The Draize eye irritation test has been in use since 1944. One eye of a conscious rabbit is instilled with the test substance, while the other eye is the control. The reactions of the cornea, conjunctiva, and iris are then evaluated in a period of one hour to three weeks. Because of the test's moral and scientific short comings, a replacement has been actively sought, but there is still none in the horizon. Today, the Draize test remains the only one that can classify a substance into one of the four categories of eye irritation: corrosive/severe, moderate, mild, and non- irritants. Specifically, none of the in viro methods can identify the moderate and mild irritants and the Draize test continues. None of the in vitro method addresses the issue of ocular inflammation, an integral part of the Draize eye irritation test brought upon by dying cells following test substance instillation. We hypothesize that an in vitro method with an end point that can predict the inflammatory potential of the injured eye can identify moderate and mild irritants, in addition to the corrosive/severe and non- irritants. Using human donor corneas not used for transplantation, our preliminary studies demonstrated in a totally animal (and animal product)-free system that the levels of corneal endothelial damage, revealed by alizarin red S staining and quantified by image analysis of the red stain in the panoramic view of the corneal endothelium, predicted with 100% accuracy the irritation potential of 18 test substances in the corrosive/severe, moderate, mild, and non- irritants categories. Our goal is to demonstrate that our method can fully replace the Draize eye irritation test.
The specific aims are to 1) use our method to classify test substances that have historical Draize test data available in public, 2) study the status of VIP in the damaged corneal endothelium in the test substance-treated human donor corneas.

Public Health Relevance

There are many chemicals in our environment and in products we use every day, such as cosmetics, hairspray, and household cleaners. One of the important safety tests required for new chemical products is to evaluate the potential to cause eye injury. This has traditionally (since 1944) been accomplished by applying chemicals to the eyes of live rabbits. This test is overwhelmingly viewed as inhumane. There is growing consensus among scientists and a federal mandate to reduce or eliminate the use of animals in chemical safety testing. Of the available in vitro methods developed over the years, only four are currently accepted by regulators to identify chemicals as severe eye irritants vs others or non-irritants vs others. Specifically, none of the in vitro methods can identify the moderate and mild irritants and the live rabbit eye test remains the only one that can identify eye irritants inall categories. We have developed a new method for chemical safety testing that uses available resources (human donor corneas not used for transplantation) and does not use animals or any animal products, which is the ultimate goal of alternative method development. Early results indicate that it is a reliable (in 100% agreement with the live rabbit eye test) and more sensitive method to classify chemicals in all categories of eye irritants.

National Institute of Health (NIH)
National Eye Institute (NEI)
Exploratory/Developmental Grants (R21)
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Special Emphasis Panel (BNVT)
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Mckie, George Ann
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University of Maryland Baltimore
Schools of Medicine
United States
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