Trimethylamine -N-oxide induces osteogenic responses in human aortic valve interstitial cells in vitro and aggravates aortic valve lesions in mice.

Aims Recent studies have shown that the choline-derived metabolite trimethylamine N-oxide (TMAO) is a biomarker that promotes cardiovascular disease through the induction of inflammation and stress. Inflammatory responses and stress are involved in the progression of calcified aortic valve disease (CAVD). Here, we examined whether TMAO induces the osteogenic differentiation of aortic valve interstitial cells (AVICs) through endoplasmic reticulum (ER) and mitochondrial stress pathways in vitro and in vivo. Methods and results Plasma TMAO levels were higher in patients with CAVD (n = 69) than in humans without CAVD (n = 263), as examined by liquid chromatography-tandem mass spectrometry. Western blot and staining probes showed that TMAO- induced an osteogenic response in human AVICs. Moreover, TMAO promoted ER stress, mitochondrial stress and NF-κB activation in vitro. Notably, the TMAO- mediated effects were reversed by the use of ER stress, mitochondrial stress and NF-κB activation inhibitors and siRNA. Mice treated with supplemental choline in a high fat diet had markedly increased TMAO levels and aortic valve thicknesses, which were reduced by 3,3-dimethyl-1-butanol (DMB, an inhibitor of trimethylamine formation) treatment. Conclusions Choline-derived TMAO promotes osteogenic differentiation through ER and mitochondrial stress pathways in vitro and aortic valve lesions in vivo. Translational perspective Trimethylamine-N-oxide (TMAO), a gut microbiota-generated metabolite, is associated with cardiovascular diseases. Here, we show that patients with calcified aortic valve disease (CAVD) have elevated circulating TMAO levels. TMAO induces osteogenic responses in human aortic valve interstitial cells via endoplasmic reticulum-mitochondrial stress in vitro and aggravates aortic valve lesions in mice. This may provide clues to the pathogenesis of CAVD and attractive potential targets for the prevention, diagnosis and treatment of this disease.