Trimethylamine-N-oxide is an important target for heart and brain diseases

Trimethylamine-N-oxide (TMAO) is a metabolite produced from dietary nutrients by the gut microbiome, which was first discovered in 2011 and used to predict the risk of cardiovascular disease [1]. The precursors of TMAO, including phosphatidylcholine [1], choline [2], and L-carnitine [3], are commonly found in cheese, red meat, seafood, egg yolks, and other foods. To date, four different microbial enzyme systems have been identified that can convert precursor substances into trimethylamine (TMA), including choline-TMA lyase (cutC/D) [4], carnitine monooxygenase (cntA/B) [5], betaine reductase, and TMAO reductase [6]. In addition, yeaW/X, which is homologous to cntA/B and can utilize a variety of substrates such as choline, betaine, carnitine and γ-butyrobetaine, can also promote the synthesis of TMA. The cooperation between yeaW/X and cutC/D was the most well studied [7].

Most of the TMA produced by gut metabolism enters the circulatory system; TMA is absorbed into the bloodstream and enters the liver, where it is converted to TMAO by flavin monooxygenases (FMOs) [8]. The FMO family contains five functional enzymes, of which FMO3 is the key restriction enzyme for the conversion of TMA to TMAO, with the highest conversion efficiency. Excess TMA is directly decomposed into dimethylamine. Previous studies have shown that TMAO can accumulate in the heart, kidneys, brain or other tissues and participate in a variety of biological processes, such as activating platelet aggregation, increasing foam cell formation, inducing inflammatory responses, and reducing reverse cholesterol transport. In most cases, most of TMAO is eliminated by the kidneys, and the rest is reduced to TMA by TMAO reductase in the intestine [9].

In 2013, based on a nontargeted metabolomics analysis, researchers established a direct association between TMAO and cardiovascular event risk. TMAO has been found to have a certain correlation with atherosclerosis, heart failure, hypertension, chronic kidney disease, tumors, obesity-related diseases, diabetes and other diseasesin nearly a decade [10], [11], [12], [13].

However, it seems that the effect of TMAO on various diseases is controversial. If there is no rule out whether there are other ways to produce TMAO, the effect of TMAO on diseases is often a process involving genes and environment, and it is obviously individualized. Multiple studies have concluded that TMAO does not exacerbate the development of atherosclerosis in mice, and a 2018 study on Apoe −/− mice showed that choline supplementation promoted an increase in TMAO levels in mice but had no effect on the absolute size of aortic plaques [14]. There are also clinical studies showing that in healthy young adults, TMAO may not significantly increase the risk of early atherosclerotic disease [15]. The probability that TMAO raises the risk of disease varies from person to person, is associated with different genotypes, and cannot be generalized to the entire population [16]. Due to large population variations, plasma TMAO is sometimes negatively correlated with the severity of the disease [17].

Notably, the prognostic value of TMAO is not corrected by traditional risk factors, which highlights its potential as a biomarker that can be used for risk stratification beyond traditional risks. While not all studies observed an association between elevated TMAO levels and the risk of new-onset cardiovascular disease (CVD) tests conducted on existing clinical studies have shown an association between elevated TMAO levels and the risk of new-onset CVD. Multiple meta-analyses concluded that there was indeed a strong correlation between elevated circulating TMAO levels and CVD risk and mortality in multiple cohorts across different continents [18, 19]. In many studies, plasma TMAO cutoffs > 6 μmol/L predicted an increased risk of adverse cardiac events [20]. In a recent meta-analysis of 25,000 participants, every 10 μmol/L increase in TMA increased all-cause mortality by 7.6% [18].

There are individual differences in metabolites of intestinal flora in both human beings and experimental animals, which requires the experimenter to pay attention to keep the collection and preservation methods of fecal samples unchanged and use relatively unified sequencing platform and data analysis method when doing research. These differences may affect the consistency of the gut microbiota and lead to changes in the gut microbiota. In human clinical cohort studies, many parameters of subjects may affect the consistency of microbiota findings, such as dietary habits, lifestyle, and medications. In animal studies, different ages, living conditions, or diets and feeding periods will also affect the intestinal flora. In a recent report by Hazen et al., the gut microbial pathways that produce TMA from γ-butyrobetaine were revealed and helped explain how diets rich in red meat led to elevated TMAO levels and increased risk of CVD [21]. But they also note that, according to a previous report on the provision of l-carnitine supplements to vegans and omnivores, mean TMAO levels increased in both groups, but TMA(O) levels did not change in a significant proportion of vegans [22]. This indicates that the metabolic capacity of gut microbes has a certain stability, which may remain unchanged over time, which can explain some doubts about the role of TMAO. The broad role of TMAO is unquestionable, so properly designing studies to make reliable discoveries is critical for future research.

In addition, because of the progress of brain-intestine axis research, TMAO and neurological-related diseases are not in the minority. In vivo studies have shown the presence of TMAO in the cerebrospinal fluid of mice and humans [23, 24]. This means that hepatic TMAO can penetrate the blood-brain barrier (BBB) to reach the nerve center, but the specific mechanism of penetration is still unclear [25]. There is another idea that TMAO found in cerebrospinal fluid is partly converted from TMA by FMOs in the brain, rather than completely from the penetration of the BBB [26].

Similarly, there is much debate about whether the effects of TMAO on the brain are positive or harmful. Clinical studies have shown that patients with Alzheimer’s disease (AD) dementia have higher levels of TMAO in cerebrospinal fluid than individuals with normal cognitive function, but the specific effects of TMAO on brain cognitive function cannot be clarified. High-circulation TMAO may also promote neuroinflammation by increasing intrabrainous nuclear factor kappa-B (NF-κB) and proinflammatory cytokines, which are recognized mediators of cognitive aging and nerve function [27].

In a mice study, TMAO can induce brain aging and aging-related cognitive dysfunction in senescence-resistant 1 (SAMR1) mice, and aggravate the brain aging process in senescence-prone 8 (SAMP8) mice, which may provide a new idea for the effect of gut microbiota on brain aging process and help to delay aging by regulating gut microbiota metabolites [28]. Regarding the relationship between TMAO and ischemic stroke, in a large Chinese cohort, elevated TMAO levels were associated with an increased risk of recurrent stroke in patients with small-artery occlusion subtypes [29]. However, TMAO appears to affect the brain in a dose-dependent manner. Some studies have shown that TMAO may exert a neuroprotective effect within the normal physiological range (plasma concentration of 0.5–5.0 μmol/L) [30].

Gut microbiota, as a complex microecosystem, constantly affects the health of the host, and the physiological and pathological processes in which it participates have gradually become the most important position for new drug discovery and research. The mechanism of TMAO’s effect on cardio-cerebral, renal, and systemic metabolic disorders still requires further study. Because intestinal flora metabolites are heavily influenced by the environment, individual variability, and heterogeneity of experimental design, this makes it necessary for experimenters to understand these highly complex systems, especially as we turn more to human studies, and need to continuously improve computational and statistical methods to obtain and implement the multiomics and longitudinal data required for integrated methods. The studies on the gut microbiota-TMA-TMAO metabolic pathway and various organ diseases are the most typical examples of gut microbiota metabolites. It is reasonable to believe that microecological agents, small molecule drugs, fecal microbiota transplantation and other intervention methods can become a new means of disease prevention and treatment in the future.