The Center for Alternatives to Animal Testing is an academic center affiliated with the Division of Toxicological Sciences in the Department of Environmental Health Sciences of the Johns Hopkins University Bloomberg School of Public Health.

 

Johns Hopkins School of Public Health

Research Grants 2001-2002

Bovine Corneal Organ Culture: An Ex Vivo Model for Chemical Toxicity Tests

Fu-Shin Yu, PhD
The Schepens Eye Research Institute, Harvard University, Boston, Massachusetts

For five decades, the Draize test has remained the accepted method for evaluating the potential of test material to cause eye irritation or injury. Criticisms of this method center around the inhumane treatment of animals and the irreproducibility of the subjective scoring procedure. There is a great demand for a mechanistic based in vitro testing system that will minimize the use of animals in chemical toxicity tests. Recently, we have adapted a simple, long-term organ culture method as an ex vivo model for chemical toxicity tests. Here, toxicity can be assessed by applying test chemicals to the surface of the cultured corneas. Since this system more closely resembles an in vivo testing system than cell culture, it should serve as an appropriate model for chemical safety tests. The corneas we use are prepared from the bovine (or porcine) eyes, economical and resourceful by-products of meat industry, no live animals are euthanized for testing. Using this system, we will examine the corneal response to chemicals in several paradigms.

  1. The activation of AP- I and NF-kB, two well-known stress-responsive proteins controlling gene expression, in response to chemical stimuli will be used as endpoints for ocular irritancy.
  2. Disruption of corneal barrier function by test chemicals will assess by opacity, epithelial fluorescein retention, and leakage of metabolic enzymes.
  3. Recovery of epithelial function after exposure of the corneal organ culture to tested chemicals will be monitored and used for assigning final ratings of chemicals.

To date, our analyses have demonstrated that corneal organ culture, in combination with the measurements of corneal epithelial permeability and DNA-binding activity of stress-response transcription factors, presents a suitable model for evaluating ocular irritation potential of the innocuous, mild, moderate, and severe irritants. This ex vivo system might be used as an alternative to in vivo Draize test.