Aims/hypothesis Mitochondrial DNA (mtDNA) mutations cause several diseases, including mitochondrial inherited diabetes and deafness (MIDD), typically associated with the mtDNA A3243G point mutation on tRNALeu gene. The common hypothesis to explain the link between the genotype and the phenotype is that the mutation might impair mitochondrial metabolism expressly required for beta cell functions. However, this assumption has not yet been tested. Methods We used clonal osteosarcoma cytosolic hybrid cells (namely cybrids) harbouring mitochondria derived from MIDD patients and containing either exclusively wild-type or mutated (A3243G) mtDNA. According to the importance of mitochondrial metabolism in beta cells, we studied the impact of the mutation on key parameters by comparing stimulation of these cybrids by the main insulin secretagogue glucose and the mitochondrial substrate pyruvate. Results Compared with control mtDNA from the same patient, the A3243G mutation markedly modified metabolic pathways leading to a high glycolytic rate (2.8-fold increase), increased lactate production (2.5-fold), and reduced glucose oxidation (−83%). We also observed impaired NADH responses (−56%), negligible mitochondrial membrane potential, and reduced, only transient ATP generation. Moreover, cybrid cells carrying patient-derived mutant mtDNA exhibited deranged cell calcium handling with increased cytosolic loads (1.4-fold higher), and elevated reactive oxygen species (2.6-fold increase) under glucose deprivation. Conclusions/interpretation The present study demonstrates that the mtDNA A3243G mutation impairs crucial metabolic events required for proper cell functions, such as coupling of glucose recognition to insulin secretion.

Weight regain after caloric restriction results in accelerated fat storage in adipose tissue. This catch-up fat phenomenon is postulated to result partly from suppressed skeletal muscle thermogenesis, but the underlying mechanisms are elusive. We investigated whether the reduced rate of skeletal muscle contraction-relaxation cycle that occurs after caloric restriction persists during weight recovery and could contribute to catch-up fat. Using a rat model of semistarvation-refeeding, in which fat recovery is driven by suppressed thermogenesis, we show that contraction and relaxation of leg muscles are slower after both semistarvation and refeeding. These effects are associated with (i) higher expression of muscle deiodinase type 3 (DIO3), which inactivates tri-iodothyronine (T3), and lower expression of T3-activating enzyme, deiodinase type 2 (DIO2), (ii) slower net formation of T3 from its T4 precursor in muscles, and (iii) accumulation of slow fibers at the expense of fast fibers. These semistarvation-induced changes persisted during recovery and correlated with impaired expression of transcription factors involved in slow-twitch muscle development. We conclude that diminished muscle thermogenesis following caloric restriction results from reduced muscle T3 levels, alteration in muscle-specific transcription factors, and fast-to-slow fiber shift causing slower contractility. These energy-sparing effects persist during weight recovery and contribute to catch-up fat.

BackgroundWe have recently demonstrated that palmitoleic acid (16:1n7) increases lipolysis, glucose uptake and glucose utilization for energy production in white adipose cells. In the present study, we tested the hypothesis that palmitoleic acid modulates bioenergetic activity in white adipocytes.MethodsFor this, 3 T3-L1 pre-adipocytes were differentiated into mature adipocytes in the presence (or absence) of palmitic (16:0) or palmitoleic (16:1n7) acid at 100 or 200 μM. The following parameters were evaluated: lipolysis, lipogenesis, fatty acid (FA) oxidation, ATP content, oxygen consumption, mitochondrial mass, citrate synthase activity and protein content of mitochondrial oxidative phosphorylation (OXPHOS) complexes.ResultsTreatment with 16:1n7 during 9 days raised basal and isoproterenol-stimulated lipolysis, FA incorporation into triacylglycerol (TAG), FA oxidation, oxygen consumption, protein expression of subunits representing OXPHOS complex II, III, and V and intracellular ATP content. These effects were not observed in adipocytes treated with 16:0.ConclusionsPalmitoleic acid, by concerted action on lipolysis, FA esterification, mitochondrial FA oxidation, oxygen consumption and ATP content, does enhance white adipocyte energy expenditure and may act as local hormone.

The Arabidopsis thaliana uncoupling protein (UCP) gene was expressed in Escherichia coli and isolated protein reconstituted into liposomes. Linoleic acid-induced H+ fluxes were sensitive to purine nucleotide inhibition with an apparent K(i) (in mM) of 0.8 (GDP), 0.85 (ATP), 0.98 (GTP), and 1.41 (ADP); the inhibition was pH-dependent. Kinetics of AtPUMP1-mediated H+ fluxes were determined for lauric, myristic, palmitic, oleic, linoleic, and linolenic acids. Properties of recombinant AtPUMP1 indicate that it represents a plant counterpart of animal UCP2 or UCP3. This work brings the functional and genetic approaches together for the first time, providing strong support that AtPUMP1 is truly an UCP.

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