IntroductionIn mammals obesity results when caloric intake exceeds energy use (1). Information on the pathways regulating both feeding and energy expenditure has expanded remarkably in the past several years, and a number of novel hypothalamic peptides that either stimulate or inhibit feeding have been described (1). Among these peptides, melanin-concentrating hormone (MCH) is known to play an important role in feeding behavior (2-4). In the brain, MCH expression is limited to the lateral hypothalamus (5) and responds to nutritional signals, including fasting and leptin deficiency (3). Thus, expression is increased with fasting and is also markedly increased in the Lep ob /Lep ob mouse, which lacks leptin (3). When administered intracerebroventricularly (ICV), MCH leads to a rapid increase in feeding in rats (3, 4). Importantly, MCH ablation in mice leads to a syndrome of leanness associated with hypophagia and a relative increase in oxygen consumption (6).The lean phenotype of MCH-ablated mice is of particular interest (6), in that genetic ablation of neuropeptide-Y (NPY), another appetite-stimulating neuropeptide (7, 8), does not affect body weight (9) unless combined with other obesity genes (10). Indeed, MCH is thus far the only known hypothalamic peptide whose ablation results in leanness. This is in contrast to the number of peptides whose mutation or ablation leads to obesity. These peptides include leptin and the melanocortin-4 receptors (11), proopiomelanocortin (POMC) (12), and the serotonin IIC receptor (13).As MCH is a significant regulator of body weight whose absence leads to decreased percentage of body adiposity, we examined the possibility that overexpression of MCH eutopically in the lateral hypothalamus might lead to a syndrome of obesity. To examine this question we generated a line of transgenic animals overexpressing MCH (MCH-OE) in the lateral hypothalamus. On the original FVB background, mice were not obese on a standard diet. However, they became obese when the gene was bred to homozygosity and the animals were fed a high-fat diet. MCH-OE animals were hyperphagic, hyperleptinemic, and had higher blood glucose levels. Animals were also significantly hyperinsulinemic and failed to respond to an insulin challenge. Several lines of investigation suggest that the hypothalamic neuropeptide melanin-concentrating hormone (MCH) regulates body weight in mammals. Obese mice lacking functional leptin overexpress the MCH message in the fed or fasted state. Acute intracerebroventricular injection of MCH increases energy intake in rats. Mice lacking the MCH gene are lean. To test the hypothesis that chronic overexpression of MCH in mice causes obesity, we produced transgenic mice that overexpress MCH (MCH-OE) in the lateral hypothalamus at approximately twofold higher levels than normal mice. On the FVB genetic background, homozygous transgenic animals fed a high-fat diet ate 10% more and were 12% heavier at 13 weeks of age than wild-type animals, and they had higher systemic leptin levels. Blood gluco...

Growth and differentiation factor 8 (GDF8) is a TGF-β superfamily member, and negative regulator of skeletal muscle mass. GDF8 inhibition results in prominent muscle growth in mice, but less impressive hypertrophy in primates, including man. Broad TGF-β inhibition suggests another family member negatively regulates muscle mass, and its blockade enhances muscle growth seen with GDF8-specific inhibition. Here we show that activin A is the long-sought second negative muscle regulator. Activin A specific inhibition, on top of GDF8 inhibition, leads to pronounced muscle hypertrophy and force production in mice and monkeys. Inhibition of these two ligands mimics the hypertrophy seen with broad TGF-β blockers, while avoiding the adverse effects due to inhibition of multiple family members. Altogether, we identify activin A as a second negative regulator of muscle mass, and suggest that inhibition of both ligands provides a preferred therapeutic approach, which maximizes the benefit:risk ratio for muscle diseases in man.

BackgroundLoss of skeletal muscle mass and function in humans is associated with significant morbidity and mortality. The role of myostatin as a key negative regulator of skeletal muscle mass and function has supported the concept that inactivation of myostatin could be a useful approach for treating muscle wasting diseases.MethodsWe generated a myostatin monoclonal blocking antibody (REGN1033) and characterized its effects in vitro using surface plasmon resonance biacore and cell-based Smad2/3 signaling assays. REGN1033 was tested in mice for the ability to induce skeletal muscle hypertrophy and prevent atrophy induced by immobilization, hindlimb suspension, or dexamethasone. The effect of REGN1033 on exercise training was tested in aged mice. Messenger RNA sequencing, immunohistochemistry, and ex vivo force measurements were performed on skeletal muscle samples from REGN1033-treated mice.ResultsThe human monoclonal antibody REGN1033 is a specific and potent myostatin antagonist. Chronic treatment of mice with REGN1033 increased muscle fiber size, muscle mass, and force production. REGN1033 prevented the loss of muscle mass induced by immobilization, glucocorticoid treatment, or hindlimb unweighting and increased the gain of muscle mass during recovery from pre-existing atrophy. In aged mice, REGN1033 increased muscle mass and strength and improved physical performance during treadmill exercise.ConclusionsWe show that specific myostatin antagonism with the human antibody REGN1033 enhanced muscle mass and function in young and aged mice and had beneficial effects in models of skeletal muscle atrophy.

Intensive insulin therapy and protein restriction delay the development of nephropathy in a variety of conditions, but few interventions are known to reverse nephropathy. Having recently observed that the ketone 3-beta-hydroxybutyric acid (3-OHB) reduces molecular responses to glucose, we hypothesized that a ketogenic diet, which produces prolonged elevation of 3-OHB, may reverse pathological processes caused by diabetes. To address this hypothesis, we assessed if prolonged maintenance on a ketogenic diet would reverse nephropathy produced by diabetes. In mouse models for both Type 1 (Akita) and Type 2 (db/db) diabetes, diabetic nephropathy (as indicated by albuminuria) was allowed to develop, then half the mice were switched to a ketogenic diet. After 8 weeks on the diet, mice were sacrificed to assess gene expression and histology. Diabetic nephropathy, as indicated by albumin/creatinine ratios as well as expression of stress-induced genes, was completely reversed by 2 months maintenance on a ketogenic diet. However, histological evidence of nephropathy was only partly reversed. These studies demonstrate that diabetic nephropathy can be reversed by a relatively simple dietary intervention. Whether reduced glucose metabolism mediates the protective effects of the ketogenic diet remains to be determined.

Nutrient-sensitive hypothalamic neurons regulate energy balance and glucose homeostasis, but the molecular mechanisms mediating hypothalamic responses to nutritional state remain incompletely characterized. To address these mechanisms, the present studies used quantitative PCR to characterize the expression of a panel of genes the hypothalamic expression by nutritional status of which had been suggested by DNA microarray studies. Although these genes regulate a variety of function, the most prominent set regulate intermediary metabolism, and the overall pattern clearly indicated that a 48-h fast produced a metabolic reprogramming away from glucose metabolism and toward the utilization of alternative fuels, particularly lipid metabolism. This general reprogramming of intermediary metabolism by fasting was observed both in cortex and hypothalamus but most prominently in hypothalamus. The effect of fasting on the expression of these genes may be mediated by reduction in plasma glucose or glucose metabolism, rather than leptin, because they were generally recapitulated by hypoglycemia even in the presence of elevated insulin and in vitro by low glucose but were not recapitulated in ob/ob mice. These studies suggest that fasting reduces glucose metabolism and thus minimizes the production of hypothalamic malonyl-coenzyme A. However, because the reprogramming of glucose metabolism by fasting was also observed in cortex, this apparent substrate competition may mediate more general responses to nutritional deprivation, including those responsible for the protective effects of dietary restriction. The present studies also provide a large panel of novel glucose-regulated genes that can be used as markers of glucose action to address mechanisms mediating hypothalamic responses to nutritional state.