Apigenin Ameliorates Dyslipidemia, Hepatic Steatosis and Insulin Resistance by Modulating Metabolic and Transcriptional Profiles in the Liver of High-Fat Diet-Induced Obese Mice

Abstract

Several in vitro and in vivo studies have reported the anti-inflammatory, anti-diabetic and anti-obesity effects of the flavonoid apigenin. However, the long-term supplementary effects of low-dose apigenin on obesity are unclear. Therefore, we investigated the protective effects of apigenin against obesity and related metabolic disturbances by exploring the metabolic and transcriptional responses in high-fat diet (HFD)-induced obese mice. C57BL/6J mice were fed an HFD or apigenin (0.005%, w/w)-supplemented HFD for 16 weeks. In HFD-fed mice, apigenin lowered plasma levels of free fatty acid, total cholesterol, apolipoprotein B and hepatic dysfunction markers and ameliorated hepatic steatosis and hepatomegaly, without altering food intake and adiposity. These effects were partly attributed to upregulated expression of genes regulating fatty acid oxidation, tricarboxylic acid cycle, oxidative phosphorylation, electron transport chain and cholesterol homeostasis, downregulated expression of lipolytic and lipogenic genes and decreased activities of enzymes responsible for triglyceride and cholesterol ester synthesis in the liver. Moreover, apigenin lowered plasma levels of pro-inflammatory mediators and fasting blood glucose. The anti-hyperglycemic effect of apigenin appeared to be related to decreased insulin resistance, hyperinsulinemia and hepatic gluconeogenic enzymes activities. Thus, apigenin can ameliorate HFD-induced comorbidities via metabolic and transcriptional modulations in the liver.

Keywords: apigenin; hepatic metabolic and transcriptional responses; hepatic steatosis; high-fat diet-induced obesity; insulin resistance.

Figure 1

Effects of apigenin on (A) food intake; (B) energy intake; (C,D) body weight; (E) fat-pad mass and (F) plasma leptin level in C57BL/6J mice fed a high-fat diet. Data are shown as the means ± S.E. HFD: high-fat diet (20% fat, 1% cholesterol); API: HFD + 0.005% apigenin.

Figure 2

Effects of apigenin on (A) fasting blood glucose level; (B) plasma insulin level; (C) homeostatic index of insulin resistance (HOMO-IR); (D) hepatic glucose metabolism-related enzyme activities; (E) hepatic glycogen content and (F) plasma pro-inflammatory marker levels in C57BL/6J mice fed a high-fat diet. Data are shown as the means ± S.E. Values are significantly different between the high-fat diet and apigenin groups according to Student’s t-test: * p < 0.05; ** p < 0.01; *** p < 0.001. HFD: high-fat diet (20% fat, 1% cholesterol); API: HFD + 0.005% apigenin.

Figure 3

Effect of apigenin on plasma lipids and apolipoproteins in C57BL/6J mice fed a high-fat diet. Data are shown as the means ± S.E. Values are significantly different between the high-fat diet and apigenin groups according to Student’s t-test: * p < 0.05. HFD: high-fat diet (20% fat, 1% cholesterol); API: HFD + 0.005% apigenin.

Figure 4

Effect of apigenin on liver weight (A), hepatic morphology (B), plasma transaminases activities (C) and activities of hepatic enzymes controlling the synthesis of triglyceride and cholesterol ester (D) in C57BL/6J mice fed a high-fat diet; ((A), (C), and (D)) Data are shown as the means ± S.E. Values are significantly different between the high-fat diet and apigenin groups according to Student’s t-test: * p < 0.05, ** p < 0.05; (B) Original magnification ×200. Bar, 50 M. HFD: high-fat diet (20% fat, 1% cholesterol); API: HFD + 0.005% apigenin.

Figure 5

The top 10 most upregulated and downregulated genes in the livers of the apigenin group compared to the control group (A); functional gene ontologies associated with the apigenin-responsive genes (B); and real-time quantitative PCR validation (C); (A,B) comparison of differentially-expressed genes in the apigenin group vs. the control group using Benjamin–Hochberg adjusted p-value < 0.05, FDR (False Discovery Rate) <5%, fold change >1; (B) functional gene ontology terms enriched among apigenin responsive genes are clustered according to biological processes (enrichment score >1) using DAVID. The heatmap shows the expression profiles of the representative apigenin responsive genes in each cluster; (C) Data are shown as the means ± S.E. Values are significantly different between the high-fat diet and apigenin groups according to Student’s t-test: * p < 0.05; ** p < 0.05. Microarray data based on pooled RNA hybridized to Illumina MouseWG-6 v2.0 BeadChips. HFD: high-fat diet (20% fat, 1% cholesterol); API: HFD + 0.005% Apigenin; Lpl: lipoprotein lipase; Pparγ: peroxisome proliferator-activated receptor γ; Srebf1: sterol regulatory element-binding transcription factor 1; Dgat2: diacylglycerol O-acyltransferase 2; Scd1: stearoyl-CoA desaturase-1; Cidea: cell death activator; ES: Enrichment Score.

Figure 6

Schematic diagram showing the mechanisms underlying the beneficial effects of apigenin on obesity-related metabolic disturbances. Apigenin decreased the activities of hepatic enzymes controlling triglyceride synthesis and cholesterol esterification and increased the expression of hepatic genes involved in fatty acid oxidation, the TCA cycle, OXPHOS, the electron transport chain and cholesterol homeostasis while decreasing the expression of hepatic lipogenic and lipolytic genes, indicating that these changes may be potential mechanisms for improving dyslipidemia and hepatic steatosis in HFD-fed mice. Moreover, apigenin decreased plasma pro-inflammatory adipocytokines levels and hepatic gluconeogenic enzyme activities, which may be partly associated with the improved hyperglycemia, hyperinsulinemia and insulin resistance.