Choline/Betaine Prognostics Depend on Trimethylamine-N-Oxide
Choline/Betaine Prognostics Depend on Trimethylamine-N-Oxide
Aims Recent metabolomics and animal model studies show trimethylamine-N-oxide (TMAO), an intestinal microbiota-dependent metabolite formed from dietary trimethylamine-containing nutrients such as phosphatidylcholine (PC), choline, and carnitine, is linked to coronary artery disease pathogenesis. Our aim was to examine the prognostic value of systemic choline and betaine levels in stable cardiac patients.
Methods and results We examined the relationship between fasting plasma choline and betaine levels and risk of major adverse cardiac events (MACE = death, myocardial infraction, stroke) in relation to TMAO over 3 years of follow-up in 3903 sequential stable subjects undergoing elective diagnostic coronary angiography. In our study cohort, median (IQR) TMAO, choline, and betaine levels were 3.7 (2.4–6.2)μM, 9.8 (7.9–12.2)μM, and 41.1 (32.5–52.1)μM, respectively. Modest but statistically significant correlations were noted between TMAO and choline (r = 0.33, P < 0.001) and less between TMAO and betaine (r = 0.09, P < 0.001). Higher plasma choline and betaine levels were associated with a 1.9-fold and 1.4-fold increased risk of MACE, respectively (Quartiles 4 vs. 1; P < 0.01, each). Following adjustments for traditional cardiovascular risk factors and high-sensitivity C-reactive protein, elevated choline [1.34 (1.03–1.74), P < 0.05], and betaine levels [1.33 (1.03–1.73), P < 0.05] each predicted increased MACE risk. Neither choline nor betaine predicted MACE risk when TMAO was added to the adjustment model, and choline and betaine predicted future risk for MACE only when TMAO was elevated.
Conclusion Elevated plasma levels of choline and betaine are each associated with incident MACE risk independent of traditional risk factors. However, high choline and betaine levels are only associated with higher risk of future MACE with concomitant increase in TMAO.
Choline is a semi-essential nutrient for humans, meaning that while it can be endogenously synthesized, additional dietary intake of choline is also required or else a deficiency state will eventually occur. A common nutrient in many animal and some plant products, choline is found in dietary sources such as egg yolk, meats, liver, fish, dairy products, nuts, and soybean. Choline is a component of phosphatidylcholine (PC, also known as lecithin), the major phospholipid in cell membranes. It also is a precursor of the neurotransmitter acetylcholine, and following oxidation to betaine, choline functions as a methyl group donor in pathways that produce S-adenosylmethionine. As a methyl donor, choline and betaine can influence DNA and histone methylation. Choline deficiency has been linked to neurological impairment, and targeting enzymes related to choline metabolism has been postulated to provide promising therapeutic opportunities for tumour growth arrest. The connection between dietary choline and betaine and heart disease in subjects is unclear. While high intake of dietary choline and betaine was reportedly associated with diminished inflammation in one study, epidemiological studies have reported a limited association between high choline and betaine intake and cardiovascular morbidity that was attenuated following adjustments for other cardiovascular risk markers.
There is a growing appreciation that intestinal microbiota can participate in metabolic phenotypes such as obesity and insulin resistance. Recently, through a series of animal and human studies, our group has identified a direct mechanistic link between intestinal microbiota-dependent metabolism of trimethylamine (TMA)-containing dietary nutrients such as choline, PC, and carnitine and production of trimethylamine-N-oxide (TMAO), a metabolite shown to both alter cholesterol and sterol metabolism in multiple compartments and to directly enhanced atherosclerosis in animal models. Gut microbiota have been shown to play an obligate role in TMAO formation from ingested choline, PC, and carnitine in both animal models and humans. Moreover, elevated levels of TMAO have been shown to predict increased prevalent cardiovascular disease (CVD) risk, and increased incident risk of major adverse cardiovascular events (MACE = myocardial infarction, stroke, and death) in large-scale human clinical studies. Betaine, an oxidation product of choline, is generally regarded as safe for dietary ingestion. Surprisingly, although a TMA-containing species, whether orally ingested betaine produces TMAO or not is unknown. Moreover, while elevated whole-blood choline levels have been reported to predict increased cardiovascular risk, the relationship between TMAO and the clinical prognostic value of choline or betaine is unknown. Herein, we show that oral ingestion of d9-betaine results in generation of circulating levels of d9-TMAO in a gut microbiota-dependent fashion. We further examine the relationship between circulating levels of choline and betaine and development of cardiovascular morbidity and mortality, and the impact of TMAO on this relationship.
Abstract and Introduction
Abstract
Aims Recent metabolomics and animal model studies show trimethylamine-N-oxide (TMAO), an intestinal microbiota-dependent metabolite formed from dietary trimethylamine-containing nutrients such as phosphatidylcholine (PC), choline, and carnitine, is linked to coronary artery disease pathogenesis. Our aim was to examine the prognostic value of systemic choline and betaine levels in stable cardiac patients.
Methods and results We examined the relationship between fasting plasma choline and betaine levels and risk of major adverse cardiac events (MACE = death, myocardial infraction, stroke) in relation to TMAO over 3 years of follow-up in 3903 sequential stable subjects undergoing elective diagnostic coronary angiography. In our study cohort, median (IQR) TMAO, choline, and betaine levels were 3.7 (2.4–6.2)μM, 9.8 (7.9–12.2)μM, and 41.1 (32.5–52.1)μM, respectively. Modest but statistically significant correlations were noted between TMAO and choline (r = 0.33, P < 0.001) and less between TMAO and betaine (r = 0.09, P < 0.001). Higher plasma choline and betaine levels were associated with a 1.9-fold and 1.4-fold increased risk of MACE, respectively (Quartiles 4 vs. 1; P < 0.01, each). Following adjustments for traditional cardiovascular risk factors and high-sensitivity C-reactive protein, elevated choline [1.34 (1.03–1.74), P < 0.05], and betaine levels [1.33 (1.03–1.73), P < 0.05] each predicted increased MACE risk. Neither choline nor betaine predicted MACE risk when TMAO was added to the adjustment model, and choline and betaine predicted future risk for MACE only when TMAO was elevated.
Conclusion Elevated plasma levels of choline and betaine are each associated with incident MACE risk independent of traditional risk factors. However, high choline and betaine levels are only associated with higher risk of future MACE with concomitant increase in TMAO.
Introduction
Choline is a semi-essential nutrient for humans, meaning that while it can be endogenously synthesized, additional dietary intake of choline is also required or else a deficiency state will eventually occur. A common nutrient in many animal and some plant products, choline is found in dietary sources such as egg yolk, meats, liver, fish, dairy products, nuts, and soybean. Choline is a component of phosphatidylcholine (PC, also known as lecithin), the major phospholipid in cell membranes. It also is a precursor of the neurotransmitter acetylcholine, and following oxidation to betaine, choline functions as a methyl group donor in pathways that produce S-adenosylmethionine. As a methyl donor, choline and betaine can influence DNA and histone methylation. Choline deficiency has been linked to neurological impairment, and targeting enzymes related to choline metabolism has been postulated to provide promising therapeutic opportunities for tumour growth arrest. The connection between dietary choline and betaine and heart disease in subjects is unclear. While high intake of dietary choline and betaine was reportedly associated with diminished inflammation in one study, epidemiological studies have reported a limited association between high choline and betaine intake and cardiovascular morbidity that was attenuated following adjustments for other cardiovascular risk markers.
There is a growing appreciation that intestinal microbiota can participate in metabolic phenotypes such as obesity and insulin resistance. Recently, through a series of animal and human studies, our group has identified a direct mechanistic link between intestinal microbiota-dependent metabolism of trimethylamine (TMA)-containing dietary nutrients such as choline, PC, and carnitine and production of trimethylamine-N-oxide (TMAO), a metabolite shown to both alter cholesterol and sterol metabolism in multiple compartments and to directly enhanced atherosclerosis in animal models. Gut microbiota have been shown to play an obligate role in TMAO formation from ingested choline, PC, and carnitine in both animal models and humans. Moreover, elevated levels of TMAO have been shown to predict increased prevalent cardiovascular disease (CVD) risk, and increased incident risk of major adverse cardiovascular events (MACE = myocardial infarction, stroke, and death) in large-scale human clinical studies. Betaine, an oxidation product of choline, is generally regarded as safe for dietary ingestion. Surprisingly, although a TMA-containing species, whether orally ingested betaine produces TMAO or not is unknown. Moreover, while elevated whole-blood choline levels have been reported to predict increased cardiovascular risk, the relationship between TMAO and the clinical prognostic value of choline or betaine is unknown. Herein, we show that oral ingestion of d9-betaine results in generation of circulating levels of d9-TMAO in a gut microbiota-dependent fashion. We further examine the relationship between circulating levels of choline and betaine and development of cardiovascular morbidity and mortality, and the impact of TMAO on this relationship.