Insights into energy balance dysregulation from a mouse model of methylmalonic aciduria.

Inherited disorders of mitochondrial metabolism, including isolated methylmalonic aciduria, present unique challenges to energetic homeostasis by disrupting energy-producing pathways. To better understand global responses to energy shortage, we investigated a hemizygous mouse model of methylmalonyl-CoA mutase (Mmut)-type methylmalonic aciduria. We found Mmut mutant mice to have reduced appetite, energy expenditure and body mass compared with littermate controls, along with a relative reduction in lean mass but increase in fat mass. Brown adipose tissue showed a process of whitening, in line with lower body surface temperature and lesser ability to cope with cold challenge. Mutant mice had dysregulated plasma glucose, delayed glucose clearance and a lesser ability to regulate energy sources when switching from the fed to fasted state, while liver investigations indicated metabolite accumulation and altered expression of peroxisome proliferator-activated receptor and Fgf21-controlled pathways. Together, these shed light on the mechanisms and adaptations behind energy imbalance in methylmalonic aciduria and provide insight into metabolic responses to chronic energy shortage, which may have important implications for disease understanding and patient management.

Integrated multi-omics reveals anaplerotic rewiring in methylmalonyl-CoA mutase deficiency.

Methylmalonic aciduria (MMA) is an inborn error of metabolism with multiple monogenic causes and a poorly understood pathogenesis, leading to the absence of effective causal treatments. Here we employ multi-layered omics profiling combined with biochemical and clinical features of individuals with MMA to reveal a molecular diagnosis for 177 out of 210 (84%) cases, the majority (148) of whom display pathogenic variants in methylmalonyl-CoA mutase (MMUT). Stratification of these data layers by disease severity shows dysregulation of the tricarboxylic acid cycle and its replenishment (anaplerosis) by glutamine. The relevance of these disturbances is evidenced by multi-organ metabolomics of a hemizygous Mmut mouse model as well as through identification of physical interactions between MMUT and glutamine anaplerotic enzymes. Using stable-isotope tracing, we find that treatment with dimethyl-oxoglutarate restores deficient tricarboxylic acid cycling. Our work highlights glutamine anaplerosis as a potential therapeutic intervention point in MMA.

Cellular and computational models reveal environmental and metabolic interactions in MMUT-type methylmalonic aciduria.

Methylmalonyl-coenzyme A (CoA) mutase (MMUT)-type methylmalonic aciduria is a rare inherited metabolic disease caused by the loss of function of the MMUT enzyme. Patients develop symptoms resembling those of primary mitochondrial disorders, but the underlying causes of mitochondrial dysfunction remain unclear. Here, we examined environmental and genetic interactions in MMUT deficiency using a combination of computational modeling and cellular models to decipher pathways interacting with MMUT. Immortalized fibroblast (hTERT BJ5ta) MMUT-KO (MUTKO) clones displayed a mild mitochondrial impairment in standard glucose-based medium, but they did not to show increased reliance on respiratory metabolism nor reduced growth or viability. Consistently, our modeling predicted MUTKO specific growth phenotypes only for lower extracellular glutamine concentrations. Indeed, two of three MMUT-deficient BJ5ta cell lines showed a reduced viability in glutamine-free medium. Further, growth on 183 different carbon and nitrogen substrates identified increased NADH (nicotinamide adenine dinucleotide) metabolism of BJ5ta and HEK293 MUTKO cells compared with controls on purine- and glutamine-based substrates. With this knowledge, our modeling predicted 13 reactions interacting with MMUT that potentiate an effect on growth, primarily those of secondary oxidation of propionyl-CoA, oxidative phosphorylation and oxygen diffusion. Of these, we validated 3-hydroxyisobutytyl-CoA hydrolase (HIBCH) in the secondary propionyl-CoA oxidation pathway. Altogether, these results suggest compensation for the loss of MMUT function by increasing anaplerosis through glutamine or by diverting flux away from MMUT through the secondary propionyl-CoA oxidation pathway, which may have therapeutic relevance.

BRAFV600E Overrides NOTCH Signaling in Thyroid Cancer.

Background: Several mechanisms likely cooperate with the mitogen-activated protein (MAP)-kinase pathway to promote cancer progression in the thyroid. One putative pathway is NOTCH signaling, which is implicated in several other malignancies. In thyroid cancer, data regarding the role of the NOTCH pathway are insufficient and even contradictory. Methods: A BRAFV600E-driven papillary thyroid carcinoma (PTC) mouse model was subjected to NOTCH pathway genetic alterations, and the tumor burden was followed by ultrasound. Further analyses were performed on PTC cell lines or noncancerous cells transfected with NOTCHIC or BRAFV600E, which were then subjected to pharmacological treatment with MAP-kinase or NOTCH pathway inhibitors. Results: The presence of the BRAFV600E mutation coupled with overexpression of the NOTCH intracellular domain led to significantly bigger thyroid tumors in mice, to a more aggressive carcinoma, and decreased overall survival. Although more cystic, the tumors did not progress into anaplastic thyroid carcinomas. On the contrary, the deletion of RBP-jκ (a major cofactor involved in NOTCH signaling) did not alter the phenotype in mice. BRAFV600E-mutated PTC cell lines were resistant to pharmacological inhibition of the NOTCH pathway. Inhibition of MEK1/2 uncovered a predominant effect on Hes1/Hey1 transcription compared with NOTCH inhibition in BRAFV600E-mutated cell lines. Finally, γ-secretase activity and γ-secretase subunit transcription levels were dependent on ERK activation. Our findings suggest that MAP-kinase activity overrides the NOTCH pathway in the context of thyroid cancer. Conclusions: The interaction between the BRAF and NOTCH pathways demonstrates that the BRAFV600E mutation might bypass NOTCH and exert a strong positive effect on NOTCH downstream targets in thyroid carcinoma.

Effects of imidazoline-like drugs on liver and adipose tissues, and their role in preventing obesity and associated cardio-metabolic disorders.

BACKGROUND/OBJECTIVES: We previously observed that selective agonists of the sympatho-inhibitory I1 imidazoline receptors (LNP ligands) have favorable effects on several cardiovascular and metabolic disorders defining the metabolic syndrome, including body weight. The objectives of this study were to explore the effects of LNPs on adiposity and the mechanisms involved, and to evaluate their impact on metabolic homeostasis. METHODS: Young Zucker fa/fa rats were treated with LNP599 (10 mg/kg/day) for 12 weeks. Effects on body weight, adiposity (regional re-distribution, morphology, and function of adipose tissues), cardiovascular and metabolic homeostasis, and liver function were evaluated. Direct effects on insulin and AMP-activated protein kinase (AMPK) signaling were studied in human hepatoma HepG2 cells. RESULTS: LNP599 treatment limited the age-dependent remodeling and inflammation of subcutaneous, epididymal, and visceral adipose tissues, and prevented total fat deposits and the development of obesity. Body-weight stabilization was not related to reduced food intake but rather to enhanced energy expenditure and thermogenesis. Cardiovascular and metabolic parameters were also improved and were significantly correlated with body weight but not with plasma norepinephrine. Insulin and AMPK signaling were enhanced in hepatic tissues of treated animals, whereas blood markers of hepatic disease and pro-inflammatory cytokine levels were reduced. In cultured HepG2 cells, LNP ligands phosphorylated AMPK and the downstream acetyl-CoA carboxylase and prevented oleic acid-induced intracellular lipid accumulation. They also significantly potentiated insulin-mediated AKT activation and this was independent from AMPK. CONCLUSIONS: Selective I1 imidazoline receptor agonists protect against the development of adiposity and obesity, and the associated cardio-metabolic disorders. Activation of I1 receptors in the liver, leading to stimulation of the cellular energy sensor AMPK and insulin sensitization, and in adipose tissues, leading to improvement of morphology and function, are identified as peripheral mechanisms involved in the beneficial actions of these ligands.