Scholarship & Research
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Item Mass spectrometry based lipidomics as a tool in the search for biomarkers and mechanisms of disease(Montana State University - Bozeman, College of Letters & Science, 2016) Willems, Daniel Lee; Chairperson, Graduate Committee: Edward Dratz; Nicholas E. Goocey and Edward A. Dratz were co-authors of the article, 'A highly reproducible and efficient lipid extraction protocol enhanced using 3D printing of centrifuge adapters for optimum glass vials' submitted to the journal 'Lipids' which is contained within this thesis.; Max Koch, Nicholas E. Goocey, Blaine R. Roberts and Edward A. Dratz were co-authors of the article, 'Lipidomic analysis of human brain cortex in alzheimer's disease reveals aberrant levels of acetylcholine precursor speices' submitted to the journal 'American journal of alzheimer's disease and other dementias' which is contained within this thesis.; Max Koch, Nicholas E. Goocey, Blaine R. Roberts and Edward A. Dratz were co-authors of the article, 'Lipidomics reveals aberrent metabolism of lipid molecules in alzheimer's disease cerebral cortex' which is contained within this thesis.Lipidomics studies a highly diverse class of compounds insoluble in water and soluble in organic solvents. Lipids are a major component of cells and tissues, take part in a rich network of metabolic reactions, and are implicated in many disease mechanisms. Lipidomics complements genomics, proteomics and the more common metabolomic analysis of hydrophilic metabolites and can provide new insights into disease mechanisms. The problem approached in this thesis was to compare different methods of sample preparation for lipidomics and apply lipidomics to the study of two major health problems: Nonalcoholic Fatty Liver Disease (NAFLD) and Alzheimer's Disease (AD). Excessive dietary intake of sucrose and fructose, common in the Western Diet, increases deposition of triacylglycerides in the liver and leads to cognitive decline in experimental animals. NAFLD increases the risk of type 2 diabetes, obesity and AD. The high diversity and hydrophobicity of lipids complicates their separation, detection and analysis. However, modern chromatography and mass spectrometry instrumentation and techniques are greatly improving the capability of lipidomic analysis. A lipid extraction protocol was optimized for reproducibility and yield, and was used to extract lipids from rat liver under sucrose stress in a model of human NAFLD and human brain cortex from Alzheimer's Disease (AD) compared to controls. The samples were analyzed using mass spectrometry. The NALFD study did not yield the expected results, instead these data provided a foundation for designing future experiments in progress and to validate the methods used in the AD study. The AD studies showed that several phosphatidylcholine species are down regulated along with acetyl-CoA, which may be the source of low levels of the neurotransmitter acetylcholine in AD. Two different chromatography methods were used to seek a higher coverage of different lipids. Differences in the lipids in AD and controls were evident in the omega-6 and omega-3 fatty acids. The precursors of long omega-3s synthesis were increased while the products EPA and DHA were decreased. In a similar fashion, precursors to long omega-6s were found to be decreased, while the products were increased. This suggests that the omega-6 synthesis pathway may be outcompeting the omega-3 synthesis.Item Fossil viruses, redox paradigms and predictive metabolism from a systems biology perspective(Montana State University - Bozeman, College of Letters & Science, 2014) Heinemann, Joshua Vance; Chairperson, Graduate Committee: Brian Bothner; Walid S. Maaty, George Gauss, Narahari Akkaladevi, Susan K. Brumfield, Vamseedhar Rayaprolu, Mark Young, C. Martin Lawrence and Brian Bothner were co-authors of the article, 'Fossil record of an HK-97-like provirus' in the journal 'Virology' which is contained within this thesis.; Timothy Hamerly, Walid S. Maaty, Navid Movahed, Joseph D. Steffens, Benjamin D. Reeves, Jonathan K. Hilmer, Jesse Therien, Paul A. Grieco, John W. Peters and Brian Bothner were co-authors of the article, 'Expanding the paradigm of thiol redox in the thermophilic root of life' in the journal 'Biochimica et biophysica acta' which is contained within this thesis.; Aurélien Mazurie, Monika Tokmina-Lukaszewska, Greg J. Beilman and Brian Bothner were co-authors of the article, 'Application of support vector machines to metabolomics experiments with limited replicates' in the journal 'Metabolomics' which is contained within this thesis.; Brigit Noon, Mohammad J. Mohigmi, Aurélien Mazurie, David L. Dickensheets and Brian Bothner were co-authors of the article, 'Real-time digitization of metabolomic patterns from a living system using mass spectrometry' submitted to the journal 'Journal of the American Chemical Society' which is contained within this thesis.One of the goals of systems biology is to develop a model which encapsulates the molecular, structural and temporal complexity of a living organism. While modern omics experiments can deliver a high resolution view of an organism's molecular complexity, methods for correlating the information from multiple biomolecular systems (i.e. genes, proteins and metabolites) and their changes over time remain greatly underdeveloped. Presented in this research are: (1) methods for understanding the inter-relation of multiple biomolecular systems correlating genomics, proteomics and metabolomics experiments; (2) techniques for machine learning based metabolic biomarker selection; (3) robotics technology for real-time measurement of changes in metabolism. The methods for correlating information from multiple biomolecular systems have provided a new perspective of biomolecular adaptation and evolutionary relationships in the thermophilic archaea. The techniques for biomarker selection have provided a method to assess the reliability of biomarkers in experiments where limited samples are available. The new technology has provided an engineered system for automated analysis of metabolic patterns and how they change over time. Together, these results have created a framework for future improvement of our understanding of biology through the use of molecular biology, machine learning and robotics.