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Darrell Robert Lewis, Mark Seamone, Christopher Seamone; The Resistance to Collagenase Dissolution of Cross-Linked Cryopreserved Donor Corneas. Invest. Ophthalmol. Vis. Sci. 2016;57(12):2380.
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© ARVO (1962-2015); The Authors (2016-present)
To determine if UV-A riboflavin cross-linking (CXL) of donor corneas after cryopreservation confers resistance to keratolysis as has been described for fresh donor corneas.
Three methods of donor cornea cryopreservation were compared to donor corneas stored in Optisol-GS at 4°C for less than 2 weeks (control). The cryopreservation arms consisted of corneas stored in Optisol-GS at -15°C, Optisol-GS at -70°C, and ≥99% anhydrous glycerol at -80°C. Within each cryopreservation method, corneas were randomized to either receive CXL or not. The primary outcome measure was the time to complete corneal dissolution in a collagenase-A solution.Donor corneas cryopreserved in Optisol-GS at -15°C (n=4), Optisol-GS at -70°C (n=4), and ≥99% glycerol at -80°C (n=4) were thawed for 30 minutes in a room temperature balanced salt solution. Corneas were mounted on artificial anterior chambers and de-epithelialized. Pre-treatment with 0.1% riboflavin in dextran solution every 2 minutes for 15 minutes was undertaken. High fluence, accelerated CXL (9 mW/cm2, 10 min) was then performed. As controls, 4 donor corneas matching each of the cryopreservation methods were thawed but did not undergo CXL. Additionally, 4 fresh donor corneas did not undergo CXL. Corneas were punched with an 8mm Baron trephine. These 8 mm corneal buttons were agitated in a 0.3% collagenase-A solution at 37°C. Corneas were examined every 30 minutes until complete dissolution occurred.Data was analysed via Mann-Whitney U-test and Kruskal-Wallis analysis where appropriate.
Donor corneas cryopreserved in ≥99% glycerol at -80°C required a statistically significantly longer time for complete corneal dissolution than fresh donor corneas (p=0.03). Dissolution times for donor corneas cryopreserved in Optisol-GS at either -15°C or -70°C did not statistically significantly differ from fresh donor corneas, p=0.13 and p=0.26 respectively.
Cross-linking donor corneas cryopreserved in ≥99% glycerol at -80°C increases their resistance to keratolysis. Donor corneas cryopreserved in Optisol-GS have a similar resistance to keratolysis as fresh donor corneas.The ability to enhance cryopreserved donor corneas via CXL may prove advantageous to reduce graft failure rates. Cryopreserved tissue may be appropriate for deep anterior lamellar keratoplasty, glaucoma tube shunt coverage, Boston keratoprosthesis carriers, and tectonic grafting for melts.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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