OBJECTIVE-Obesity is associated with endocrine abnormalities that predict the progression of insulin resistance to type 2 diabetes. Because skeletal muscle has been shown to secrete proteins that could be used as biomarkers, we characterized the secreted protein profile of muscle cells derived from extremely obese (BMI 48.8 Ϯ 14.8 kg/m 2 ; homeostasis model assessment [HOMA] 3.6 Ϯ 1.0) relative to lean healthy subjects (BMI 25.7 Ϯ 3.2 kg/m 2 ; HOMA 0.8 Ϯ 0.2).RESEARCH DESIGN AND METHODS-We hypothesized that skeletal muscle would secrete proteins that predict the severity of obesity. To test this hypothesis, we used a "bottom-up" experimental design using stable isotope labeling by amino acids in culture (SILAC) and liquid chromatography/mass spectometry/ mass spectometry (LC-MS/MS) to both identify and quantify proteins secreted from cultured myotubes derived from extremely obese compared with healthy nonobese women.RESULTS-Using SILAC, we discovered a 2.9-fold increase in the secretion of myostatin from extremely obese human myotubes. The increased secretion and biological activity of myostatin were validated by immunoblot (3.16 Ϯ 0.18, P Ͻ 0.01) and a myoblast proliferation assay using conditioned growth medium. Myostatin was subsequently shown to increase in skeletal muscle (23%, P Ͻ 0.05) and plasma (35%, P Ͻ 0.05) and to correlate (r 2 ϭ 0.6, P Ͻ 0.05) with the severity of insulin resistance.CONCLUSIONS-Myostatin is a potent antianabolic regulator of muscle mass that may also play a role in energy metabolism. These findings show that increased expression of myostatin in skeletal muscle with obesity and insulin resistance results in elevated circulating myostatin. This may contribute to systemic metabolic deterioration of skeletal muscle with the progression of insulin resistance to type 2 diabetes. Diabetes 58: [30][31][32][33][34][35][36][37][38] 2009 O besity and type 2 diabetes are associated with endocrine abnormalities that are either precipitated by or precede the onset of peripheral insulin resistance (1). These include changes in circulating proteins and peptides that produce endothelial dysfunction, low-grade inflammation, and a prothrombotic state, all of which contribute to increased cardiovascular risk (2-4). Secreted proteins or the "secretome" constitute an important class of biologically active molecules that are released into circulation where they facilitate cross-talk between organ systems. Because secreted proteins are also involved in the progression of cardiovascular disease and cancer, there is significant interest in mining the secretome for novel biological markers (5). Whereas endocrine organs specialize in the secretion of proteins into circulation, there is mounting evidence that adipose tissue and skeletal muscle constitutively or intermittently secrete bioactive proteins (6,7). In this study, we hypothesized that skeletal muscle of extremely obese and insulinresistant women would secrete proteins into circulation that act as prognostic or diagnostic biomarkers of obesityass...

Aspartame consumption is implicated in the development of obesity and metabolic disease despite the intention of limiting caloric intake. The mechanisms responsible for this association remain unclear, but may involve circulating metabolites and the gut microbiota. Aims were to examine the impact of chronic low-dose aspartame consumption on anthropometric, metabolic and microbial parameters in a diet-induced obese model. Male Sprague-Dawley rats were randomized into a standard chow diet (CH, 12% kcal fat) or high fat (HF, 60% kcal fat) and further into ad libitum water control (W) or low-dose aspartame (A, 5–7 mg/kg/d in drinking water) treatments for 8 week (n = 10–12 animals/treatment). Animals on aspartame consumed fewer calories, gained less weight and had a more favorable body composition when challenged with HF compared to animals consuming water. Despite this, aspartame elevated fasting glucose levels and an insulin tolerance test showed aspartame to impair insulin-stimulated glucose disposal in both CH and HF, independently of body composition. Fecal analysis of gut bacterial composition showed aspartame to increase total bacteria, the abundance of Enterobacteriaceae and Clostridium leptum. An interaction between HF and aspartame was also observed for Roseburia ssp wherein HF-A was higher than HF-W (P<0.05). Within HF, aspartame attenuated the typical HF-induced increase in the Firmicutes:Bacteroidetes ratio. Serum metabolomics analysis revealed aspartame to be rapidly metabolized and to be associated with elevations in the short chain fatty acid propionate, a bacterial end product and highly gluconeogenic substrate, potentially explaining its negative affects on insulin tolerance. How aspartame influences gut microbial composition and the implications of these changes on the development of metabolic disease require further investigation.

Purpose There is mounting evidence that skeletal muscle produces and secretes biologically active proteins or “myokines” that facilitate metabolic cross talk between organ systems. The increased expression of myostatin, a secreted anabolic inhibitor of muscle growth and development, has been associated with obesity and insulin resistance. Despite these intriguing findings, there have been few studies linking myostatin and insulin resistance. Methods To explore this relationship in more detail, we quantified myostatin protein in muscle and plasma from 10 insulin-resistant, middle aged (53.1 ± 5.5 years) men before and after 6 months of moderate aerobic exercise training (1200 kcal/wk at 40–55% peak VO2). To establish a case-effect relationship we also injected C57/Bl6 male mice with high-physiologic levels of recombinant myostatin protein. Results Myostatin protein levels were shown to decrease in muscle (37%, P=0.042, n=10) and matching plasma samples (28.7 pre-training to 22.8 ng/ml post-training, P=0.003, n=9) with aerobic exercise. Furthermore, the strong correlation between plasma myostatin levels and insulin sensitivity (R2 = 0.82, P<0.001, n=9) suggested a cause-effect relationship that was subsequently confirmed by inducing insulin resistance in myostatin-injected mice. A modest increase (44%) in plasma myostatin levels was also associated with significant reductions in the insulin-stimulated phosphorylation of AKT (Thr308) in both muscle and liver of myostatin treated animals. Conclusions These findings indicate that both muscle and plasma myostatin protein levels are regulated by aerobic exercise and furthermore, that myostatin is in the causal pathway of acquired insulin resistance with physical inactivity.

BackgroundGastrointestinal dysfunction and gut microbial composition disturbances have been widely reported in autism spectrum disorder (ASD). This study examines whether gut microbiome disturbances are present in the BTBRT + tf/j (BTBR) mouse model of ASD and if the ketogenic diet, a diet previously shown to elicit therapeutic benefit in this mouse model, is capable of altering the profile.FindingsJuvenile male C57BL/6 (B6) and BTBR mice were fed a standard chow (CH, 13 % kcal fat) or ketogenic diet (KD, 75 % kcal fat) for 10–14 days. Following diets, fecal and cecal samples were collected for analysis. Main findings are as follows: (1) gut microbiota compositions of cecal and fecal samples were altered in BTBR compared to control mice, indicating that this model may be of utility in understanding gut-brain interactions in ASD; (2) KD consumption caused an anti-microbial-like effect by significantly decreasing total host bacterial abundance in cecal and fecal matter; (3) specific to BTBR animals, the KD counteracted the common ASD phenotype of a low Firmicutes to Bacteroidetes ratio in both sample types; and (4) the KD reversed elevated Akkermansia muciniphila content in the cecal and fecal matter of BTBR animals.ConclusionsResults indicate that consumption of a KD likely triggers reductions in total gut microbial counts and compositional remodeling in the BTBR mouse. These findings may explain, in part, the ability of a KD to mitigate some of the neurological symptoms associated with ASD in an animal model.Electronic supplementary materialThe online version of this article (doi:10.1186/s13229-016-0099-3) contains supplementary material, which is available to authorized users.

Myostatin is a member of the transforming growth factor-beta/bone morphogenetic protein (TGF-β/BMP) super-family of secreted factors that functions as a potent inhibitor of skeletal muscle growth. Moreover, considerable evidence has accumulated that myostatin also regulates metabolism and that its inhibition can significantly attenuate the progression of obesity and diabetes. While at least part of these effects on metabolism can be attributable to myostatin’s influence over skeletal muscle growth and therefore on the total volume of metabolically active lean body mass, there is mounting evidence that myostatin affects the growth and metabolic state of other tissues, including the adipose and the liver. In addition, recent work has explored the role of myostatin in substrate mobilization, uptake and/or utilization of muscle independent of its effects on body composition. Finally, the effects of both endurance and resistance exercise on myostatin expression, as well as the potential role of myostatin in the beneficial metabolic adaptations occurring in response to exercise, have also begun to be delineated in greater detail. The purpose of this review is to summarize the work to date on the expression and function of myostatin in obesity, diabetes, and exercise adaptation.

The predictive ability of metabolic profiling to detect obesity-induced perturbations in metabolism has not been clearly established. Complex aetiologies interacting with environmental factors highlight the need to understand how specific manipulations alter metabolite profiles in this state. The aim of this study was to determine if targeted metabolomic profiling could be employed as a reliable tool to detect dietary-induced insulin resistance in a small subset of experimental animals (n = 10/treatment). Following weaning, male C57BL/6J littermates were randomly divided into two dietary groups: chow and high fat. Following 12 weeks of dietary manipulation, mice were fasted for 5 h prior to serum collection. The resultant high fat-fed animals were obese and insulin resistant as shown by a euglycaemic-hyperinsulinaemic clamp. Sera were analysed by proton nuclear magnetic resonance spectroscopy, and 46 known compounds were identified and quantified. Multivariate analysis by orthogonal partial least squares discriminant analysis, a projection method for class separation, was then used to establish models of each treatment. Models were able to predict class separation between diets with 90% accuracy. Variable importance plots revealed the most important metabolites in this discrimination to include lysine, glycine, citrate, leucine, suberate and acetate. These metabolites are involved in energy metabolism and may be representative of the perturbations taking place with insulin resistance. Results show metabolomics to reliably describe the metabolic effects of insulin resistance in a small subset of samples and are an initial step in establishing metabolomics as a tool to understand the biochemical signature of insulin resistance.

Watts R, Johnsen VL, Shearer J, Hittel DS. Myostatin-induced inhibition of the long noncoding RNA Malat1 is associated with decreased myogenesis.

Objective-Statins are safe medications but have side effects including myalgia and rhabdomyolysis. How statins provoke muscle damage is not known, but this effect is exacerbated by exercise. Methods and Results-Healthy subjects took Atorvastatin (80 mg/daily) or placebo for 4 weeks. Biopsies of both vastus lateralis muscles were performed 8 hours after eccentric exercise (known to result in muscle soreness and damage) of the left leg at baseline and the right leg after statin/placebo treatment. Gene expression was determined using Affymetrix GeneChips, and selected genes confirmed by polymerase chain reaction (qRT-PCR). Atorvastatin had little effect on gene expression at rest. When combined with exercise, 56 genes were differentially expressed with 18% involved in the ubiquitin proteasome pathway (UPP) and 20% involved in protein folding and catabolism, and apoptosis. Conclusion-This is the first investigation to our knowledge to implicate involvement of the UPP in skeletal muscle in response to combined exercise and statin treatment, possibly explaining the onset of myalgia with exertion.