Two regulatory T cell populations in the visceral adipose tissue shape systemic metabolism.

Visceral adipose tissue (VAT) is an energy store and endocrine organ critical for metabolic homeostasis. Regulatory T (Treg) cells restrain inflammation to preserve VAT homeostasis and glucose tolerance. Here, we show that the VAT harbors two distinct Treg cell populations: prototypical serum stimulation 2-positive (ST2+) Treg cells that are enriched in males and a previously uncharacterized population of C-X-C motif chemokine receptor 3-positive (CXCR3+) Treg cells that are enriched in females. We show that the transcription factors GATA-binding protein 3 and peroxisome proliferator-activated receptor-γ, together with the cytokine interleukin-33, promote the differentiation of ST2+ VAT Treg cells but repress CXCR3+ Treg cells. Conversely, the differentiation of CXCR3+ Treg cells is mediated by the cytokine interferon-γ and the transcription factor T-bet, which also antagonize ST2+ Treg cells. Finally, we demonstrate that ST2+ Treg cells preserve glucose homeostasis, whereas CXCR3+ Treg cells restrain inflammation in lean VAT and prevent glucose intolerance under high-fat diet conditions. Overall, this study defines two molecularly and developmentally distinct VAT Treg cell types with unique context- and sex-specific functions.

Dysferlin deficiency alters lipid metabolism and remodels the skeletal muscle lipidome in mice.

Defects in the gene coding for dysferlin, a membrane-associated protein, affect many tissues, including skeletal muscles, with a resultant myopathy called dysferlinopathy. Dysferlinopathy manifests postgrowth with a progressive loss of skeletal muscle function, early intramyocellular lipid accumulation, and a striking later replacement of selective muscles by adipocytes. To better understand the changes underpinning this disease, we assessed whole-body energy homeostasis, skeletal muscle fatty acid metabolism, lipolysis in adipose tissue, and the skeletal muscle lipidome using young adult dysferlin-deficient male BLAJ mice and age-matched C57Bl/6J WT mice. BLAJ mice had increased lean mass and reduced fat mass associated with increased physical activity and increased adipose tissue lipolysis. Skeletal muscle fatty acid metabolism was remodeled in BLAJ mice, characterized by a partitioning of fatty acids toward storage rather than oxidation. Lipidomic analysis identified marked changes in almost all lipid classes examined in the skeletal muscle of BLAJ mice, including sphingolipids, phospholipids, cholesterol, and most glycerolipids but, surprisingly, not triacylglycerol. These observations indicate that an early manifestation of dysferlin deficiency is the reprogramming of skeletal muscle and adipose tissue lipid metabolism, which is likely to contribute to the progressive adverse histopathology in dysferlinopathies.

Pilot investigations into the mechanistic basis for adverse effects of glucocorticoids in dysferlinopathy.

BACKGROUND: Dysferlinopathies are a clinically heterogeneous group of muscular dystrophies caused by gene mutations resulting in deficiency of the membrane-associated protein dysferlin. They manifest post-growth and are characterised by muscle wasting (primarily in the limb and limb-gridle muscles), inflammation, and replacement of myofibres with adipose tissue. The precise pathomechanism for dysferlinopathy is currently unclear; as such there are no treatments currently available. Glucocorticoids (GCs) are widely used to reduce inflammation and treat muscular dystrophies, but when administered to patients with dysferlinopathy, they have unexpected adverse effects, with accelerated loss of muscle strength. METHODS: To investigate the mechanistic basis for the adverse effects of GCs in dysferlinopathy, the potent GC dexamethasone (Dex) was administered for 4-5 weeks (0.5-0.75 µg/mL in drinking water) to dysferlin-deficient BLA/J and normal wild-type (WT) male mice, sampled at 5 (Study 1) or 10 months (Study 2) of age. A wide range of analyses were conducted. Metabolism- and immune-related gene expression was assessed in psoas muscles at both ages and in quadriceps at 10 months of age. For the 10-month-old mice, quadriceps and psoas muscle histology was assessed. Additionally, we investigated the impact of Dex on the predominantly slow and fast-twitch soleus and extensor digitorum longus (EDL) muscles (respectively) in terms of contractile function, myofibre-type composition, and levels of proteins related to contractile function and metabolism, plus glycogen. RESULTS: At both ages, many complement-related genes were highly expressed in BLA/J muscles, and WT mice were generally more responsive to Dex than BLA/J. The effects of Dex on BLA/J mice included (i) increased expression of inflammasome-related genes in muscles (at 5 months) and (ii) exacerbated histopathology of quadriceps and psoas muscles at 10 months. A novel observation was pronounced staining for glycogen in many myofibres of the damaged quadriceps muscles, with large pale vacuolated myofibres, suggesting possible myofibre death by oncosis. CONCLUSION: These pilot studies provide a new focus for further investigation into the adverse effects of GCs on dysferlinopathic muscles.

Metallothionein 2a gene expression is increased in subcutaneous adipose tissue of type 2 diabetic patients.

STUDY BACKGROUND: Insulin resistance plays an important role in the pathogenesis of type 2 diabetes and the metabolic syndrome. Many of the genes and pathways involved have been identified but some remain to be defined. Metallothioneins (Mts) are a family of anti-oxidant proteins and metallothionein 2a (Mt2a) polymorphims have been recently associated with type 2 diabetes and related complications. Our objective was to determine the Mt2a gene expression levels in adipose tissues from diabetic patients and the effect of Mt treatment on adipocyte insulin sensitivity. METHODS: Samples of subcutaneous and visceral adipose tissues from lean, type 2 diabetic and non-diabetic obese patients were analysed using RT-qPCR for Mt2a mRNA abundance. The regulation of Mt2a expression was further studied in 3T3-L1 adipocytes treated or not with TNFα (10 ng/ml, 72 h) to induce insulin resistance. The effects of Mt on glucose uptake were investigated in cultured adipocytes treated with recombinant Mt protein. RESULTS: We found that the Mt2a gene expression was significantly higher in adipose tissue of type 2 diabetic patients in comparison to that of lean (p=0.003) subjects. In 3T3-L1 adipocytes, insulin resistance induced by TNFα increased Mt2a mRNA levels (p=3×10(-4)) and insulin-stimulated glucose uptake was significantly inhibited by 53% (p=8×10(-4)) compared to vehicle, when 3T3-L1 adipocytes were treated with Mt protein. CONCLUSIONS: These data suggest that Mt2a might be involved in insulin resistance through the up-regulation of Mt gene expression, which may lead to the modulation of insulin action in fat cells. These results suggest the concept of considering Mt proteins as markers and potential targets in type 2 diabetes.

Phosphoproteomics of three exercise modalities identifies canonical signaling and C18ORF25 as an AMPK substrate regulating skeletal muscle function.

Exercise induces signaling networks to improve muscle function and confer health benefits. To identify divergent and common signaling networks during and after different exercise modalities, we performed a phosphoproteomic analysis of human skeletal muscle from a cross-over intervention of endurance, sprint, and resistance exercise. This identified 5,486 phosphosites regulated during or after at least one type of exercise modality and only 420 core phosphosites common to all exercise. One of these core phosphosites was S67 on the uncharacterized protein C18ORF25, which we validated as an AMPK substrate. Mice lacking C18ORF25 have reduced skeletal muscle fiber size, exercise capacity, and muscle contractile function, and this was associated with reduced phosphorylation of contractile and Ca2+ handling proteins. Expression of C18ORF25 S66/67D phospho-mimetic reversed the decreased muscle force production. This work defines the divergent and canonical exercise phosphoproteome across different modalities and identifies C18ORF25 as a regulator of exercise signaling and muscle function.

A Neural basis for Octanoic acid regulation of energy balance.

OBJECTIVES: Nutrient sensing by hypothalamic neurons is critical for the regulation of food intake and energy expenditure. We aimed to identify long- and medium-chain fatty acid species transported into the brain, their effects on energy balance, and the mechanisms by which they regulate activity of hypothalamic neurons. METHODS: Simultaneous blood and cerebrospinal fluid (CSF) sampling was undertaken in rats and metabolic analyses using radiolabeled fatty acid tracers were performed on mice. Electrophysiological recording techniques were used to investigate signaling mechanisms underlying fatty acid-induced changes in activity of pro-opiomelanocortin (POMC) neurons. RESULTS: Medium-chain fatty acid (MCFA) octanoic acid (C8:0), unlike long-chain fatty acids, was rapidly transported into the hypothalamus of mice and almost exclusively oxidized, causing rapid, transient reductions in food intake and increased energy expenditure. Octanoic acid differentially regulates the excitability of POMC neurons, activating these neurons directly via GPR40 and inducing inhibition via an indirect non-synaptic, purine, and adenosine receptor-dependent mechanism. CONCLUSIONS: MCFA octanoic acid is a central signaling nutrient that targets POMC neurons via distinct direct and indirect signal transduction pathways to instigate changes in energy status. These results could explain the beneficial health effects that accompany MCFA consumption.

Mas Receptor Activation Slows Tumor Growth and Attenuates Muscle Wasting in Cancer.

Cancer cachexia is a multifactorial syndrome characterized by a progressive loss of skeletal muscle mass associated with significant functional impairment. Cachexia robs patients of their strength and capacity to perform daily tasks and live independently. Effective treatments are needed urgently. Here, we investigated the therapeutic potential of activating the "alternative" axis of the renin-angiotensin system, involving ACE2, angiotensin-(1-7), and the mitochondrial assembly receptor (MasR), for treating cancer cachexia. Plasmid overexpression of the MasR or pharmacologic angiotensin-(1-7)/MasR activation did not affect healthy muscle fiber size in vitro or in vivo but attenuated atrophy induced by coculture with cancer cells in vitro. In mice with cancer cachexia, the MasR agonist AVE 0991 slowed tumor development, reduced weight loss, improved locomotor activity, and attenuated muscle wasting, with the majority of these effects dependent on the orexigenic and not antitumor properties of AVE 0991. Proteomic profiling and IHC revealed that mechanisms underlying AVE 0991 effects on skeletal muscle involved miR-23a-regulated preservation of the fast, glycolytic fibers. MasR activation is a novel regulator of muscle phenotype, and AVE 0991 has orexigenic, anticachectic, and antitumorigenic effects, identifying it as a promising adjunct therapy for cancer and other serious muscle wasting conditions. SIGNIFICANCE: These findings demonstrate that MasR activation has multiple benefits of being orexigenic, anticachectic, and antitumorigenic, revealing it as a potential adjunct therapy for cancer.Graphical Abstract: http://cancerres.aacrjournals.org/content/canres/79/4/706/F1.large.jpg.See related commentary by Rupert et al., p. 699.

Suppressing fatty acid uptake has therapeutic effects in preclinical models of prostate cancer.

Metabolism alterations are hallmarks of cancer, but the involvement of lipid metabolism in disease progression is unclear. We investigated the role of lipid metabolism in prostate cancer using tissue from patients with prostate cancer and patient-derived xenograft mouse models. We showed that fatty acid uptake was increased in human prostate cancer and that these fatty acids were directed toward biomass production. These changes were mediated, at least partly, by the fatty acid transporter CD36, which was associated with aggressive disease. Deleting Cd36 in the prostate of cancer-susceptible Pten-/- mice reduced fatty acid uptake and the abundance of oncogenic signaling lipids and slowed cancer progression. Moreover, CD36 antibody therapy reduced cancer severity in patient-derived xenografts. We further demonstrated cross-talk between fatty acid uptake and de novo lipogenesis and found that dual targeting of these pathways more potently inhibited proliferation of human cancer-derived organoids compared to the single treatments. These findings identify a critical role for CD36-mediated fatty acid uptake in prostate cancer and suggest that targeting fatty acid uptake might be an effective strategy for treating prostate cancer.