Froukje Vanweert^1*, Anne Gemmink¹, Gert Schaart¹, Nima Gharahdaghi³, Kenneth Smith³, Daniel J. Wilkinson³, Philip J. Atherton³, Ludwig J. Dubois², Matthijs K.C. Hesselink¹, and Esther Phielix¹

¹Department of Nutrition and Movement Sciences, NUTRIM Institute of Nutrition and Translational Research in Metabolism, Maastricht University, Maastricht, The Netherlands

²Medical Research Council-Versus Arthritis Centre for Musculoskeletal Ageing Research, National Institute for Health Research Nottingham Biomedical Research Centre, Clinical, Metabolic and Molecular Physiology, University of Nottingham, Royal Derby Hospital, Derby, United Kingdom

³Department of Precision Medicine, GROW Research Institute for Oncology and Reproduction, Maastricht University, Maastricht, The Netherlands

*e-mail: f.vanweert@maastrichtuniversity.nl

Background: There is evidence that type 2 diabetes (T2D) patients feature elevated branched-chain amino acids (BCAA) plasma levels, possibly due to a compromised BCAA catabolism. We recently showed in T2D patients that pharmacologically lowering plasma BCAA levels improved whole-body insulin sensitivity by 27%. Exactly how BCAA is linked to insulin sensitivity is currently unknown. We hypothesize that (1)elevated BCAA levels impede insulin signalling and (2)compromised BCAA catabolism reduces TCA cycle intermediates. This 2-hit approach leads to compromised substrate oxidation and insulin sensitivity.

Methods: Mass spectrometry (insulin-stimulated glucose uptake), Western blotting (insulin signalling) and SeaHorse experiments (mitochondrial respiration and glycolytic metabolism) were conducted to investigate the impact of incremental BCAA concentrations in C2C12 and human primary myotubes.

Results: In insulin-stimulated C2C12 myotubes, exposed to high BCAA concentrations (1.6mM) S6K phosphorylation was induced (0.4mM: 0.55±0.18 vs 1.6mM: 1.93±0.50 RAU, p<0.05), while Akt phosphorylation was blunted (0.4mM: 1.00±0.05 vs 1.6 mM: 0.43±0.06 RAU, p<0.01). The anticipated decreased insulin-stimulated glucose uptake was, however, not detected. In human primary myotubes, a progressive increase in BCAA concentration (from 0.4 to 20 mM) correlated with a blunted mitochondrial respiration (r=-0.65, p=0.007) and tended to lower glycolytic metabolism (r=-0.42; p=0.10) (Fig1).

Conclusion: These preliminary results suggest that elevated BCAA levels impede insulin signalling through S6K and reduce both mitochondrial respiration and glycolytic metabolism in vitro.

Supported by: Kootstra Talent Fellowship Programme (Maastricht UMC+), EMBO Scientific Exchange Grant (number 9142) and Foundation “De Drie Lichten”