Diabetes Metab J.  2023 Nov;47(6):771-783. 10.4093/dmj.2022.0265.

Effects Of Exercise Training And Chlorogenic Acid Supplementation On Hepatic Lipid Metabolism In Prediabetes Mice

• Shirkhani S
• Marandi SM
• Nasr-Esfahani MH
• Kim SK

• KMID: 2548154
• DOI: http://doi.org/10.4093/dmj.2022.0265

Abstract

Background
Since prediabetes is a risk factor for metabolic syndromes, it is important to promote a healthy lifestyle to prevent prediabetes. This study aimed to determine the effects of green coffee (GC), chlorogenic acid (CGA) intake, and exercise training (EX) on hepatic lipid metabolism in prediabetes male C57BL/6 mice.
Methods
Forty-nine mice were randomly divided into two groups feeding with a normal diet (n=7) or a high-fat diet (HFD, n=42) for 12 weeks. Then, HFD mice were further divided into six groups (n=7/group): control (pre-D), GC, CGA, EX, GC+EX, and CGA+EX. After additional 10 weeks under the same diet, plasma, and liver samples were obtained.
Results
HFD-induced prediabetes conditions with increases in body weight, glucose, insulin, insulin resistance, and lipid profiles were alleviated in all treatment groups. Acsl3, a candidate gene identified through an in silico approach, was lowered in the pre-D group, while treatments partly restored it. HFD induced adverse alterations of de novo lipogenesis- and β oxidation-associated molecules in the liver. However, GC and CGA supplementation and EX reversed or ameliorated these changes. In most cases, GC or CGA supplementation combined with EX has no synergistic effect and the GC group had similar results to the CGA group.
Conclusion
These findings suggest that regular exercise is an effective non-therapeutic approach for prediabetes, and CGA supplementation could be an alternative to partially mimic the beneficial effects of exercise on prediabetes.

Keyword

Chlorogenic acid; Coffee; Exercise; Lipogenesis; Liver; Pre-Diabetic state

Figure

• 

  Fig. 1. Treatments mitigated high-fat diet (HFD)-induced prediabetic phenotypes. At the first stage of the study, mice were treated with two different diets for the first 12 weeks: a normal diet (ND, n=7) and HFD (n=42). Then, in the second stage, HFD mice were further divided into six groups (n=7/group): no treatment (pre-D), treated with green coffee (GC), chlorogenic acid (CGA), exercise training (EX), GC+EX, and CGA+EX. Treatments were applied for 10 weeks under the same diet. At the end of the second stage (22nd week), (A) glucose tolerance test, (B) the area under the curve (AUC), (C) the homeostatic model assessment for insulin resistance (HOMA-IR) index, (D) plasma insulin, (E) total cholesterol (TC), (F) triglyceride (TG), (G) low-density lipoprotein (LDL), (H) high-density lipoprotein (HDL), (I) alanine transaminase (ALT), and (J) aspartate transaminase (AST) levels were assessed. aP<0.05 vs. ND, bP<0.01 vs. ND, cP<0.05 vs. pre-D, dP<0.01 vs. pre-D.

• 

  Fig. 2. Acyl-CoA synthetase long chain family member 3 (Acsl3) as a candidate gene is predicted to be differentially expressed with a high-fat diet (HFD), green coffee (GC), and/or exercise training (EX) in mouse livers and its responses to treatments. (A) Volcano plot showing differentially expressed genes with HFD compared to normal diet (ND) groups based on the data from GSE53131. (B) Volcano plot showing genes differentially expressed with GC intake in the setting of HFD based on the data from GSE53131. (C) Volcano plot showing differentially expressed genes with exercise compared to control groups based on the data from GSE104079. The red color represents the genes (|log fold change [FC]| >0.5 and false discovery rate [FDR] <0.5) significantly altered by each comparison. (D) The Venn diagram presents the distribution of significantly altered genes in three different comparisons. At the first stage of the study, mice were treated with two different diets for the first 12 weeks: a ND (n=7) and HFD (n=42). Then, in the second stage, HFD mice were further divided into six groups (n=7/group): no treatment (pre-D), treated with GC, chlorogenic acid (CGA), EX, GC+EX, and CGA+EX. Treatments were applied for 10 weeks under the same diet. At the end of the second stage (22nd week), Acsl3 gene (E) and protein (F) expression levels in the liver were measured using quantitative reverse transcription polymerase chain reaction and immunoblotting, respectively. 18s and β-actin were used to normalize gene and protein expression, respectively. aP<0.05 vs. ND, bP<0.01 vs. ND, cP<0.05 vs. pre-D, dP<0.01 vs. pre-D.

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  Fig. 3. Treatments attenuated high-fat diet (HFD)-induced increases in triglyceride production in the liver. At the first stage of the study, mice were treated with two different diets for the first 12 weeks: a normal diet (ND, n=7) and HFD (n=42). Then, in the second stage, HFD mice were further divided into six groups (n=7/group): no treatment (pre-D), treated with green coffee (GC), chlorogenic acid (CGA), exercise training (EX), GC+EX, and CGA+EX. Treatments were applied for 10 weeks under the same diet. At the end of the second stage (22nd week), (A) sterol regulatory element-binding protein-1c (Srebp-1c), (B) fatty acid synthase (Fasn), (C) acetyl-CoA carboxylase 1 (Acc1), and (E) glycerol-3-phosphate acyltransferase 1 (Gpat1) gene expression levels in the liver were measured using quantitative reverse transcription polymerase chain reaction. (D) ACC1 and (F) GPAT1 protein expression levels in the liver were assessed by immunoblotting. 18s and β-actin were used to normalize gene and protein expression, respectively. aP<0.05 vs. ND, bP<0.01 vs. ND, cP<0.05 vs. pre-D, dP<0.01 vs. pre-D.

• 

  Fig. 4. Treatments mitigated high-fat diet (HFD)-induced decreases in β-oxidation in the liver. At the first stage of the study, mice were treated with two different diets for the first 12 weeks: a normal diet (ND, n=7) and HFD (n=42). Then, in the second stage, HFD mice were further divided into six groups (n=7/group): no treatment (pre-D), treated with green coffee (GC), chlorogenic acid (CGA), exercise training (EX), GC+EX, and CGA+EX. Treatments were applied for 10 weeks under the same diet. At the end of the second stage (22nd week), (A) peroxisome proliferator-activated receptor alpha (Ppar-α), (B) acetyl-CoA carboxylase 2 (Acc2), and (D) carnitine palmitoyl transferase I (Cpt1) gene expression levels in the liver were measured using quantitative reverse transcription polymerase chain reaction. (C) ACC2 and (E) CPT1 protein expression levels in the liver were assessed by immunoblotting. 18s and β-actin were used to normalize gene and protein expression, respectively. aP<0.05 vs. ND, bP<0.01 vs. ND, cP<0.05 vs. pre-D, dP<0.01 vs. pre-D.

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  Fig. 5. Graphical abstract. A prediabetic hepatic cell representing de novo lipogenesis (DNL) (left) associated with a high-fat diet (HFD) and the preventive effects of treatments (right). HFD elevates plasma free fatty acids (FFAs) which enter hepatic cells and contribute to the activation of acyl-CoA synthetase (ACSL) known to convert fatty acids (FAs) into fatty acyl-CoAs. Acyl-CoAs then proceed with de novo lipogenesis (acetyl-CoA carboxylase 1 [ACC1]) and/or β-oxidation inhibition (ACC2). Consequently, with the involvement of fatty acid synthase (Fasn) and glycerol-3-phosphate acyltransferase 1 (GPAT1), triglyceride (TG) can be formed. In this study, lipogenic molecules were found to increase with HFD (red color) which can lead to an increase in lipid droplets inside the cell owing to increased TG production (left). However, exercise training (EX), green coffee (GC), and chlorogenic acid (CGA) treatments mitigated HFD-induced changes in the lipogenic molecules (green color). Thus, β-oxidation via carnitine palmitoyl transferase I (CPT1) can be increased, and FA levels and the lipid droplet size inside the hepatic cell can be decreased.

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