Mass spectrometry based lipidomics as a tool in the search for biomarkers and mechanisms of disease

Abstract

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.