20181112

Development, Assessment, and Expansion of Models for 2H/13C-Metabolic Flux Analysis In Vivo

Dr. Jamey D Young

Chemical & Biomolecular Engineering Molecular Physiology & Biophysics Vanderbilt University

【要旨】

Isotope-based modeling of liver citric acid cycle and gluconeogenic fluxes in vivo is performed through a parsimonious balance of measurements and assumptions. Our lab previously developed a novel microscale method to quantitatively assess hepatic metabolism in conscious, unrestrained mice through simultaneous intravenous infusions of three stable isotopes. Metabolic fluxes were determined through GC-MS analysis of a 40μL plasma glucose sample, followed by model-enabled flux regression. This was the first study that demonstrated the ability to estimate hepatic fluxes in vivo based on plasma sample volumes that can be readily collected from a conscious mouse. The methodology has been applied to investigate the effects of AMPK knockout, exercise, and fatty liver disease progression in mice. However, recent publications by leading groups have debated the validity of key in vivo assumptions regarding isotope-specific assessments of liver metabolism. These groups contend that metabolic perturbations associated with the administration of 13C-labeled lactate or propionate tracers give rise to different estimates of liver citric acid cycle (CAC) and anaplerotic fluxes. Therefore, we examined the controversy surrounding these flux estimates using our flexible INCA modeling platform that enables rigorous testing of model assumptions. Fasted C57Bl/6J mice were infused with either [13C3]lactate or [13C3]propionate isotopes, and hepatic fluxes were regressed using models with gradually increasing complexity and relaxed assumptions. We confirmed that liver pyruvate cycling fluxes were incongruent between different 13C tracers in models with conventional assumptions. We then constructed in vivo flux models that included physiological cross-talk between the liver and other tissues. By expanding these models to include increased metabolite labeling information and fewer constraining assumptions, we demonstrated that liver pyruvate cycling estimates were significant using either [13C3]lactate or [13C3]propionate and that inconsistencies in hepatic flux estimates emanate, in part, from peripheral metabolism. To our knowledge, this represents the most rigorous attempt so far to consider inter-organ metabolite trafficking in the analysis of data from in vivo isotope labeling experiments.

日時: 2018年11月12日(月) 17:00~18:30
場所: 理学部3号館4F 412室
連絡先: 理学系研究科 生物科学専攻 生物情報科学科
黒田 真也(skuroda AT bs.s.u-tokyo.ac.jp)