Bofutsushosan improves gut barrier function with a bloom of Akkermansia muciniphila and improves glucose metabolism in mice with diet-induced obesity - PubMed (original) (raw)

. 2020 Mar 26;10(1):5544.

doi: 10.1038/s41598-020-62506-w.

Isao Usui 2 3, Allah Nawaz 2 4, Yoshiko Igarashi 2, Keisuke Okabe 2 5, Yukihiro Furusawa 6, Shiro Watanabe 7, Seiji Yamamoto 8, Masakiyo Sasahara 8, Yoshiyuki Watanabe 2, Yoshinori Nagai 9, Kunimasa Yagi 2, Takashi Nakagawa 4, Kazuyuki Tobe 10

Affiliations

Bofutsushosan improves gut barrier function with a bloom of Akkermansia muciniphila and improves glucose metabolism in mice with diet-induced obesity

Shiho Fujisaka et al. Sci Rep. 2020.

Abstract

Obesity and insulin resistance are associated with dysbiosis of the gut microbiota and impaired intestinal barrier function. Herein, we report that Bofutsushosan (BFT), a Japanese herbal medicine, Kampo, which has been clinically used for constipation in Asian countries, ameliorates glucose metabolism in mice with diet-induced obesity. A 16S rRNA sequence analysis of fecal samples showed that BFT dramatically increased the relative abundance of Verrucomicrobia, which was mainly associated with a bloom of Akkermansia muciniphila (AKK). BFT decreased the gut permeability as assessed by FITC-dextran gavage assay, associated with increased expression of tight-junction related protein, claudin-1, in the colon. The BFT treatment group also showed significant decreases of the plasma endotoxin level and expression of the hepatic lipopolysaccharide-binding protein. Antibiotic treatment abrogated the metabolic effects of BFT. Moreover, many of these changes could be reproduced when the cecal contents of BFT-treated donors were transferred to antibiotic-pretreated high fat diet-fed mice. These data demonstrate that BFT modifies the gut microbiota with an increase in AKK, which may contribute to improving gut barrier function and preventing metabolic endotoxemia, leading to attenuation of diet-induced inflammation and glucose intolerance. Understanding the interaction between a medicine and the gut microbiota may provide insights into new pharmacological targets to improve glucose metabolism.

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Conflict of interest statement

The authors declare no competing interests.

Figures

Figure 1

Figure 1

Bofutsushosan (BFT) improved glucose metabolism without affecting body weight. (A) Body weight of mice treated with either saline (white) or BFT (black) on an high fat-diet (HFD) (n = 8–9). Average food (B) and water (C) intake of mice between 11 and 14 weeks of age (n = 8–9). Weight of epididymal adipose tissue (eWAT) (D) inguinal adipose tissue (iWAT) (E), brown adipose tissue (BAT) (F) and cecum (G) of mice treated with either saline or BFT (18–19 weeks of age, n = 8–11). (H) Oral glucose tolerance test (OGTT) and (I) AUC of HFD-fed mice treated with saline (white) or BFT (black) for 5 weeks (n = 18). (J) Plasma insulin levels in 2-hour-fasted state (n = 9–11, 18 weeks of age). (K) Insulin tolerance test (ITT) of HFD-fed mice treated with saline (white) or BFT (black) for 8 weeks (n = 18–19). *p < 0.05, **p < 0.01 by unpaired, 2-tailed t test.

Figure 2

Figure 2

BFT modulates microbiota composition and increases Akkermansia muciniphila. (A) Migration rate of charcoal meal in the small intestine. (B) Principal component analysis (PCA) of fecal 16S rRNA sequencing data for mice treated with either saline or BFT on an HFD at 19 weeks of age (n = 4). (C) Representation of individual (left) and average (right) bacterial phyla in the fecal microbiota at 19 weeks of age (n = 4). (D) Fecal Akkermansia DNA levels normalized to eubacteria levels measured by qPCR (n = 4). (E) Relative fecal Akkermansia DNA levels normalized to eubacteria levels with or without BFT under antibiotic treatment (abx+) and withdrawal of antibiotics for 3 weeks (abx −) (n = 8). *p < 0.05, **p < 0.01 by unpaired, 2-tailed t test.

Figure 3

Figure 3

BFT increases tight-junction related protein in the colon and prevents from metabolic endotoxemia in HF-fed mice. (A) Relative short-chain fatty acid levels in the cecum of HFD (white) or HFD + BFT (black). (B) Fecal mucin content in mice treated either chow, HFD or HFD + BFT. (C) FITC levels in serum of mice after gavage administration of FITC-dextran. The Figure in the upper square is relative Akkermansia DNA levels of mice on a HFD or HFD + BFT used for the experiments (n = 7–8). (D) Western blots for claudin1 in the colon of mice on a chow or an HFD with or without BFT (18 weeks of age, 12 weeks on the HFD, 8 weeks on BFT) and quantitation of claudin1 protein normalized to β-actin (n = 3 per group). (E) Lipopolysaccharide (LPS) levels in serum of mice fed on an HFD with either saline or BFT for 12 weeks (n = 6–7). (F) Lbp mRNA levels in the liver (n = 7). (G) Lyz1 and F4/80 mRNA levels in the colon (n = 8–13). *p < 0.05 and **p < 0.01, by unpaired, 2-tailed t test for (A,C,E,F,G) or ANOVA, followed by Turkey-Kramer post-hoc for (B,D).

Figure 4

Figure 4

BFT decreases adipose tissue inflammation and improved steatohepatitis in HFD-fed mice. qPCR of macrophage and inflammation related genes in epididymal adipose tissues (A) and inguinal adipose tissue (B) of mice treated with either saline (white) or BFT (black) on an HFD for 8 weeks (n = 11–22). (C) Immunohistochemistry of epididymal adipose tissue for F4/80 and collagen I. (D) Quantification of crown like structures (CLS). (E) H&E staining and Sirius Red staining of the liver of chow, HFD or HFD + BFT treated mice. The arrows indicate Sirius Red positive area. (F) Liver weight of chow, HFD or HFD + BFT fed mice. *p < 0.05 and **p < 0.01, by unpaired, 2-tailed t test for (A,B), or ANOVA, followed by Turkey-Kramer post-hoc for (D,F).

Figure 5

Figure 5

Improvement of glucose metabolism with decreased adipose tissue inflammation by BFT– modified gut microbiota is transferable. (A) OGTT of HFD-fed mice treated with or without BFT before (left) (15 weeks old) and after (right) antibiotic treatment for 3 weeks. (B) OGTT of the HFD-fed recipient mice performed before (triangle) (17 weeks old) and after (circle) (26 weeks old) bacterial transfer from mice treated with saline (left) or BFT for 2 weeks (right) (n = 8). (C) ITT of the HFD-fed recipient mice performed before (triangle) and after (circle) bacterial transfer from mice treated with saline (left) or BFT (right) (n = 8). (D) Cecum weight of recipient mice. (E) qPCR for inflammatory gene expressions in the epididymal adipose tissue of recipient mice (n = 8). *p < 0.05 and **p < 0.01, by unpaired, 2-tailed t test.

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