The long-term goal of this application is to elucidate the role of oxidative stress in the biological effects of DEPs in the respiratory tract. We hypothesize that generation of oxidative stress by organic chemicals in DEPs (DEP xenobiotics) initiates a stratified pulmonary response that commences with the: 1) activation of the ARE pathway, which transitions to 2) excitation of inflammation and 3) cell injury in the lung, with all three responses being critical to enhancement of asthma. To elucidate how DEP chemicals interact with oxidative stress pathways in the lung, we will employ a very sensitive system for analyzing oxidative stress by oxidative xenobiotics, including those in DEP. This pathway, which utilizes the transcription factor Nrf2, induces the expression of the oxidative stress enzyme, HO-1 via AREs. HO-1, in turn, has potent antioxidant, anti-inflammatory and anti-apoptotic effects in the lung.
In Specific Aim 1, we will test the hypothesis that the effect of DEP on HO-1 expression in vitro is mediated by redox cycling DEP chemicals that operate via Nrf2 and AREs in the promoter of that gene. We will test the effect of HO-1 for its ability to protect epithelial cells and macrophages against the oxidative stress effects of DEP, using drugs that modulate HO-1 expression or HO-1 activity.
In Specific Aim 2 we will test the hypothesis that the pro-inflammatory and pro-allergic effects of DEP in the murine lung are related to oxidative stress and can be modified by modulating HO-1 expression through genetic or pharmacological means. This will be accomplished by testing intratracheal DEP instillation on HO-1 expression, along with other markers for oxidative stress, inflammation and apoptosis. We will also determine whether pharmacological modification of HO-1 activity and transgenic HO-1 overexpression influence the adjuvant effects of DEP in the murine asthma model. We will use the same model, in collaboration with Project 4, to determine the effects of Nrf2 knockout on the pro-oxidative and allergic/inflammatory effects of DEP.
In Specific Aim 3, we will test the hypothesis that interference in HO-1 expression by a known poly-(GT) polymorphism in the human HO-1 gene enhances the pro-allergic effects of DEP in humans. This will be accomplished in collaboration with Projects 1 and 3, and will involve genotyping and intranasal DEP challenge followed nasal lavage and studying the parameters for oxidative stress and allergic inflammation.
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