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Voss B.J.,Microbiology and Immunology | McDonald W.H.,Mass Spectrometry Research Center | Cover T.L.,Vanderbilt University
Journal of Proteomics | Year: 2016

Helicobacter pylori colonizes the human stomach and is associated with an increased risk of gastric cancer and peptic ulcer disease. Analysis of H. pylori protein secretion is complicated by the occurrence of bacterial autolysis. In this study, we analyzed the exoproteome of H. pylori at multiple phases of bacterial growth and identified 74 proteins that are selectively released into the extracellular space. These include proteins known to cause alterations in host cells, antigenic proteins, and additional proteins that have not yet been studied in any detail. The composition of the H. pylori exoproteome is dependent on the phase of bacterial growth. For example, the proportional abundance of the vacuolating toxin VacA in culture supernatant is higher during late growth phases than early growth phases, whereas the proportional abundance of many other proteins is higher during early growth phases. We detected marked variation in the subcellular localization of putative secreted proteins within soluble and membrane fractions derived from intact bacteria. By providing a comprehensive view of the H. pylori exoproteome, these results provide new insights into the array of secreted H. pylori proteins that may cause alterations in the gastric environment. © 2015 Published by Elsevier B.V. Source


Adler L.,Medical University of South Carolina | Boyer N.P.,Medical University of South Carolina | Anderson D.M.,Mass Spectrometry Research Center | Spraggins J.M.,Mass Spectrometry Research Center | And 5 more authors.
Photochemical and Photobiological Sciences | Year: 2015

The bis-retinoid N-retinylidene-N-retinylethanolamine (A2E) is one of the major components of lipofuscin, a fluorescent material that accumulates with age in the lysosomes of the retinal pigment epithelium (RPE) of the human eye. Lipofuscin, as well as A2E, exhibit a range of cytotoxic properties, which are thought to contribute to the pathogenesis of degenerative diseases of the retina such as Age-related Macular Degeneration. Consistent with such a pathogenic role, high levels of lipofuscin fluorescence are found in the central area of the human RPE, and decline toward the periphery. Recent reports have however suggested a surprising incongruence between the distributions of lipofuscin and A2E in the human RPE, with A2E levels being lowest in the central area and increasing toward the periphery. To appraise such a possibility, we have quantified the levels of A2E in the central and peripheral RPE areas of 10 eyes from 6 human donors (ages 75-91 years) with HPLC and UV/VIS spectroscopy. The levels of A2E in the central area were on average 3-6 times lower than in peripheral areas of the same eye. Furthermore, continuous accumulation of selected ions (CASI) imaging mass spectrometry showed the presence of A2E in the central RPE, and at lower intensities than in the periphery. We have therefore corroborated that in human RPE the levels of A2E are lower in the central area compared to the periphery. We conclude that the levels of A2E cannot by themselves provide an explanation for the higher lipofuscin fluorescence found in the central area of the human RPE. © 2015 The Royal Society of Chemistry and Owner Societies. Source


Rahman S.M.J.,Vanderbilt University | Seeley E.H.,Mass Spectrometry Research Center | Manier M.L.,Vanderbilt University | Manier M.L.,Mass Spectrometry Research Center | And 11 more authors.
Cancer Research | Year: 2011

Early detection may help improve survival from lung cancer. In this study, our goal was to derive and validate a signature from the proteomic analysis of bronchial lesions that could predict the diagnosis of lung cancer. Using previously published studies of bronchial tissues, we selected a signature of nine matrix-assisted laser desorption ionization mass spectrometry (MALDI MS) mass-to-charge ratio features to build a prediction model diagnostic of lung cancer. The model was based on MALDI MS signal intensity (MALDI score) from bronchial tissue specimens from our 2005 published cohort of 51 patients. The performance of the prediction model in identifying lung cancer was tested in an independent cohort of bronchial specimens from 60 patients. The probability of having lung cancer based on the proteomic analysis of the bronchial specimens was characterized by an area under the receiver operating characteristic curve of 0.77 (95% CI 0.66-0.88) in this validation cohort. Eight of the nine features were identified and validated by Western blotting and immunohistochemistry. These results show that proteomic analysis of endobronchial lesions may facilitate the diagnosis of lung cancer and the monitoring of high-risk individuals for lung cancer in surveillance and chemoprevention trials. © 2011 American Association for Cancer Research. Source


Voziyan P.A.,Mass Spectrometry Research Center | Voziyan P.A.,Vanderbilt University | Hudson B.G.,Microbiology and Immunology | Caprioli R.M.,Mass Spectrometry Research Center
Journal of Lipid Research | Year: 2014

Diabetic nephropathy (DN) is a major lifethreatening complication of diabetes. Renal lesions affect glomeruli and tubules, but the pathogenesis is not completely understood. Phospholipids and glycolipids are molecules that carry out multiple cell functions in health and disease, and their role in DN pathogenesis is unknown . We employed high spatial resolution MALDI imaging MS to determine lipid changes in kidneys of eNOS db/db mice, a robust model of DN. Phospholipid and glycolipid structures, localization patterns, and relative tissue levels were determined in individual renal glomeruli and tubules without disturbing tissue morphology. A signifi cant increase in the levels of specifi c glomerular and tubular lipid species from four different classes, i.e., gangliosides, sulfoglycosphingolipids, lysophospholipids, and phosphatidylethanolamines, was detected in diabetic kidneys compared with nondiabetic controls. Inhibition of nonenzymatic oxidative and glycoxidative pathways attenuated the increase in lipid levels and ameliorated renal pathology, even though blood glucose levels remained unchanged. Our data demonstrate that the levels of specifi c phospho- and glycolipids in glomeruli and/or tubules are associated with diabetic renal pathology. We suggest that hyperglycemia- induced DN pathogenic mechanisms require intermediate oxidative steps that involve specifi c phospholipid and glycolipid species. -Grove, K. J., P. A. Voziyan, J. M. Spraggins, S. Wang, P. Paueksakon, R. C. Harris, B. G. Hudson, and R. M. Caprioli. Diabetic nephropathy induces alterations in the glomerular and tubule lipid profi les. J. Lipid Res. 2014. 55: 1375 - 1385 © 2014 by the American Society for Biochemistry and Molecular Biology, Inc. Source


Molina M.,University of Sao Paulo | Steinbach S.,Ruhr University Bochum | Park Y.M.,Korea Basic Science Institute | Park Y.M.,Mass Spectrometry Research Center | And 10 more authors.
Journal of Neural Transmission | Year: 2015

Brain function in normal aging and neurological diseases has long been a subject of interest. With current technology, it is possible to go beyond descriptive analyses to characterize brain cell populations at the molecular level. However, the brain comprises over 100 billion highly specialized cells, and it is a challenge to discriminate different cell groups for analyses. Isolating intact neurons is not feasible with traditional methods, such as tissue homogenization techniques. The advent of laser microdissection techniques promises to overcome previous limitations in the isolation of specific cells. Here, we provide a detailed protocol for isolating and analyzing neurons from postmortem human brain tissue samples. We describe a workflow for successfully freezing, sectioning and staining tissue for laser microdissection. This protocol was validated by mass spectrometric analysis. Isolated neurons can also be employed for western blotting or PCR. This protocol will enable further examinations of brain cell-specific molecular pathways and aid in elucidating distinct brain functions. © 2015, Springer-Verlag Wien. Source

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