Ceramide analog C2-cer induces a loss in insulin sensitivity in muscle cells through the salvage/recycling pathway.

Ceramides have been shown to play a major role in the onset of skeletal muscle insulin resistance and therefore in the prevalence of type 2 diabetes. However, many of the studies involved in the discovery of deleterious ceramide actions used a nonphysiological, cell-permeable, short-chain ceramide analog, the C2-ceramide (C2-cer). In the present study, we determined how C2-cer promotes insulin resistance in muscle cells. We demonstrate that C2-cer enters the salvage/recycling pathway and becomes deacylated, yielding sphingosine, re-acylation of which depends on the availability of long chain fatty acids provided by the lipogenesis pathway in muscle cells. Importantly, we show these salvaged ceramides are actually responsible for the inhibition of insulin signaling induced by C2-cer. Interestingly, we also show that the exogenous and endogenous monounsaturated fatty acid oleate prevents C2-cer to be recycled into endogenous ceramide species in a diacylglycerol O-acyltransferase 1-dependent mechanism, which forces free fatty acid metabolism towards triacylglyceride production. Altogether, the study highlights for the first time that C2-cer induces a loss in insulin sensitivity through the salvage/recycling pathway in muscle cells. This study also validates C2-cer as a convenient tool to decipher mechanisms by which long-chain ceramides mediate insulin resistance in muscle cells and suggests that in addition to the de novo ceramide synthesis, recycling of ceramide could contribute to muscle insulin resistance observed in obesity and type 2 diabetes.

Impact of Smoking on Salivary Lipid Profile and Oxidative Stress in Young Adults: A Comparative Analysis between Traditional Cigarettes, E-Cigarettes, and Heat-Not-Burn Products.

BACKGROUND Smoking nicotine is considered to be one of the most harmful addictions, leading to the development of a number of health complications, including many pathologies in the oral cavity. The aim of this study was to examine the effect of smoking traditional cigarettes, e-cigarettes, and heat-not-burn products on profiles of salivary lipids and lipid peroxidation products in the unstimulated and stimulated saliva of healthy young adults with a smoking habit of up to 3 years. MATERIAL AND METHODS We enrolled 3 groups of 25 smoking patients each and a control group matched for age, gender, and oral status. In saliva collected from patients from the study groups and participants from the control group, the concentrations of sphingolipids: sphingosine, sphinganine, sphingosine-1-phosphate, ceramides, and salivary lipid peroxidation products - malondialdehyde (MDA) and 4-hydroxynonenal (HNE) - were measured. The normality of distribution was assessed using the Shapiro-Wilk test. For comparison of the results, one-way analysis of variance (ANOVA) followed by post hoc Tukey test was used. RESULTS We demonstrated that each type of smoking causes a decrease in the concentration of salivary lipids, and there was an increased concentration of salivary MDA and 4-HNE. CONCLUSIONS Smoking in the initial period of addiction leads to an increase in the concentration of lipid peroxidation products through increased oxidative stress, leading to disturbance of the lipid balance of the oral cavity (eg, due to damage to cell membranes).

Diverse Sphingolipid Profiles in Rectal and Colon Cancer.

Colorectal cancer is a heterogenous group of neoplasms showing a variety of clinical and pathological features depending on their anatomical location. Sphingolipids are involved in the formation and progression of cancers, and their changes are an important part of the abnormalities observed during carcinogenesis. Because the course of rectal and colonic cancer differs, the aim of the study was to assess whether the sphingolipid profile is also different in tumors of these two regions. Using a combination of ultra-high-performance liquid chromatography combined with triple quadrupole mass spectrometry, differences in the amounts of cellular sphingolipids were found in colorectal cancer. Sphingosine content was higher in rectal cancer than in adjacent healthy tissue, while the content of two ceramides (C18:0-Cer and C20:0-Cer) was lower. In colon cancer, a higher content of sphingosine, sphinganine, sphingosine-1-phosphate, and two ceramides (C14:0-Cer and C24:0-Cer) was found compared to healthy tissue, but there was no decrease in the amount of any of the assessed sphingolipids. In rectal cancer, the content of sphinganine and three ceramides (C16:0-Cer, C22:0-Cer, C24:0-Cer), as well as the entire pool of ceramides, was significantly lower compared to colon cancer. The S1P/Cer ratio in rectal cancer (S1P/C18:1-Cer, S1P/C20:0-Cer, S1P/C22:0-Cer, S1P/C24:1-Cer) and in colon cancer (S1P/C18:0-Cer, S1P/C18:1-Cer, S1P/C20:0-Cer) was higher than in adjacent healthy tissue and did not differ between the two sites (rectal cancer vs. colonic cancer). It seems that the development of colorectal cancer is accompanied by complex changes in the metabolism of sphingolipids, causing not only qualitative shifts in the ceramide pool of cancer tissue but also quantitative disturbances, depending on the location of the primary tumor.

The Effect of Silencing the Genes Responsible for the Level of Sphingosine-1-phosphate on the Apoptosis of Colon Cancer Cells.

Sphingosine-1-phosphate (S1P) and ceramides (Cer) are engaged in key events of signal transduction, but their involvement in the pathogenesis of colorectal cancer is not conclusive. The aim of our study was to investigate how the modulation of sphingolipid metabolism through the silencing of the genes involved in the formation (SPHK1) and degradation (SGPL1) of sphingosine-1-phosphate would affect the sphingolipid profile and apoptosis of HCT-116 human colorectal cancer cells. Silencing of SPHK1 expression decreased S1P content in HCT-116 cells, which was accompanied by an elevation in sphingosine, C18:0-Cer, and C18:1-Cer, increase in the expression and activation of Caspase-3 and -9, and augmentation of apoptosis. Interestingly, silencing of SGLP1 expression increased cellular content of both the S1P and Cer (C16:0-; C18:0-; C18:1-; C20:0-; and C22:0-Cer), yet inhibited activation of Caspase-3 and upregulated protein expression of Cathepsin-D. The above findings suggest that modulation of the S1P level and S1P/Cer ratio regulates both cellular apoptosis and CRC metastasis through Cathepsin-D modulation. The cellular ratio of S1P/Cer seems to be a crucial component of the above mechanism.

Adipose-Derived Exosomes as Possible Players in the Development of Insulin Resistance.

Adipose tissue (AT) is an endocrine organ involved in the management of energy metabolism via secretion of adipokines, hormones, and recently described secretory microvesicles, i.e., exosomes. Exosomes are rich in possible biologically active factors such as proteins, lipids, and RNA. The secretory function of adipose tissue is affected by pathological processes. One of the most important of these is obesity, which triggers adipose tissue inflammation and adversely affects the release of beneficial adipokines. Both processes may lead to further AT dysfunction, contributing to changes in whole-body metabolism and, subsequently, to insulin resistance. According to recent data, changes within the production, release, and content of exosomes produced by AT may be essential to understand the role of adipose tissue in the development of metabolic disorders. In this review, we summarize actual knowledge about the possible role of AT-derived exosomes in the development of insulin resistance, highlighting methodological challenges and potential gains resulting from exosome studies.

Role of sphingolipids in the pathogenesis of intrahepatic cholestasis.

BACKGROUND & AIMS: Intrahepatic cholestasis of pregnancy (ICP) is the most common pregnancy-related liver disorder that affects from 0.2% to 15.6% pregnant women. The disease is connected with increased risk of fetal morbidity and mortality, but is unfortunately detected quite late. The diagnosis of ICP is based on only one manifestation: pruritus which mainly affects soles and palms. METHODS: Twenty intrahepatic cholestasis of pregnancy (ICP) women and twenty healthy pregnant women (control group) took part in the study. In the study group, blood sampling for baseline measurements was performed on the first day of hospital stay - before the commencement of treatment with ursodeoxycholic acid (UDCA) - and repeated after 7 days of 900 mg UDCA per day. An additional blood sample was collected on the second day after childbirth. In the control group, blood samples were collected directly after hospital admission. We compared plasma sphingolipids in samples of the subjects from ICP and ICP + UDCA-treated groups as well as the ICP group after delivery with the healthy controls. RESULTS: Of all sphingolipids, the median values of C16-Cer and C18-Cer were significantly higher in the plasma of cholestasis patients not treated with UDCA as compared to the control. Following 7 days of UDCA treatment, a considerable decrease in C16-Cer, C18-Cer and the total concentration of bile acids was noted as compared to the baseline. CONCLUSION: It is known that sphingolipids serve as modulators of liver regeneration. We assume these substances could be potential markers for detecting early onsets of intrahepatic cholestasis of pregnancy.

The influence of patient's age on metabolic and bariatric results of laparoscopic sleeve gastrectomy in 2-year observation.

BACKGROUND: The incidence of obesity has been constantly growing and bariatric procedures are considered to be the most effective treatment solution for morbidly obese patients. The results of laparoscopic sleeve gastrectomy (LSG) may differ depending on patient's age, gender, preoperative body mass index (BMI) and physical activity. METHODS: The aim of this study was to evaluate age-related differences in the outcome of LSG in terms of weight loss parameters, lipid and carbohydrate profile. The retrospective analysis of 555 patients who had undergone LSG was performed to compare the metabolic outcomes of surgery in individuals < 45 and ≥ 45 years old. Evaluation of weight loss parameters along with selected laboratory data was performed to demonstrate the results of LSG in 2 years follow-up. RESULTS: Overall, 238 males and 317 females (43%/57%) with median age of 43 years and median preoperative BMI of 46.41 (42.06-51.02) kg/m2 were analyzed. Patients in both groups presented significant weight loss at 24 months after the surgery with comparable percentage of total weight loss (40.95% in < 45 years old group and 40.44% in ≥ 45 years old group). The percentage of excess weight loss (78.52% vs. 74.53%) and percentage of excess BMI loss (91.95% vs. 88.01%) were higher in patients < 45 years old. However, the differences were not statistically significant (p = 0.662, p = 0.788 respectively). Patients under 45 years old experienced faster decrease in fasting glucose level that was observed after only 3 months (109 mg/dl to 95 mg/dl in < 45 years old group vs. 103.5 mg/dl to 99.5 mg/dl in ≥ 45 years old group, p < 0.001). Both groups presented improvement of lipid parameters during the observation. However, patients < 45 years old achieved lower values of LDL at 3 and 12 months follow-up (115 mg/dl vs. 126 mg/dl, p = 0.010; 114.8 mg/dl vs. 122 mg/dl, p = 0.002). Younger group of patients also showed superior improvement of triglycerides level. CONCLUSIONS: LSG results in significant weight loss in all patients regardless age. In turn, superior and faster improvement in lipid and carbohydrate profile is achieved in patients under 45 years old.

GPAT Gene Silencing in Muscle Reduces Diacylglycerols Content and Improves Insulin Action in Diet-Induced Insulin Resistance.

Skeletal muscle is an important tissue responsible for glucose and lipid metabolism. High-fat diet (HFD) consumption is associated with the accumulation of bioactive lipids: long chain acyl-CoA, diacylglycerols (DAG) and ceramides. This leads to impaired insulin signaling in skeletal muscle. There is little data on the involvement of DAG in the development of these disorders. Therefore, to clarify this enigma, the gene encoding glycerol-3-phosphate acyltransferase enzyme (GPAT, responsible for DAG synthesis) was silenced through shRNA interference in the gastrocnemius muscle of animals with diet-induced insulin resistance. This work shows that HFD induces insulin resistance, which is accompanied by an increase in the concentration of plasma fatty acids and the level of bioactive lipids in muscle. The increase in these lipids inhibits the insulin pathway and reduces muscle glucose uptake. GPAT silencing through electroporation with shRNA plasmid leads to a reduction in DAG and triacylglycerol (TAG) content, an increase in the activity of the insulin pathway and glucose uptake without a significant effect on ceramide content. This work clearly shows that DAG accumulation has a significant effect on the induction of muscle insulin resistance and that inhibition of DAG synthesis through GPAT modulation may be a potential target in the treatment of insulin resistance.

The effect of high-fat diet and inhibition of ceramide production on insulin action in liver.

Liver, as one of the most important organs involved in lipids and glucose metabolism, is perceived as a key tissue for pharmacotherapy of insulin resistance (IRes) and type 2 diabetes. Ceramides (Cer) are biologically active lipids, which accumulation is associated with the induction of muscle IRes. We sought to determine the role of intrahepatic bioactive lipids production on insulin action in liver of insulin-resistant rats and after myriocin administration. The experiments were conducted on male Wistar rats divided into three groups: Control, fed high-fat diet (HFD), and fed HFD and treated with myriocin (HFD/Myr). Before sacrifice, the animals were infused with a [U-13 C]palmitate to calculate lipid synthesis rate by means of tracer incorporation technique in particular lipid groups. Liver Cer, diacylglycerols (DAG), acyl-carnitine concentration, and isotopic enrichment were analyzed by LC/MS/MS. Proteins involved in lipid metabolism and insulin pathway were analyzed by western blot analysis. An OGTT and ITT was also performed. HFD-induced IRes and increased both the synthesis rate and the content of DAG and Cer, which was accompanied by inhibition of an insulin pathway. Interestingly, myriocin treatment reduced synthesis rate not only of Cer but also DAG and improved insulin sensitivity. We conclude that the insulin-sensitizing action of myriocin in the liver is a result of the lack of inhibitory effect of lipids on the insulin pathway, due to the reduction of their synthesis rate. This is the first study showing how the synthesis rate of individual lipid groups in liver changes after myriocin administration.

The Relationship between the Concentration of Cathepsin A, D, and E and the Concentration of Copper and Zinc, and the Size of the Aneurysmal Enlargement in the Wall of the Abdominal Aortic Aneurysm.

BACKGROUND: Despite advances in diagnostics and treatment, aortic aneurysms are an important clinical problem, mainly due to the accompanying complications that may lead to direct loss of life, also the number of diagnosed and operated aneurysms is constantly increasing. The aim of this study is to determine the relationship between the concentration of lysosomal peptidases cathepsin A, D, and E in the wall of the abdominal aortic aneurysm and the concentration of copper and zinc, and the size of the aneurysm widening in the wall of the abdominal aortic aneurysm. METHODS: The study included 27 patients with abdominal aortic aneurysm from the Department of Vascular Surgery and Transplantation of the University Clinical Hospital in Bialystok. The research material was the wall of the abdominal aortic aneurysm collected intraoperatively. The control material consisted of fragments of the abdominal aorta obtained from organ donors for transplantation. The concentration of cathepsin A, D, and E was determined using enzyme-linked immunosorbent assays. Concentrations of copper and zinc were determined by flame atomic absorption spectrometry after prior mineralization of the samples. All patients were interviewed and asked about basic demographic data, comorbidities, and risk factors for cardiovascular disease to which they were exposed in the past. The statistical analysis was performed using Statistica 10 statistical package. Mann-Whitney U-tests were used and also Spearman's r correlation assuming a significance level of P < 0.05. RESULTS: The concentration of cathepsin A, D, and E was higher in the aortic wall altered by the aneurysm than in the wall of the control aorta (P < 0.05). The analysis of the data showed that there was a positive correlation between the concentration of cathepsin A and D and the width of the aneurysmal widening (r = 0.699 and 0.750, respectively). There was no correlation between cathepsin E concentration and aneurysm width. CONCLUSIONS: The higher contents of cathepsin A, D, and E in the wall of the aortic aneurysm than in the normal aortic wall, as well as a positive correlation between the concentration of cathepsin A and D and the width of the aneurysmal widening, allow to assume the participation of these enzymes in the pathogenesis of the aneurysm.

A Critical Review of Electroporation as A Plasmid Delivery System in Mouse Skeletal Muscle.

The gene delivery to skeletal muscles is a promising strategy for the treatment of both muscular disorders (by silencing or overexpression of specific gene) and systemic secretion of therapeutic proteins. The use of a physical method like electroporation with plate or needle electrodes facilitates long-lasting gene silencing in situ. It has been reported that electroporation enhances the expression of the naked DNA gene in the skeletal muscle up to 100 times and decreases the changeability of the intramuscular expression. Coelectransfer of reporter genes such as green fluorescent protein (GFP), luciferase or beta-galactosidase allows the observation of correctly performed silencing in the muscles. Appropriate selection of plasmid injection volume and concentration, as well as electrotransfer parameters, such as the voltage, the length and the number of electrical pulses do not cause long-term damage to myocytes. In this review, we summarized the electroporation methodology as well as the procedure of electrotransfer to the gastrocnemius, tibialis, soleus and foot muscles and compare their advantages and disadvantages.

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.

Effects of meal ingestion on intramyocellular ceramide concentrations and fractional de novo synthesis in humans.

We investigated the effects of meal ingestion on intramyofibrillar (IMF) and subsarcolemmal (SS) ceramide metabolism in volunteers ranging from lean to obese. Thirty-eight women and men underwent a steady-state meal ingestion protocol that included a 6.5-h infusion of [U-13C]palmitate and muscle biopsies 1.5 and 6.5 h after starting the tracer infusion. We measured IMF and SS sphingolipid concentrations and the contribution of plasma palmitate to intramyocellular C16:0 ceramide by use of LC-MS-MS. In response to meal ingestion SS C24 ceramide concentrations, but not C14-C20 concentrations, increased significantly. IMF ceramide concentrations did not change. The increases in SS C24 ceramides were negatively related to parameters of insulin resistance. The fractional contribution of plasma palmitate to intramyocellular C16:0 ceramides in both IMF and SS fractions was inversely related to overweight status (ß = -0.432, P = 0.0095 and ß = -0.443, P = 0.0058, respectively). These data indicate that meal ingestion has differing effects on SS ceramide subspecies and suggest that the fractional de novo synthesis of intramyocellular ceramide from plasma palmitate in the postprandial condition is reduced in those who are overweight.

Inhibition of Ceramide De Novo Synthesis Affects Adipocytokine Secretion and Improves Systemic and Adipose Tissue Insulin Sensitivity.

Ceramide accumulation in muscle and in liver is implicated in the induction of insulin resistance. Much less in known about the role of ceramide in adipose tissue. The aim of the present study was to elucidate the role of ceramide in adipose tissue and to clarify whether lipids participate in the regulation of adipocytokine secretion. The experiments were performed on male Wistar rats divided into three groups: 1. Control, 2. fed high fat diet (HFD), and 3. fed HFD and treated with myriocin. Ceramide (Cer) and diacylglycerol (DAG) content were analyzed by LC/MS/MS. Hormone sensitive lipase (HSL) phosphorylation was analyzed by Western Blot. Plasma adiponectin and tumor necrosis factor alpha (TNF-α) concentration were measured by enzyme-linked immunosorbent assay. An oral glucose tolerance test (OGTT) and insulin tolerance test (ITT) was also performed. In HFD group, total DAG and Cer content was elevated in both subcutaneous and visceral adipose tissue, which was accompanied by increased glucose, insulin, and HOMA-IR value. Myriocin treatment restored HOMA-IR as well as glucose and insulin concentration to control values. Moreover, myriocin decreased not only Cer, but also DAG levels in both fat depots. Furthermore, we observed a strong correlation between adiponectin (negative) and TNF-α (positive) and Cer in both fat tissues, which suggests that Cer is involved in the regulation of adipocytokine secretion.

Sustained Action of Ceramide on the Insulin Signaling Pathway in Muscle Cells: IMPLICATION OF THE DOUBLE-STRANDED RNA-ACTIVATED PROTEIN KINASE.

In vivo, ectopic accumulation of fatty acids in muscles leads to alterations in insulin signaling at both the IRS1 and Akt steps. However, in vitro treatments with saturated fatty acids or their derivative ceramide demonstrate an effect only at the Akt step. In this study, we adapted our experimental procedures to mimic the in vivo situation and show that the double-stranded RNA-dependent protein kinase (PKR) is involved in the long-term effects of saturated fatty acids on IRS1. C2C12 or human muscle cells were incubated with palmitate or directly with ceramide for short or long periods, and insulin signaling pathway activity was evaluated. PKR involvement was assessed through pharmacological and genetic studies. Short-term treatments of myotubes with palmitate, a ceramide precursor, or directly with ceramide induce an inhibition of Akt, whereas prolonged periods of treatment show an additive inhibition of insulin signaling through increased IRS1 serine 307 phosphorylation. PKR mRNA, protein, and phosphorylation are increased in insulin-resistant muscles. When PKR activity is reduced (siRNA or a pharmacological inhibitor), serine phosphorylation of IRS1 is reduced, and insulin-induced phosphorylation of Akt is improved. Finally, we show that JNK mediates ceramide-activated PKR inhibitory action on IRS1. Together, in the long term, our results show that ceramide acts at two distinct levels of the insulin signaling pathway (IRS1 and Akt). PKR, which is induced by both inflammation signals and ceramide, could play a major role in the development of insulin resistance in muscle cells.

Short Term Palmitate Supply Impairs Intestinal Insulin Signaling via Ceramide Production.

The worldwide prevalence of metabolic diseases is increasing, and there are global recommendations to limit consumption of certain nutrients, especially saturated lipids. Insulin resistance, a common trait occurring in obesity and type 2 diabetes, is associated with intestinal lipoprotein overproduction. However, the mechanisms by which the intestine develops insulin resistance in response to lipid overload remain unknown. Here, we show that insulin inhibits triglyceride secretion and intestinal microsomal triglyceride transfer protein expression in vivo in healthy mice force-fed monounsaturated fatty acid-rich olive oil but not in mice force-fed saturated fatty acid-rich palm oil. Moreover, when mouse intestine and human Caco-2/TC7 enterocytes were treated with the saturated fatty acid, palmitic acid, the insulin-signaling pathway was impaired. We show that palmitic acid or palm oil increases ceramide production in intestinal cells and that treatment with a ceramide analogue partially reproduces the effects of palmitic acid on insulin signaling. In Caco-2/TC7 enterocytes, ceramide effects on insulin-dependent AKT phosphorylation are mediated by protein kinase C but not by protein phosphatase 2A. Finally, inhibiting de novo ceramide synthesis improves the response of palmitic acid-treated Caco-2/TC7 enterocytes to insulin. These results demonstrate that a palmitic acid-ceramide pathway accounts for impaired intestinal insulin sensitivity, which occurs within several hours following initial lipid exposure.

Plasma and ovarian tissue sphingolipids profiling in patients with advanced ovarian cancer.

PURPOSE: The role of lipids in carcinogenesis through induction of abnormal cell lines in the human body is currently undisputable. Based on the literature, bioactive sphingolipids play an essential role in the development and progression of cancer and are involved in the metastatic process. The aim of this study was to determine the concentration of selected sphingolipids in patients with advanced ovarian cancer (AOC, FIGO III/IV, high grade ovarian cancer). METHODS: Seventy-four patients with ovarian cancer were enrolled. Plasma concentrations of C16-Cer, C18:1-Cer and C18-Cer were assessed by LC/MS/MS. The content of tissue sphingolipids was measured using a UHPLC/MS/MS. RESULTS: Plasma concentration of 3 ceramides: C16-Cer, C18:1-Cer and C18-Cer was significantly elevated in women with advanced ovarian cancer compared to control group (P=0.031; 0.022; 0.020; respectively). There were increases in concentration of 5 ceramides: C16-Cer, C18:1-Cer, C18-Cer, C24:1-Cer, C24-Cer (P=0.025; 0.049; 0.032; 0.005; 0.013, respectively) and S1P (P=0.004) in ovarian tissue of women with advanced ovarian cancer compared to healthy individuals. Importantly, significantly higher risk of ovarian cancer when the plasma concentration of C16-Cer>311.88ng/100µl (AUC: 0.76, P=0.0261); C18:1-Cer>4.75ng/100µl (AUC: 0.77, P=0.0160) and C18-Cer>100.76ng/100µl (AUC:0.77, P=0.0136) was noticed. CONCLUSIONS: Bioactive sphingolipids play an essential role in the development and progression of cancer and they also take part in the process of metastasizing. This study suggests that some sphingolipids can be used as potential biomarkers of advanced ovarian cancer and that they can play an important role in the pathogenesis of this disease.

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.

Plasma C16-Cer levels are increased in patients with preterm labor.

INTRODUCTION: The pathogenesis of preterm labor is fragmentarily explained. The most widely accepted theory points out to infection and inflammation as possible causes, which can be mediated by potentially different factors, including sphingolipid mediators. Sphingolipids are a class of lipids that have been shown as important mediators in various cell processes such as: proliferation, growth, apoptosis, stress response, necrosis and inflammation. The aim of the study was to assess plasma concentrations of selected sphingolipids in patients with preterm labor. MATERIAL AND METHODS: We used ultra-high performance liquid chromatography with triple mass spectrometry (UHPLC-ESI-MS/MS) to assess plasma concentrations of the 11 sphingolipids in patients presenting with symptoms of preterm labor (n=61) and threatened preterm labor (n=40). RESULTS: We observed a statistically significant increase (p-value<0.004) in plasma concentrations of C16-Cer in patients with preterm labor as compared to the control group. We also found C16-Cer to be the best predictor of preterm labor in the group of patients with symptoms occurring after 32 weeks of gestation. CONCLUSIONS: Our findings show a possible involvement of selected sphingolipids, especially C16-Cer, in the pathogenesis of preterm labor. Their role as predictors of preterm delivery needs to be validated in the future on larger group of patients.

A mitochondrial-targeted ubiquinone modulates muscle lipid profile and improves mitochondrial respiration in obesogenic diet-fed rats.

The prevalence of the metabolic syndrome components including abdominal obesity, dyslipidaemia and insulin resistance is increasing in both developed and developing countries. It is generally accepted that the development of these features is preceded by, or accompanied with, impaired mitochondrial function. The present study was designed to analyse the effects of a mitochondrial-targeted lipophilic ubiquinone (MitoQ) on muscle lipid profile modulation and mitochondrial function in obesogenic diet-fed rats. For this purpose, twenty-four young male Sprague-Dawley rats were divided into three groups and fed one of the following diets: (1) control, (2) high fat (HF) and (3) HF+MitoQ. After 8 weeks, mitochondrial function markers and lipid metabolism/profile modifications in skeletal muscle were measured. The HF diet was effective at inducing the major features of the metabolic syndrome--namely, obesity, hepatic enlargement and glucose intolerance. MitoQ intake prevented the increase in rat body weight, attenuated the increase in adipose tissue and liver weights and partially reversed glucose intolerance. At the muscle level, the HF diet induced moderate TAG accumulation associated with important modifications in the muscle phospholipid classes and in the fatty acid composition of total muscle lipid. These lipid modifications were accompanied with decrease in mitochondrial respiration. MitoQ intake corrected the lipid alterations and restored mitochondrial respiration. These results indicate that MitoQ protected obesogenic diet-fed rats from some features of the metabolic syndrome through its effects on muscle lipid metabolism and mitochondrial activity. These findings suggest that MitoQ is a promising candidate for future human trials in the metabolic syndrome prevention.