Carotid baroreceptor stimulation attenuates obesity-related hypertension through sympathetic-driven IL- 22 restoration of intestinal homeostasis

Figures

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  Fig. 1. CBS controls the body weight gain and lowers blood pressure in obesity-related hypertensive rats. a Schematic representation of the study design. SD rats were fed with an HFHFD, or ND and sacrificed at 13 weeks. Fecal contents were collected at 0, 4 and 12 weeks. The CBS was implanted at 5 weeks. b Representative images of rat body size. c Comparison of visceral fat mass among the 4 groups. d Body weight change in rats. e Systolic blood pressure. f Diastolic blood pressure. g Mean arterial pressure. C-sham group, n = 7; C-CBS group, n = 6; OH-sham group, n = 7; OH-CBS group, n = 7. Results are expressed as mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001 versus OH-Sham; #P < 0.05, ##P < 0.01, ###P < 0.001 versus C-sham. FC: Fecal Collection; CBS: Carotid Baroreceptor Stimulation; HFHFD: High-Fat, High-Fructose Diet; ND: Normal Diet

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  Fig. 2. Effects of HFHFD on gut microbial diversity and composition. a, b α-Diversity Chao1 index and Shannon index (n = 8–14). c Principal coordinate analysis (PCoA) score plot based on unweighted UniFrac metrics. d Relative abundances of the top 20 dominant phyla. e Relative abundances of the top 20 dominant genera. f Heatmap of the relative abundance of the top 50 dominant genera for each sample. g Cladogram generated from LEfSe analysis

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  Fig. 3. CBS affects the gut microbial diversity and composition of obesity-related hypertensive rats. a, b α-Diversity Chao1 index and Shannon index (n = 4–7). c Principal coordinate analysis (PCoA) score plot based on unweighted UniFrac metrics. d Relative abundances of the top 20 dominant phyla. e Relative abundances of the top 20 dominant genera. f Heatmap of the relative abundance of the top 50 dominant genera for each sample. g Cladogram generated from LEfSe analysis. h Bar graph of the linear discriminant analysis (LDA) score distribution showing genera with LDA scores greater than the set value of 3. i Heatmap of Spearman correlation analysis of the 30 microbes associated with obesity-related parameters and blood pressure. The Spearman rank correlation coefficient (r) ranged from − 1 (blue) to 1 (red). *P < 0.05, **P < 0.01, ***P < 0.001 A, B Statistical comparison: Kruskal–Wallis test

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  Fig. 4. CBS exerts beneficial effects on gut microbiota and related metabolites in obesity-related hypertensive rats. a Concentration of SCFAs in the serum of each group (n = 4–6). b Concentration of TMAO and its precursors in the serum of each group (n = 4–6). c Heatmap of the Spearman correlation analysis of the top 50 microbes associated with SCFAs and TMAO. The Spearman rank correlation coefficient (r) ranged from − 0.6 (blue) to 0.6 (red). d Relative abundance of genera between groups. *P < 0.05, **P < 0.01, versus OH-Sham; #P < 0.05, ##P < 0.01, versus C-sham. A, B, D statistical comparison: Kruskal–Wallis test

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  Fig. 5. CBS attenuates gut pathological alterations and protects intestinal barrier function in obesity-related hypertensive rats. a, e Representative Masson trichrome staining images of small intestinal tissue sections and quantification of the percent fibrotic area (scale bar = 50 µm, images were captured at X400 magnification, n = 6–7). b, f Representative H&E-stained images of small intestinal tissue and quantification of muscular thickness (scale bar = 50 µm, images were captured at X400 magnification, n = 6–7). c, g Representative H&E-stained images of small intestinal tissue and quantification of villus length (scale bar = 100 µm, images were captured at X200 magnification, n = 6–7). d, h Representative AB-PAS staining images of small intestinal tissue sections and quantification of the number of goblet cells per villus (scale bar = 100 µm, images were captured at X200 magnification, n = 6–7). i, j, k Immunoblot analysis of tight junction proteins Claudin- 1 and Occludin in small intestinal tissue tissues (n = 4). l Plasma concentrations of intestinal fatty acid-binding protein (I-FABP). Results are expressed as mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001

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  Fig. 6. CBS alleviates intestinal autonomic imbalance and reduces intestinal inflammation in obesity-related hypertensive rats. a Representative images of tyrosine hydroxylase immunofluorescence (red) and acetyltransferase immunofluorescence (green), scale bar = 50 µm, images were captured at X400 magnification, n = 6. b, c Quantitative analysis of the immunofluorescence results (n = 6, two visual fields were randomly selected from each section for statistical analysis). d Small intestinal norepinephrine (NE) concentration (n = 6–7). e–j Relative mRNA expression of IL- 22, IL- 17 A, IL- 5, TNF-α, IL- 6, and IL- 10 in the small intestine (n = 6). k, l Heatmap of the correlation network between inflammatory factor mRNA content and genus, TMAO, and SCFAs levels. Results are expressed as mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001

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  Fig. 7. CBS increases IL- 22 production by ILC3 s, improves intestinal mucus barrier, and reduces lipid absorption. a Logical plot of the flow cytometry gate (CD45+, CD3−, CD90+, RORγt+). b, c Ratio of ILC3 and IL- 22+ILC3 in the intestine was detected by flow cytometry (n = 5). d IL- 22 protein levels in small intestinal tissue (n = 5). e Representative images of MUC2 immunofluorescence (green), scale bar = 50 µm, images were captured at X400 magnification, n = 6. f–i Relative mRNA expression of MUC2, RegIIIγ, CD36, Fabp1 in small intestinal tissue (n = 6). Results are expressed as mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001