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.
复制标题
TMAVA 是一种肠道微生物的代谢产物，肝脏脂肪变性患者的血浆中含量增加，抑制 γ-丁甜菜碱羟化酶，并加剧小鼠的脂肪肝
DOI：
10.1053/j.gastro.2020.02.033
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发表时间：
2020-02
期刊：
影响因子：
29.4
通讯作者：
Lemin Zheng
中科院分区：
文献类型：
--
作者：
Mingming Zhao;Lin Zhao;Xuelian Xiong;Yuan He;Wei Huang;Zihao Liu;Liang Ji;Bing Pan;Xuefeng Guo;Leibo Wang;Si Cheng;Ming Xu;Hongyuan Yang;Yuxin Yin;Minerva T Garcia-Barrio;Y Eugene Chen;Xiangbao Meng;Lemin Zheng
背景与目标
非酒精性脂肪性肝病的特征在于甘油三酯在肝脏中的过度积累。我们的目的是鉴定脂肪肝患者与健康个体（对照）肝组织中不同的代谢物，并研究这些代谢物可能导致小鼠脂肪肝的机制。
方法
我们从中国的一个中心（发现队列）的15名肝脂肪变性患者和15名对照者中获得血液样本。我们对血浆进行了非靶向液相色谱-质谱分析，以鉴定与肝脂肪变性相关的分析物。然后，我们对来自2个独立队列的血液样本进行了靶向代谢组学分析，这些队列的个体在中国接受了年度健康检查（1157例有或无糖尿病的受试者和766例有或无肝脏脂肪变性的受试者;重复队列）。我们对来自C57 BL/6 J小鼠、无菌小鼠和给予抗生素的小鼠的血浆进行质谱分析。将CRISPR介导的编码γ-丁酰甜菜碱羟化酶的基因破坏的C57 BL/6 J小鼠（BBOX敲除小鼠）在其饮用水中给予0.325%（m/v）N，N，N-三甲基-5-氨基戊酸（TMAVA），并置于45%高脂肪饮食（HFD）2个月。采集血浆、肝组织和粪便样本;通过16 S rRNA基因测序分析粪便样本。通过测量[3 H]棕榈酸释放的3 H2O来确定肝组织中的肝脂肪酸氧化（FAO）。通过电子显微镜分析肝组织，观察线粒体和蛋白质组学分析。我们使用表面等离子体共振分析来定量TMAVA对BBOX的亲和力。
结果
在发现和复制队列中，与对照组相比，肝脏脂肪变性受试者血浆中TMAVA（被认为是肠道微生物的代谢产物）的水平升高。在1个重复队列中，肝脏TMAVA水平升高的受试者发生脂肪肝的比值比为1.82（95% CI，1.14-2.90; P= 0.012）。与对照组小鼠相比，给予抗生素的小鼠或无菌小鼠的血浆TMAVA显著降低。我们发现肠道细菌粪肠球菌和铜绿假单胞菌代谢三甲基赖氨酸TMAVA;三甲基赖氨酸水平显着高于血浆脂肪变性患者比对照组。我们发现TMAVA结合并抑制BBOX，减少肉毒碱的合成。给予TMAVA的小鼠的粪便微生物组发生改变，耐冷性降低;与仅给予HFD的小鼠相比，其血浆和肝组织的肉毒碱和酰基肉毒碱水平显著降低，其肝细胞的线粒体FAO减少。在HFD上给予TMAVA的小鼠发生肝脏脂肪变性，其通过补充肉毒碱而减少。BBOX基因敲除小鼠肉毒碱缺乏，减少FAO，增加摄取和肝脏积累的游离脂肪酸和加剧HFD诱导的脂肪肝。
结论
TMAVA的水平在患有肝脂肪变性的受试者的血浆中增加。在小鼠中，肠道微生物将三甲基赖氨酸代谢为TMAVA，其减少肉毒碱合成和FAO以促进脂肪变性。
BACKGROUND & AIMS
Non-alcoholic fatty liver disease is characterized by excessive hepatic accumulation of triglycerides. We aimed to identify metabolites that differ in liver tissues 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 766 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 with CRISPR-mediated disruption of the gene encoding γ-butyrobetaine hydroxylase (BBOX-knockout 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 rRNA gene sequencing. 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 levels of TMAVA was 1.82 (95% 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 on a HFD. Mice given TMAVA on a 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.
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