The long term goal of this application is to elucidate the in vivo significance and molecular mechanisms of integrin-mediated activation of transforming growth factor beta (TGFbeta). In current and previous cycles of this grant we have shown that the integrin, alphavbetaS, activates TGFbeta and that this process plays a central role in models of pulmonary fibrosis and acute lung injury in mice. We have also identified three stimuli, thrombin, LPA and S1P that can each activate this process be initiating signals from G protein coupled receptors. In this application we propose to first determine the importance of this pathway in a more clinically relevant model of acute lung injury that combines low dose LPS with normal tidal volume ventilation. For these experiments we will also determine whether any protection seen with loss or blockade of the integrin is specifically due to loss of the integrin from alveolar epithelium and/or to loss of TGFbeta activation using mice we have generated expressing either the integrin or active TGFbeta as alveolar epithelial transgenes. In parallel, we will explore the mechanisms underlying indcution of this process through activation of G protein coupled receptors. We will use cells from mice lacking Galpha12, Galpha13 or Galphai and shRNA designed to knockdown each subunit to identify the G proteins involved in these responses. We will take a similar approach to identify the relevant LPA and S1P receptors. To genetically determine the relevance of actin-myosin mediated contraction, and to identify the specific myosin isoforms invovled, we will also use shRNA-mediated knockdown of each of the three myosin II isoforms we have identified in lung epithelial cells. To determine the significance of cell contraction we will also quantify alphavbeta6-medated TGFbeta activation by cells plated on substrates of varying stiffness. Finally, we will use labeled forms of the integrin and a TGFbeta receptor to perform live videomicroscopy and test the idea that this process involves the formation of a TGFbeta activation synapse across adjacent cells. Lay summary: This application seeks to understand a key process that controls pulmonary fibrosis and acute lung injury, two common and currently largely untreatable diseases. By determining the mechansims by which cells in the lung control this process, we should be able to identify and validate novel targets for design of effectiive treatments for these diseases.

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
Method to Extend Research in Time (MERIT) Award (R37)
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Special Emphasis Panel (NSS)
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Eu, Jerry Pc
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University of California San Francisco
Internal Medicine/Medicine
Schools of Medicine
San Francisco
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Erle, David J; Sheppard, Dean (2014) The cell biology of asthma. J Cell Biol 205:621-31
Sheppard, Dean (2013) ROCKing pulmonary fibrosis. J Clin Invest 123:1005-6
Friedman, Scott L; Sheppard, Dean; Duffield, Jeremy S et al. (2013) Therapy for fibrotic diseases: nearing the starting line. Sci Transl Med 5:167sr1
Su, George; Atakilit, Amha; Li, John T et al. (2013) Effective treatment of mouse sepsis with an inhibitory antibody targeting integrin *v*5. Crit Care Med 41:546-53
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Hogmalm, Anna; Sheppard, Dean; Lappalainen, Urpo et al. (2010) beta6 Integrin subunit deficiency alleviates lung injury in a mouse model of bronchopulmonary dysplasia. Am J Respir Cell Mol Biol 43:88-98
Melton, Andrew C; Bailey-Bucktrout, Samantha L; Travis, Mark A et al. (2010) Expression of *v*8 integrin on dendritic cells regulates Th17 cell development and experimental autoimmune encephalomyelitis in mice. J Clin Invest 120:4436-44
Atabai, Kamran; Jame, Sina; Azhar, Nabil et al. (2009) Mfge8 diminishes the severity of tissue fibrosis in mice by binding and targeting collagen for uptake by macrophages. J Clin Invest 119:3713-22
Singh, Purva; Chen, Chun; Pal-Ghosh, Sonali et al. (2009) Loss of integrin alpha9beta1 results in defects in proliferation, causing poor re-epithelialization during cutaneous wound healing. J Invest Dermatol 129:217-28
Xu, Ming Yan; Porte, Joanne; Knox, Alan J et al. (2009) Lysophosphatidic acid induces alphavbeta6 integrin-mediated TGF-beta activation via the LPA2 receptor and the small G protein G alpha(q). Am J Pathol 174:1264-79

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