Abstract
A mechanistic link between trimethylamine N-oxide (TMAO) and atherogenesis has been reported. TMAO is generated enzymatically in the liver by the oxidation of trimethylamine (TMA), which is produced from dietary choline, carnitine and betaine by gut bacteria. It is known that certain members of methanogenic archaea (MA) could use methylated amines such as trimethylamine as growth substrates in culture. Therefore, we investigated the efficacy of gut colonization with MA on lowering plasma TMAO concentrations. Initially, we screened for the colonization potential and TMAO lowering efficacy of five MA species in C57BL/6 mice fed with high choline/TMA supplemented diet, and found out that all five species could colonize and lover plasma TMAO levels, although with different efficacies. The top performing MA, Methanobrevibacter smithii, Methanosarcina mazei, and Methanomicrococcus blatticola, were transplanted into Apoe−/− mice fed with high choline/TMA supplemented diet. Similar to C57BL/6 mice, following initial provision of the MA, there was progressive attrition of MA within fecal microbial communities post-transplantation during the initial 3 weeks of the study. In general, plasma TMAO concentrations decreased significantly in proportion to the level of MA colonization. In a subsequent experiment, use of antibiotics and repeated transplantation of Apoe−/− mice with M. smithii, led to high engraftment levels during the 9 weeks of the study, resulting in a sustained and significantly lower average plasma TMAO concentrations (18.2 ± 19.6 μM) compared to that in mock-transplanted control mice (120.8 ± 13.0 μM, p < 0.001). Compared to control Apoe−/− mice, M. smithii-colonized mice also had a 44% decrease in aortic plaque area (8,570 μm [95% CI 19587–151821] vs. 15,369 μm [95% CI [70058–237321], p = 0.34), and 52% reduction in the fat content in the atherosclerotic plaques (14,283 μm [95% CI 4,957–23,608] vs. 29,870 μm [95% CI 18,074–41,666], p = 0.10), although these differences did not reach significance. Gut colonization with M. smithii leads to a significant reduction in plasma TMAO levels, with a tendency for attenuation of atherosclerosis burden in Apoe−/− mice. The anti-atherogenic potential of MA should be further tested in adequately powered experiments.
Highlights
Atherosclerotic vascular disease is the leading cause of death in the US1
M. luminyensis and M. portucalensis had the lowest engraftment levels ranging from 0.4 × 103 to 1.3 × 103 copies/mg stool on day 2 post-transplant, which decreased on the 30th day
On day 2 post-transplant, plasma TMAO concentrations were significantly lower in mice colonized with M. smithii (14.8 ± 15.7 μM, p = 0.003), M.mazei (6.9 ± 10.6 μM, p < 0.001), M. blatticola (5.9 ± 3.8 μM, p < 0.001), M. luminyensis (102.3 ± 27.3 μM, p = 0.03), and M. portucalensis (45.57 ± 16.8 μM, p < 0.01), compared to the positive control mice (Fig. 1B)
Summary
Atherosclerotic vascular disease is the leading cause of death in the US1. The gut microbiome is recognized as a mediator of numerous host physiological processes. Changes in the gut microbiome have been causally linked to several metabolic, inflammatory, and cardiovascular diseases, including atherosclerosis[2,3]. Systemic concentrations of the gut microbe-derived metabolite, trimethylamine-N-oxide (TMAO), is associated with atherosclerosis and major adverse cardiovascular events[4,5,6]. Choline diet dependent enhancement in atherosclerosis could be prevented by either gut microbiota suppression with broad spectrum antibiotics[6], or in more recent studies, administration of an inhibitor of microbial choline trimethylamine (TMA) lyase activity and TMAO generation (4,4-dimethyl-1-butanol)[7], indicating a causal link between TMAO and atherosclerosis. FMO3 gene mutations cause the inherited disorder primary trimethylaminuria (TMAU)[14], known as the fish odor syndrome, which severely reduces the ability to convert TMA to TMAO. We tested the effect of MA colonization on blood TMAO level and atherosclerosis burden in the atherosclerosis prone Apoe−/− mouse model
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