Pistachio Consumption Alleviates Inflammation and Improves Gut Microbiota Composition in Mice Fed a High-Fat Diet

Abstract

High-fat diet (HFD) induces inflammation and microbial dysbiosis, which are components of the metabolic syndrome. Nutritional strategies can be a valid tool to prevent metabolic and inflammatory diseases. The aim of the present study was to evaluate if the chronic intake of pistachio prevents obesity-associated inflammation and dysbiosis in HFD-fed mice. Three groups of male mice (four weeks old; n = 8 per group) were fed for 16 weeks with a standard diet (STD), HFD, or HFD supplemented with pistachios (HFD-P; 180 g/kg of HFD). Serum, hepatic and adipose tissue inflammation markers were analyzed in HFD-P animals and compared to HFD and STD groups. Measures of inflammation, obesity, and intestinal integrity were assessed. Fecal samples were collected for gut microbiota analysis. Serum TNF-α and IL-1β levels were significantly reduced in HFD-P compared to HFD. Number and area of adipocytes, crown-like structure density, IL-1β, TNF-α, F4-80, and CCL-2 mRNA expression levels were significantly reduced in HFD-P subcutaneous and visceral adipose tissues, compared to HFD. A significant reduction in the number of inflammatory foci and IL-1β and CCL-2 gene expression was observed in the liver of HFD-P mice compared with HFD. Firmicutes/Bacteroidetes ratio was reduced in HFD-P mice in comparison to the HFD group. A pistachio diet significantly increased abundance of healthy bacteria genera such as Parabacteroides, Dorea, Allobaculum, Turicibacter, Lactobacillus, and Anaeroplasma, and greatly reduced bacteria associated with inflammation, such as Oscillospira, Desulfovibrio, Coprobacillus, and Bilophila. The intestinal conductance was lower in HFD-P mice than in the HFD mice, suggesting an improvement in the gut barrier function. The results of the present study showed that regular pistachio consumption improved inflammation in obese mice. The positive effects could be related to positive modulation of the microbiota composition.

Keywords: HFD mice; adipose tissue; gut microbiota; obesity-related inflammation; pistachio intake.

Figure 1

Effects of pistachio consumption on pro-inflammatory cytokines. Serum circulating levels of IL-1β (A) and TNF-α (B) in the lean, HFD, and HFD-P groups. Data are expressed as mean ± SEM; (n = 8/group). * p < 0.05 compared with lean; # p < 0.05 compared with HFD.

Figure 2

Effects of pistachio consumption on adipocyte morphology. (A) Adipocyte size distribution (%) and (B) adipocyte mean area (μm²) of the epididymal visceral adipose tissues (VAT) and subcutaneous adipose tissue (SAT) in lean, HFD, and HFD-P mice. (C) Adipose tissue staining (H&E staining, magnification 10×) in the lean, HFD, and HFD-P mice. Data are expressed as mean ± SEM; (n = 8/group). Compared to the lean mice (** p < 0.01; *** p < 0.001); Compared to the HFD mice (### p < 0.001).

Figure 3

Effects of pistachio consumption on Crown Like Structures CLS density. (A) Representative results of the density of MAC-2 positive CLS stained in epididymal visceral adipose tissue (VAT) and subcutaneous adipose tissue (SAT) of the three groups of animals (CLS number/10.000 adipocytes). (B) VAT and SAT immunohistochemistry (IHC) analysis for MAC-2 positive macrophages forming CLS (arrows) in the lean, HFD, and HFD-P animals (magnification 10×). (C) Effect of Pistachio consumption on IL-1β, TNF-α, F4-80, and CCL2 mRNA expression in VAT and SAT of the lean, HFD, and HFD-P mice. Data are expressed as mean ± SEM; (n = 8/group). * p < 0.05 compared to the lean mice (* p < 0.05; ** p < 0.01; *** p < 0.001); # p < 0.05 compared to the HFD mice (# p < 0.05; ## p < 0.01; ### p < 0.001).

Figure 4

Effect of pistachio consumption on liver inflammation. (A) Liver histology of the lean, HFD, and HFD-P mice was examined by H&E staining. Arrows indicate the points of inflammatory foci (magnification 10×). (B) Quantification of inflammatory foci per 5 random fields under 20× magnification. (C) mRNA levels of IL-1β, TNF-α, F4-80, and CCL2 in the livers of the lean, HFD, and HFD-P mice (B). Data are represented by the means ± SEM. (n = 8/group). * p < 0.05 compared to the lean mice (** p < 0.01; *** p < 0.001); # p < 0.05 compared to the HFD mice (# p < 0.05; ## p < 0.01).

Figure 5

16S rDNA sequencing of bacterial DNA in the feces of the lean, HFD, and HFD-P mice, in order to discriminate the intestinal microbial profile. (A) Graphic representation of the relative abundance (%) of the gut microbiota phyla composition of the three groups of animals. (B) Ratio of Firmicutes to Bacteroidetes in the lean, HFD, and HFD-P. Data are expressed as mean ± SEM; (n = 8/group). (*** p < 0.001); hash denotes significant difference when compared to the HFD group (# p < 0.05; ## p < 0.01; ### p < 0.001).

Figure 6

Genus level taxonomic distributions of the microbial communities in the feces of the lean, HFD, and HFD-P mice. (A) Genera abundance (%) was significantly modified by the pistachio intake. (B) Genera abundance (%) was not modified by the pistachio intake. Data are expressed as means ± SEM; (n = 8/group). (* p < 0.05; ** p < 0.01; *** p < 0.001); hash denotes significant difference compared to the HFD group (# p < 0.05; ## p < 0.01; ### p < 0.001).

Figure 7

Effect of pistachio consumption on the conductance of isolated duodenal sections from the lean, HFD, and HFD-P mice through the Ussing chambers technique. Data are expressed as mean ± SEM; (n = 8/group). (** p < 0.01); hash denotes significant difference compared with the HFD (## p < 0.01).