Quantitative Mass Spectrometry

Environmental Health Sciences

Investigators

Linked publications, trials & patents

Abstract

Quantitative Mass Spectrometry incorporates collaborative projects in which the Mass Spectrometry Group provides quantitative information about, typically, small molecules by GCMS, LCMS and LCMSMS or a combination. An example of the types of projects this includes is the isoprostane analyses undertaken as part of the NIEHS led study of biomarkers of oxidative stress. We currently have several major collaborations underway. [unreadable] 1) with the Zeldin Lab quantitating arachidonic acid metabolites that are relevant to inflammatory, vasodilatory, endothelial protective and post-ischemic cardioprotective effects. We have provide prostaglandin quantitation for a study on the pharmacological inhibition or genetic disruption of cyclooxygenase (COX)-1 or COX-2 exacerbates the inflammatory and functional responses of the lung to environmentally relevant stimuli. In this study, to further examine the contribution of COX-derived eicosanoids to basal lung function and to allergic lung inflammation, transgenic (Tr) mice were generated in which overexpression of human COX-1 was targeted to airway epithelium. Although no differences in basal respiratory or lung mechanical parameters were observed, COX-1 Tr mice had increased bronchoalveolar lavage fluid PGE2 content compared with wild-type littermates (23.0 3.6 vs 8.4 1.4 pgml; p < 0.05) and exhibited decreased airway responsiveness to inhaled methacholine. In an OVA-induced allergic airway inflammation model, comparable up-regulation of COX-2 protein was observed in the lungs of allergic wild-type and COX-1 Tr mice. Furthermore, no genotype differences were observed in allergic mice in total cell number, eosinophil content (70 vs 76% of total cells, respectively), and inflammatory cytokine content of bronchoalveolar lavage fluid, or in airway responsiveness to inhaled methacholine (p > 0.05). To eliminate the presumed confounding effects of COX-2 up-regulation, COX-1 Tr mice were bred into a COX-2 null background. In these mice, the presence of the COX-1 transgene did not alter allergen-induced inflammation but significantly attenuated allergen-induced airway hyperresponsiveness, coincident with reduced airway leukotriene levels. Collectively, these data indicate that COX-1 overexpression attenuates airway responsiveness under basal conditions but does not influence allergic airway inflammation. [unreadable] Jeffrey W. Card, Michelle A. Carey, J. Alyce Bradbury, Joan P. Graves, Fred B. Lih, Michael P. Moorman, Daniel L. Morgan, Laura M. DeGraff, Yun Zhao, Julie F. Foley, and Darryl C. Zeldin[unreadable] Cyclooxygenase-1 Overexpression Decreases Basal Airway Responsiveness but Not Allergic Inflammation[unreadable] J. Immunol., Oct 2006; 177: 4785 - 4793. [unreadable] [unreadable] Cyclooxygenase (COX)-derived eicosanoids have been implicated in the pathogenesis of pulmonary fibrosis. Uncertainty regarding the influence of COX-2 on experimental pulmonary fibrosis prompted the Zeldin Lab to clarify the fibrotic and functional effects of intratracheal bleomycin administration in mice genetically deficient in COX-2. Further, the effects of airway-specific COX-1 overexpression on fibrotic and functional outcomes in wild type and COX-2 knockout mice were assessed. Equivalent increases in airway cell influx, lung collagen content and histopathological evidence of fibrosis were observed in wild type and COX-2 knockout mice 21 days following bleomycin treatment, suggesting that COX-2 deficiency did not alter the extent or severity of fibrosis in this model. However, bleomycin-induced alterations in respiratory mechanics were more severe in COX-2 knockout mice than in wild type mice as illustrated by a greater decrease in static compliance compared to genotype-matched, saline-treated control mice (26 3% vs. 11 4% decreases for COX-2 knockout and wild type mice, respectively; p<0.05). The influence of COX-1 overexpression in airway Clara cells was also examined. Whereas the fibrotic effects of bleomycin were not altered in wild type or COX-2 knockout mice overexpressing COX-1, the exaggerated lung function decrement in bleomycin-treated COX-2 knockout mice was prevented by COX-1 overexpression and coincided with decreased airway cysteinyl leukotriene levels. Collectively, these data suggest an important regulatory role for COX-2 in the maintenance of lung function in the setting of lung fibrosis, but not in the progression of the fibrotic process per se.[unreadable] Am. J. Respir. Cell Mol. Biol. 2007, in press.[unreadable] Cyclooxygenase-2 Deficiency Exacerbates Bleomycin-induced Lung Dysfunction but not Fibrosis [unreadable] Jeffrey W Card, James W Voltz, Michelle A Carey, J. Alyce Bradbury, Laura M DeGraff, Fred B Lih, James C Bonner, Daniel L Morgan, Gordon P Flake, and Darryl C Zeldin [unreadable] [unreadable] 2) a collaboration with James MohlerMark Titus on a Program Project (UNCRoswell Cancer Institute) on Interference with the Androgen Receptor and its Ligands in Recurrent Prostate Cancer.[unreadable] [unreadable] 3) with the Roberts Lab on the affect of a diet high in linoleic acid on cnacer metastasis.[unreadable] Several studies have shown that dietary changes can alter the progression and risk of numerous cancers, including pancreatic, intestinal, and breast cancer. A recent study from the Roberts Lab has reported that linoleic acid (LA), a dietary omega-6 polyunsaturated fatty acid (PUFA), enhances the invasion of peritoneal metastasis of gastric carcinoma cells and stimulates carcinogenesis. The current study expands further on this by studying the fatty acid profile of duodenum and serum in relation to dietary LA levels and the effect on cancer progression in mice receiving OCUM-2MD3 cells, a highly metastatic human gastric carcinoma cell line. The mice were divided into two groups, both of which were fed a high fat diet (23%ww). One group was fed a diet containing a normal proportion of LA (2%w LA) and the other fed a diet high in LA (12%w LA). Fatty acid content of samples was analyzed by gas chromatographyelectron impactmass spectrometry (GCEIMS). Lipids were isolated, hydrolyzed, and free fatty acids were converted to methyl esters prior to analysis. Fatty acid profiling revealed an increase in LA concentrations in serum and duodenum of mice treated with 12% LA diet when compared to mice treated with 2% LA diet. Lauric (12:0), myristic (14:0), palmitic (16:0), and oleic (18:1n9), however, showed a decrease in concentrations in serum of mice fed the 12% LA diet compared to those fed 2% LA diet. The data show that circulating and tissue fatty acid profiles are affected by dietary intake of linoleic acid and that this dietary change also promotes cancer growth and invasion. The mechanism through which these effects manifest is currently being investigated.