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In the obesity-resistant SJL mouse strain, we previously identified a naturally occurring loss-of-function mutation in the gene for Tbc1d1. Characterization of recombinant inbred mice that carried the Tbc1d1(SJL) allele on a C57BL/6J background indicated that loss of TBC1D1 protects from obesity, presumably by increasing the use of fat as energy source. To provide direct functional evidence for an involvement of TBC1D1 in energy substrate metabolism, we generated and characterized conventional Tbc1d1 knockout mice. TBC1D1-deficient mice showed moderately reduced body weight, decreased respiratory quotient, and an elevated resting metabolic rate. Ex vivo analysis of intact isolated skeletal muscle revealed a severe impairment in insulin- and AICAR-stimulated glucose uptake in glycolytic extensor digitorum longus muscle and a substantially increased rate of fatty acid oxidation in oxidative soleus muscle. Our results provide direct evidence that TBC1D1 plays a major role in glucose and lipid utilization, and energy substrate preference in skeletal muscle.

Leptin administration within the subphysiological to physiological range diminishes atherosclerotic lesions. Leptin appears to mediate its antiatherogenic effects indirectly through reduction of hypercholesterolemia and liver steatosis, as well as upregulation of insulin-sensitizing and atheroprotective adiponectin.

Transgenic (UCP1-TG) mice with ectopic expression of UCP1 in skeletal muscle (SM) show a phenotype of increased energy expenditure, improved glucose tolerance and increase substrate metabolism in SM. To investigate the potential role of skeletal muscle AMPKα2 activation in the metabolic phenotype of UCP1-TG mice we generated double transgenic (DTG) mice, by crossing of UCP1-TG mice with DN-AMPKα2 mice overexpressing a dominant negative α2 subunit of AMPK in SM which resulted in an impaired AMPKα2 activity by 90±9% in SM of DTG mice. Biometric analysis of young male mice showed decreased body weight, lean and fat mass for both UCP1-TG and DTG compared to WT and DN-AMPKα2 mice. Energy intake and weight-specific total energy expenditure were increased, both in UCP1-TG and DTG mice. Moreover, glucose tolerance, insulin sensitivity and fatty acid oxidation were not altered in DTG compared to UCP1-TG. Also uncoupling induced induction and secretion of fibroblast growth factor 21 (FGF21) from SM was preserved in DTG mice. However, voluntary physical cage activity as well as ad libitum running wheel access during night uncovered a severe activity intolerance of DTG mice. Histological analysis showed a progressive degenerative morphology in SM of DTG mice which was not observed in SM of UCP1-TG mice. Moreover, ATP-depletion related cellular stress response via heat shock protein 70 was highly induced, whereas capillarization regulator VEGF was suppressed in DTG muscle. In addition, AMPKα2-mediated induction of mitophagy regulator ULK1 was suppressed in DTG mice, as well as mitochondrial respiratory capacity and content. In conclusion, we demonstrate that AMPKα2 is dispensable for SM mitochondrial uncoupling induced metabolic effects on whole body energy balance, glucose homeostasis and insulin sensitivity. But strikingly, activation of AMPKα2 seems crucial for maintaining SM function, integrity and the ability to compensate chronic metabolic stress induced by SM mitochondrial uncoupling.

Macrophages are essential for innate immunity and inflammatory responses and differentiate into various functional phenotypes. Tribbles homolog 1 (), a member of the mammalian Tribbles homolog pseudokinase family, has been implicated in regulation of cell differentiation, proliferation, and metabolism, but its role in macrophage biology has not been fully elucidated. Here, we investigated the consequences of deficiency on macrophage functions and M1/M2 polarization. Bone marrow-derived macrophages (BMDMs) fromdeficient () mice exhibited elevated phagocytic capacity, correlating with up-regulation of several scavenger receptors. Concomitantly, uptake of modified low-density lipoprotein was increased in BMDMs. macrophages also exhibited diminished migration in the presence of the chemokine MCP-1, associated with reduced expression of the MCP-1 receptor Furthermore, deficiency attenuated the response of BMDMs to both M1 and M2 stimuli; induction of the M1-marker genes ,, and upon LPS/IFNγ stimulation and of the M2-marker genes, , and upon IL-4 stimulation was reduced. Functionally, deficiency decreased secretion of proinflammatory cytokines (IL-6, TNFα, IL-1β, and CXCL1) and reduced nitric oxide and reactive oxygen species production in M1-polarized macrophages. Supporting the attenuated M2 phenotype, IL-4-stimulated macrophages secreted less IL-10 and TGFβ. Mechanistically, BMDMs displayed lower levels of Janus kinase 1 (JAK1), resulting in reduced activation of LPS/IFNγ-mediated STAT1 signaling. Likewise, decreased levels of JAK1 along with lower activation of STAT6 and STAT3 were observed in M2-polarized BMDMs. Our findings suggest that extensively controls macrophage M1/M2 polarization via the JAK/STAT signaling pathway.

In obese leptin-deficient mice, lack of TBC1D1 has no impact on feeding behavior or energy intake but results in increased energy expenditure, altered energy substrate preference with increased fatty acid oxidation and suppression of obesity. TBC1D1 may have an evolutionary conserved role in regulating energy homeostasis in vertebrates.

BACKGROUND: Genetic variants at the TRIB1 gene locus are strongly associated with plasma lipid traits and the risk of coronary artery disease in humans. Here, we analyzed the consequences of Trib1 deficiency on lipid metabolism and atherosclerotic lesion formation in atherosclerosis-susceptible Ldlr −/ − . METHODS: Trib1 −/− mice were crossed onto the Ldlr −/− background to generate double-knockout mice ( Trib1 −/− Ldlr −/− ) and fed a semisynthetic, modified AIN76 diet (0.02% cholesterol and 4.3% fat) until 20 weeks of age. RESULTS: Trib1 −/− Ldlr −/− mice had profoundly larger (5.8-fold) and more advanced atherosclerotic lesions at the aortic root as compared with Trib1 +/+ Ldlr −/− controls. Further, we observed significantly elevated plasma total cholesterol and triglyceride levels in Trib1 −/− Ldlr −/− mice, resulting from higher VLDL (very-low-density lipoprotein) secretion. Lipidomics analysis revealed that loss of Trib1 altered hepatic lipid composition, including the accumulation of cholesterol and proinflammatory ceramide species, which was accompanied by signs of hepatic inflammation and injury. Concomitantly, we detected higher plasma levels of IL (interleukin)-6 and LCN2 (lipocalin 2), suggesting increased systemic inflammation in Trib1 −/− Ldlr −/− mice. Hepatic transcriptome analysis demonstrated significant upregulation of key genes controlling lipid metabolism and inflammation in Trib1 −/− Ldlr −/− mice. Further experiments suggested that these effects may be mediated through pathways involving a C/EPB-PPARγ axis and JNK signaling. CONCLUSIONS: We provide experimental evidence that Trib1 deficiency promotes atherosclerotic lesion formation in a complex manner that includes the modulation of lipid metabolism and inflammation.

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