• 1. Rehman AA, Ahsan H, Khan FH. α-2-macroglobulin: a physiological guardian. J Cell Physiol 2013;228:16651675.

  • 2. Borth W. α2-macroglobulin, a multifunctional binding protein with targeting characteristics. FASEB J 1992;6:33453353.

  • 3. Haffner JC, Fecteau KA, Eiler E. Inhibition of collagenase breakdown of equine corneas by tetanus antitoxin, equine serum and acetylcysteine. Vet Ophthalmol 2003;6:6772.

    • Search Google Scholar
    • Export Citation
  • 4. Ollivier FJ, Brooks DE, Kallberg ME, et al. Evaluation of various compounds to inhibit activity of matrix metalloproteinases in the tear film of horses with ulcerative keratitis. Am J Vet Res 2003;64:10811087.

    • Search Google Scholar
    • Export Citation
  • 5. Burns FR, Stack MS, Gray RD, et al. Inhibition of purified collagenase from alkali-burned rabbit corneas. Invest Ophthalmol Vis Sci 1989;30:15691575.

    • Search Google Scholar
    • Export Citation
  • 6. Conway ED, Stiles J, Townsend WM, et al. Comparison of the in vitro anti-collagenase efficacy of homologous serum and plasma on degradation of corneas of cats, dogs, and horses. Am J Vet Res 2016;77:627633.

    • Search Google Scholar
    • Export Citation
  • 7. Anitua E, Muruzabal F, Tayebba A, et al. Autologous serum and plasma rich in growth factors in ophthalmology: preclinical and clinical studies. Acta Ophthalmol 2015;93:e605e614.

    • Search Google Scholar
    • Export Citation
  • 8. Lopez-Garcia JS, Murube del Castillo J, Garcia Lozano I, et al. Autologous serum and blood derivatives in ophthalmology. Arch Soc Esp Oftalmol 2012;87:376377.

    • Search Google Scholar
    • Export Citation
  • 9. Brion M, Lambs L, Berthon G. Metal ion-tetracycline interactions in biological fluids. Part 5. Formation of zinc complexes with tetracycline and some of its derivatives and assessment of their biological significance. Agents Actions 1985;17:229242.

    • Search Google Scholar
    • Export Citation
  • 10. Baker A, Plummer CE, Szabo NJ, et al. Doxycycline levels in preocular tear film of horses following oral administration. Vet Ophthalmol 2008;11:381385.

    • Search Google Scholar
    • Export Citation
  • 11. Monk CS, Jeong SY, Gibson DJ, et al. The presence of minocycline in the tear film of normal horses following oral administration and its anticollagenase activity. Vet Ophthalmol 2018;21:5865.

    • Search Google Scholar
    • Export Citation
  • 12. Collins SP, Labelle AL, Dirikolu L, et al. Tear film concentrations of doxycycline following oral administration in ophthalmologically normal dogs. J Am Vet Med Assoc 2016;249:508514.

    • Search Google Scholar
    • Export Citation
  • 13. Chandler HL, Gemensky-Metzler AJ, Bras ID, et al. In vivo effects of adjunctive tetracycline treatment on refractory corneal ulcers in dogs. J Am Vet Med Assoc 2010;237:378386.

    • Search Google Scholar
    • Export Citation
  • 14. Berman MB. Collagenase inhibitors: rationale for their use in treating corneal ulceration. Int Ophthalmol Clin 1975;15:4966.

  • 15. Epstein SP, Ahdoot M, Marcus E, et al. Comparative toxicity of preservatives on immortalized corneal and conjunctival epithelial cells. J Ocul Pharmacol Ther 2009;25:113119.

    • Search Google Scholar
    • Export Citation
  • 16. Furrer P, Mayer JM, Plazonnet B, et al. Ocular tolerance of preservatives on the murine cornea. Eur J Pharm Biopharm 1999;47:105112.

  • 17. Hook CW, Brown SI, Iwanij W, et al. Characterization and inhibition of corneal collagenase. Invest Ophthalmol 1971;10:496503.

  • 18. Ramaesh T, Ramaesh K, Riley SC, et al. Effects of N-acetylcysteine on matrix metalloproteinase-9 secretion and cell migration of human corneal epithelial cells. Eye (Lond) 2012;26:11381144.

    • Search Google Scholar
    • Export Citation
  • 19. Thermes F, Molon-Noblot S, Grove J. Effects of acetylcysteine on rabbit conjunctival and corneal surfaces. A scanning electron microscopy study. Invest Ophthalmol Vis Sci 1991;32:29582963.

    • Search Google Scholar
    • Export Citation
  • 20. Aldavood SJ, Behyar R, Sarchahi AA, et al. Effect of acetylcysteine on experimental corneal wounds in dogs. Ophthalmic Res 2003;35:319323.

    • Search Google Scholar
    • Export Citation
  • 21. Angleton EL, Van Wart HE. Preparation and reconstitution with divalent metal ions of class I and class II Clostridium histolyticum apocollagenases. Biochemistry 1988;27:74067412.

    • Search Google Scholar
    • Export Citation

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Comparison of the efficacy of various concentrations and combinations of serum, ethylenediaminetetraacetic acid, tetracycline, doxycycline, minocycline, and N-acetylcysteine for inhibition of collagenase activity in an in vitro corneal degradation model

Beth A. Kimmitt DVM1, George E. Moore DVM, PhD2, and Jean Stiles DVM, MS3
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  • 1 Department of Veterinary Clinical Sciences, College of Veterinary Medicine, Purdue University, West Lafayette, IN 47907.
  • | 2 Veterinary Administration, College of Veterinary Medicine, Purdue University, West Lafayette, IN 47907.
  • | 3 Department of Veterinary Clinical Sciences, College of Veterinary Medicine, Purdue University, West Lafayette, IN 47907.

Abstract

OBJECTIVE To compare the efficacy of various concentrations and combinations of serum, EDTA, 3 tetracyclines, and N-acetylcysteine (NAC) for collagenase inhibition in an in vitro corneal degradation model.

SAMPLE Grossly normal corneas from recently euthanized dogs and horses and fresh serum from healthy dogs and horses.

PROCEDURES Serum was pooled by species for in vitro use. For each species, sections of cornea were dried, weighed, and incubated with clostridial collagenase (800 U/mL) in 5 mL of a 5mM calcium chloride-saline (0.9% NaCl) incubation solution and 500 μL of 1 of 19 treatments (homologous serum; 0.3%, 1.0%, or 2% EDTA; 0.1%, 0.5%, or 1.0% tetracycline, doxycycline, or minocycline; 0.5%, 1.0%, or 5.0% NAC; serum with 0.5% tetracycline; serum with 1.0% EDTA; or 1.0% EDTA with 0.5% tetracycline). Positive and negative control specimens were incubated with 5 mL of incubation solution with and without collagenase, respectively. Each control and treatment was replicated 4 times for each species. Following incubation, corneal specimens were dried and reweighed. The percentage corneal degradation was calculated and compared among treatments within each species.

RESULTS Treatments with tetracyclines at concentrations ≥ 0.5%, with EDTA at concentrations ≥ 0.3%, and with NAC at concentrations ≥ 0.5% were more effective at preventing corneal degradation than serum in both species. The efficacy of each combination treatment was equal to or less than that of its components.

CONCLUSIONS AND CLINICAL RELEVANCE Results suggested EDTA, tetracyclines, and NAC may be beneficial for topical treatment of keratomalacia, but in vivo studies are required.

Contributor Notes

Address correspondence to Dr. Stiles (stilesj@purdue.edu).