Nopal (Opuntia ficus indica) protects from metabolic endotoxemia by modifying gut microbiota in obese rats fed high fat/sucrose diet

Abstract

Current efforts are directed to reducing the gut dysbiosis and inflammation produced by obesity. The purpose of this study was to investigate whether consuming nopal, a vegetable rich in dietary fibre, vitamin C, and polyphenols can reduce the metabolic consequences of obesity by modifying the gut microbiota and preventing metabolic endotoxemia in rats fed a high fat and sucrose diet. With this aim, rats were fed a high fat diet with 5% sucrose in the drinking water (HFS) for 7 months and then were fed for 1 month with HFS + 5% nopal (HFS + N). The composition of gut microbiota was assessed by sequencing the 16S rRNA gene. Nopal modified gut microbiota and increased intestinal occludin-1 in the HFS + N group. This was associated with a decrease in metabolic endotoxemia, glucose insulinotropic peptide, glucose intolerance, lipogenesis, and metabolic inflexibility. These changes were accompanied by reduced hepatic steatosis and oxidative stress in adipose tissue and brain, and improved cognitive function, associated with an increase in B. fragilis. This study supports the use of nopal as a functional food and prebiotic for its ability to modify gut microbiota and to reduce metabolic endotoxemia and other obesity-related biochemical abnormalities.

Conflict of interest statement

The authors declare that they have no competing interests.

Figures

Figure 1

Effect of nopal on body weight gain, serum glucose and insulin concentrations. (A) Body weight gain of rats fed control diet (C) or high fat diet +5% sucrose in drinking water (HFS) during 7 months (pretreatment period). After this period, obese rats were switched to one of the experimental diets for 1 month (treatment period): HFS, HFS + N, HFS-C, and HFS-C + N. (B) Fasting serum glucose and (D) insulin after the pretreatment and (C), (E) treatment periods respectively. Values are shown as means ± SEM, n = 5 rats per group. Statistical analysis of the pretreatment period was assessed by student t-test. Comparisons among groups after the treatment period were analyzed by two-way ANOVA followed by Fisher’s post-hoc test. Different letters indicate significant differences among groups, a > b > c > d, P < 0.05. Comparisons between 2 groups were analyzed by t-student test.

Figure 2

Effect of nopal consumption on blood lipids. (A) Fasting serum triglycerides, (C) total cholesterol and (E) LDL cholesterol of rats fed control diet (C) or high fat diet +5% sucrose in drinking water (HFS) during 7 months (pretreatment period). After this period, obese rats were switched to one of the experimental diets for 1 month (treatment period): HFS, HFS + N, HFS-C, HFS-C + N; (B), (D and F). Values are shown as means ± SEM, n = 5. Statistical analysis of the pretreatment period was assessed by student t-test. Comparisons among groups after the treatment period were analyzed by two-way ANOVA followed by Fisher´s post-hoc test. Different letters indicate significant differences among groups, a > b > c > d, P < 0.05. Comparisons between 2 groups were analyzed by student t test.

Figure 3

Effect of nopal on Energy expenditure and energy intake. (A) O2 consumption, (C) respiratory exchange ratio of Wistar rats fed a control diet or a high fat diet +5% sucrose in drinking water. water (HFS) during 7 months (pretreatment period). After this period, obese rats were switched to one of the experimental diets for 1 month (treatment period): HFS, HFS + N, HFS-C, HFS-C + N; (B), (D). Food intake, water consumption and energy intake in all the experimental groups. Values are shown as means ± SEM, n = 5 rats per group. Comparisons among groups were analyzed by two-way ANOVA. Comparisons between 2 groups were analyzed by student t test.

Figure 4

Nopal consumption modifies the intestinal microbiota. (A) Alpha diversity and rarefaction curves. (B) Principal components analysis of the (C), HFS, HFS + N, HFS-C + N and HFS-C groups, n = 5 per group, (C) Relative abundance of Firmicutes and Bacteriodetes, (D) relative abundance of gut microbiota at the genus level, (E) and at species level and (F) heatmap of the ten bacterial species with greatest differences among groups.

Figure 5

Effect of nopal consumption on histological morphology and expression of occludin-1 in the colon of Wistar rats fed different dietary treatments. (A–E) Hematoxilin-Eosin staining (HE), (F) Mucosal layer quantitative analysis. Values are shown as means ± SEM, n = 5 rats per group and (G–K). Immunohistochemical analysis of occludin-1 in the colon of rats fed control diet (C), high fat diet with 5% sucrose in their drinking water (HFS), obese rats consuming HFS + 5% nopal (HFS + N), and obese rats switched to C diet with (HFS-C + N) or without (HFS-C) 5% nopal at the end of the treatment period 400X magnification.

Figure 6

Nopal consumption decreases metabolic endotoxemia, prevents the hypersecretion of glucose insulinotropic peptide (GIP) and improves glucose tolerance. Fasting serum lipopolysaccharide (LPS) after the pretreatment (A) and treatments (B) periods. Serum GIP after the pretreatment (C) and treatments (D) periods. ipGTT and area under curve (AUC) after the pretreatment (E) and treatments (F) periods. Wistar rats were fed one of the following diets: control diet (C), high fat diet with 5% sucrose in their drinking water (HFS), HFS + 5% nopal (HFS + N), and obese rats switched to C diet with (HFS-C + N) or without (HFS-C) 5% nopal. Data are expressed as mean ± SEM (n = 5/group). Comparisons among groups were analysed by 2-way ANOVA followed by Fisher’s PLSD test. Different letters indicate significant differences among groups (a > b > c > d). P < 0.05. Comparisons between two groups were analysed by student t test.

Figure 7

Effect of nopal consumption on serum leptin and expression of genes of inflammation and oxidative stress in adipose tissue. Panel (A) and (B) show the concentration of fasting serum leptin after the pretreatment and treatment periods respectively. Relative mRNA abundance of (C) leptin, (D) Tnf-α, (E) NADPH oxidase (Nox) and (F) amyloid precursor protein (App) in adipose tissue of Wistar rats fed one of the following diets: control diet (C), high fat diet with 5% sucrose in drinking water (HFS), HFS + 5% nopal (HFS + N), and obese rats switched to C diet with (HFS-C + N) or without (HFS-C) 5% nopal. Data are expressed as mean ± SEM (n = 5/group). Comparisons among groups were analysed by 2-way ANOVA followed by Fisher’s PLSD test. Different letters indicate significant differences among groups (a > b > c > d), P < 0.05. Comparisons between two groups were analysed by student t-test.

Figure 8

Nopal consumption decreases oxidative stress and improves cognitive function. (A) Malondialdehyde (MDA) levels in the prefrontal cortex, (B) abeta1–40 peptide in the ventral CA1 region, (C) spontaneous alternations and latency time, (D) glial fibrillary acidic protein (GFAP) in the stratum oriens and the stratum radiatum, (E) spine density in dendrites of the CA1 region, and (F) morphology of dendritic spines in the CA1 region. Data are expressed as mean ± SEM (n = 5/group). Comparisons among groups were analysed by 2-way ANOVA followed by Fisher’s PLSD test. Different letters indicate significant differences among groups (a > b > c > d).

Figure 9

Nopal consumption decreases lipogenesis, hepatic steatosis and inflammation. Relative expression of (A) Srebp-1, (B) Acetyl-CoA carboxylase Acc, (C) Fas, (D) Ppar-α and (E) Cpt-1 in liver of Wistar rats fed one of the following diets: control diet (C), high fat diet with 5% sucrose in drinking water (HFS), HFS + 5% nopal (HFS + N), and obese rats switched to C diet with (HFS-C + N) or without (HFS-C) 5% nopal. (F–J) Hematoxilin-Eosin staining (HE) and (K–O) immunohistochemistry of TNF-α in liver of these rats. Data are expressed as mean ± SEM (n = 5/group). Comparisons among groups were analysed by 2-way ANOVA followed by Fisher’s PLSD test. Different letters indicate significant differences among groups (a > b > c > d). 400X magnification.

Figure 10

Consumption of nopal, a vegetable rich in soluble and insoluble fibres, polyphenols, vitamin C and with a low glycemic index, reduces gut dysbiosis and increases the abundance of occludin-1 improving intestinal permeability associated with an increase in B fragilis which in turn reduces paracellular transport of lipopolysaccharide (LPS) by the intestinal epithelium reducing metabolic endotoxemia. Nopal consumption also decreases the hypersecretion of glucose insulinotropic peptide (GIP). The metabolic consequences are a reduction in insulin resistance, lipogenesis, hepatic steatosis and cognitive damage. This figure was created using the software ChemBio Draw v. 13.0.2.3020 (www.cambridgesoft.com).