1. Academic Validation
  2. TMAVA, a Metabolite of Intestinal Microbes, Is Increased in Plasma From Patients With Liver Steatosis, Inhibits γ-Butyrobetaine Hydroxylase, and Exacerbates Fatty Liver in Mice

TMAVA, a Metabolite of Intestinal Microbes, Is Increased in Plasma From Patients With Liver Steatosis, Inhibits γ-Butyrobetaine Hydroxylase, and Exacerbates Fatty Liver in Mice

  • Gastroenterology. 2020 Jun;158(8):2266-2281.e27. doi: 10.1053/j.gastro.2020.02.033.
Mingming Zhao 1 Lin Zhao 2 Xuelian Xiong 2 Yuan He 3 Wei Huang 4 Zihao Liu 5 Liang Ji 1 Bing Pan 1 Xuefeng Guo 5 Leibo Wang 6 Si Cheng 7 Ming Xu 8 Hongyuan Yang 9 Yuxin Yin 10 Minerva T Garcia-Barrio 11 Y Eugene Chen 11 Xiangbao Meng 6 Lemin Zheng 12
Affiliations

Affiliations

  • 1 The Institute of Cardiovascular Sciences and Institute of Systems Biomedicine, School of Basic Medical Sciences, Key Laboratory of Molecular Cardiovascular Sciences of Ministry of Education, NHC Key Laboratory of Cardiovascular Molecular Biology and Regulatory Peptides, Health Science Center, Peking University, Beijing, China.
  • 2 Department of Endocrinology and Metabolism, Fudan Institute of Metabolic Diseases, Zhongshan Hospital, Fudan University, Shanghai, China.
  • 3 National Research Institute for Health and Family Planning, Beijing, China.
  • 4 Gene Therapy Center and the Institute of Hypertension, Internal Medicine Department and Cardiovascular Division, Department of Internal Medicine, Tongji Hospital, Tongji Medical College Huazhong University of Science and Technology, Wuhan, China.
  • 5 Beijing National Laboratory for Molecular Sciences, State Key Laboratory for Structural Chemistry of Unstable and Stable Species, College of Chemistry and Molecular Engineering, Peking University, Beijing, China.
  • 6 State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing, China.
  • 7 China National Clinical Research Center for Neurological Diseases, Tiantan Hospital, Advanced Innovation Center for Human Brain Protection, The Capital Medical University, Beijing, China.
  • 8 Department of Cardiology and Institute of Vascular Medicine, Peking University Third Hospital, Beijing, China.
  • 9 School of Biotechnology and Biomolecular Sciences, The University of New South Wales, Sydney, New South Wales, Australia.
  • 10 The Institute of Systems Biomedicine, Peking University, Beijing, China.
  • 11 Cardiovascular Center, Department of Internal Medicine, University of Michigan Medical Center, Ann Arbor, Michigan.
  • 12 The Institute of Cardiovascular Sciences and Institute of Systems Biomedicine, School of Basic Medical Sciences, Key Laboratory of Molecular Cardiovascular Sciences of Ministry of Education, NHC Key Laboratory of Cardiovascular Molecular Biology and Regulatory Peptides, Health Science Center, Peking University, Beijing, China; China National Clinical Research Center for Neurological Diseases, Tiantan Hospital, Advanced Innovation Center for Human Brain Protection, The Capital Medical University, Beijing, China. Electronic address: zhengl@bjmu.edu.cn.
Abstract

Background & aims: Nonalcoholic fatty liver disease is characterized by excessive hepatic accumulation of triglycerides. We aimed to identify metabolites that differ in plasma of patients with liver steatosis vs healthy individuals (controls) and investigate the mechanisms by which these might contribute to fatty liver in mice.

Methods: We obtained blood samples from 15 patients with liver steatosis and 15 controls from a single center in China (discovery cohort). We performed untargeted liquid chromatography with mass spectrometry analysis of plasma to identify analytes associated with liver steatosis. We then performed targeted metabolomic analysis of blood samples from 2 independent cohorts of individuals who underwent annual health examinations in China (1157 subjects with or without diabetes and 767 subjects with or without liver steatosis; replication cohorts). We performed mass spectrometry analysis of plasma from C57BL/6J mice, germ-free, and mice given Antibiotics. C57BL/6J mice were given 0.325% (m/v) N,N,N-trimethyl-5-aminovaleric acid (TMAVA) in their drinking water and placed on a 45% high-fat diet (HFD) for 2 months. Plasma, liver tissues, and fecal samples were collected; fecal samples were analyzed by 16S ribosomal RNA gene Sequencing. C57BL/6J mice with CRISPR-mediated disruption of the gene encoding γ-butyrobetaine hydroxylase (BBOX-knockout mice) were also placed on a 45% HFD for 2 months. Hepatic fatty acid oxidation (FAO) in liver tissues was determined by measuring liberation of 3H2O from [3H] palmitic acid. Liver tissues were analyzed by electron microscopy, to view mitochondria, and proteomic analyses. We used surface plasmon resonance analysis to quantify the affinity of TMAVA for BBOX.

Results: Levels of TMAVA, believed to be a metabolite of intestinal microbes, were increased in plasma from subjects with liver steatosis compared with controls, in the discovery and replication cohorts. In 1 replication cohort, the odds ratio for fatty liver in subjects with increased liver plasma levels of TMAVA was 1.82 (95% confidence interval [CI], 1.14-2.90; P = .012). Plasma from mice given Antibiotics or germ-free mice had significant reductions in TMAVA compared with control mice. We found the intestinal bacteria Enterococcus faecalis and Pseudomonas aeruginosa to metabolize trimethyllysine to TMAVA; levels of trimethyllysine were significantly higher in plasma from patients with steatosis than controls. We found TMAVA to bind and inhibit BBOX, reducing synthesis of carnitine. Mice given TMAVA had alterations in their fecal microbiomes and reduced cold tolerance; their plasma and liver tissue had significant reductions in levels of carnitine and acyl-carnitine and their hepatocytes had reduced mitochondrial FAO compared with mice given only an HFD. Mice given TMAVA on an HFD developed liver steatosis, which was reduced by carnitine supplementation. BBOX-knockout mice had carnitine deficiency and decreased FAO, increasing uptake and liver accumulation of free fatty acids and exacerbating HFD-induced fatty liver.

Conclusions: Levels of TMAVA are increased in plasma from subjects with liver steatosis. In mice, intestinal microbes metabolize trimethyllysine to TMAVA, which reduces carnitine synthesis and FAO to promote steatosis.

Keywords

Gut; Microbiota; NASH; Peroxisome.

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