Traditional Chinese Medicine Formula Jian Pi Tiao Gan Yin Reduces Obesity in Mice by Modulating the Gut Microbiota and Fecal Metabolism

Abstract

The current study employed the high-fat diet (HFD) induced murine model to assess the relationship between the effect of Jian Pi Tiao Gan Yin (JPTGY) and the alterations of gut microbiota and fecal metabolism. C57BL/6 mice were used to establish an animal model of obesity via HFD induce. Serum biochemical indicators of lipid metabolism were used to evaluate the pharmacodynamics of JPTGY in obese mice. Bacterial communities and metabolites in the feces specimens from the controls, the Group HFD, and the JPTGY-exposed corpulency group were studied by 16s rDNA genetic sequence in combination with liquid chromatography-mass spectrometry (LC-MS) based untargeted fecal metabolomics techniques. Results revealed that JPTGY significantly decreased the levels of total cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C), and elevated high-density lipoprotein cholesterol (HDL-C). Moreover, JPTGY could up-regulate the abundance and diversity of fecal microbiota, which was characterized by the higher phylum of proteobacteria. Consistently, at the genus levels, JPTGY supplementation induced enrichments in Lachnospiraceae NK4A136 group, Oscillibacter, Turicibacter, Clostridium sensu stricto 1, and Intestinimonas, which were intimately related to 14 pivotal fecal metabolins in respond to JPTGY therapy were determined. What is more, metabolomics further analyses show that the therapeutic effect of JPTGY for obesity involves linoleic acid (LA) metabolism paths, alpha-linolenic acid (ALA) metabolism paths, glycerophospholipid metabolism paths, arachidonic acid (AA) metabolism paths, and pyrimidine metabolism paths, which implied the potential mechanism of JPTGY in treating obesity. It was concluded that the linking of corpulency phenotypes with intestinal flora and fecal metabolins unveils the latent causal link of JPTGY in the treatment of hyperlipidemia and obesity.

Figure 1

Effects of High-fat diet and JPTGY on serum lipid profile. (a) TC; (b) TG; (c) LDL-C; and (d) HDL-C. Data were displayed as average ± SD (n = 10) and studied via one-way ANOVA ( ^∗p < 0.05,  ^∗ ∗p < 0.01 and  ^{∗ ∗ ∗}p < 0.001).

Figure 2

Rarefaction curve and Shannon index analysis of the bacterial community in different treatments. The curves of every group tend to be flat with the increase of extracted sequence number, revealing that the sequence identification data amount of samples is acceptable, and more data amount will merely generate a little novel OTU.

Figure 3

The bacterial alpha diversity index in each group. (a) richness; (b) Shannon diversity; (c) Chao1 index; and (d) Phylogenetic diversity. The letters denote if there are remarkable diversities amongst the three treatments at p < 0.05. The identical letters denote no remarkable diversities and diverse letters, remarkable diversities. (e) Venn diagrams display core and shared intestinal flora species of the mice in diverse groups.

Figure 4

Bacterial community composition of three different treatments. (a) The β-diversity displaying as non-metric multi-dimensional scaling (NMDS) at the OTU level. (b) Hierarchical clustering analysis of UPGMA samples based on binary Jaccard distance.

Figure 5

Phylum composition of each treatment and the Firmicute/Bacteroidetes ratio in the phylum level. (a) The relative richness of the top 10 ranked phyla was displayed. (b) The ratio of F/B. The letters denote if there are remarkable diversities amongst the three treatments at p < 0.05. The identical letters denote no remarkable diversities and diverse letters denote remarkable diversities. Bar charts reflect the mean value and standard deviation for each group.

Figure 6

Representative microbes in different groups. (a) LDA scoring was calculated for taxa with differential richness in the feces flora of the Group NC, HFD, and JPTGY. The LDA scoring revealed the effect size and ranking of every differentially abundant taxon (LDA >3). (b) Cladogram reveals the phylogenesis distribution of function markers in response to JPTGY therapy. The area of the circle is associated with taxon richness in a positive manner.

Figure 7

Prediction of gut microbiota functional pathway in mice taking JPTGY. The heatmap showed distinguishingly expressed potential functions.

Figure 8

Metabolomics analysis among three groups. (a) The PLS-DA score plot; (b) volcano map of “HFD vs. JPTGY”; (c) volcano map of “HFD vs. NC”; (d) volcano map of “JPTGY vs. NC”; and (e) Pearson correlative analyses between the intestinal flora phyla and varied feces metabolins. Positive and negative associations are presented as red and blue in the heatmap, separately. The metabolins in correspondence to the metabolism path are displayed on the right. (f) Metabolic pathway analysis of Group JPTGY.