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.

Assessment of dietary-lifestyle patterns and adherence to the USDA recommendations in Lebanese pregnant women amid the economic crisis: Findings from a national representative cross-sectional study.

As Lebanon's economic crisis become uncontrollable, Lebanese pregnant women face malnutrition, with many having to skip meals and switch to resort to cheap and unhealthy alternatives altogether. The objectives of the study were to assess the dietary and lifestyle patterns of Lebanese pregnant women and to evaluate their diets compliance with the United States Department of Agriculture (USDA) pregnancy recommendations, before and during the Lebanese escalating economic crisis. A cross-sectional study was conducted between April 2021 and January 2022. A validated self-administrated questionnaire was administered during the first, second and third trimesters of pregnancy among 363 women in all Lebanese governorates. Most of the pregnant women were free of diseases. While the majority did not smoke, 14.1% smoked hookah / shisha during pregnancy. The adherence to the USDA recommendations in our sample did not significantly vary prior to and throughout the socioeconomic crisis, and it was generally low. Only the mean consumption of vegetables increased during the socioeconomic crisis (p<0.05). Regarding physical activity, while the proportion of active women slightly decreased during the socioeconomic crisis, around 55% were still active. In conclusion, higher attention should be given to the dietary habits and health of this critical population, through effective interventions that increase awareness and achieve measurable improvements.

Ceramide Transporter CERT Is Involved in Muscle Insulin Signaling Defects Under Lipotoxic Conditions.

One main mechanism of insulin resistance (IR), a key feature of type 2 diabetes, is the accumulation of saturated fatty acids (FAs) in the muscles of obese patients with type 2 diabetes. Understanding the mechanism that underlies lipid-induced IR is an important challenge. Saturated FAs are metabolized into lipid derivatives called ceramides, and their accumulation plays a central role in the development of muscle IR. Ceramides are produced in the endoplasmic reticulum (ER) and transported to the Golgi apparatus through a transporter called CERT, where they are converted into various sphingolipid species. We show that CERT protein expression is reduced in all IR models studied because of a caspase-dependent cleavage. Inhibiting CERT activity in vitro potentiates the deleterious action of lipotoxicity on insulin signaling, whereas overexpression of CERT in vitro or in vivo decreases muscle ceramide content and improves insulin signaling. In addition, inhibition of caspase activity prevents ceramide-induced insulin signaling defects in C2C12 muscle cells. Altogether, these results demonstrate the importance of physiological ER-to-Golgi ceramide traffic to preserve muscle cell insulin signaling and identify CERT as a major actor in this process.

Targeting sphingolipid metabolism in the treatment of obesity/type 2 diabetes.

INTRODUCTION: Obesity is a major factor that is linked to the development of type 2 diabetes (T2D). Excess circulating fatty acids (FAs), which characterize obesity, induce insulin resistance, steatosis, ß cells dysfunction and apoptosis. These deleterious effects have been defined as lipotoxicity. AREAS COVERED: FAs are metabolized to different lipid species, including ceramides which play a crucial role in lipotoxicity. The action of ceramides on tissues, such as muscle, liver, adipose tissue and pancreatic ß cells, during the development of T2D will also be reviewed. In addition, the potential antagonist action of other sphingolipids, namely sphingoid base phosphates, on lipotoxicity in skeletal muscle and ß cells will be addressed. EXPERT OPINION: Ceramide is a critical mediator to the development of T2D linked to obesity. Targeting proteins involved in ceramide's deleterious action has not been possible due to their involvement in many other intracellular signaling pathways. A possible means of counteracting ceramide action would be to prevent the accumulation of the specific ceramide species involved in both insulin resistance and ß-cell apoptosis/dysfunction. Another possibility would be to adjust the dynamic balance between ceramide and sphingoid base phosphate, both known to display opposing properties on the development of T2D-linked obesity.

Characterising the inhibitory actions of ceramide upon insulin signaling in different skeletal muscle cell models: a mechanistic insight.

Ceramides are known to promote insulin resistance in a number of metabolically important tissues including skeletal muscle, the predominant site of insulin-stimulated glucose disposal. Depending on cell type, these lipid intermediates have been shown to inhibit protein kinase B (PKB/Akt), a key mediator of the metabolic actions of insulin, via two distinct pathways: one involving the action of atypical protein kinase C (aPKC) isoforms, and the second dependent on protein phosphatase-2A (PP2A). The main aim of this study was to explore the mechanisms by which ceramide inhibits PKB/Akt in three different skeletal muscle-derived cell culture models; rat L6 myotubes, mouse C2C12 myotubes and primary human skeletal muscle cells. Our findings indicate that the mechanism by which ceramide acts to repress PKB/Akt is related to the myocellular abundance of caveolin-enriched domains (CEM) present at the plasma membrane. Here, we show that ceramide-enriched-CEMs are markedly more abundant in L6 myotubes compared to C2C12 myotubes, consistent with their previously reported role in coordinating aPKC-directed repression of PKB/Akt in L6 muscle cells. In contrast, a PP2A-dependent pathway predominantly mediates ceramide-induced inhibition of PKB/Akt in C2C12 myotubes. In addition, we demonstrate for the first time that ceramide engages an aPKC-dependent pathway to suppress insulin-induced PKB/Akt activation in palmitate-treated cultured human muscle cells as well as in muscle cells from diabetic patients. Collectively, this work identifies key mechanistic differences, which may be linked to variations in plasma membrane composition, underlying the insulin-desensitising effects of ceramide in different skeletal muscle cell models that are extensively used in signal transduction and metabolic studies.