CerS1 but Not CerS5 Gene Silencing, Improves Insulin Sensitivity and Glucose Uptake in Skeletal Muscle

Skeletal muscle is perceived as a major tissue in glucose and lipid metabolism. High fat diet (HFD) lead to the accumulation of intramuscular lipids, including: long chain acyl-CoA, diacylglycerols, and ceramides. Ceramides are considered to be one of the most important lipid groups in the generation of skeletal muscle insulin resistance. So far, it has not been clearly established whether all ceramides adversely affect the functioning of the insulin pathway, or whether there are certain ceramide species that play a pivotal role in the induction of insulin resistance. Therefore, we designed a study in which the expression of CerS1 and CerS5 genes responsible for the synthesis of C18:0-Cer and C16:0-Cer, respectively, was locally silenced in the gastrocnemius muscle of HFD-fed mice through in vivo electroporation-mediated shRNA plasmids. Our study indicates that HFD feeding induced both, the systemic and skeletal muscle insulin resistance, which was accompanied by an increase in the intramuscular lipid levels, decreased activation of the insulin pathway and, consequently, a decrease in the skeletal muscle glucose uptake. CerS1 silencing leads to a reduction in C18:0-Cer content, with a subsequent increase in the activity of the insulin pathway, and an improvement in skeletal muscle glucose uptake. Such effects were not visible in case of CerS5 silencing, which indicates that the accumulation of C18:0-Cer plays a decisive role in the induction of skeletal muscle insulin resistance.

Feeding Rhythm-Induced Hypothalamic Agouti-Related Protein Elevation via Glucocorticoids Leads to Insulin Resistance in Skeletal Muscle

Circadian phase shifts in peripheral clocks induced by changes in feeding rhythm often result in insulin resistance. However, whether the hypothalamic control system for energy metabolism is involved in the feeding rhythm-related development of insulin resistance is unknown. Here, we show the physiological significance and mechanism of the involvement of the agouti-related protein (AgRP) in evening feeding-associated alterations in insulin sensitivity. Evening feeding during the active dark period increased hypothalamic AgRP expression and skeletal muscle insulin resistance in mice. Inhibiting AgRP expression by administering an antisense oligo or a glucocorticoid receptor antagonist mitigated these effects. AgRP-producing neuron-specific glucocorticoid receptor-knockout (AgRP-GR-KO) mice had normal skeletal muscle insulin sensitivity even under evening feeding schedules. Hepatic vagotomy enhanced AgRP expression in the hypothalamus even during ad-lib feeding in wild-type mice but not in AgRP-GR-KO mice. The findings of this study indicate that feeding in the late active period may affect hypothalamic AgRP expression via glucocorticoids and induce skeletal muscle insulin resistance.

Cell models for studying muscle insulin resistance

Type 2 diabetes is one of the most prevalent chronic diseases in the world today. Insulin resistance - a reduced responsiveness of tissues to insulin - is a hallmark of type 2 diabetes pathology. Skeletal muscle plays a pivotal role in glucose homeostasis - it is responsible for the majority of insulin-mediated glucose disposal and thus is one of the tissues most affected by insulin resistance. To study the molecular mechanisms of a disease, researchers often turn to cell models - they are inexpensive, easy to use, and exist in a controlled environment with few unknown variables. Cell models for exploring muscle insulin resistance are constructed using primary cell cultures or immortalised cell lines and treating them with fatty acids, high insulin or high glucose concentrations. The choice of cell culture, concentration and duration of the treatment and the methods for measuring insulin sensitivity, in order to confirm the model, are rarely discussed. Choosing an appropriate and physiologically relevant model for a particular topic of interest is required in order for the results to be reproducible, relevant, comparable and translatable to more complex biological systems. Cell models enable research that would otherwise be inaccessible but, especially when studying human disease, they do not serve a purpose if they are not in line with the biological reality. This review aims to summarise and critically evaluate the most commonly used cell models of muscle insulin resistance: the rationale for choosing these exact treatments and conditions, the protocols for constructing the models and the measurable outcomes used for confirming insulin resistance in the cells.