The overall objective of this proposed study is to further understanding of how inflammatory immune factors interact with and impact the natural healing response of host epithelial cell surfaces to infectious damage. Knowledge of mechanisms driving the airway epithelial reparative function in response to damage caused by infectious disease has important implications for development of therapies for inadequate respiratory function caused not only by infectious disease but also common respiratory disorders such as asthma, which involves frequent interaction with inflammatory factors and airway epithelial remodeling. Based on previous studies, we hypothesize that IL-1beta (IL-1?), a pleiotropie cytokine produced downstream of the cytosolic viral sensing pathway components NLRP3 and Caspase-1, is critical for the proper airway epithelial surface healing response to necrotic and apoptotic damage caused by influenza A virus PRS (H1N1) infection.
Aims I and II will use in vivo mouse models to characterize the timing and source of """"""""healing"""""""" IL-1?, while Aim III involves in vitro primary mouse tracheal epithelial cell (mTEC) culture to study the direct effect of lL-1? on the reparative response of influenza-damaged cell layers. For in vivo studies, the timing of """"""""healing"""""""" IL-1? will be determined using two complementary methods. First, anti-lL1? will be used to temporally remove IL-1? from the respiratory system during early-stage or late-stage influenza infection in wild-type BL/6 mice. Second, recombinant IL-1? (rIL-1?) will be added to the lung environment at the stated stages during infection in mice deficient in IL-1beta signaling;these mice include IL-1? -/-, NLRP3-/-, and Caspase-1-/- mice. Determination of impact on lung epithelial healing responses will be done by histological examination of lung tissues for signs of pneumonia, damage, and attempts at healing functions such as collagen deposition and fibrosis. Impact of IL-1? on lung function will be evaluated by blood gas measurements on mice at peak infection and post-recovery stages. Cellular sources of """"""""healing"""""""" 1L-1? will be determined by evaluation of stated epithelial reparative functions in infected bone marrow chimeric mice in which one cell compartment or the other lacks the ability to produce IL-1?.
Aim III in vitro studies will use make use of anti-lL-1? Ab and rtL-1? to remove or add in lL-1? to BL/6 or IL-1? signaling pathway-deficient cell types during the response to influenza infection. Measures of epithelial recovery include BrdU incorporation for proliferation and TEER measurements and tight junction staining to measure cell layer integrity. Public health: Influenza infection causes damage to the cells that line the lungs. This proposal seeks to understand the role of a specific natural protein in promoting the lung cells'healing.

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
Postdoctoral Individual National Research Service Award (F32)
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Special Emphasis Panel (ZRG1-F07-K (20))
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Colombini-Hatch, Sandra
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St. Jude Children's Research Hospital
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Sanders, Catherine J; Johnson, Brian; Frevert, Charles W et al. (2013) Intranasal influenza infection of mice and methods to evaluate progression and outcome. Methods Mol Biol 1031:177-88
Sanders, Catherine J; Vogel, Peter; McClaren, Jennifer L et al. (2013) Compromised respiratory function in lethal influenza infection is characterized by the depletion of type I alveolar epithelial cells beyond threshold levels. Am J Physiol Lung Cell Mol Physiol 304:L481-8
Askovich, Peter S; Sanders, Catherine J; Rosenberger, Carrie M et al. (2013) Differential host response, rather than early viral replication efficiency, correlates with pathogenicity caused by influenza viruses. PLoS One 8:e74863
Sanders, Catherine J; Doherty, Peter C; Thomas, Paul G (2011) Respiratory epithelial cells in innate immunity to influenza virus infection. Cell Tissue Res 343:13-21