Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
, 59 (1), 143-53

Hepatic cannabinoid-1 Receptors Mediate Diet-Induced Insulin Resistance by Increasing De Novo Synthesis of Long-Chain Ceramides

Affiliations

Hepatic cannabinoid-1 Receptors Mediate Diet-Induced Insulin Resistance by Increasing De Novo Synthesis of Long-Chain Ceramides

Resat Cinar et al. Hepatology.

Abstract

Obesity is associated with increased activity of two lipid signaling systems (endocannabinoids [ECs] and ceramides), with both being implicated in insulin resistance. Cannabinoid-1 receptor (CB1 R) antagonists reverse obesity and insulin resistance, but have psychiatric side effects. Here we analyzed the role of ceramide in CB1 R-mediated insulin resistance in C57Bl6/J mice with high-fat diet-induced obesity (DIO), using JD5037, a peripherally restricted CB1 R inverse agonist. Chronic JD5037 treatment of DIO mice reduced body weight and steatosis and improved glucose tolerance and insulin sensitivity. Peripheral CB1 R blockade also attenuated the diet-induced increase in C14:0, C16:0, C18:0, and C20:0 ceramide species with either C16 or C18 sphingosine-base in the liver. Decreased ceramide levels reflected their reduced de novo synthesis, due to inhibition of the activity of serine-palmitoyl transferase (SPT) and the expression of its SPTLC3 catalytic subunit, as well as reduced ceramide synthase (CerS) activity related to reduced expression of CerS1 and CerS6. JD5037 treatment also increased ceramide degradation due to increased expression of ceramidases. In primary cultured mouse hepatocytes and HepG2 cells, the EC anandamide increased ceramide synthesis in an eIF2α-dependent manner, and inhibited insulin-induced akt phosphorylation by increased serine phosphorylation of IRS1 and increased expression of the serine/threonine phosphatase Phlpp1. These effects were abrogated by JD5037 or the SPT inhibitor myriocin. Chronic treatment of DIO mice with myriocin or JD5037 similarly reversed hepatic insulin resistance, as verified using a euglycemic/hyperinsulinemic clamp.

Conclusion: ECs induce CB1 R-mediated, endoplasmic reticulum stress-dependent synthesis of specific ceramide subspecies in the liver, which plays a key role in obesity-related hepatic insulin resistance.

Figures

Fig. 1
Fig. 1
HFD-induced elevation of the hepatic level of ceramides is reversed by peripheral blockade (A) or genetic deletion of CB1R (B). Levels of ceramides in liver were measured 22h after the last of 28 daily i.p. doses of JD5037 (3 mg/kg), myriocin (0.3 mg/kg) or vehicle in DIO mice and in age-matched lean controls (n=6/group)(A), or in the liver of CB1+/+, CB1−/− and hCB1−/− mice maintained on STD (solid columns) or HFD (open columns) for 16 weeks (n=6/group)(B). Animals were 22-week old at time of sacrifice. *P<0.05 relative to Vehicle-STD; #P<0.05 relative to Vehicle-HFD.
Fig. 2
Fig. 2
Effects of short-term CB1R blockade on hepatic ceramides and body weight in DIO (A) and db/db mice (B). In both groups, mice were treated for 7 days with JD5037, 3 mg/kg/day, n=6. *P<0.05 relative to Vehicle-STD; #P<0.05 relative to Vehicle-HFD (A) or vehicle-db/db (B).
Fig. 3
Fig. 3
Glucose homeostasis in DIO mice is similarly improved by blockade of CB1R or de novo ceramide synthesis, as tested by ipGTT and ipIST (A), fasting plasma glucose and insulin (B), insulin clamp (C), and 2-deoxy-glucose uptake by muscle and fat (D) and plasma levels of human insulin during clamp (E). Data represent mean ± SEM from 5–6 mice per group.
Fig. 4
Fig. 4
Peripheral CB1R antagonism reverses increased SPT activity and SPTLC3 gene expression in DIO mice. A: Enzyme activity of SPT; B: gene expression of SPT subunits; C: SPT activity in liver microsomes from lean mice in the absence or presence of myriocin or JD5037 in vitro; D: gene expression of ceramide synthase subtypes; E: ceramide synthase activity.
Fig. 5
Fig. 5
Peripheral CB1R antagonism promotes degradation of ceramides in DIO mice liver. Gene expression of sphingomyelinase isozymes (A); activity of total sphingomyelinase (B) and acid sphingomyelinase (C); gene expression of ceramidase subtypes and sphingosine kinase in liver (D); levels of hepatic sphingosine and sphingosine-1-phosphate (S1P) from vehicle- or JD5037-treated (3 mg/kg, i.p.) DIO (HFD) or lean control (STD) mice (n=6/group) (E).
Fig. 6
Fig. 6
HFD-induced robust increase in C16 sphingoid-base ceramides in liver microsomes is reversible by CB1R blockade. Levels of ceramide subspecies with C16 (d16:1) (A) or C18 sphingoid-base (d18:1) (B), and SPT activity in liver microsomes using myristoyl CoA as substrate (C).
Fig. 7
Fig. 7
CB1R-mediated increase in de novo ceramide synthesis and inhibition of insulin signaling are reversed by myriocin treatment and are dependent on ER stress. Ion chromatogram of [2H3]C16 dihydroceramide (d18:0/16:0) generated by mouse primary hepatocytes (3×106 cells/well) from [2H3]serine, in the absence or presence of appropriate combinations of anandamide (AEA), rimonabant (1 μM, SR), or myriocin (n=3/group) (A); preincubation of Hep G2 cells with myriocin or JD5037 for 1h occluded both AEA inhibition of insulin-induced akt phosphorylation (B) and anandamide-induced IRS1 ser307 phosphorylation and Phlpp1 expression (C). Each blot in B–C has been replicated 3 times with similar results. (D) siRNA- or shRNA-mediated knockdown of eIF2α proportionally reduces or eliminates AEA-induced ceramide synthesis.

Similar articles

See all similar articles

Cited by 43 articles

See all "Cited by" articles

Publication types

MeSH terms

Feedback