Protective role of the ELOVL2/docosahexaenoic acid axis in glucolipotoxicity-induced apoptosis in rodent beta cells and human islets.

AIMS/HYPOTHESIS: Dietary n-3 polyunsaturated fatty acids, especially docosahexaenoic acid (DHA), are known to influence glucose homeostasis. We recently showed that Elovl2 expression in beta cells, which regulates synthesis of endogenous DHA, was associated with glucose tolerance and played a key role in insulin secretion. The present study aimed to examine the role of the very long chain fatty acid elongase 2 (ELOVL2)/DHA axis on the adverse effects of palmitate with high glucose, a condition defined as glucolipotoxicity, on beta cells. METHODS: We detected ELOVL2 in INS-1 beta cells and mouse and human islets using quantitative PCR and western blotting. Downregulation and adenoviral overexpression of Elovl2 was carried out in beta cells. Ceramide and diacylglycerol levels were determined by radio-enzymatic assay and lipidomics. Apoptosis was quantified using caspase-3 assays and poly (ADP-ribose) polymerase cleavage. Palmitate oxidation and esterification were determined by [U-14C]palmitate labelling. RESULTS: We found that glucolipotoxicity decreased ELOVL2 content in rodent and human beta cells. Downregulation of ELOVL2 drastically potentiated beta cell apoptosis induced by glucolipotoxicity, whereas adenoviral Elovl2 overexpression and supplementation with DHA partially inhibited glucolipotoxicity-induced cell death in rodent and human beta cells. Inhibition of beta cell apoptosis by the ELOVL2/DHA axis was associated with a decrease in ceramide accumulation. However, the ELOVL2/DHA axis was unable to directly alter ceramide synthesis or metabolism. By contrast, DHA increased palmitate oxidation but did not affect its esterification. Pharmacological inhibition of AMP-activated protein kinase and etomoxir, an inhibitor of carnitine palmitoyltransferase 1 (CPT1), the rate-limiting enzyme in fatty acid ß-oxidation, attenuated the protective effect of the ELOVL2/DHA axis during glucolipotoxicity. Downregulation of CPT1 also counteracted the anti-apoptotic action of the ELOVL2/DHA axis. By contrast, a mutated active form of Cpt1 inhibited glucolipotoxicity-induced beta cell apoptosis when ELOVL2 was downregulated. CONCLUSIONS/INTERPRETATION: Our results identify ELOVL2 as a critical pro-survival enzyme for preventing beta cell death and dysfunction induced by glucolipotoxicity, notably by favouring palmitate oxidation in mitochondria through a CPT1-dependent mechanism.

The Crucial Role of C18-Cer in Fat-Induced Skeletal Muscle Insulin Resistance.

BACKGROUND/AIMS: Muscle bioactive lipids accumulation leads to several disorder states. The most common are insulin resistance (IR) and type 2 diabetes. There is an ongoing debate which of the lipid species plays the major role in induction of muscle IR. Our aim was to elucidate the role of particular lipid group in induction of muscle IR. METHODS: The analyses were performed on muscle from the following groups of rats: 1. Control, fed standard diet, 2 HFD, fed high fat diet, 3. HFD/Myr, fed HFD and treated with myriocin (Myr), an inhibitor of ceramide de novo synthesis. We utilized [U13C] palmitate isotope tracer infusion and mass spectrometry to measure content and synthesis rate of muscle long-chain acyl-CoA (LCACoA), diacylglycerols (DAG) and ceramide (Cer). RESULTS: HFD led to intramuscular accumulation of LCACoA, DAG and Cer and skeletal muscle IR. Myr-treatment caused decrease in Cer (most noticeable for stearoyl-Cer and oleoyl-Cer) and accumulation of DAG, possibly due to re-channeling of excess of intramuscular LCACoA towards DAG synthesis. An improvement in insulin sensitivity at both systemic and muscular level coincided with decrease in ceramide, despite elevated intramuscular DAG. CONCLUSION: The improved insulin sensitivity was associated with decreased muscle stearoyl- and oleoyl-ceramide content. The results indicate that accumulation of those ceramide species has the greatest impact on skeletal muscle insulin sensitivity in rats.

The role of adipose tissue and excess of fatty acids in the induction of insulin resistance in skeletal muscle.

Skeletal muscle is the main tissue responsible for insulin-stimulated glucose uptake. Consumption of a high-fat diet rich in saturated fats (HFD) and obesity are associated with accumulation of intramuscular lipids that leads to several disorders, e.g. insulin resistance (IRes) and type 2 diabetes (T2D). The mechanism underlying the induction of IRes is still unknown. It was speculated that accumulation of intramuscular triacylglycerols (TAG) is linked to induction of IRes. Now, research focuses on bioactive lipids: long-chain acyl-CoA (LCACoA), diacylglycerols (DAG) and ceramides (Cer). It has been demonstrated that accumulation of each of the above-mentioned lipid classes negatively affects the insulin signaling pathway. It is not clear which of those lipids play the most important role in HFD-induced skeletal muscle IRes. The aim of the present work is to present the current knowledge of the role of adipose tissue and excess of fatty acids in the induction of insulin resistance.

Sources, metabolism, and regulation of circulating sphingosine-1-phosphate.

Sphingosine-1-phosphate (S1P) is a bioactive sphingolipid that acts either as an intracellular messenger or as a ligand for its membrane receptors. S1P is a normal constituent of blood, where it is found both in plasma and blood cells. Compared with other cell types, sphingolipid metabolism in erythrocytes and platelets has unique features that allow the erythrocytes and platelets to accumulate S1P. In plasma, S1P is bound mainly to HDLs and albumin. Of note, metabolism and biological activity of S1P is to a large extent affected by the type of its carrier. Plasma S1P is characterized by a short half-life, indicating rapid clearance by degradative enzymes and the presence of high-capacity sources involved in maintaining its high concentration. These sources include blood cells, vascular endothelium, and hepatocytes. However, the extent to which each of these contributes to the plasma pool of S1P is a matter of debate. Circulating S1P plays a significant physiological role. It was found to be the key regulator of lymphocyte trafficking, endothelial barrier function, and vascular tone. The purpose of this review is to summarize the present state of knowledge on the metabolism, transport, and origin of plasma S1P, and to discuss the mechanisms regulating its homeostasis in blood.

Obesity, Bioactive Lipids, and Adipose Tissue Inflammation in Insulin Resistance.

Obesity is a major risk factor for the development of insulin resistance and type 2 diabetes. The exact mechanism by which adipose tissue induces insulin resistance is still unclear. It has been demonstrated that obesity is associated with the adipocyte dysfunction, macrophage infiltration, and low-grade inflammation, which probably contributes to the induction of insulin resistance. Adipose tissue synthesizes and secretes numerous bioactive molecules, namely adipokines and cytokines, which affect the metabolism of both lipids and glucose. Disorders in the synthesis of adipokines and cytokines that occur in obesity lead to changes in lipid and carbohydrates metabolism and, as a consequence, may lead to insulin resistance and type 2 diabetes. Obesity is also associated with the accumulation of lipids. A special group of lipids that are able to regulate the activity of intracellular enzymes are biologically active lipids: long-chain acyl-CoAs, ceramides, and diacylglycerols. According to the latest data, the accumulation of these lipids in adipocytes is probably related to the development of insulin resistance. Recent studies indicate that the accumulation of biologically active lipids in adipose tissue may regulate the synthesis/secretion of adipokines and proinflammatory cytokines. Although studies have revealed that inflammation caused by excessive fat accumulation and abnormalities in lipid metabolism can contribute to the development of obesity-related insulin resistance, further research is needed to determine the exact mechanism by which obesity-related insulin resistance is induced.

The Impact of OMEGA-3 Fatty Acids Supplementation on Insulin Resistance and Content of Adipocytokines and Biologically Active Lipids in Adipose Tissue of High-Fat Diet Fed Rats.

It has been established that OMEGA-3 polyunsaturated fatty acids (PUFAs) may improve lipid and glucose homeostasis and prevent the "low-grade" state of inflammation in animals. Little is known about the effect of PUFAs on adipocytokines expression and biologically active lipids accumulation under the influence of high-fat diet-induced obesity. The aim of the study was to examine the effect of fish oil supplementation on adipocytokines expression and ceramide (Cer) and diacylglycerols (DAG) content in visceral and subcutaneous adipose tissue of high-fat fed animals. The experiments were carried out on Wistar rats divided into three groups: standard diet-control (SD), high-fat diet (HFD), and high-fat diet + fish oil (HFD+FO). The fasting plasma glucose and insulin concentrations were examined. Expression of carnitine palmitoyltransferase 1 (CPT1) protein was determined using the Western blot method. Plasma adipocytokines concentration was measured using ELISA kits and mRNA expression was determined by qRT-PCR reaction. Cer, DAG, and acyl-carnitine (A-CAR) content was analyzed by UHPLC/MS/MS. The fish oil supplementation significantly decreased plasma insulin concentration and Homeostatic Model Assesment for Insulin Resistance (HOMA-IR) index and reduced content of adipose tissue biologically active lipids in comparison with HFD-fed subjects. The expression of CPT1 protein in HFD+FO in both adipose tissues was elevated, whereas the content of A-CAR was lower in both HFD groups. There was an increase of adiponectin concentration and expression in HFD+FO as compared to HFD group. OMEGA-3 fatty acids supplementation improved insulin sensitivity and decreased content of Cer and DAG in both fat depots. Our results also demonstrate that PUFAs may prevent the development of insulin resistance in response to high-fat feeding and may regulate the expression and secretion of adipocytokines in this animal model.

The effect of high fat diet and metformin treatment on liver lipids accumulation and their impact on insulin action.

We sought to determine whether metformin treatment reverses a high-fat diet (HFD)-induced hepatic insulin resistance (IRes) and to identify lipid intermediates involved in induction of liver IRes. The experiments were conducted on male Wistar rats divided into three groups: 1. Control, 2. fed HFD and 3. fed HFD and treated with metformin. The animals were infused with a [U-13C]palmitate to measure fractional lipid synthesis rate. This allowed for the calculation of fractional synthesis rate of signaling lipids (FSR) through the estimation of their isotopic enrichment. Liver ceramide (Cer), diacylglycerol (DAG) and acyl-carnitine concentration and enrichment were analyzed by LC/MS/MS. The content of proteins involved in lipid metabolism and insulin signaling were analyzed by Western Blot. HFD treatment increased the content and FSR of DAG and Cer in the liver which was accompanied by systemic insulin resistance and inhibition of hepatic insulin signaling pathway under insulin stimulation. Metformin treatment ameliorated systemic insulin resistance and augmented the hepatic insulin signaling cascade. It reduced both the concentration and FSR of Cer, DAG, and increased acyl-carnitine content and the expression of mitochondrial markers. We postulate, that in liver, the insulin sensitizing effect of metformin depends on augmentation of mitochondrial ß-oxidation, which protects from hepatic accumulation of both the Cer and DAG and preserves insulin sensitivity under HFD consumption. Moreover, we showed that hepatic content of Cer and DAG corresponds with their respective FSR.

Effect of metformin on bioactive lipid metabolism in insulin-resistant muscle.

Intramuscular accumulation of bioactive lipids leads to insulin resistance and type 2 diabetes (T2D). There is lack of consensus concerning which of the lipid mediators has the greatest impact on muscle insulin action in vivo Our aim was to elucidate the effects of high-fat diet (HFD) and metformin (Met) on skeletal muscle bioactive lipid accumulation and insulin resistance (IR) in rats. We employed a [U-13C]palmitate isotope tracer and mass spectrometry to measure the content and fractional synthesis rate (FSR) of intramuscular long-chain acyl-CoA (LCACoA), diacylglycerols (DAG) and ceramide (Cer). Eight weeks of HFD-induced intramuscular accumulation of LCACoA, DAG and Cer accompanied by both systemic and skeletal muscle IR. Metformin treatment improved insulin sensitivity at both systemic and muscular level by the augmentation of Akt/PKB and AS160 phosphorylation and decreased the content of DAG and Cer and their respective FSR. Principal component analysis (PCA) of lipid variables revealed that altered skeletal muscle IR was associated with lipid species containing 18-carbon acyl-chain, especially with C18:0-Cer, C18:1-Cer, 18:0/18:2-DAG and 18:2/18:2-DAG, but not palmitate-derived lipids. It is concluded that the insulin-sensitizing action of metformin in skeletal muscle is associated with decreased 18-carbon acyl-chain-derived bioactive lipids.