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2002 Progress Report: Genetic Mechanisms of Susceptibility to Inhaled Pollutants
EPA Grant Number: R826724C003Subproject: this is subproject number 003 , established and managed by the Center Director under grant R826724
(EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).
Center: CECEHDPR - Johns Hopkins University Hospital
Center Director: Eggleston, Peyton A.
Title: Genetic Mechanisms of Susceptibility to Inhaled Pollutants
Investigators: Kleeberger, Steven R.
Institution: Johns Hopkins University
EPA Project Officer: Fields, Nigel
Project Period: January 1, 1998 through January 1, 2002
Project Period Covered by this Report: January 1, 2001 through January 1, 2002
Project Amount: Refer to main center abstract for funding details.
RFA: Centers for Children's Environmental Health and Disease Prevention Research (1998)
Research Category: Children's Health , Health Effects
Description:
Objective:The goals and objectives have not changed.
The primary objective of this project is to utilize proven positional cloning techniques to identify the gene or genes that determine differential susceptibility to Ozone (03)-induced pulmonary inflammation and injury in inbred mice, and search for homologues in the human genome. Four specific aims were developed to accomplish this objective:
a. To generate high-resolution linkage maps for susceptibility to 03-induced
airways inflammation/epithelial injury.
b. To construct high-resolution long-range physical maps of the regions
of mouse chromosomes 17 and carrying the 03 susceptibility loci.
c. To develop congenic strains of mice that contain the genomic region
that confers differential susceptibility to 03-induced inflammation and
epithelial
injury.
d. To characterize the kinetics of lung response to 03 exposure in 03-resistant
and susceptible congenic mouse strains.
Previously, the Investigators have demonstrated the importance of innate immune defense mechanisms, especially the role of TLR4, in the response to oxidants, such as ozone (O3) in the lung. To better understand the role of TLR4 and its downstream effector mechanisms, in collaboration with Steven Kleeberger at NIEHS, they have performed CDNA microarray analysis to determine the candidate genes that determine differential susceptibility to ozone. They previously demonstrated that Tlr4 plays an important role in responsiveness to ozone. Therefore, they have used susceptible C3H/HeOuJ (OuJ or wildtype) and resistant C3 (TLR4 mutant) mice as a model. Both strains of mice were exposed to either 0.3 PPM ozone or filtered air, RNA was isolated from lungs and reverse transcribed to CDNA and microarray analysis was carried out to identify candidate effector genes that mediate differential responses in these strains. Comparative analysis of gene expression profiles revealed a differential expression of several candidate genes. Briefly, exposure of OuJ, which does not have mutations in TLR4 gene, to ozone caused an up regulation of expression of 17 genes and down regulation of expression of 78 genes in the lungs. In contrast, exposure of HeJ (Tlr4 mutant) mice to ozone resulted up regulation of message levels of 5 genes, while 8 genes message levels were down regulated. These results indicate that a TLR4 mediates regulation of a significant number of genes in the lungs of mice upon exposure to ozone. Some of the genes that are differentially expressed in HeJ and OuJ mice following exposure to ozone are shown below:
Differential gene expression analysis in the lungs of ozone susceptible C3H/HeOuJ (OuJ or wildtype; left panel) and ozone-resistant C3H-HeJ (TLR4 mutant) mice exposed to filter air or ozone (0.3 parts/million) for 72h hr. Differential gene expression profiles were determined by GeneChip analysis in collaboration with the Allied Genomics Center at the Johns Hopkins Bayview campus using Affymetrix arrays. Up or down-regulated genes were determined by standard protocol and software provided by Affymetxix.
Ongoing studies have been designed to further characterize candidate genes that are differentially regulated by TLR4. We have also initiated a study to further investigate the mechanisms through which TLR4 is activated following O3 exposure. Specially, they are especially interested in investigating the ligands that bind to TLR4 following O3 exposure. We are utilizing cell culture approach to understand two candidate proteins, heat shock proteins (Hsp 60 or and 70) and fibronectin, which are found in the broncboalveloar lavage fluid upon exposure to ozone in human and animal models. Moreover, these proteins have been shown to bind to TLR4 in other model systems.
Future Activities:This work with TLR4 is of particular interest and relevance to the field of air pollution effects on the lung and the mechanisms of susceptibility. The studies reported above are the first to unequivocally demonstrate that innate immunity genes have an important role in the pulmonary response to oxidants and particulates. These studies may also provide some understanding to the mechanisms through which particulates and oxidants lead to the morbidity and mortality in human populations reported in epidemiological studies.
Although the goals of the project have not been changed, in the next granting period project will mainly focuses on characterizing the TLR4-responsive genes and mechanisms of TLR4 activation by ozone.
Supplemental Keywords:Allergens, Asthma, Biochemistry, Children's Health, Epidemiology, Human Health Risk Assessment, age-related differences, airway disease, community-based intervention, genetics, Ozone. , Air, Scientific Discipline, Health, RFA, Susceptibility/Sensitive Population/Genetic Susceptibility, Biology, Risk Assessments, genetic susceptability, Health Risk Assessment, Chemistry, Children's Health, particulate matter, Environmental Chemistry, Allergens/Asthma, Genetics, environmental hazard exposures, allergen, health effects, inhalation, allergens, air quality, ozone, ozone induced airway dysfunction, assessment of exposure, childhood respiratory disease, environmental health effects, susceptibility, toxics, community-based intervention, human health effects, particulates, respiratory, sensitive populations, biological response, genetic mechanisms, air pollution, airway disease, children, inhaled, cellular biology, analytical chemistry, community-based studies, disease, exposure, airway epithelial cells, environmental health hazard, asthma, genetic predisposition, human exposure, Human Health Risk Assessment, morbidity, PM, airway inflammation, community based
Progress and Final Reports:
2000 Progress Report
Original Abstract
Main Center Abstract and Reports:
R826724 CECEHDPR - Johns Hopkins University Hospital
Subprojects under this Center:
(EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).
R826724C001 A Randomized, Controlled Trial of Home Exposure Control in Asthma
R826724C002 Mechanisms Of Particulate-Induced Allergic Asthma
R826724C003 Genetic Mechanisms of Susceptibility to Inhaled Pollutants
R826724C004 The Relationship Of Airborne Pollutants And Allergens To Asthma Morbidity