GLP-1 receptor activation inhibits VLDL production and reverses hepatic steatosis by decreasing hepatic lipogenesis in high-fat-fed APOE*3-Leiden mice

Abstract

Objective: In addition to improve glucose intolerance, recent studies suggest that glucagon-like peptide-1 (GLP-1) receptor agonism also decreases triglyceride (TG) levels. The aim of this study was to evaluate the effect of GLP-1 receptor agonism on very-low-density lipoprotein (VLDL)-TG production and liver TG metabolism.

Experimental approach: The GLP-1 peptide analogues CNTO3649 and exendin-4 were continuously administered subcutaneously to high fat diet-fed APOE*3-Leiden transgenic mice. After 4 weeks, hepatic VLDL production, lipid content, and expression profiles of selected genes involved in lipid metabolism were determined.

Results: CNTO3649 and exendin-4 reduced fasting plasma glucose (up to -30% and -28% respectively) and insulin (-43% and -65% respectively). In addition, these agents reduced VLDL-TG production (-36% and -54% respectively) and VLDL-apoB production (-36% and -43% respectively), indicating reduced production of VLDL particles rather than reduced lipidation of apoB. Moreover, they markedly decreased hepatic content of TG (-39% and -55% respectively), cholesterol (-30% and -55% respectively), and phospholipids (-23% and -36% respectively), accompanied by down-regulation of expression of genes involved in hepatic lipogenesis (Srebp-1c, Fasn, Dgat1) and apoB synthesis (Apob).

Conclusion: GLP-1 receptor agonism reduces VLDL production and hepatic steatosis in addition to an improvement of glycemic control. These data suggest that GLP-receptor agonists could reduce hepatic steatosis and ameliorate dyslipidemia in patients with type 2 diabetes mellitus.

Figure 1. GLP-1 receptor agonism reduces body weight and fasting plasma glucose and insulin levels.

APOE*3-Leiden (E3L) mice were fed a high fat diet (HFD) for 22 weeks. The last 4 weeks, mice were treated with either vehicle (HFD control), CNTO3649 (1.0 or 3.0 mg/kg/day) or exendin-4 (15 or 50 μg/kg/day). As a control for HFD feeding, an additional group of mice was included fed a chow diet that received vehicle (chow control). Blood was collected by tail bleeding after 7 h of fasting. Just before drug treatment (week 18) and after treatment (week 22), body weight (A), plasma glucose (B) and plasma insulin (C) levels were determined. Values are means ± SEM for at least 6 mice per group. *P<0.05, **P<0.01 compared to HFD controls.

Figure 2. GLP-1 receptor agonism reduces hepatic VLDL-TG and VLDL-apoB production without affecting VLDL particle composition.

E3L mice were fed a HFD for 22 weeks. The last 4 weeks, mice were treated with either vehicle (HFD control), CNTO3649 (1.0 or 3.0 mg/kg/day) or exendin-4 (15 or 50 μg/kg/day). As a control for HFD feeding, an additional group of mice fed a chow diet was included that received vehicle (chow control). After 7 h fasting, mice were injected with Tran³⁵S label (t = −30 min) and Triton WR-1339 (t = 0 min). Blood was drawn at the indicated time points and plasma TG concentrations were determined (A, B). VLDL-TG production rate was calculated as µmol/h from the slopes of the TG-time curves of the individual mice (C). At t = 120 min, mice were exsanguinated, and VLDL was isolated by density gradient ultracentrifugation. ³⁵S-activity was determined, and VLDL-apoB production rate was calculated as dpm.h⁻¹ (D). The VLDL-TG production rate to VLDL-apoB production rate ratio was calculated as nmol/dpm (E). The content of triglycerides, cholesterol, phospholipids and protein in VLDL was determined and calculated as % of total mass (F). Values are means ± SEM for at least 6 mice per group. *P<0.05, **P<0.01, ***P<0.001 compared to HFD controls. TG: triglycerides; TC: total cholesterol; PL: phospholipids; Pro: protein.

Figure 3. GLP-1 receptor agonism reverses high fat diet-induced hepatic steatosis.

E3L mice were fed HFD for 13 weeks. The last 4 weeks, mice were treated with either vehicle (HFD control), CNTO3649 (0.3 or 1.0 mg/kg/day) or exendin-4 (15 or 50 μg/kg/day). As a control for HFD feeding, an additional group of mice was included fed a chow diet that received vehicle (chow control). Livers were isolated from 7 h fasted mice, liver pieces were homogenized, and triglycerides, cholesterol and phospholipids were determined as nmol per mg protein. Values are means ± SEM for at least 6 mice per group. *P<0.05, **P<0.01, ***P<0.001 compared to HFD controls.

Figure 4. GLP-1 receptor agonism affects hepatic expression of genes involved in VLDL production, lipogenesis, and lipid homeostasis.

E3L mice were fed HFD for 13 weeks. The last 4 weeks, mice were treated with either vehicle (HFD control), CNTO3649 (0.3 or 1.0 mg/kg/day) or exendin-4 (15 or 50 μg/kg/day). As a control for HFD feeding, an additional group of mice was included fed a chow diet that received vehicle (chow control). Livers were isolated from 7 h fasted mice, and mRNA was extracted from liver pieces. mRNA values of indicated genes were normalized to Cyclo and Hprt mRNA levels. Data were calculated as fold difference as compared with the HFD control group. Values are means ± SEM for at least 6 mice per group. *P<0.05, **P<0.01, ***P<0.001 compared to HFD controls.