Heme oxygenase-1 gene expression as a stress index to ocular irritation

Purpose. Predicting the toxic potential of compounds to the ocular surface has depended on the Draize test for the past half century. Alternatives to Draize testing have recently been sought for a number of reasons. Stress ge ne expression has emerged as a means of quantifying cellular reaction and, thus, the toxic potential of the compound in question. This study examines the expression of the major stress response gene heme oxygenase-1 (HO-1) in a human corneal epithelial cell line (HCE-T) following challenges with a n umber of known ocular irritants. Methods. HCE-T was used to investigate the effect of ocular irritants on ce ll viability and HO-1 expression. Irritants tested included hydrogen peroxi de, isopropyl alcohol, sodium hydroxide and trichloroacetic acid. HCE-T cel ls were grown to 80% confluency and treated with the listed irritants at a concentration range of 10-100 mu M Cell viability and northern blot analysi s were performed following a 24 and 48 hr incubation period. Results. HCE-T cells expressed HO-1 mRNA and HO activity similar to other h uman cell lines. Northern blot analysis demonstrated that levels of HO-1 mR NA transcripts increased regularly after exposure to the irritants in a con centration-dependent manner. Studies on the effect of various inhibitors an d inducers of HO-1 on cell viability showed that inhibition of HO-1 potenti ates the cytotoxic effect of ocular irritants. In contrast, pre-induction o f HO-1 in HCE-T decreases the effect of various irritants on cell viability . Conclusions. These results are consistent with the idea that HO-1 mRNA leve ls may be used as an indicator of toxicity resulting from ocular irritants and that HCE-T cells respond to stress in a fashion similar to other human cell lines. This strategy for testing may be important in the development o f an alternative to Draize testing. The results of this study also suggest that HO-1 may constitute a part of the protective defense mechanism against chemical injury.