Aims/hypothesis While it is well known that diet-induced obesity causes insulin resistance, the precise mechanisms underpinning the initiation of insulin resistance are unclear. To determine factors that may cause insulin resistance, we have performed a detailed time-course study in mice fed a high-fat diet (HFD). Methods C57Bl/6 mice were fed chow or an HFD from 3 days to 16 weeks and glucose tolerance and tissue-specific insulin action were determined. Tissue lipid profiles were analysed by mass spectrometry and inflammatory markers were measured in adipose tissue, liver and skeletal muscle. Results Glucose intolerance developed within 3 days of the HFD and did not deteriorate further in the period to 12 weeks. Whole-body insulin resistance, measured by hyperinsulinaemic-euglycaemic clamp, was detected after

BackgroundObesity and type 2 diabetes (T2DM) are associated with increased circulating free fatty acids and triacylglycerols. However, very little is known about specific molecular lipid species associated with these diseases. In order to gain further insight into this, we performed plasma lipidomic analysis in a rodent model of obesity and insulin resistance as well as in lean, obese and obese individuals with T2DM.Methodology/Principal FindingsLipidomic analysis using liquid chromatography coupled to mass spectrometry revealed marked changes in the plasma of 12 week high fat fed mice. Although a number of triacylglycerol and diacylglycerol species were elevated along with of a number of sphingolipids, a particularly interesting finding was the high fat diet (HFD)-induced reduction in lysophosphatidylcholine (LPC) levels. As liver, skeletal muscle and adipose tissue play an important role in metabolism, we next determined whether the HFD altered LPCs in these tissues. In contrast to our findings in plasma, only very modest changes in tissue LPCs were noted. To determine when the change in plasma LPCs occurred in response to the HFD, mice were studied after 1, 3 and 6 weeks of HFD. The HFD caused rapid alterations in plasma LPCs with most changes occurring within the first week. Consistent with our rodent model, data from our small human cohort showed a reduction in a number of LPC species in obese and obese individuals with T2DM. Interestingly, no differences were found between the obese otherwise healthy individuals and the obese T2DM patients.ConclusionIrrespective of species, our lipidomic profiling revealed a generalized decrease in circulating LPC species in states of obesity. Moreover, our data indicate that diet and adiposity, rather than insulin resistance or diabetes per se, play an important role in altering the plasma LPC profile.

SUMMARYEpithelial invagination is a common feature of embryogenesis. An example of invagination morphogenesis occurs during development of the early eye when the lens placode forms the lens pit. This morphogenesis is accompanied by a columnar-toconical cell shape change (apical constriction or AC) and is known to be dependent on the cytoskeletal protein Shroom3. Because Shroom3-induced AC can be Rock1/2 dependent, we hypothesized that during lens invagination, RhoA, Rock and a RhoA guanine nucleotide exchange factor (RhoA-GEF) would also be required. In this study, we show that Rock activity is required for lens pit invagination and that RhoA activity is required for Shroom3-induced AC. We demonstrate that RhoA, when activated and targeted apically, is sufficient to induce AC and that RhoA plays a key role in Shroom3 apical localization. Furthermore, we identify Trio as a RhoA-GEF required for Shroom3-dependent AC in MDCK cells and in the lens pit. Collectively, these data indicate that a Trio-RhoA-Shroom3 pathway is required for AC during lens pit invagination.

Interleukin-6 (IL-6) plays a paradoxical role in inflammation and metabolism. The pro-inflammatory effects of IL-6 are mediated via IL-6 "trans-signaling," a process where the soluble form of the IL-6 receptor (sIL-6R) binds IL-6 and activates signaling in inflammatory cells that express the gp130 but not the IL-6 receptor. Here we show that trans-signaling recruits macrophages into adipose tissue (ATM). Moreover, blocking trans-signaling with soluble gp130Fc protein prevents high-fat diet (HFD)-induced ATM accumulation, but does not improve insulin action. Importantly, however, blockade of IL-6 trans-signaling, unlike complete ablation of IL-6 signaling, does not exacerbate obesity-induced weight gain, liver steatosis, or insulin resistance. Our data identify the sIL-6R as a critical chemotactic signal for ATM recruitment and suggest that selectively blocking IL-6 trans-signaling may be a more favorable treatment option for inflammatory diseases, compared with current treatments that completely block the action of IL-6 and negatively impact upon metabolic homeostasis.

The sphingolipids sphingosine-1-phosphate (S1P) and ceramide are important bioactive lipids with many cellular effects. Intracellular ceramide accumulation causes insulin resistance, but sphingosine kinase 1 (SphK1) prevents ceramide accumulation, in part, by promoting its metabolism into S1P. Despite this, the role of SphK1 in regulating insulin action has been largely overlooked. Transgenic (Tg) mice that overexpress SphK1 were fed a standard chow or high-fat diet (HFD) for 6 weeks before undergoing several metabolic analyses. SphK1 Tg mice fed an HFD displayed increased SphK activity in skeletal muscle, which was associated with an attenuated intramuscular ceramide accumulation compared with wild-type (WT) littermates. This was associated with a concomitant reduction in the phosphorylation of c-jun amino-terminal kinase, a serine threonine kinase associated with insulin resistance. Accordingly, skeletal muscle and whole-body insulin sensitivity were improved in SphK1 Tg, compared with WT mice, when fed an HFD. We have identified that the enzyme SphK1 is an important regulator of lipid partitioning and insulin action in skeletal muscle under conditions of increased lipid supply.

The original version of this article included a misprint of the Gene Expression Omnibus (GEO) number for the microarray data reported in the paper. The actual accession number is GEO: GSE63761. In addition, the Supplemental Information was published with the Supplemental Experimental Procedures missing. Both of these errors have since been corrected online. The journal apologizes for any inconvenience.

FTY720 is a sphingosine-1-phosphate analog that has been shown to inhibit ceramide synthesis in vitro. Because ceramide accumulation in muscle is associated with insulin resistance, we aimed to examine whether FTY720 would prevent muscle ceramide accumulation in high fat-fed mice and subsequently improve glucose homeostasis. Male C57Bl/6 mice were fed either a chow or high fat-diet (HFD) for 6 wk, after which they were treated with vehicle or FTY720 (5 mg/kg) daily for a further 6 wk. The ceramide content of muscle was examined and insulin action was assessed. Whereas the HFD increased muscle ceramide, this was prevented by FTY720 treatment. This was not associated with alterations in the expression of genes involved in sphingolipid metabolism. Interestingly, the effects of FTY720 on lipid metabolism were not limited to ceramide because FTY720 also prevented the HFD-induced increase in diacylglycerol and triacylglycerol in muscle. Furthermore, the increase in CD36 mRNA expression induced by fat feeding was prevented in muscle of FTY720-treated mice. This was associated with an attenuation of the HFD-induced increase in palmitate uptake and esterification. In addition, FTY720 improved glucose homeostasis as demonstrated by a reduction in plasma insulin, an improvement in whole-body glucose tolerance, an increase in insulin-stimulated glucose uptake, and Akt phosphorylation in muscle. In conclusion, FTY720 exerts beneficial effects on muscle lipid metabolism that prevent lipid accumulation and improve glucose tolerance in high fat-fed mice. Thus, FTY720 and other compounds that target sphingosine-1-phosphate signaling may have therapeutic potential in treating insulin resistance.

Diabetes mellitus is a serious threat to human health. Tea is cultivated around the world, and its polysaccharide components are reported to be an effective approach for managing type 2 diabetes with fewer adverse effects than medication. To examine the therapeutic effect of tea polysaccharides on diabetes, a type 2 diabetic rat model was generated. We showed that tea polysaccharides remarkably decreased fasting blood glucose and the levels of total cholesterol, total triglyceride, low-density lipoprotein cholesterol, and free fatty acid of type 2 diabetic rats. 16S rRNA sequencing and metabolomics were used to investigate the variation of gut microbiota and the metabolites profiles of diabetic rats after intervention of tea polysaccharides. We found that tea polysaccharides maintained the diversity of gut microbiota and restored the relative abundance of some bacterial genera (Lachnospira, Victivallis, Roseburia, and Fluviicola) which was reduced by diabetes. According to metabolomics analysis, we found that amino acid and other related metabolites was influenced by tea polysaccharides intervention. Correlation analysis among metabolites, gut microbiota, and parameters of hypoglycemic indicated that tea polysaccharides had hypoglycemic and hypolipidemic effect on type 2 diabetes via the modulation of gut microbiota and the improvement of host metabolism.