Many human diseases are caused by a dysregulation of lipid metabolism, including atherosclerosis, cancer, neurodegeneration, diabetes, and fatty liver. The development of effective treatments for lipid related disorders is hinder by a lack of modern in vivo biochemistry techniques for studying lipid metabolism. The overall goal of this proposal is to develop tools and protocol to measure the rates of lipid biosynthesis and remodeling by stable isotope labeling with sensitivity comparable to radio-isotope tracing with the specificity and broad coverage of modern mass spectrometry based lipidomics. This is enabled by an ultra-high resolution orbitrap mass spectrometer I developed in collaboration of Thermo Scientific, now commercially available as the Lumos 1M. This instrument has sufficient resolution to resolve the natural abundance 13C from a tracer isotope, for example 2H, in intact lipid ions. By resolving the dominant natural abundance ions from tracer isotopes will improve the signal to noise ratio by at least 2 orders of magnitude (1:1 vs >1:100) and increase the dynamic range. This advancement will allow in vivo analysis of lipid metabolism to study a variety of disease, and will ultimately lead to lipid fluxomics analysis that is translatable to human studies. By measuring lipid flux in patients we will be able to directly studying the progression of metabolic syndrome, potentially circumventing the need for animal models, and measure the effectiveness of therapies and interventions. To facilitate the development and widespread implementation of this technology, I will address the fundamental roadblocks to adapting this technology. Firstly, the commercial instrument is engineered for proteomics applications, in particular the electrospray ionization source. By working with the manufacturer and translating my lipidomics experience to this new platform I will overcome these issue. Secondly, I will develop novel data collection approaches for both chromatography and direct infusion based applications to accommodate the long transient time and coalescence issues associated with ultra-high resolution resonance based mass spectrometry. Thirdly, software tools will be developed to extract ultra-high resolution data in a time efficient manner, convert the data to physically interpretable parameters, and map data onto biochemical pathways. Lastly, I will develop protocols and platforms for stable isotope labeling by deuterium labeled water (D2O) and other isotope labeled metabolic tracers in mouse models of metabolic syndrome relevant to my lab?s research program studying the mechanism for fat accumulation. By accomplishing these aims this technology will be accessible to the biomedical research community. My multi-disciplinary training in engineering, physical, analytical and biochemistry, and mouse genetics makes me well-suited to develop this technology and the lipid centric research environment at UT Southwestern is the ideal location for the initial application.
The diagnosis and treatment of diseases caused by dysregulation of lipid metabolism, including diabetes, cardiovascular disease, cancer, neurodegeneration, and fatty liver, is hinder by a lack of tools for measuring lipid biochemistry in vivo. Here, we develop tools for measuring the flux of lipids with stable isotopes using a novel ultra-high resolution mass spectrometer that can resolve natural abundance isotopes from tracer isotopes. We will overcome fundamental barriers to the implementation of this technology so that it can be accessed by the biomedical research community.
