Aorta- and liver-generated TMAO enhances trained immunity for increased inflammation via ER stress/mitochondrial ROS/glycolysis pathways
Fatma Saaoud, … , Hong Wang, Xiaofeng Yang
Fatma Saaoud, … , Hong Wang, Xiaofeng Yang
Published November 17, 2022
Citation Information: JCI Insight. 2023;8(1):e158183. https://doi.org/10.1172/jci.insight.158183.
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Research Article Immunology Inflammation

Aorta- and liver-generated TMAO enhances trained immunity for increased inflammation via ER stress/mitochondrial ROS/glycolysis pathways

Abstract

We determined whether gut microbiota-produced trimethylamine (TMA) is oxidized into trimethylamine N-oxide (TMAO) in nonliver tissues and whether TMAO promotes inflammation via trained immunity (TI). We found that endoplasmic reticulum (ER) stress genes were coupregulated with MitoCarta genes in chronic kidney diseases (CKD); TMAO upregulated 190 genes in human aortic endothelial cells (HAECs); TMAO synthesis enzyme flavin-containing monooxygenase 3 (FMO3) was expressed in human and mouse aortas; TMAO transdifferentiated HAECs into innate immune cells; TMAO phosphorylated 12 kinases in cytosol via its receptor PERK and CREB, and integrated with PERK pathways; and PERK inhibitors suppressed TMAO-induced ICAM-1. TMAO upregulated 3 mitochondrial genes, downregulated inflammation inhibitor DARS2, and induced mitoROS, and mitoTEMPO inhibited TMAO-induced ICAM-1. β-Glucan priming, followed by TMAO restimulation, upregulated TNF-α by inducing metabolic reprogramming, and glycolysis inhibitor suppressed TMAO-induced ICAM-1. Our results have provided potentially novel insights regarding TMAO roles in inducing EC activation and innate immune transdifferentiation and inducing metabolic reprogramming and TI for enhanced vascular inflammation, and they have provided new therapeutic targets for treating cardiovascular diseases (CVD), CKD-promoted CVD, inflammation, transplantation, aging, and cancer.

Authors

Fatma Saaoud, Lu Liu, Keman Xu, Ramon Cueto, Ying Shao, Yifan Lu, Yu Sun, Nathaniel W. Snyder, Sheng Wu, Ling Yang, Yan Zhou, David L. Williams, Chuanfu Li, Laisel Martinez, Roberto I. Vazquez-Padron, Huaqing Zhao, Xiaohua Jiang, Hong Wang, Xiaofeng Yang

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Figure 9

TMAO induced immune metabolic reprogramming — including increased acetyl-CoA, glycolysis, and proton efflux rates — and glycolysis inhibitor suppressed TMAO-induced ICAM-1 expression.

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TMAO induced immune metabolic reprogramming — including increased acetyl...
(A) TMAO upregulated 2 glucose metabolism/TCA cycle genes (OGDHL and OMA1). (B) OGDHL is a key step in the TCA cycle and glycolysis pathways. (C) TMAO increased acetyl-CoA generation. HAECs were treated with TMAO (600 μM) for 24 hours, and acetyl CoA production was detected using acetyl-CoA assay kit (n = 3) following the manufacturer’s instruction. (D) The principle and profile of Seahorse glycolytic rate assay. In the cells, energy is produced by 2 different pathways, including mitochondrial respiration and glycolysis. In the glycolysis pathway, glucose is converted into lactate, and the protons are extruded into the extracellular media and detected as extracellular acidification rate (ECAR). In addition, CO2 produced by the mitochondrial TCA cycle extruded into the extracellular space and increases ECAR. First, the inhibition of mitochondrial complex I and III by rotenone and antimycin A (Rot/AA) resulted in the reduced rate of proton efflux from respiration, which is calculated and removed from the total proton efflux rate results in glycolytic proton efflux rate (glycoPER). Second, glycolysis inhibitor 2-DG (hexokinase, HK2 inhibitor) is injected to stop glycolytic acidification and confirm pathway specificity. (E) Our experimental design (n = 16 wells/group). The 6 glycolysis parameters were measured in this assay, including basal glycolysis, basal proton efflux rate, percentage of PER from glycolysis, compensatory glycolysis, post–2-DG acidification, and mitoOCR/glycoPER. (F) Inhibition of glycolysis reduced TMAO-induced EC activation (n = 3; the experiment was repeated 3 times). Data are represented as the mean ± SEM (t test; *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001).