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. 2020 Jun;63(6):1211-1222.
doi: 10.1007/s00125-020-05128-1. Epub 2020 Mar 17.

One-leg inactivity induces a reduction in mitochondrial oxidative capacity, intramyocellular lipid accumulation and reduced insulin signalling upon lipid infusion: a human study with unilateral limb suspension

Lena Bilet  1   2 , Esther Phielix  1   2 , Tineke van de Weijer  1   2   3 , Anne Gemmink  1   2 , Madeleen Bosma  1   2 , Esther Moonen-Kornips  1   2 , Johanna A Jorgensen  1   2 , Gert Schaart  2 , Dongyan Zhang  4 , Kenneth Meijer  1   2 , Maria Hopman  5 , Matthijs K C Hesselink  1   2 , D Margriet Ouwens  6 , Gerald I Shulman  4   7 , Vera B Schrauwen-Hinderling  1   2   3 , Patrick Schrauwen  8   9
Affiliations

One-leg inactivity induces a reduction in mitochondrial oxidative capacity, intramyocellular lipid accumulation and reduced insulin signalling upon lipid infusion: a human study with unilateral limb suspension

Lena Bilet et al. Diabetologia. 2020 Jun.

Abstract

Aims/hypothesis: Physical inactivity, low mitochondrial function, increased intramyocellular lipid (IMCL) deposition and reduced insulin sensitivity are common denominators of chronic metabolic disorders, like obesity and type 2 diabetes. Yet, whether low mitochondrial function predisposes to insulin resistance in humans is still unknown.

Methods: Here we investigated, in an intervention study, whether muscle with low mitochondrial oxidative capacity, induced by one-legged physical inactivity, would feature stronger signs of lipid-induced insulin resistance. To this end, ten male participants (age 22.4 ± 4.2 years, BMI 21.3 ± 2.0 kg/m2) underwent a 12 day unilateral lower-limb suspension with the contralateral leg serving as an active internal control.

Results: In vivo, mitochondrial oxidative capacity, assessed by phosphocreatine (PCr)-recovery half-time, was lower in the inactive vs active leg. Ex vivo, palmitate oxidation to 14CO2 was lower in the suspended leg vs the active leg; however, this did not result in significantly higher [14C]palmitate incorporation into triacylglycerol. The reduced mitochondrial function in the suspended leg was, however, paralleled by augmented IMCL content in both musculus tibialis anterior and musculus vastus lateralis, and by increased membrane bound protein kinase C (PKC) θ. Finally, upon lipid infusion, insulin signalling was lower in the suspended vs active leg.

Conclusions/interpretation: Together, these results demonstrate, in a unique human in vivo model, that a low mitochondrial oxidative capacity due to physical inactivity directly impacts IMCL accumulation and PKCθ translocation, resulting in impaired insulin signalling upon lipid infusion. This demonstrates the importance of mitochondrial oxidative capacity and muscle fat accumulation in the development of insulin resistance in humans.

Trial registration: ClinicalTrial.gov NCT01576250.

Funding: PS was supported by a 'VICI' Research Grant for innovative research from the Netherlands Organization for Scientific Research (Grant 918.96.618).

Keywords: Fat oxidation; Insulin resistance; Intramyocellular lipid content; Mitochondrial function; Mitochondrial oxidative capacity; Physical inactivity; Unilateral lower-limb suspension.

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Figures

Fig. 1
Fig. 1
(a) Study design and (b) ULLS set up. Image in (b) from Berg et al. [23], reprinted with permission from the American Physiological Society. DEXA, dual energy x-ray absorptiometry
Fig. 2
Fig. 2
Participant compliance to the intervention as measured with (a) activity monitors (accelerometers) (n = 4–9) and (b) temperature sensors (n = 9). *p < 0.05. Data expressed as mean ± SEM
Fig. 3
Fig. 3
Mitochondrial oxidative capacity and incorporation of labelled palmitate into DAG and TAG in the active vs the suspended leg, post-suspension, in the overnight fasted state. (a–c) Mitochondrial oxidative capacity expressed as (a) PCr-recovery T1/2 in vivo (n = 10), and (b,c) ex vivo [14C]palmitate oxidation to CO2 (b; n = 9) and ASMs (c; n = 9). (d, e (both n = 9) [14C]palmitate incorporation into TAGs (d) and DAGs (e). *p < 0.05
Fig. 4
Fig. 4
(a) In vivo IMCL in the musculus tibialis anterior, measured by 1H-MRS, in the active vs the suspended leg, post-suspension, in the overnight fasted state (n = 9). (b) Ex vivo IMCL in musculus vastus lateralis, measured with Oil Red O staining, in the overnight fasted state (n = 6). *p < 0.05, **p < 0.01
Fig. 5
Fig. 5
Ratio of PKCθ in the muscle membrane:cytosol (a measure of PKCθ activation) in the overnight fasted state. Representative blots are also shown (a, active; s, suspended). n = 8; *p < 0.05
Fig. 6
Fig. 6
Plasma concentrations of (a) NEFA (n = 10; data expressed as mean ± SEM) and (b) glucose during the insulin+lipid infusion (n = 10). ***p < 0.001, compared with baseline (time [t]0)
Fig. 7
Fig. 7
The expression level of (a) insulin receptor, (b) p-IRS1Ser1101, (c) p-AktThr308, (d) p-AktSer473, (e) PDK4, (f) p-GSSer641, (g) p-GSK3βSer9, (h) p-FOXO (FOXO1Thr24/FOXO3aThr32), (i) p-AMPKThr172 and (j) p-ACCSer79 in the active vs the suspended leg, in the insulin-stimulated state. Representative blots also shown (a, active; s, suspended). n = 8; *p < 0.05
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