Superfund Research Program
Effects Related Biomarkers of Toxic Exposures
Project Leader: Terrance J. Kavanagh
Grant Number: P42ES004696
Funding Period: 1995 - 2006
Oxidative stress plays an important role in a number of diseases including cardiovascular disease, pulmonary fibrosis, diabetes, Parkinson's disease and Alzheimer’s disease. Because the antioxidant glutathione can be altered in a number of these diseases, Dr. Kavanagh has been examining the role of the rate-limiting enzyme in glutathione synthesis, glutamate-cysteine ligase, in their pathophysiology. The researchers have investigated the role of a GAG trinucleotide repeat polymorphism in GCLC in a number of diseases including cystic fibrosis, Type 1 diabetes mellitus (T1DM) and Parkinson's disease (PD). In CF and T1DM, they have found associations between the numbers of repeats and the severity of disease or the age of onset. In a collaboration with colleagues in UW Pulmonary Medicine and at the University of British Columbia, Dr. Kavanagh’s research team has extended their original observations made in a Seattle cohort of CF patients, and found that this association still holds. A manuscript describing results in CF patients has been submitted for publication. In addition, a manuscript describing results in T1DM patients has also been submitted for publication and is now in revision.
Using homologous recombination and embryonic stem cell technology, project investigators generated transgenic mice that are null for the Gclm gene. The investigators target the exon 1 using a b-galactodosidase-neomycin (b-GEO) cDNA gene in the forward orientation, which can be used as a convenient in vivo reporter of the native Gclm promoter activity. This activity is easily ascertained using either colorometric (Xgal) or chemiluminescence assays. These mice have been used to assess the effects of oxidative stress from acetaminophen exposure. They are highly sensitive to these exposures. A manuscript describing the results in CF patients has been submitted for publication. In addition, in collaboration with Drs. Dan Luchtel and Joel Kaufman, the investigators have found that the expression of the b-gal reporter gene in mice exposed to diesel exhaust is enhanced. This suggests that these mice will be useful as a screen for environmental agents that cause oxidative stress.
In collaboration with Dr. Hooper (Project 7), the researchers have assessed expression of Gcl and metallothionein genes as biomarkers of heavy metal exposure in small mammals (deer mice) captured at a Superfund site using quantitative real-time PCR. Analysis of RT-PCR data indicates significant correlation between transcriptional gene expression for Gclc and cadmium liver concentrations in both males and females; and between both Gcl subunits and copper concentrations in females only. Project investigators have also used mRNA samples from the livers of these mice for global gene expression analysis using 16K Mus musculus cDNA microarrays. Initial analyses of these data find a suppression of genes associated with copper transport and lymphocyte differentiation. These studies will be extended to reveal which gene expression changes are most closely related to metal contamination at this site. Such data will be useful for identifying gene expression biomarkers of heavy metal exposure at concentrations present at a Superfund hazardous waste site.