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IDENTIFICATION OF INTELLIGENT BIOMARKERS OF EXPOSURE AND HARM IN THE RESPIRATORY EPITHELIAL TO TOBACCO SMOKE COMPONENTS.
BéruBé, K., Sexton, K., Jones, T.
School of Biosciences, Cardiff University, Museum Avenue, Cardiff, CF10 3US, Wales, UK.
Keywords: 3-Dimensional model, Air liquid interface, Biomarkers, Cadmium, EpiAirway, Formaldehyde, Growth factors, Human respiratory epithelia, Inflammation, Inflammatory, Intelligent Biomarkers, Lung epithelia, Metabolism, Nicotine, Tobacco smoke components (TSC), Toxicological tool, Urethane, Xenobiotic metabolism
Endpoints: 2-D SDS-PAGE, Gene expression, MALDI-TOF MS, Microarray, Protein expression
Materials Tested: Cadmium, Formaldehyde, Nicotine, Urethane
Summary:
A novel toxicological tool, which consists of a differentiated, 3-D, in vitro model of human respiratory epithelia, (EpiAirway-100 cells; MatTek Corp., USA), will be utilized by researchers at Cardiff University to examine the early gene response(s) following exposure to tobacco smoke components (TSC).
EpiAirway tissues will be exposed at the air/liquid interface to representative particle and vapour phase components of cigarette smoke. The TSC selected represent an overview of the compounds found in cigarette smoke that induce thrombogenic events (nicotine), cell toxicity (cadmium) and produce reactive metabolites during xenobiotic metabolism (formaldehyde and urethane).
Surrogate solutions of TSC will be tested for their capacity to up- and/or down-regulate genes in the respiratory epithelia following acute exposure, as a means to identifying intelligent biomarkers of exposure and harm. Conventional toxicological analysis will be used to establish the dose of the various TSC needed to cause changes in epithelial resistance, secreted surface proteins and release of inflammatory markers.
Following establishment of the dose required to achieve these different biological endpoints, toxicogenomic investigations will be initiated. Macroarray technology will be employed to compare the patterns of mRNA expression of human genes associated with stress, simultaneously from control and TSC treated lung tissue. Major candidate genes will be classified (growth factors, inflammation, xenobiotic metabolism) and associated with the biological endpoints.
It is anticipated that stringent lists of candidate genes associated with these changes will be generated, thereby providing data on the mechanisms of the biological endpoints, i.e. intelligent biomarkers.
Proteomic analysis will be used to correlate candidate gene response with a specific protein. The interest will be to study the proteins involved in the early stress phase of toxicant challenge. Once protein function has been identified, this will provide insight into the mechanism of action of the responsive genes and hence, the “intelligent biomarkers” of TSC exposure in lung epithelia.
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