Numerous drugs which are systemically administered manifest their oculotoxicities more often in the anterior tissues (e.g. increased intraocular pressure/glaucoma, cataracts/corneal opacity) than in the posterior tissues (e.g. retinopathy and optic neuropathy). Since cytotoxic drug metabolites reach the nonvascular target tissues (cornea and lens) via the aqueous humor, it is important that the drug metabolizing activities of the ciliary body function efficiently for the production of """"""""drug-free"""""""" aqueous humor. Of various drug metabolizing activities, cytochrome P450-mediated activities which metabolize many of the systemic drugs are of primary importance for this function of the ciliary body. The focus of this application, therefore, is to investigate cytochrome P450-mediated drug metabolizing activities and their genetic expression in the ciliary body. There are four specific aims. 1. We have found that the lack of genetic responsiveness to inducers of P450-mediated aryl hydrocarbon hydroxylase in two inbred mouse strains, i.e. responsive C57BL/6 and nonresponsive DBA/2, is inherited in the eye as a Mendelian autosomal recessive trait. Expression of P450 genes and their gene products in different ocular tissues of inducer-treated mice is investigated with appropriate probes by in-situ hybridization and immunocytochemistry. 2. We have previously studied an acetaminophen model of oculotoxicity in inducer-responsive mice. Using this model it is investigated why the ciliary body and other anterior tissues are more susceptible to drug toxicity than posterior tissues in the eye. 3. We have developed a method to isolate ciliary nonpigmented epithelial cells and pigmented epithelial cells from porcine eye and showed that induction of P450-mediated activities by treatment with inducers occurs mainly in primary cultures of the nonpigmented epithelial cells. It is investigated why there are marked differences in the inducibility of P450 enzymes between the two juxtaposed epithelial cells. 4. Using cell cultures of ciliary epithelium we have shown induction by clofibrate of peroxisomal acyl CoA oxidase activity and microsomal P450- mediated activities involved in eicosanoid metabolism. It is investigated whether induction of these enzyme activities associated with different cellular organelles is mediated by the same clofibrate receptor (or receptor complex) and what the nature of the receptor is.
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