The long term objective is to understand the mechanisms by which lung tumor growth is enhanced by non-genotoxic compounds and to identify the genes which regulate susceptibility to this process. The food additive, butylated hydroxytoluene (BHT), encourages the development of tumors from previously initiated cells. The quinone methide, QM-OH, is derived from a hydroxylated metabolite of BHT, BHT-OH, and is the probable reactive species that promotes tumors. These metabolic reactions occur in the bronchiolar nonciliated Clara cells, one of the cell types from which these tumors arise. Inbred strains vary in their susceptibility to BHT- induced lung tumor promotion. Chronic administration of BHT to inbred mice down-regulates the pulmonary concentrations of the alpha isozyme of protein kinase C (PKCa) and calpain in promotion-sensitive strains but not in promotion-resistant strains. PKC content is inversely correlated with lung growth; low PKC concentrations are associated with rapid lung cell proliferation. We will identify susceptibility genes by high resolution mapping using simple sequence length polymorphism analysis, and apply this technique to support the hypothesis that genes which regulate BHT metabolism, PKCa, calpain, and K-ras are candidate genes for susceptibility. We will test whether the CYPIIB gene is also a likely candidate gene. We will study respiratory epithelial cells in vivo and Clara cells isolated from promotion-sensitive vs. -resistant strains following chronic BHT exposure to: analyze the kinetics and mechanism of calpain and PKCa down-regulation; examine the phosphorylation status of key proteins in the K-ras signal transduction pathway known to be regulated by PKC and calpain; and test whether BHT treatment affects the susceptibility to programmed cell death and ability to undergo gap junctional intercellular communication.
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