Toxicoproteomics is the use of global protein expression technologies in toxicology to gain a better understanding of environmental and genetic factors in toxicant exposure and in long term development of disease. The formulation of a strategy to integrate transcript, protein and toxicology data is a major objective for the field of Toxicogenomics. The National Center for Toxicogenomics is pursuing a strategy of conducting parallel DNA microarray and proteomic analyses on the same tissues from each Toxicogenomics study. The advantage of this approach is to bring more information to bear on toxicological problems in identifying affected, biochemical and regulatory pathways that can lead to biomarker and toxicity signature discovery. Both DNA microarray and proteomic technologies are driven by measuring differential expression of transcripts and proteins after toxicant exposure. The high level of information density and gene discovery potential in microarray analysis can be enhanced by proteomics technologies, which allow identification of toxicant-altered proteins based on subcellular localization as well as characterization of post-translational modifications that occur in signaling pathway proteins. Further, proteomic platforms appear particularly well suited for biomarker development in blood in that the appearance of new polypeptides in the serum proteome may reveal early stage disease and organ toxicity. Intramural toxicoproteomic research is a coordinated effort between two research groups, the Proteomics Group within the NCT and the Mass Spectrometry Group in the Laboratory of Structural Biology. Proteomics technologies used in this integrated program involve two-dimensional (2D) gel electrophoresis and mass spectrometry for global protein separation and protein identification, respectively. Surface retentate mass spectrometry or surface enhanced laser desorption ionization, SELDI, is being used by the Proteomics Group for serum biomarker development and classification of control and toxicant-exposed experimental animals and also in clinical studies with the epidemiology branch to classify control and diseased populations.Progress in toxicoproteomics occurred in four areas during the year in 2003 to 2004. A primary goal has been biomarker development in liver and serum using proteomic approaches in response to hepatotoxic chemicals. A second area of progress has been in conducting parallel genomic and proteomic studies to compare gene and protein expression. A third area of development has been in serum biomarker development using the SELDI platform in experimental animal studies and in patients with documented clinical disease. Fourth, the use of antibody arrays is being explored as a complementary platform to provide information on a focused set of gene expression responses. Proteomics Resource Contract: The contract to assist the NCT in applying proteomics to toxiciology began in July, 2002 and was terminated in December, 2004 for financial and personnel retention issues. During the functioning 1.5 years of the contract, the contractor worked upon proteomic analysis of two major experimental studies in rats that were conducted by my group. The first project involved acetaminophen hepatotoxicity and the second project focused upon the model inflammagen, lipopolysacharide, and its ability to simulate certain aspects of endotoxemia and hepatotoxicity. Fortunately, all toxicoproteomics data generated by the proteomics recourse contract were captured by the NCT and the bioinformatics contractor, SAIC, for inclusion into the toxicogenomics database, CEBS. Data from these two projects were received by the NCT and will be published. Alex Merrick and Ken Tomer served as technical project officers on this contract. Intramural Research: Past years accomplishments have focused upon the subcellular effects of toxicants upon liver tissue and these efforts have continued. Progress was made on proteomic mapping of liver nuclei with the MS/microchemistry core facility, led by Ken Tomer. Collaborative work with Rick Fannin and Rick Paules of the NIEHS Microarray Group has led to submission of a manuscript on the whole blood transcript changes in response to the inflammagen, lipopolysaccharide (LPS), to establish blood-born biomarkers of LPS treated rats. SELDI: Surface enhanced laser desorption ionization (SELDI) is a relatively new proteomic platform that has been best applied to protein profiling of serum from diseased patients. A proof of principle study was performed using serum from rats acutely exposed to the TLR4 agonist, lipopolysaccharide (LPS). A second SELDI study has been completed on human serum samples from controal and diagnosed patients with amyelotrophic lateral sclerosis (ALS) in collaboration with Freya Kamel and Jack Taylor of the Epidemiology branch. A third SELDI study is in progress on patient serums obtained from a controlled study of acutely exposed male and female volunteers to the analgesic, acetaminophen, in the clinical studies unit at UNC Chapel Hill Hospital. This study is being conducted in collaboration with Drs. Russo and Watkins at UNC and the NIEHS Microarray Center.
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