Comprehensive evaluation of the bactericidal activities of free bile acids in the large intestine of humans and rodents - PubMed (original) (raw)

Comprehensive evaluation of the bactericidal activities of free bile acids in the large intestine of humans and rodents

Masamichi Watanabe et al. J Lipid Res. 2017 Jun.

Abstract

In addition to functioning as detergents that aid digestion of dietary lipids in the intestine, some bile acids have been shown to exhibit antimicrobial activity. However, detailed information on the bactericidal activities of the diverse molecular species of bile acid in humans and rodents is largely unknown. Here, we investigated the toxicity of 14 typical human and rodent free bile acids (FBAs) by monitoring intracellular pH, membrane integrity, and viability of a human intestinal bacterium, Bifidobacterium breve Japan Collection of Microorganisms (JCM) 1192T, upon exposure to these FBAs. Of all FBAs evaluated, deoxycholic acid (DCA) and chenodeoxycholic acid displayed the highest toxicities. Nine FBAs common to humans and rodents demonstrated that α-hydroxy-type bile acids are more toxic than their oxo-derivatives and β-hydroxy-type epimers. In five rodent-specific FBAs, β-muricholic acid and hyodeoxycholic acid showed comparable toxicities at a level close to DCA. Similar trends were observed for the membrane-damaging effects and bactericidal activities to Blautia coccoides JCM 1395T and Bacteroides thetaiotaomicron DSM 2079T, commonly represented in the human and rodent gut microbiota. These findings will help us to determine the fundamental properties of FBAs and better understand the role of FBAs in the regulation of gut microbiota composition.

Keywords: bile acids and salts; bile acids and salts/metabolism; bile acids and salts/physical chemistry; deoxycholic acid; gut microbiota; hyodeoxycholic acid; obesity; β-muricholic acid.

Copyright © 2017 by the American Society for Biochemistry and Molecular Biology, Inc.

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Figures

Fig. 1.

Fig. 1.

Effects of FBAs on the intracellular pH of B. breve JCM 1192T. Internal pH measurements were conducted by fluorescence measurement of cFSE. Cells were energized by 10 mM glucose (indicated as “Glc”) in buffer 2 (pH 6.5) to generate the ΔpH. The respective FBAs were then added at the indicated final concentrations. The bile acids evaluated are listed in Table 1. Precipitation of bile acids was observed using 7-oxo-LCA and UDCA, which prevented complete dissipation of the ΔpH. Valinomycin (Val, 20 nM) and nigericin (Nig, 200 nM) were added to completely dissipate the residual ΔΨ and ΔpH, respectively. The data are representative of at least two experiments that yielded similar results.

Fig. 2.

Fig. 2.

Selected ion-recording (SIR) chromatograms for the FBA standard mixture solution obtained from UPLC/ESI-MS analysis. See Table 1 for the abbreviated names of the FBAs. The standard mixture solution contains 50 μM of each FBA.

Fig. 3.

Fig. 3.

Double logarithmic plots for the UPLC retention factors versus the concentrations of FBAs common to humans and rodents required for inducing loss of viability. See Table 1 for the abbreviated names of the FBAs. k′, UPLC retention factor.

Fig. 4.

Fig. 4.

Classification of human and rodent FBAs according to their bactericidal activities. See Table 1 for the abbreviated names of the FBAs. “Viability loss-inducing concentration (mM)” was defined as the FBA concentration that caused a reduction in viability to <20% of that of the untreated cells. The asterisk indicates that the bactericidal activities of UCA were not tested at concentrations above 6 mM (Table 2), but were predicted to be above 6 mM from their extremely high hydrophilicities (Table 1) and the decreasing internal pH profiles (Fig. 1).

Fig. 5.

Fig. 5.

Two models of the interaction between CA molecules and the phospholipid bilayer in the bacterial cell membrane. The closed ovals and open/closed circles represent the α-hydroxy groups and carboxyl group in a CA molecule, respectively. The open circle corresponds to the protonated form, while closed circles correspond to the dissociated form of the carboxyl group. The number of carbon atoms is indicated. See text for description. IN, inside of the cell; OUT, outside of the cell.

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