Hyperoxia is frequently used in the treatment of pulmonary insufficiency in premature infants. However, hyperoxia contributes to the development of bronchopulmonary dysplasia (BPD). The molecular mechanisms of oxygen-mediated lung injury are not understood, but reactive oxygen species (ROS) are likely to play important roles. The central hypothesis of the research proposed in the present application is that cytochrome P450 1A (CYP1A) enzymes, which have been implicated in the formation and/or detoxication of ROS, contribute significantly to the mechanisms governing hyperoxic lung injury, with oxidative DNA lesions playing a critical role in this process. In order to test this hypothesis, we propose the following Specific Aims: 1. To test the hypothesis that mouse and human CYP1A enzymes play important mechanistic roles in the protective effects of 2-naphthoflavone (BNF) against hyperoxic lung injury. (ii) To test the hypothesis that humanized CYP1A1 mice [hCYP1A1-Cyp1a1 (-/-)], or hCYP1A1_1A2-Cyp1a1/1a2 (-/-)] mice on will be more tolerant to hyperoxic injury, and that BNF pre-treatment will further ameliorate oxygen toxicity in these mice. (iii) To test the hypothesis that liver CYP1As contribute to hyperoxic lung injury via the action of pro- inflammatory cytokines, which are released from the hyperoxic lung. 2. To test the hypothesis that mice lacking the gene encoding AHR, nrf2 or NQO1 will be more susceptible to hyperoxic lung injury, and that these mice will be rescued by pre-treatment with BNF. 3. To test the hypothesis that oxidative DNA damage is a critical contributor to lung injury, and that CYP1A1 and/or phase II enzymes play a key role in the detoxification of DNA-reactive ROS, which in turn attenuates hyperoxic lung injury.
This aim has two sub-aims: (i) Specifically, we will determine if pulmonary oxidative DNA lesions would be higher in mice that have greater susceptibility to hyperoxic lung injury. (ii) We will test the hypothesis that tracheal aspirates of premature infants who developed BPD will have higher oxidative DNA lesions, especially those derived from cylcopurines and F2-isoprostane adducts, compared to those who did not. 4.To test the hypothesis that CYP1A1 enzymes play a protective role against cell toxicity in cultured cells, and that oxidative DNA adducts play a mechanistic role in contributing to oxygen mediated toxicities leading to apoptosis and/or necrosis. We propose two sets of experiments in this Specific Aim. (i) To test the hypothesis that pulmonary cells lacking the genes for CYP1A1 would be more susceptible, and cells overexpressing human CYP1A1 (hCYP1A1) gene would be less susceptible to oxygen-mediated toxicity than similarly exposed wild type cells. (ii) To test the hypothesis that oxidative DNA lesions play a mechanistic role in mediating cell toxicity, eventually leading to apoptosis and/or necrosis. The proposed studies should provide conceptual foundation(s) for achieving our long-term goals, which are the development of rational strategies for the prevention and treatment of lung diseases (e.g. BPD and ARDS) associated with hyperoxia.
Hyperoxia is frequently used in the treatment of pulmonary insufficiency in premature infants. However, hyperoxia contributes to the development of bronchopulmonary dysplasia (BPD). Using animal and cell culture models, this project is aimed at developing rational strategies for the prevention and treatment of lung diseases (e.g. BPD and ARDS) associated with hyperoxia.
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