Novel role of Quercetin in ameliorating metabolic syndrome via VDR mediated activation of adiponectin/AdipoR2 signaling

Abstract

A sedentary lifestyle and physical inactivity leads to metabolic syndrome-associated comorbidities involving abdominal obesity, type 2 diabetes, hyperlipidaemia associated Cardiovascular Diseases (CVDs), and Metabolic dysfunction-associated fatty liver disease (MAFLD). In this study, we evaluated the novel hepato/cardio/adipo-protective role of Quercetin via Vitamin D Receptor, and elucidated its underlying mechanisms in reducing lipotoxicity, inflammation and fibrosis in high calorie diet induced metabolic syndrome. Male Swiss albino mice were fed with western diet and sugar water for multiple time intervals. Anti-lipotoxicity, anti-inflammatory, and anti-fibrotic effect of Quercetin was assessed by Oil Red O, H&E and TMS staining at different time points. The lipid profile, mRNA expression of inflammatory markers (TNF- α, IL-1β, IL-6 and MCP-1), fibrotic markers (α-SMA, COL1A1, COL1A2), adiponectin, AdipoR2, and VDR expression levels were measured from RNA pools of adipose, liver and heart tissues. Also, lipid-lowering and anti-steatohepatitic effects of Quercetin was assessed using mouse 3T3-L1 adipocytes, rat H9c2 cardiac cells, and human HepG2 hepatocytes. Our results indicate that, western diet fed mice with Quercetin ameliorated lipid profile and lipotoxicity. Histopathological examination and gene expression data revealed that Quercetin reduced hepatic and cardiac inflammation and fibrosis-associated markers. Interestingly, Quercetin treatment increased the serum levels of adiponectin and mRNA expressions of AdipoR2 and VDR. In-vitro experiments revealed the reduction in lipid accumulation of 3T3-L1 and fatty-acid-treated hepatic and cardiac cells following Quercetin treatment. These findings indicate that Quercetin exhibits a protective role on multiple organs through VDR activation and subsequent Adipo/AdipoR2 signaling in metabolic syndrome associated obesity, hepatic injury, and cardiac dysfunction.

Keywords: Adiponectin; Adiponectin receptor 2; Cardiac fibrosis; Metabolic associated steatohepatitis; Obesity; Quercetin; Vitamin D receptor.

Fig. 1

Effect of Quercetin on mice body weight and serum glucose level in WDSW diet induced in Swiss albino mice. (A) Outline representation of experimental design. (B) Photographs represent the changes in body weight of 20 week mice group. (C) Graph represent the mice body weight. (D) Insulin Tolerance Test (ITT). (E) Glucose Tolerance Test (GTT). Data are represented as the mean ± SEM from 5 mice in each group.

Fig. 2

Quercetin inhibited differentiation and lipid accumulation in the 3T3-L1 mature adipocytes. (A) Schematic representation of 3T3-L1 differentiation. (B) Oil Red O staining. Images were obtained at 20× magnification and scale bar: 100 μm). (C) Adipolysis assay. (D) Intracellular triglyceride content in 3T3-L1. Data are represented as the mean ± SEM from 5 mice in each group.

Fig. 3

Quercetin treatment decreased adipocyte size and alleviated hyperlipidaemia in WDSW diet induced Swiss albino mice. (A) Representative morphology of adipose tissue. (B) Adipose tissue weight. (C) Serum cholesterol. (D) Serum triglycerides. (E) Serum High density lipoproteins. (F) H&E staining of adipose tissue. Data are represented as the mean ± SEM from 5 mice in each group.

Fig. 4

Lipid accumulation suppressing and cytoprotective effect of Quercetin on fatty acid exposed HepG2 and H9C2 cell lines. (A–B) Oil Red O staining of steatosis induced by Oleic acid. Images were obtained at 20× magnification and scale bar: 100 μm). (C-D) Lipid content of steatosis induced by oleic acid. Data are represented as the mean ± SEM from 5 mice in each group.

Fig. 5

Quercetin improved hepatic injury in WDSW fed mice. (A) Morphology images of the liver. (B) Liver weight. (C) ALP. (D) AST. (E) ALT. (F) Oil Red O staining of liver tissue. (G) H&E staining, (H) TMS staining, (I–K) Relative mRNA expression of fibrosis target genes in liver tissue. For all images, magnification: 40× and Scale bar: 50 μm. Data are represented as the mean ± SEM from 5 mice in each group.

Fig. 6

Quercetin ameliorates cardiac inflammation and fibrosis in WDSW diet fed mice. (A) H& E staining, (B) TMS staining, Relative mRNA expression of inflammatory markers in heart tissue. (C) MCP-1. (D) IL-6. (E) TNF-α relative mRNA expression of fibrosis target genes in heart tissue. (F) COL1A1. (G) COLA2A1. (H) α-SMA. For all images, magnification: 40× and Scale bar: 50 μm. Data are represented as the mean ± SEM from 5 mice in each group.

Fig. 7

Quercetin intake suppresses NF-κB activity in adipose tissue of WDSW diet induced Swiss albino mice. (A) Adipokine array (B) Relative mRNA expression of inflammatory markers in adipose tissue. (G) IL-1β. (H) MCP-1. (I) TNF-α. (J) IL-6.

Fig. 8

Quercetin regulates MAFLD and cardiac lipotoxicity through Adiponectin/AdipoR2 signalling pathway activation. (A) Adiponectin level in 3T3-L1 condition media. (B) Serum adiponectin level. (C) Relative hepatic AdipoR2 mRNA expression in liver tissue. (E) Relative cardiac AdipoR2 mRNA expression in heart tissues. Relative mRNA expression of PPAR-α target genes. (E–F) HepG2, (G-H) H9c2, (I-J) Hepatic (K-L) Cardiac. Data are represented as the mean ± SEM from 5 mice in each group.

Fig. 9

Quercetin treatment leads to Adiponectin/AdipoR2 activation via VDR. (A) VDR expression in adipose tissue at 20 weeks. (B) VDR expression in liver tissue at 20 weeks. Relative mRNA expression of VDR gene. (C) Adipose tissue. (B) Liver. For all images, magnification: 40× and Scale bar: 50 μm. Data are represented as the mean ± SEM from 5 mice in each group.

Fig. 10

Schematic representation depicting the cross-talk mechanisms involved in western diet induced metabolic syndrome and therapeutic effect of Quercetin. Western diet induced obesity associated MAFLD and cardiac fibrosis finally causes metabolic syndrome. Quercetin treatment activates VDR in adipose tissue and aid in the inhibition of the activity of NF-κB. Quercetin treatment with WDSW diet up to 20 weeks ameliorated obesity associated MAFLD and cardiac fibrosis by inhibiting the lipid accumulation, inflammation and fibrosis through Adipo/AdipoR2 signalling via VDR activation. This provides experimental evidences that Quercetin has multifaceted potential as a therapeutic drug to treat metabolic syndrome.

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