorsal or ventral retinal periphery. Multiple lesions within a focal region of retina resulted in a greater density of GFAP reactive end feet with a corresponding greater spread. However, when five to ten lesions were made in a horizontal row, the Müller cells over the entire retina became GFAP immunoreactive. This pan-retinal reaction took several days to spread, peaked at 7-14 days, and contracted back to the primary lesion sites by 2 months. This spread of Müller cell reactivity may be triggered by the diffusion of substances released by injury or it may be due to direct cellular communication. The extensive indirect effect on Müller cells of laser irradiation might be an important component of the clinical effect of laser photocoagulation and indicates a long distance communication mechanism between retinal glia which is poorly understood. This study also shows the importance of the time at which the Müller cell response is assessed.

Biological evidence of genetic exchange in Entamoeba histolytica.

The demonstration of a new zymodeme of Entamoeba histolytica produced in culture from cloned isolates suggested possible genetic exchange in this parasite. We have attempted to substantiate that finding by using rats as biological hosts. Clones were made from 3 separate isolates of E. histolytica, each established in culture from liver pus or faeces. After enzyme characterization these clones, of zymodemes II, XIV and XIX, were paired in each of the 3 possible combinations and the mixtures injected into the caecum of rats. Clones of new or hybrid zymodemes were produced as well as the original parents, with one exception, the mixture of XIV and XIX, from which only one of the parents was recovered. The hybrids produced included zymodeme XX, observed previously, and zymodeme XI, a naturally occurring zymodeme that has in the past been recovered only from subjects with invasive amoebiasis.