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  • Author or Editor: Gregory S. Schultz x
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Abstract

OBJECTIVE

To develop and assess a novel ex vivo corneal culture technique involving an agarose-based dome scaffold (ABDS) for use as a model of in vivo corneal wound healing in dogs and rabbits.

SAMPLE

Corneas from clinically normal dogs (paired corneas from 8 dogs and 8 single corneas) and rabbits (21 single corneas).

PROCEDURES

8 single dog corneas (DCs), 1 DC from each pair, and 10 rabbit corneas (RCs) were wounded with an excimer laser; 1 DC from each pair and 11 RCs remained unwounded. Corneas were cultured for 21 days on ABDSs (8 pairs of DCs and all RCs) or on flat-topped scaffolds (8 single DCs). The surface area of corneal fluorescein retention was measured every 6 (DCs) or 12 (RCs) hours until full corneal epithelialization was detected. Changes in corneal clarity were evaluated at 0, 7, 14, and 21 days.

RESULTS

Median time to full epithelialization for wounded dog and rabbit corneas was 48 and 60 hours, respectively; among wounded DCs, time to full epithelization did not differ by scaffold type. After 21 days of culture on ABDSs, all DCs and RCs that epithelialized developed a circular, diffuse, cloud-like pattern of optical haze, whereas DCs cultured on flat-topped scaffolds developed a focal, crater-like region of optical haze. All corneas on the ABDSs maintained convex curvature throughout the study.

CONCLUSIONS AND CLINICAL RELEVANCE

Wounded ex vivo DCs and RCs cultured on ABDSs reliably epithelialized, formed optical haze (consistent with in vivo wound healing), and maintained convex curvature. This culture technique may be adaptable to other species.

Full access
in American Journal of Veterinary Research

Abstract

Objective—To examine in vitro effects of various antiproteolytic compounds on activity of matrix metalloproteinase (MMP)-2 and -9 in the tear film of horses with active corneal ulcers.

Sample Population—Samples of tear film obtained from the eyes of 34 horses with active ulcerative keratitis.

Procedure—Horses were sedated, and tear samples were collected from the lower fornix of 34 ulcerated eyes by use of capillary tubes. The protease inhibitors 0.2% EDTA, 0.1% doxycycline, 10% N-acetylcysteine (NAC), 0.1% solution of a modified dipeptide that contains hydroxamic acid (ie, ilomostat), 0.1% α1-proteinase inhibitor (PI), 0.5% α1-PI, and 100% fresh equine serum (ES) were used to treat pooled samples. Amount of latent and active MMP-2 and -9 was measured by optical density scanning of gelatin zymograms of treated and untreated tear samples.

Results—Pooled tear samples obtained from ulcerated eyes contained the latent and active forms of MMP-2 and -9. Compared with MMP activity in untreated samples, total MMP activity (sum of all bands detected) observed on the gelatin zymogram gels was reduced by 99.4% by EDTA, 96.3% by doxycycline, 98.8% by NAC, 98.9% by ilomostat, 52.4% by 0.1% α1-PI, 93.6% by 0.5% α1-PI, and 90.0% by ES.

Conclusions and Clinical Relevance—We documented that EDTA, doxycycline, NAC, ilomostat, α1- PI, and ES inhibited MMP activity in vitro. Because these compounds use different mechanisms to inhibit various families of proteases in the tear film of horses, a combination of these protease inhibitors may be beneficial for treatment of corneal ulcers in horses. (Am J Vet Res 2003;64:1081–1087)

Full access
in American Journal of Veterinary Research