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, 9 (1), 14150

Chitin-glucan and Pomegranate Polyphenols Improve Endothelial Dysfunction

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Chitin-glucan and Pomegranate Polyphenols Improve Endothelial Dysfunction

Audrey M Neyrinck et al. Sci Rep.

Abstract

The vascular dysfunction is the primary event in the occurrence of cardio-vascular risk, and no treatment exists until now. We tested for the first time the hypothesis that chitin-glucan (CG) - an insoluble fibre with prebiotic properties- and polyphenol-rich pomegranate peel extract (PPE) can improve endothelial and inflammatory disorders in a mouse model of cardiovascular disease (CVD), namely by modulating the gut microbiota. Male Apolipoprotein E knock-out (ApoE-/-) mice fed a high fat (HF) diet developed a significant endothelial dysfunction attested by atherosclerotic plaques and increasing abundance of caveolin-1 in aorta. The supplementation with CG + PPE in the HF diet reduced inflammatory markers both in the liver and in the visceral adipose tissue together with a reduction of hepatic triglycerides. In addition, it increased the activating form of endothelial NO-synthase in mesenteric arteries and the heme-nitrosylated haemoglobin (Hb-NO) blood levels as compared with HF fed ApoE-/- mice, suggesting a higher capacity of mesenteric arteries to produce nitric oxide (NO). This study allows to pinpoint gut bacteria, namely Lactobacillus and Alistipes, that could be implicated in the management of endothelial and inflammatory dysfunctions associated with CVD, and to unravel the role of nutrition in the modulation of those bacteria.

Conflict of interest statement

The authors declare no competing interests.

Figures

Figure 1
Figure 1
Body weight evolution (a), body weight gain (b) and fat mass gain (c) of ApoE−/− mice fed a low fat diet (CT), a high fat (HF) diet or a HF diet supplemented with 5% chitin-glucan (CG) or a combination of 5% CG and 0.5% pomegranate peel extracts (CG + PPE) for 8 weeks. Data with different superscript letters are significantly different at p < 0.05 (one-way ANOVA); ns = non significant and *p < 0.05 (two-way ANOVA).
Figure 2
Figure 2
Lipid accumulation in the liver tissue. Hepatic content of lipids (a), cholesterol (b) and triglycerides (c) of ApoE−/− mice fed a low fat diet (CT), a high fat (HF) diet or a HF diet supplemented with 5% chitin-glucan (CG) or a combination of 5% CG and 0.5% pomegranate peel extracts (CG + PPE) for 8 weeks. Lipid fraction area (d) and mean size of lipid droplets (e) were automatically analysed by ImageJ from Oil red O staining of frozen section of the main lobe of liver, bar = 100 µm (f). Data with different superscript letters are significantly different at p < 0.05 (ANOVA).
Figure 3
Figure 3
Expression of inflammatory markers in the visceral adipose tissue (a) and the liver (b). ApoE−/− mice were fed a high fat (HF) diet or a HF diet supplemented with 5% chitin-glucan (CG) or a combination of 5% CG and 0.5% pomegranate peel extracts (CG + PPE) for 8 weeks. Data are expressed as the mean ± SEM. The dotted line depicts the relative values observed in ApoE−/− mice fed a low fat diet (set at 1). *p < 0.05 versus HF group (ANOVA).
Figure 4
Figure 4
Western blot analyses on aorta (a,b) and mesenteric arteries (c) with anti-caveolin-1 (a) or anti-phosphorylated endothelial NOS (eNOS)ser1177 (b,c). ApoE−/− mice were fed a high fat (HF) diet or a HF diet supplemented with 5% chitin-glucan (CG) or a combination of 5% CG and 0.5% pomegranate peel extracts (CG + PPE) for 8 weeks. Data are expressed as the mean ± SEM. Values are expressed relative to ApoE−/− mice fed a low fat diet (set at 1). *p < 0.05 versus HF group (ANOVA).
Figure 5
Figure 5
Endothelium-dependent relaxation of preconstricted mesenteric arteries and level of Hb-NO in venous blood. Endothelium-dependent relaxation was evaluated by cumulative addition of acetylcholine (Ach) on pre-contracted arteries with a high KCl-solution, in the absence (a) or the presence of nitric oxide synthase inhibitor Nω-Nitro-L-arginine methyl ester (b) or a cyclooxygenase inhibitor (c). Level of Hb-NO in venous blood (d). ApoE−/− mice were fed a low fat diet (CT), a high fat (HF) diet or a HF diet supplemented with 5% chitin-glucan (CG) or a combination of 5% CG and 0.5% pomegranate peel extracts (CG + PPE) for 8 weeks. Data with different superscript letters are significantly different at p < 0.05 (one-way ANOVA); ns = non significant and *p < 0.05 (two-way ANOVA).
Figure 6
Figure 6
Microbiota structure assessed by non-metric dimensional scaling (NMDS) and bacterial diversity. A four dimensional model, built on a species level dissimilarity matrix has been obtained with a stress value of 0.08 (a). Reciprocal Simpson alpha-diversity, Chao richness and Simpson evenness (b) assessed from amplicon sequencing data of microbial cecal content of ApoE−/−mice fed, a low fat diet (CT), a high fat (HF) diet or a HF diet supplemented with 5% chitin-glucan (CG) or a combination of 5% CG and 0.5% pomegranate peel extracts (CG + PPE) for 8 weeks (p > 0.05 ANOVA, Kruskal-Wallis test).
Figure 7
Figure 7
Cecal bacteria assessed by qPCR. Total bacteria (a), Bifidobacterium spp. (b), Akkermansia muciniphila (c), Roseburia spp. (d), Alistipes spp. (e), Lactobacillus spp. (f) in the cecal content of ApoE−/−mice fed, a low fat diet (CT), a high fat (HF) diet or a HF diet supplemented with 5% chitin-glucan (CG) or a combination of 5% CG and 0.5% pomegranate peel extracts (CG + PPE) for 8 weeks. Data are Whiskers plots with minimum and maximum. Data with different superscript letters are significantly different at p < 0.05 (ANOVA, Kruskal-Wallis test).

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