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Gut flora metabolism of phosphatidylcholine promotes cardiovascular disease

Abstract

Metabolomics studies hold promise for the discovery of pathways linked to disease processes. Cardiovascular disease (CVD) represents the leading cause of death and morbidity worldwide. Here we used a metabolomics approach to generate unbiased small-molecule metabolic profiles in plasma that predict risk for CVD. Three metabolites of the dietary lipid phosphatidylcholine—choline, trimethylamine N-oxide (TMAO) and betaine—were identified and then shown to predict risk for CVD in an independent large clinical cohort. Dietary supplementation of mice with choline, TMAO or betaine promoted upregulation of multiple macrophage scavenger receptors linked to atherosclerosis, and supplementation with choline or TMAO promoted atherosclerosis. Studies using germ-free mice confirmed a critical role for dietary choline and gut flora in TMAO production, augmented macrophage cholesterol accumulation and foam cell formation. Suppression of intestinal microflora in atherosclerosis-prone mice inhibited dietary-choline-enhanced atherosclerosis. Genetic variations controlling expression of flavin monooxygenases, an enzymatic source of TMAO, segregated with atherosclerosis in hyperlipidaemic mice. Discovery of a relationship between gut-flora-dependent metabolism of dietary phosphatidylcholine and CVD pathogenesis provides opportunities for the development of new diagnostic tests and therapeutic approaches for atherosclerotic heart disease.

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Figure 1: Strategy for metabolomics studies to identify plasma analytes associated with cardiovascular risk.
Figure 2: Identification of metabolites of dietary PC and an obligatory role for gut flora in generation of plasma analytes associated with CVD risks.
Figure 3: Plasma levels of choline, TMAO and betaine are associated with atherosclerosis risks in humans and promote atherosclerosis in mice.
Figure 4: Hepatic Fmo genes are linked to atherosclerosis and dietary PC metabolites enhance macrophage scavenger receptor expression.
Figure 5: Obligatory role of gut flora in dietary choline enhanced atherosclerosis.
Figure 6: Gut-flora-dependent metabolism of dietary PC and atherosclerosis.

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Acknowledgements

We wish to thank E. Sehayek for discussions, L. W. Castellani for help with lipoprotein profile analysis, and F. McNally, M. Berk and M. Pepoy for technical assistance. This research was supported by National Institutes of Health grants R01 HL103866, P01 HL098055, P01HL087018-020001, P01 HL28481 and P01 HL30568. B.J.B. was supported by NIH training grant T32-DK07789. The clinical study GeneBank was supported in part by P01 HL076491-055328, R01 HL103931 and the Cleveland Clinic Foundation General Clinical Research Center of the Cleveland Clinic/Case Western Reserve University CTSA (1UL1RR024989). Some of the laboratory studies (haemaglobin A1C, fasting glucose) in GeneBank were supported by R01 DK080732 and Abbott Diagnostics provided supplies for performance of some of the fasting lipid profile, glucose, creatinine, troponin I and hsCRP measured in GeneBank.

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Z.W. performed metabolomics analyses, and biochemical, cellular, animal model and mass spectrometry studies. He assisted with statistical analyses, and assisted in both drafting and critical review of the manuscript. E.K., B.D. and J.D.S. assisted with performance of animal models and their analyses. B.S.L. synthesized d9-DPPC and assisted in metabolomics/mass spectrometry analyses. B.J.B., H.A. and A.J.L. performed the mouse eQTL experiments and analyses, and assisted in both drafting and critical review of the manuscript. A.J.L. provided some funding for the study. R.K., E.B.B., X.F. and Y.-M.C. performed mass spectrometry analyses of clinical samples. Y.W. performed statistical analysis. A.E.F. and P.S. helped with collection of human liver biopsy material and interpretation of biochemical and pathological examination of animal liver for steatosis. W.H.W.T. assisted in GeneBank study design and enrolment, as well as analyses of clinical studies and critical review of the manuscript. J.A.D. assisted in clinical laboratory testing for human clinical studies, animal model experimental design, and critical review of the manuscript. S.L.H. conceived of the idea, designed experiments, assisted in data analyses, the drafting and critical review of the manuscript, and provided funding for the study.

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Correspondence to Stanley L. Hazen.

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Wang, Z., Klipfell, E., Bennett, B. et al. Gut flora metabolism of phosphatidylcholine promotes cardiovascular disease. Nature 472, 57–63 (2011). https://doi.org/10.1038/nature09922

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