H2O2 is the most important ROS involved in cell signaling in the lung. PMA has been used to study H2O2-mediated signaling, but asbestosis is a good model to explore the role of H2O2 in cell signaling because asbestos is biologically relevant and is known to induce high levels of H2O2. The release of TNF-alpha by alveolar macrophages in asbestosis patients is critical in the development of the disease;however, the mechanism linking H2O2 generation to TNF-alpha production is not known. Another feature of asbestosis is that alveolar macrophages from patients resemble monocytes. Our data show that monocytes produce TNF-alpha when stimulated, in vitro, with asbestos unlike alveolar macrophages. Thus, human monocytes provide a good model to explore TNF-alpha gene expression, as it relates to H2O2 generation. In this regard, our preliminary data show that H2O2 is necessary for optimal production of TNF-alpha in monocytes and that the source of H2O2 is primarily from Cu,Zn-SOD. Our preliminary data also demonstrate that Cu,Zn-SOD is highly expressed in alveolar macrophages obtained from patients with asbestosis. Since our data shows that alveolar macrophages from asbestosis patients have a high level p38 MAP kinase activity and absent ERK activity and that H2O2 activates p38 and inhibits ERK, we hypothesize that H2O2 plays a pivotal role in differentially modulating MAP kinase activity, and Cu,Zn-SOD enhances differential MAP kinase activation and TNF-alpha production in human monocytes. We will use PMA in the initial studies to understand the role of H2O2 in cell signaling in addition to performing studies using asbestos, which is more biologically relevant.
In Aim 1 we will determine if Cu,Zn-SOD-induced H2O2 modulates MAP kinase activation. Since MAP kinase activity is often controlled by phosphatases, we will investigate if H2O2 is oxidizing a phosphatase, MKP-1, and inducing expression of PP2A, which results in p38 activation and ERK inactivation, respectively. The comparison of H2O2 generation and MAP activation in wild-type and Cu,Zn- SOD KO mice will provide biological relevance.
In Aim 2 we will determine if Cu,Zn-SOD-induced H2O2 increases TNF-alpha production in monocytes and if optimal TNF-alpha expression depends on differential MAP kinase activity. We will compare MAP kinase and Cu,Zn-SOD activity and H2O2 generation in alveolar macrophages obtained from asbestosis patients to normal macrophages. Additional biological relevance will be provided by investigating the development of interstitial fibrosis in wild-type and Cu,Zn- SOD KO mice. Project Narrative: Exposure to asbestos and other environmental agents, such as diesel exhaust and particulate matter, increase hydrogen peroxide generation in the lung. The goal of the project is to understand the mechanisms by which hydrogen peroxide causes lung injury and fibrosis. Although the studies in this proposal relate to asbestosis, they will also apply to other inflammatory and fibrotic lung diseases.

Agency
National Institute of Health (NIH)
Institute
National Institute of Environmental Health Sciences (NIEHS)
Type
Research Project (R01)
Project #
5R01ES014871-05
Application #
8197540
Study Section
Innate Immunity and Inflammation Study Section (III)
Program Officer
Nadadur, Srikanth
Project Start
2007-12-01
Project End
2012-11-30
Budget Start
2011-12-01
Budget End
2012-11-30
Support Year
5
Fiscal Year
2012
Total Cost
$183,770
Indirect Cost
$61,257
Name
University of Iowa
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
062761671
City
Iowa City
State
IA
Country
United States
Zip Code
52242
He, Chao; Ryan, Alan J; Murthy, Shubha et al. (2013) Accelerated development of pulmonary fibrosis via Cu,Zn-superoxide dismutase-induced alternative activation of macrophages. J Biol Chem 288:20745-57
Murthy, Shubha; Ryan, Alan J; Carter, A Brent (2012) SP-1 regulation of MMP-9 expression requires Ser586 in the PEST domain. Biochem J 445:229-36
Osborn-Heaford, Heather L; Ryan, Alan J; Murthy, Shubha et al. (2012) Mitochondrial Rac1 GTPase import and electron transfer from cytochrome c are required for pulmonary fibrosis. J Biol Chem 287:3301-12
Scott, Jason A; Xie, Litao; Li, Hui et al. (2012) The multifunctional Ca2+/calmodulin-dependent kinase II regulates vascular smooth muscle migration through matrix metalloproteinase 9. Am J Physiol Heart Circ Physiol 302:H1953-64
He, Chao; Murthy, Shubha; McCormick, Michael L et al. (2011) Mitochondrial Cu,Zn-superoxide dismutase mediates pulmonary fibrosis by augmenting H2O2 generation. J Biol Chem 286:15597-607
Murthy, Shubha; Ryan, Alan; He, Chao et al. (2010) Rac1-mediated mitochondrial H2O2 generation regulates MMP-9 gene expression in macrophages via inhibition of SP-1 and AP-1. J Biol Chem 285:25062-73
Williard, Deborah E; Twait, Erik; Yuan, Zuobiao et al. (2010) Nuclear factor kappa B-dependent gene transcription in cholecystokinin- and tumor necrosis factor-alpha-stimulated isolated acinar cells is regulated by p38 mitogen-activated protein kinase. Am J Surg 200:283-90
Murthy, Shubha; Adamcakova-Dodd, Andrea; Perry, Sarah S et al. (2009) Modulation of reactive oxygen species by Rac1 or catalase prevents asbestos-induced pulmonary fibrosis. Am J Physiol Lung Cell Mol Physiol 297:L846-55
Tephly, Linda A; Carter, A Brent (2008) Asbestos-induced MKP-3 expression augments TNF-alpha gene expression in human monocytes. Am J Respir Cell Mol Biol 39:113-23
Cassel, Suzanne L; Eisenbarth, Stephanie C; Iyer, Shankar S et al. (2008) The Nalp3 inflammasome is essential for the development of silicosis. Proc Natl Acad Sci U S A 105:9035-40