angela s benton, bas 1 , alan m watson, phd 1,2 , zuyi wang, phd 1,3 ,
DESCRIPTION
Angela S Benton, BAS 1 , Alan M Watson, PhD 1,2 , Zuyi Wang, PhD 1,3 , Mary C Rose, PhD 1,2 , and Robert J Freishtat, MD, MPH 1,2 1 Children's National Medical Center, Washington, DC 2 The George Washington University, Washington, DC - PowerPoint PPT PresentationTRANSCRIPT
Angela S Benton, BAS1, Alan M Watson, PhD1,2, Zuyi Wang, PhD1,3,Mary C Rose, PhD1,2, and Robert J Freishtat, MD, MPH1,2 1Children's National Medical Center, Washington, DC2The George Washington University, Washington, DC3Virginia Polytechnic Institute and State University, Arlington, VA
Delineation of a TIMP-1 Network Underlying the Response to Cigarette Smoke and Oxidative Stress in Asthma
Disclosure Statement The authors have documented that
they have nothing to disclose.
Tobacco Smoke in Childhood Asthma Asthma = syndrome with genetic and
environmental components (FD Martinez, 1997, 2007)
Tobacco smoke Common environmental trigger Associated with diagnosis and increased
morbidity(Y Chen et al, 2005; J Cunningham et al, 1996; FD Gilliland et al, 2006; MK Selgrade et al, 2006; JJ Sturm et al, 2004)
Tobacco Smoke Effects onBronchial Epithelial Cigarette smoke
Respiratory epithelial response Mediated in part by oxidative stress Asthmatic bronchial epithelium more
susceptible to this stress (Bucchieri et al, 2002)
Hypothesis Tobacco smoke exposure in
asthmatic bronchial epithelium initiates an oxidative stress response Different from non-asthmatic bronchial
epithelium
Data Integration Four publicly-available data series
NCBI: Gene Expression Omnibus (GEO) Asthma- and cigarette smoke-relevant N = 153 arrays Unsupervised clustering Adequate signal/noise levels Genespring GX10
Venn Diagram Analysis
1N=4
GSE1301Lung tissue from HDM vs. PBS in vivo airway challenge
N=238 N=8
3N=2 N=99N=3
N=9
2N=6
N=29
GSE3183A549 cells
in vitro treatment with IL13 vs. PBS
N=367
GSE3184Lung tissue from HDM vs. PBS in vivo airway challenge in AJ and C3H mice
N=2,034
N=236
GSE994Epithelial brushings
from smokers vs. nonsmokers
N=130
4N=1 5N=17
Pathways Analysis
Inge
nuity
Pat
hway
s Ana
lysis
™
Tissue Inhibitor of Metalloproteinase (TIMP) -1 Secreted 31 kDA glycoprotein Inhibits protease activity of all MMPs (DE Gomez et
al, 1997) MMP-9 major lung MMP (WC Parks and SD Shapiro, 2001; H
Tanaka et al, 2000) Asthmatic BALF and sputum levels lower than
control(W Mattos et al, 2002; AM Vignola et al, 1998)
MMP-9:TIMP-1 hypothesized to drive extracellular lung remodeling (JJ Atkinson et al, 2003)
Phosgene-Induced Oxidative Stress Mice exposed to phosgene at time 0 Sacrificed at eight post-exposure time
points (0.5, 1, 4, 8, 12, 24, 48, 72 hours) Lung tissue isolated RNA extracted Whole genome expression profiling
Coordinated Expression
Highly Regulated Genes
Fold Changes in Focus Network
1 hour
8 hours
24 hours
72 hours
0 hours
Primary human differentiated respiratory epithelium (HBE) 1 hour exposure: cigarette smoke
condensate (CSC) or hydrogen peroxide 23 hour: incubation ELISA: measure protein levels of TIMP-1 and
MMP-9 in the apical and basal secretions
HBE Cell Donors
In Vitro Validation
*p=0.017 *p=0.011
A B
Summary 4-way Venn diagram approach
Publicly-available microarray data Identified TIMP-1 nucleated network of
proteins Key in response to tobacco smoke and
oxidative stress in asthma In vitro validation
Potential for subepithelial conditions known to favor airway remodeling in chronic asthma
Conclusions TIMP-1 network
Oxidative stress-induced pathway Underlies bronchial epithelial response to
cigarette smoke and oxidative stress in asthma
New Questions ETS-exposed children with asthma
Blunted TIMP-1 response? More susceptible to metalloproteinase-
mediated airway remodeling?
Acknowledgements Research Center
for Genetic Medicine @ Children’s National Medical Center Freishtat Lab
K23-RR-020069 Rose Lab
Mary Rose, PhD Alan Watson, PhD
Hoffman Lab Eric Hoffman, PhD Zuyi Wang, PhD Jinwook Seo, PhD K12-RR core laboratories NCMRR integrated
molecular core laboratories (www.ncmrr.org)
GCRC genetics core laboratories Genetic counseling Microarray