Dose-response effect of berberine on bile acid profile and gut microbiota in mice

Abstract

Background: Berberine (BBR) is a traditional antimicrobial herbal medicine. Recently, BBR has gained popularity as a supplement to lower blood lipids, cholesterol and glucose. Bile acids (BAs) are known to regulate blood levels of triglycerides, cholesterol, glucose and energy homeostasis, and gut flora play an important role in BA metabolism. However, whether BBR alters BAs metabolism or dose-response effect of BBR on gut flora is unknown.

Methods: In this study, the effects of various doses of BBR on the concentrations of BAs in liver and serum of male C57BL/6 mice were determined by UPLC-MS/MS, and the expression of BA-related genes, as well as the amount of 32 of the most abundant gut bacterial species in the terminal ileum and large intestine of male C57BL/6 mice were quantified by RT-PCR and Quantigene 2.0 Reagent System, respectively.

Results: Unconjugated BAs and total BAs were significantly altered by BBR in serum but not in liver. Increased primary BAs (βMCA, TβMCA and TUDCA) and decreased secondary BAs (DCA, LCA and the T-conjugates) were observed in livers and serum of mice fed BBR. The expression of BA-synthetic enzymes (Cyp7a1 and 8b1) and uptake transporter (Ntcp) increased 39-400 % in liver of mice fed the higher doses of BBR, whereas nuclear receptors and efflux transporters were not markedly altered. In addition, Bacteroides were enriched in the terminal ileum and large bowel of mice treated with BBR.

Conclusion: The present study indicated that various doses of BBR have effects on BA metabolism and related genes as well as intestinal flora, which provides insight into many pathways of BBR effects.

Keywords: Berberine; Bile acids; Gut Microbiota; Mice.

Fig. 1

Schematic diagram of BAs EHC. This is the recirculation of BAs between the intestine and the liver. BAs are synthesized in the hepatocyte (from cholesterol) by Cyp7a1, 27a1, 7b1 and 8b1, released into bile, transformed by the gut microbiota, reabsorbed in the small intestine, and returned to the liver to be recycled. The major genes involved in the EHC are shown

Fig. 2

Concentrations of T-conjugated BAs, unconjugated BAs and total BAs in control and BBR-treated mice. T-conjugated, unconjugated BAs and total BAs in liver and serum were quantified and the amounts are showed in a and b, respectively. All values are expressed as mean ± S.E.M. ANOVA followed by Duncan’s post hoc test were used, and * means P < 0.05 when compared with controls

Fig. 3

Concentrations of individual BAs in livers of control and BBR-treated mice. Twenty individual BAs were quantified in livers of 8-week-old control and BBR-treated mice. a showed the amounts of 10 primary BAs, and b showed the amount of 10 secondary BAs. All values are expressed as mean ± S.E.M. ANOVA followed by Duncan’s post hoc test were used, and * means P < 0.05 when compared with controls

Fig. 4

Concentrations of individual BAs in serum of control and BBR-treated mice. Eight primary (a) and four secondary BAs (b) were detected in serum of control and BBR-treated mice. All values are expressed as mean ± S.E.M. ANOVA followed by Duncan’s post hoc test were used, and * means P < 0.05 when compared with controls

Fig. 5

mRNA expression of major BA-related genes in livers of control and BBR-treated mice were examined (Fig. 5). BA synthetic enzymes involved in classical and alternative pathway were quantified in liver of BBR-treated mice (a). The mRNA expression of major BAs regulating factors in liver including FXR, SHP and Fgfr4 is demonstrated in (b). c illustrates the mRNA expression of major uptake transporters on the basolateral membrane of hepatocyte, which include Ntcp, Oatp1b2. The mRNA expression of major efflux transporters on the basolateral and canalicular membrane of hepatocyte are shown in (d). The left panel in d demonstrates the alteration of major efflux transporters on the canalicular membrane. The right panel of d shows the change of major efflux transporters on the basolateral membrane transporting the bile acids back to the blood after various doses of BBR. All values are expressed as mean ± S.E.M. ANOVA followed by Duncan’s post hoc test were used, and * means P < 0.05 when compared with controls

Fig. 6

mRNA expression of major regulating factors and transporters including FXR, Fgf15, Ostα, Ostβ and Asbt in ileum of control and BBR-treated mice were quantified with the 3 higher doses groups. All values are expressed as mean ± S.E.M. ANOVA followed by Duncan’s post hoc test were used, and * means P < 0.05 when compared with controls

Fig. 7

Bacteria in terminal ileum and large intestinal contents of control and BBR-treated mice. The relative amounts of 32 of the most abundant gut bacterial species in mice were quantified in the pooled samples of control, 100 and 300 mg/kg BBR treated groups (a), and validated by 7 bacteria in individual samples (b). All values are expressed as mean ± S.E.M. ANOVA followed by Duncan’s post hoc test were used, and * means P < 0.05 when compared with controls