The long-term objectives of this research are to define molecular mechanisms underlying interactions of dietary antioxidants with cellular and environmental oxidants in vivo by the application of global gene expression analysis. The availability of transgenic mice with selective deletion of the gene for alpha-tocopherol (a-TC) transfer protein, and the resulting depletion of tissue a-TC (vitamin E), offers a unique opportunity to rigorously examine the role of a major dietary antioxidant of biological membranes and lipoproteins in normal physiology. In addition, its potential role as a dietary supplement to intervene in the pathogenesis of chronic diseases triggered by oxidant pollutants in the environment can also be tested.
The specific aims of this proposal are to: (1) define and compare the mRNA expression profiles of lungs and livers from a-tocopherol transfer protein knockout (TTP-KO) and wild type mice; (2) assay the changes in mRNA expression profiles of lungs and livers in response to a photochemically-generated environmental oxidant, ozone, in TTP-KO and wild type mice; and (3) evaluate the changes in the expression of proteins encoded by selected mRNAs, identified through specific aims 1 and 2, by immunoblot and immuno- histochemical analysis. The proposed research will identify, for the first time, the genomic response in vivo to silencing of a gene whose product plays a significant role in the homeostasis of a primary antioxidant in membranes lipoproteins. In addition, global gene expression analysis will also identify the molecular targets of a- TC that may not be directly related to the scavenging of reactive oxygen species and the molecular response to depletion of TTP not directly related to the transfer of a-TC. Gene expression analysis following exposure to an environmental oxidant, ozone, whose primary target is the membrane milieu, will identify the """"""""transcriptional"""""""" response in vivo to oxidative stress inflicted by an environmental pollutant and the role of a dietary antioxidant, vitamin E, on oxidant-induced changes in gene expression in the target tissues. The analysis of proteins encoded by selected mRNAs will establish the potential functional implications of changes in the mRNAs detected by analysis with gene arrays. These molecular data may provide the basis for designing more effective protocols for dietary interventions with antioxidants that might ameliorate lung diseases induced by environmental oxidants.
