Recent studies have revealed strong associations between systemic trimethylamine N-oxide (TMAO) levels, atherosclerosis and cardiovascular risk. In addition, plasma L-carnitine levels in patients with high TMAO concentrations predicted an increased risk for cardiovascular disease and incident major adverse cardiac events. The aim of the present study was to investigate the relation between TMAO and L-carnitine plasma levels and diabetes. Blood plasma samples were collected from 12 and 20 weeks old db/db mice and patients undergoing percutaneous coronary intervention. Diabetic compared to non-diabetic db/L mice presented 10-fold higher TMAO, but lower L-carnitine plasma concentrations at 12 weeks of age. After 8 weeks of observation, diabetic db/db mice had significantly increased body weight, insulin resistance and TMAO concentration in comparison to non-diabetic control. In 191 patients undergoing percutaneous coronary intervention the median (interquartile range) plasma concentration of TMAO was 1.8 (1.2-2.6) µmol/L. Analysis of the samples showed a bivariate association of TMAO level with age, total cholesterol and L-carnitine. The multivariate linear regression analysis revealed that, in addition to L-carnitine as the strongest predictor of log transformed TMAO (p<0.001), the parameters of age, diabetes status and body mass index (BMI) were independently associated with increased log transformed TMAO levels (p<0.01).Our data provide evidence that age, diabetes and BMI are associated with higher TMAO levels independently of L-carnitine. These data support the hypothesis of TMAO as a cardiovascular risk marker and warrant further investigation of TMAO for diabetes research applications.

Background:The bacterial glycyl radical enzyme CutC converts choline to trimethylamine, a metabolite involved in pathogenesis of several diseases. Results: The structures of substrate-bound and substrate-free CutC revealed significant differences. Conclusion: Choline binding to the active site triggers a conformational change from the open to closed form. Significance: A novel substrate-driven conformational mechanism and a potential target for drug design have been identified.

Increased plasma concentration of trimethylamine N-oxide (TMAO), a proatherogenic metabolite, has been linked to adverse cardiovascular outcomes; however, it remains unclear whether TMAO is a biomarker or whether it induces direct detrimental cardiovascular effects. Because altered cardiac energy metabolism and mitochondrial dysfunction play crucial roles in the development of cardiovascular diseases, we hypothesized that increased TMAO concentration may alter mitochondrial energy metabolism. The aim of the present study was to determine the effects of TMAO on cardiac mitochondrial energy metabolism. Acute exposure of cardiac fibers to TMAO decreased LEAK (substrate-dependent) and OXPHOS (oxidative phosphorylation-dependent) mitochondrial respiration with pyruvate and impaired substrate flux via pyruvate dehydrogenase. The administration of TMAO at a dose of 120mg/kg for 8 weeks increased TMAO concentration in plasma and cardiac tissues 22-23 times to about 15μM and 11nmol/g, respectively. Long-term TMAO administration decreased mitochondrial LEAK state respiration with pyruvate by 30% without affecting OXPHOS state respiration. However, no significant changes in mitochondrial reactive oxygen species production were observed after acute exposure of cardiac fibers to TMAO under physiological conditions. In addition, both long-term TMAO administration and acute exposure to TMAO decreased respiration with palmitoyl-CoA indicating impaired β-oxidation. Taken together, our results demonstrate that increased TMAO concentration impairs pyruvate and fatty acid oxidation in cardiac mitochondria. Thus, the accumulation of TMAO in cardiac tissues leads to disturbances in energy metabolism that can increase the severity of cardiovascular events.

Background and purpose: Mildronate [3-(2,2,2-trimethylhydrazinium) propionate] is an anti-ischaemic drug whose mechanism of action is based on its inhibition of L-carnitine biosynthesis and uptake. As L-carnitine plays a pivotal role in the balanced metabolism of fatty acids and carbohydrates, this study was carried out to investigate whether long-term mildronate treatment could influence glucose levels and prevent diabetic complications in an experimental model of type 2 diabetes in Goto-Kakizaki (GK) rats. Experimental approach: GK rats were treated orally with mildronate at doses of 100 and 200 mg·kg -1 daily for 8 weeks. Plasma metabolites reflecting glucose and lipids, as well as fructosamine and b-hydroxybutyrate, were assessed. L-carnitine concentrations were measured by ultra performance liquid chromatography with tandem mass spectrometry. An isolated rat heart ischaemia-reperfusion model was used to investigate possible cardioprotective effects. Pain sensitivity was measured with a tail-flick latency test. Key results: Mildronate treatment significantly decreased L-carnitine concentrations in rat plasma and gradually decreased both the fed-and fasted-state blood glucose. Mildronate strongly inhibited fructosamine accumulation and loss of pain sensitivity and also ameliorated the enhanced contractile responsiveness of GK rat aortic rings to phenylephrine. In addition, in mildronate-treated hearts, the necrosis zone following coronary occlusion was significantly decreased by 30%. Conclusions and implications:These results demonstrate for the first time that in GK rats, an experimental model of type 2 diabetes, mildronate decreased L-carnitine contents and exhibited cardioprotective effects, decreased blood glucose concentrations and prevented the loss of pain sensitivity. These findings indicate that mildronate treatment could be beneficial in diabetes patients with cardiovascular problems.

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