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Sphingolipid changes do not underlie fatty acid-evoked GLUT4 insulin resistance nor inflammation signals in muscle cells.

ABSTRACT: Ceramides contribute to obesity-linked insulin resistance and inflammation in vivo, but whether this is a cell-autonomous phenomenon is debated, particularly in muscle, which dictates whole-body glucose uptake. We comprehensively analyzed lipid species produced in response to fatty acids and examined the consequence to insulin resistance and pro-inflammatory pathways. L6 myotubes were incubated with BSA-adsorbed palmitate or palmitoleate in the presence of myriocin, fenretinide, or fumonisin B1. Lipid species were determined by lipidomic analysis. Insulin sensitivity was scored by Akt phosphorylation and glucose transporter 4 (GLUT4) translocation, while pro-inflammatory indices were estimated by IκBα degradation and cytokine expression. Palmitate, but not palmitoleate, had mild effects on Akt phosphorylation but significantly inhibited insulin-stimulated GLUT4 translocation and increased expression of pro-inflammatory cytokines Il6 and Ccl2 Ceramides, hexosylceramides, and sphingosine-1-phosphate significantly heightened by palmitate correlated negatively with insulin sensitivity and positively with pro-inflammatory indices. Inhibition of sphingolipid pathways led to marked changes in cellular lipids, but did not prevent palmitate-induced impairment of insulin-stimulated GLUT4 translocation, suggesting that palmitate-induced accumulation of deleterious lipids and insulin resistance are correlated but independent events in myotubes. We propose that muscle cell-endogenous ceramide production does not evoke insulin resistance and that deleterious effects of ceramides in vivo may arise through ancillary cell communication.

SUBMITTER: Pillon NJ

PROVIDER: S-EPMC6027910 | biostudies-literature | 2018 Jul

REPOSITORIES: biostudies-literature

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Sphingolipid changes do not underlie fatty acid-evoked GLUT4 insulin resistance nor inflammation signals in muscle cells.

Pillon Nicolas J NJ Frendo-Cumbo Scott S Jacobson Maya R MR Liu Zhi Z Milligan Paul L PL Hoang Bui Hai H Zierath Juleen R JR Bilan Philip J PJ Brozinick Joseph T JT Klip Amira A

Journal of lipid research 20180523 7

Ceramides contribute to obesity-linked insulin resistance and inflammation in vivo, but whether this is a cell-autonomous phenomenon is debated, particularly in muscle, which dictates whole-body glucose uptake. We comprehensively analyzed lipid species produced in response to fatty acids and examined the consequence to insulin resistance and pro-inflammatory pathways. L6 myotubes were incubated with BSA-adsorbed palmitate or palmitoleate in the presence of myriocin, fenretinide, or fumonisin B1. ...[more]

PMID: 29794037

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Project description:Insulin stimulates glucose transport in fat and skeletal muscle cells primarily by inducing the translocation of GLUT4 (glucose transporter isoform 4) to the PM (plasma membrane) from specialized GSVs (GLUT4 storage vesicles). Glycosphingolipids are components of membrane microdomains and are involved in insulin-regulated glucose transport. Cellular glycosphingolipids decrease during adipocyte differentiation and have been suggested to be involved in adipocyte function. In the present study, we investigated the role of glycosphingolipids in regulating GLUT4 translocation. We decreased glycosphingolipids in 3T3-L1 adipocytes using glycosphingolipid synthesis inhibitors and investigated the effects on GLUT4 translocation using immunocytochemistry, preparation of PM sheets, isolation of GSVs and FRAP (fluorescence recovery after photobleaching) of GLUT4-GFP (green fluorescent protein) in intracellular structures. Glycosphingolipids were located in endosomal vesicles in pre-adipocytes and redistributed to the PM with decreased expression at day 2 after initiation of differentiation. In fully differentiated adipocytes, depletion of glycosphingolipids dramatically accelerated insulin-stimulated GLUT4 translocation. Although insulin-induced phosphorylation of IRS (insulin receptor substrate) and Akt remained intact in glycosphingolipid-depleted cells, both in vitro budding of GLUT4 vesicles and FRAP of GLUT4-GFP on GSVs were stimulated. Glycosphingolipid depletion also enhanced the insulin-induced translocation of VAMP2 (vesicle-associated membrane protein 2), but not the transferrin receptor or cellubrevin, indicating that the effect of glycosphingolipids was specific to VAMP2-positive GSVs. Our results strongly suggest that decreasing glycosphingolipid levels promotes the formation of GSVs and, thus, GLUT4 translocation. These studies provide a mechanistic basis for recent studies showing that inhibition of glycosphingolipid synthesis improves glycaemic control and enhances insulin sensitivity in animal models of Type 2 diabetes.

Dataset Information

Sphingolipid changes do not underlie fatty acid-evoked GLUT4 insulin resistance nor inflammation signals in muscle cells.

Publications

Sphingolipid changes do not underlie fatty acid-evoked GLUT4 insulin resistance nor inflammation signals in muscle cells.

Similar Datasets

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