Diabetologie und Stoffwechsel 2012; 7 - LB_16
DOI: 10.1055/s-0032-1314780

13C-labelled palmitate and metabolomics/lipidomics analyses reveal the fate of free fatty acids in fasting mice

M Hoene 1, 2, S Chen 3, J Li 3, E Schleicher 1, 2, HU Häring 1, 2, G Xu 3, C Weigert 1, 2, R Lehmann 1, 2
  • 1Internal Medicine IV, Division of Endocrinology, Diabetology, Vascular Medicine, Nephrology and Clinical Chemistry, University Hospital Tübingen, Tübingen, Germany
  • 2Paul Langerhans Institute Tübingen, Member of the German Diabetes Centre (DZD e.V.), Institute for Diabetes Research and Metabolic Diseases of the Helmholtz Centre Munich at the University of Tuebingen, Tübingen, Germany
  • 3CAS Key Laboratory of Separation Science for Analytical Chemistry, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, China

Aims: During fasting, free fatty acids (FFA) released from adipose triglyceride stores become the major source of energy. Because lipolysis is in excess over utilization, plasma FFA are considerably elevated despite an increased utilization by skeletal muscle and other peripheral tissues. The plasma concentration of acylcarnitines, intermediates of fatty acid oxidation, also increases during fasting and exercise, but it is unclear if they originate from skeletal muscle or from the liver, the key metabolic regulator and short-term buffer for excess circulating FFA. Using a stable 13C-isotope labelled fatty acid tracer in mice, we tested the hypothesis that excess palmitate is incorporated into acylcarnitines in the muscle and into triglycerides and other lipids in the liver in the fasted state.

Methods: Male C57BL/6N mice were fasted for 15h. A bolus of 20 nmol/kg body weight of [U-13C]-palmitate was applied into the caudal vein of the anaesthetised mice. After 10 minutes, plasma, liver and gastrocnemius muscles were obtained. Lipids, acylcarnitines and free fatty acids were extracted and analyzed by UPLC-mass spectrometry for [U-13C]-palmitate-derived metabolites. Values are means±SD from n=7 mice.

Results: The amount of [U-13C]-palmitate injected was aimed to correspond to the increase in plasma FFA typically caused by 15h fasting or 1h of moderately intense running in mice, to a calculated maximal plasma concentration of 300µmol/L. Ten minutes after the bolus injection, 2.5±0.5µmol/L free tracer were left in plasma and 39±12 and 14±4 nmol/gprotein in liver and muscle, respectively. Acylcarnitines derived from the tracer reached a plasma concentration of 0.82±0.18 nmol/L and were considerably higher in muscle than in the liver, 0.95±0.47 versus 0.002±0.001 nmol/gprotein. Lipids incorporating palmitate tracer were only detectable in the liver, a total of 511±160 nmol/gprotein as triglycerides and 58±9 nmol/gprotein as phosphatidylcholine.

Conclusions: By using 13C-labelled palmitate as a tracer, we could show that compared to skeletal muscle, the production of acylcarnitines from long-chain FFA in the liver is negligible. Thus, it can be concluded that the muscle and not the liver is responsible for the increase in plasma long-chain acylcarnitines during fasting. In addition, this tracer study confirmed the central role of the liver as buffering system for the storage of excess fatty acids present in the circulation during fasting and revealed that the bulk of labelled palmitate is incorporated into hepatic triglyceride and phosphatidylcholine.