Living with trimethylaminuria and body and breath malodour: personal perspectives.

BACKGROUND: Many people suffer from body and breath malodour syndromes. One of these is trimethylaminuria, a condition characterized by excretion in breath and bodily fluids of trimethylamine, a volatile and odorous chemical that has the smell of rotting fish. Trimethylaminuria can be primary, due to mutations in the gene encoding flavin-containing monooxygenase 3, or secondary, due to various causes. To gain a better understanding of problems faced by United Kingdom residents affected by body and breath malodour conditions, we conducted a survey. METHODS: Two anonymous online surveys, one for adults and one for parents/guardians of affected children, were conducted using the Opinio platform. Participants were invited via a trimethylaminuria advisory website. Questions were a mix of dropdown, checkbox and open-ended responses. Forty-four adults and three parents/guardians participated. The dropdown and checkbox responses were analysed using the Opinio platform. RESULTS: All participants reported symptoms of body/breath odour. However, not all answered every question. Twenty-three respondents experienced difficulties in being offered a diagnostic test for trimethylaminuria. Problems encountered included lack of awareness of the disorder by medical professionals and reluctance to recognise symptoms. Of those tested, 52% were diagnosed with trimethylaminuria. The main problems associated with living with body/breath malodours were bullying, harassment and ostracism in either the workplace (90%) or in social settings (88%). All respondents thought their condition had disadvantaged them in their daily lives. Open-ended responses included loss of confidence, stress, exclusion, isolation, loneliness, depression and suicidal thoughts. Respondents thought their lives could be improved by greater awareness and understanding of malodour conditions by medical professionals, employers and the general public, and appreciation that the malodour was due to a medical condition and not their fault. CONCLUSIONS: Breath and body malodour conditions can cause immense hardship and distress, both mentally and socially, having devastating effects on quality of life. It would be advantageous to establish a standardised pathway from primary care to a specialist unit with access to a robust and reliable test and diagnostic criteria. There is a need to recognise malodour disorders as a disability, giving affected individuals the same rights as those with currently recognised disabilities.

Metabolomic and transcriptomic analyses of Fmo5-/- mice reveal roles for flavin-containing monooxygenase 5 (FMO5) in NRF2-mediated oxidative stress response, unfolded protein response, lipid homeostasis, and carbohydrate and one-carbon metabolism.

Flavin-containing monooxygenase 5 (FMO5) is a member of the FMO family of proteins, best known for their roles in the detoxification of foreign chemicals and, more recently, in endogenous metabolism. We have previously shown that Fmo5-/- mice display an age-related lean phenotype, with much reduced weight gain from 20 weeks of age. The phenotype is characterized by decreased fat deposition, lower plasma concentrations of glucose, insulin and cholesterol, higher glucose tolerance and insulin sensitivity, and resistance to diet-induced obesity. In the present study we report the use of metabolomic and transcriptomic analyses of livers of Fmo5-/- and wild-type mice to identify factors underlying the lean phenotype of Fmo5-/- mice and gain insights into the function of FMO5. Metabolomics was performed by the Metabolon platform, utilising ultrahigh performance liquid chromatography-tandem mass spectroscopy. Transcriptomics was performed by RNA-Seq and results analysed by DESeq2. Disruption of the Fmo5 gene has wide-ranging effects on the abundance of metabolites and expression of genes in the liver. Metabolites whose concentration differed between Fmo5-/- and wild-type mice include several saturated and monounsaturated fatty acids, complex lipids, amino acids, one-carbon intermediates and ADP-ribose. Among the genes most significantly and/or highly differentially expressed are Apoa4, Cd36, Fitm1, Hspa5, Hyou1, Ide, Me1 and Mme. The results reveal that FMO5 is involved in upregulating the NRF2-mediated oxidative stress response, the unfolded protein response and response to hypoxia and cellular stress, indicating a role for the enzyme in adaptation to oxidative and metabolic stress. FMO5 also plays a role in stimulating a wide range of metabolic pathways and processes, particularly ones involved in lipid homeostasis, the uptake and metabolism of glucose, the generation of cytosolic NADPH, and in one-carbon metabolism. The results predict that FMO5 acts by stimulating the NRF2, XBP1, PPARA and PPARG regulatory pathways, while inhibiting STAT1 and IRF7 pathways.

Treatment of wild-type mice with 2,3-butanediol, a urinary biomarker of Fmo5 -/- mice, decreases plasma cholesterol and epididymal fat deposition.

We previously showed that Fmo5 -/- mice exhibit a lean phenotype and slower metabolic ageing. Their characteristics include lower plasma glucose and cholesterol, greater glucose tolerance and insulin sensitivity, and a reduction in age-related weight gain and whole-body fat deposition. In this paper, nuclear magnetic resonance (NMR) spectroscopy-based metabolite analyses of the urine of Fmo5 -/- and wild-type mice identified two isomers of 2,3-butanediol as discriminating urinary biomarkers of Fmo5 -/- mice. Antibiotic-treatment of Fmo5 -/- mice increased plasma cholesterol concentration and substantially reduced urinary excretion of 2,3-butanediol isomers, indicating that the gut microbiome contributed to the lower plasma cholesterol of Fmo5 -/- mice, and that 2,3-butanediol is microbially derived. Short- and long-term treatment of wild-type mice with a 2,3-butanediol isomer mix decreased plasma cholesterol and epididymal fat deposition but had no effect on plasma concentrations of glucose or insulin, or on body weight. In the case of long-term treatment, the effects were maintained after withdrawal of 2,3-butanediol. Short-, but not long-term treatment, also decreased plasma concentrations of triglycerides and non-esterified fatty acids. Fecal transplant from Fmo5 -/- to wild-type mice had no effect on plasma cholesterol, and 2,3-butanediol was not detected in the urine of recipient mice, suggesting that the microbiota of the large intestine was not the source of 2,3-butanediol. However, 2,3-butanediol was detected in the stomach of Fmo5 -/- mice, which was enriched for Lactobacillus genera, known to produce 2,3-butanediol. Our results indicate a microbial contribution to the phenotypic characteristic of Fmo5 -/- mice of decreased plasma cholesterol and identify 2,3-butanediol as a potential agent for lowering plasma cholesterol.

Material jet printing of transparent ceramic Yb:YAG planar waveguides.

A new, to the best of our knowledge, 3D additive manufacturing technique utilizing particle-loaded ink jet printing to fabricate transparent ceramic Yb:YAG planar waveguides for laser gain media was demonstrated. Rheological optimization of YAG particle-loaded inks resulted in successful droplet formation and printing resolution. Planar waveguides composed of a Yb:YAG guide encased in undoped YAG cladding were printed with guide thicknesses ranging between 25 and 350 µm and consolidated to high optical quality via solid-state sintering. Sufficiently low optical (1-3%/cm) and intermodal scatter allowed single-mode propagation with a core/clad index difference of $\Delta {n}\sim{5.0} \times {{10}^{- 4}}$ (corresponding to 3 at.% Yb). The waveguides were cladding-pumped longitudinally with a 940 nm diode array resulting in 23.6% slope efficiency in 2 ms pulsed operation.

Flavin-Containing Monooxygenase 1 Catalyzes the Production of Taurine from Hypotaurine.

Taurine is one of the most abundant amino acids in mammalian tissues. It is obtained from the diet and by de novo synthesis from cysteic acid or hypotaurine. Despite the discovery in 1954 that the oxygenation of hypotaurine produces taurine, the identification of an enzyme catalyzing this reaction has remained elusive. In large part, this is due to the incorrect assignment, in 1962, of the enzyme as an NAD-dependent hypotaurine dehydrogenase. For more than 55 years, the literature has continued to refer to this enzyme as such. Here we show, both in vivo and in vitro, that the enzyme that oxygenates hypotaurine to produce taurine is flavin-containing monooxygenase (FMO) 1. Metabolite analysis of the urine of Fmo1-null mice by 1H NMR spectroscopy revealed a buildup of hypotaurine and a deficit of taurine in comparison with the concentrations of these compounds in the urine of wild-type mice. In vitro assays confirmed that human FMO1 catalyzes the conversion of hypotaurine to taurine, utilizing either NADPH or NADH as cofactor. FMO1 has a wide substrate range and is best known as a xenobiotic- or drug-metabolizing enzyme. The identification that the endogenous molecule hypotaurine is a substrate for the FMO1-catalyzed production of taurine resolves a long-standing mystery. This finding should help establish the role FMO1 plays in a range of biologic processes in which taurine or its deficiency is implicated, including conjugation of bile acids, neurotransmitter, antioxidant and anti-inflammatory functions, and the pathogenesis of obesity and skeletal muscle disorders. SIGNIFICANCE STATEMENT: The identity of the enzyme that catalyzes the biosynthesis of taurine from hypotaurine has remained elusive. Here we show, both in vivo and in vitro, that flavin-containing monooxygenase 1 catalyzes the oxygenation of hypotaurine to produce taurine.

Flavin-containing monooxygenase 3 (FMO3): genetic variants and their consequences for drug metabolism and disease.

The review focuses on genetic variants of human flavin-containing monooxygenase 3 (FMO3) and their impact on enzyme activity, drug metabolism and disease.The majority of FMO-mediated metabolism in adult human liver is catalyzed by FMO3. Some drugs are metabolized in human liver predominantly by FMO3, but most drug substrates of FMO3 are metabolized also by other enzymes, particularly cytochromes P-450, and the FMO3-catalyzed reaction is not the major route of metabolism.Rare variants that severely affect production or activity of FMO3 cause the disorder trimethylaminuria and impair metabolism of drug substrates of FMO3. More common variants, particularly p.[(Glu158Lys);(Glu308Gly)], can moderately affect activity of FMO3 in vitro and reduce metabolism of drug substrates in vivo, in some cases increasing drug efficacy or toxicity.Common variants of FMO3 have been associated with a number of disorders, but additional studies are needed to confirm or refute such associations.Elevated plasma concentrations of trimethylamine N-oxide, a product of an FMO3-catalyzed reaction, have been implicated in certain diseases, particularly cardiovascular disease. However, the evidence is often contradictory and additional work is required to establish whether trimethylamine N-oxide is a cause, effect or biomarker of the disease.Genetic variants of other FMOs are also briefly discussed.

Biochar amendment and water stress alter rhizosphere carbon and nitrogen budgets in bauxite-processing residue sand under rehabilitation.

Nitrogen (N) bioavailability is one of the main limiting factors for microbial activity and vegetation establishment in bauxite-processing residue sand (BRS). Although beneficial effects of biochar on reducing N loss in the early stages of BRS rehabilitation have been observed previously, the underlying mechanisms of this complicated process, particularly the interactions between applied biochar and the plant rhizosphere is largely unknown. This glasshouse study (116 days), investigated the coupled effects of biochar and water stress on N bioavailability in the rhizosphere of ryegrass (Lolium rigidum) grown in BRS amended with di-ammonium phosphate (DAP) fertiliser (at rates of 0 or 2.7 t ha-1) with and without biochar amendment. The applied biochar was characterised as either aged acidic (AC) or alkaline pine (PC) and was mixed with BRS at a rate of 5% v/v under four moisture regimes (50%, 40%, 20% and 7.5% water holding capacity). Amending BRS with AC and PC biochars increased NH4+ retention and decreased cumulative NH3 volatilization within both the rhizosphere and root-free zones compared with fertiliser only treatment. These effects were more pronounced for the AC than PC biochar, suggesting that aged acidic biochar has the great potential for use in rapid establishment of vegetation in BRS disposal areas. The biochar amendment increased cumulative nitrous oxide emissions compared with DAP only treatment, with no significant differences among different moisture regimes. The Control and 20% water holding capacity (WHC) treatment showed the highest dissolved organic carbon (DOC) concentrations compared with other treatments and moisture regimes in the ryegrass rhizosphere, while the highest dissolved organic N concentration were observed in the DAP + AC treatment. Reducing moisture levels below 20% WHC generally decreased microbial biomass carbon (MBC) concentrations and activity in both the rhizosphere and root-free zones of all treatments, while total N generally decreased as moisture levels decreased from 50% to 7.5% WHC. Plant took up more N in the DAP + AC treatment compared with DAP + PC and DAP only treatments, while increasing water stress generally resulted in decreased aboveground biomass.

Metabolic Biomarkers of Ageing in C57BL/6J Wild-Type and Flavin-Containing Monooxygenase 5 (FMO5)-Knockout Mice.

It was recently demonstrated in mice that knockout of the flavin-containing monooxygenase 5 gene, Fmo5, slows metabolic ageing via pleiotropic effects. We have now used an NMR-based metabonomics approach to study the effects of ageing directly on the metabolic profiles of urine and plasma from male, wild-type C57BL/6J and Fmo5-/- (FMO5 KO) mice back-crossed onto the C57BL/6J background. The aim of this study was to identify metabolic signatures that are associated with ageing in both these mouse lines and to characterize the age-related differences in the metabolite profiles between the FMO5 KO mice and their wild-type counterparts at equivalent time points. We identified a range of age-related biomarkers in both urine and plasma. Some metabolites, including urinary 6-hydroxy-6-methylheptan-3-one (6H6MH3O), a mouse sex pheromone, showed similar patterns of changes with age, regardless of genetic background. Others, however, were altered only in the FMO5 KO, or only in the wild-type mice, indicating the impact of genetic modifications on mouse ageing. Elevated concentrations of urinary taurine represent a distinctive, ageing-related change observed only in wild-type mice.

Effect of Flavin-Containing Monooxygenase Genotype, Mouse Strain, and Gender on Trimethylamine N-oxide Production, Plasma Cholesterol Concentration, and an Index of Atherosclerosis.

The objectives of the study were to determine the contribution, in mice, of members of the flavin-containing monooxygenase (FMO) family to the production of trimethylamine (TMA) N-oxide (TMAO), a potential proatherogenic molecule, and whether under normal dietary conditions differences in TMAO production were associated with changes in plasma cholesterol concentration or with an index of atherosclerosis (Als). Concentrations of urinary TMA and TMAO and plasma cholesterol were measured in 10-week-old male and female C57BL/6J and CD-1 mice and in mouse lines deficient in various Fmo genes (Fmo1-/- , 2-/- , 4-/- , and Fmo5-/- ). In female mice most TMA N-oxygenation was catalyzed by FMO3, but in both genders 11%-12% of TMA was converted to TMAO by FMO1. Gender-, Fmo genotype-, and strain-related differences in TMAO production were accompanied by opposite effects on plasma cholesterol concentration. Plasma cholesterol was negatively, but weakly, correlated with TMAO production and urinary TMAO concentration. Fmo genotype had no effect on Als. There was no correlation between Als and either TMAO production or urinary TMAO concentration. Our results indicate that under normal dietary conditions TMAO does not increase plasma cholesterol or act as a proatherogenic molecule.

A highly sensitive liquid chromatography electrospray ionization mass spectrometry method for quantification of TMA, TMAO and creatinine in mouse urine.

Our method describes the quantification in mouse urine of trimethylamine (TMA), trimethylamine N-oxide (TMAO) and creatinine. The method combines derivatization of TMA, with ethyl bromoacetate, and LC chromatographic separation on an ACE C18 column. The effluent was continuously electrosprayed into the linear ion trap mass spectrometer (LTQ), which operated in selective ion monitoring (SIM) modes set for targeted analytes and their internal standards (IS). All validation parameters were within acceptable ranges of analytical method validation guidelines. Intra- and inter-day assay precision and accuracy coefficients of variation were <3.1%, and recoveries for TMA and TMAO were 97-104%. The method developed uses a two-step procedure. Firstly, TMA and TMAO are analyzed without a purification step using a 5-min gradient cap-LC- SIMs analysis, then creatinine is analyzed using the same experimental conditions. The method is robust, highly sensitive, reproducible and has the high-throughput capability of detecting TMA, TMAO and creatinine at on-column concentrations as low as 28 pg/mL, 115 pg/mL and 1 ng/mL, respectively. The method is suitable for analysis of TMA, TMAO and creatinine in both male and female mouse urine. The key benefits of the method are: •The small sample volume of urine required, which overcomes the difficulties of collecting sufficient volumes of urine at defined times.•No sample pre-treatment is necessary.•The quantification of TMA, TMAO and creatinine using the same cap-LC-MS method.

Identification of Flavin-Containing Monooxygenase 5 (FMO5) as a Regulator of Glucose Homeostasis and a Potential Sensor of Gut Bacteria.

We have previously identified flavin-containing monooxygenase 5 (FMO5) as a regulator of metabolic aging. The aim of the present study was to investigate the role of FMO5 in glucose homeostasis and the impact of diet and gut flora on the phenotype of mice in which the Fmo5 gene has been disrupted (Fmo5-/- mice). In comparison with wild-type (WT) counterparts, Fmo5-/- mice are resistant to age-related changes in glucose homeostasis and maintain the higher glucose tolerance and insulin sensitivity characteristic of young animals. When fed a high-fat diet, they are protected against weight gain and reduction of insulin sensitivity. The phenotype of Fmo5-/- mice is independent of diet and the gut microbiome and is determined solely by the host genotype. Fmo5-/- mice have metabolic characteristics similar to those of germ-free mice, indicating that FMO5 plays a role in sensing or responding to gut bacteria. In WT mice, FMO5 is present in the mucosal epithelium of the gastrointestinal tract where it is induced in response to a high-fat diet. In comparison with WT mice, Fmo5-/- mice have fewer colonic goblet cells, and they differ in the production of the colonic hormone resistin-like molecule ßFmo5-/- mice have lower concentrations of tumor necrosis factor α in plasma and of complement component 3 in epididymal white adipose tissue, indicative of improved inflammatory tone. Our results implicate FMO5 as a regulator of body weight and of glucose disposal and insulin sensitivity and, thus, identify FMO5 as a potential novel therapeutic target for obesity and insulin resistance.

Drug metabolism by flavin-containing monooxygenases of human and mouse.

INTRODUCTION: Flavin-containing monooxygenases (FMOs) play an important role in drug metabolism. Areas covered: We focus on the role of FMOs in the metabolism of drugs in human and mouse. We describe FMO genes and proteins of human and mouse; the catalytic mechanism of FMOs and their significance for drug metabolism; differences between FMOs and CYPs; factors contributing to potential underestimation of the contribution of FMOs to drug metabolism; the developmental and tissue-specific expression of FMO genes and differences between human and mouse; and factors that induce or inhibit FMOs. We discuss the contribution of FMOs of human and mouse to the metabolism of drugs and how genetic variation of FMOs affects drug metabolism. Finally, we discuss the utility of animal models for FMO-mediated drug metabolism in humans. Expert opinion: The contribution of FMOs to drug metabolism may be underestimated. As FMOs are not readily induced or inhibited and their reactions are generally detoxifications, the design of drugs that are metabolized predominantly by FMOs offers clinical advantages. Fmo1(-/-),Fmo2(-/-),Fmo4(-/-) mice provide a good animal model for FMO-mediated drug metabolism in humans. Identification of roles for FMO1 and FMO5 in endogenous metabolism has implications for drug therapy and initiates an exciting area of research.

Trimethylamine and Trimethylamine N-Oxide, a Flavin-Containing Monooxygenase 3 (FMO3)-Mediated Host-Microbiome Metabolic Axis Implicated in Health and Disease.

Flavin-containing monooxygenase 3 (FMO3) is known primarily as an enzyme involved in the metabolism of therapeutic drugs. On a daily basis, however, we are exposed to one of the most abundant substrates of the enzyme trimethylamine (TMA), which is released from various dietary components by the action of gut bacteria. FMO3 converts the odorous TMA to nonodorous TMA N-oxide (TMAO), which is excreted in urine. Impaired FMO3 activity gives rise to the inherited disorder primary trimethylaminuria (TMAU). Affected individuals cannot produce TMAO and, consequently, excrete large amounts of TMA. A dysbiosis in gut bacteria can give rise to secondary TMAU. Recently, there has been much interest in FMO3 and its catalytic product, TMAO, because TMAO has been implicated in various conditions affecting health, including cardiovascular disease, reverse cholesterol transport, and glucose and lipid homeostasis. In this review, we consider the dietary components that can give rise to TMA, the gut bacteria involved in the production of TMA from dietary precursors, the metabolic reactions by which bacteria produce and use TMA, and the enzymes that catalyze the reactions. Also included is information on bacteria that produce TMA in the oral cavity and vagina, two key microbiome niches that can influence health. Finally, we discuss the importance of the TMA/TMAO microbiome-host axis in health and disease, considering factors that affect bacterial production and host metabolism of TMA, the involvement of TMAO and FMO3 in disease, and the implications of the host-microbiome axis for management of TMAU.

Shifts in leaf N:P stoichiometry during rehabilitation in highly alkaline bauxite processing residue sand.

Large quantities of sodic and alkaline bauxite residue are produced globally as a by-product from alumina refineries. Ecological stoichiometry of key elements [nitrogen (N) and phosphorus (P)] plays a critical role in establishing vegetation cover in bauxite residue sand (BRS). Here we examined how changes in soil chemical properties over time in rehabilitated sodic and alkaline BRS affected leaf N to P stoichiometry of native species used for rehabilitation. Both Ca and soil pH influenced the shifts in leaf N:P ratios of the study species as supported by consistently significant positive relationships (P < 0.001) between these soil indices and leaf N:P ratios. Shifts from N to P limitation were evident for N-fixing species, while N limitation was consistently experienced by non-N-fixing plant species. In older rehabilitated BRS embankments, soil and plant indices (Ca, Na, pH, EC, ESP and leaf N:P ratios) tended to align with those of the natural ecosystem, suggesting improved rehabilitation performance. These findings highlight that leaf N:P stoichiometry can effectively provide a meaningful assessment on understanding nutrient limitation and productivity of native species used for vegetating highly sodic and alkaline BRS, and is a crucial indicator for assessing ecological rehabilitation performance.

Isolation and Culture of Mouse Hepatocytes: Gender-Specific Gene Expression Responses to Chemical Treatments.

In this chapter, the isolation of primary mouse hepatocytes and their response to chemical treatment are described. We show that it is important to consider, in the experimental design, the sex of the animals to be used. We demonstrate this by measuring the effect of sex hormones or xenobiotics on the expression of flavin-containing monooxygenase 5 in cultures of primary hepatocytes isolated from male and female mice.

The phenotype of a knockout mouse identifies flavin-containing monooxygenase 5 (FMO5) as a regulator of metabolic ageing.

We report the production and metabolic phenotype of a mouse line in which the Fmo5 gene is disrupted. In comparison with wild-type (WT) mice, Fmo5(-/-) mice exhibit a lean phenotype, which is age-related, becoming apparent after 20 weeks of age. Despite greater food intake, Fmo5(-/-) mice weigh less, store less fat in white adipose tissue (WAT), have lower plasma glucose and cholesterol concentrations and enhanced whole-body energy expenditure, due mostly to increased resting energy expenditure, with no increase in physical activity. An increase in respiratory exchange ratio during the dark phase, the period in which the mice are active, indicates a switch from fat to carbohydrate oxidation. In comparison with WT mice, the rate of fatty acid oxidation in Fmo5(-/-) mice is higher in WAT, which would contribute to depletion of lipid stores in this tissue, and lower in skeletal muscle. Five proteins were down regulated in the liver of Fmo5(-/-) mice: aldolase B, ketohexokinase and cytosolic glycerol 3-phosphate dehydrogenase (GPD1) are involved in glucose or fructose metabolism and GPD1 also in production of glycerol 3-phosphate, a precursor of triglyceride biosynthesis; HMG-CoA synthase 1 is involved in cholesterol biosynthesis; and malic enzyme 1 catalyzes the oxidative decarboxylation of malate to pyruvate, in the process producing NADPH for use in lipid and cholesterol biosynthesis. Down regulation of these proteins provides a potential explanation for the reduced fat deposits and lower plasma cholesterol characteristic of Fmo5(-/-) mice. Our results indicate that disruption of the Fmo5 gene slows metabolic ageing via pleiotropic effects.

The phenotype of a flavin-containing monooyxgenase knockout mouse implicates the drug-metabolizing enzyme FMO1 as a novel regulator of energy balance.

Flavin-containing monooxygenases (FMOs) of mammals are thought to be involved exclusively in the metabolism of foreign chemicals. Here, we report the unexpected finding that mice lacking Fmos 1, 2 and 4 exhibit a lean phenotype and, despite similar food intake, weigh less and store less triglyceride in white adipose tissue (WAT) than wild-type mice. This is a consequence of enhanced whole-body energy expenditure, due mostly to increased resting energy expenditure (REE). This is fuelled, in part, by increased fatty acid ß-oxidation in skeletal muscle, which would contribute to depletion of lipid stores in WAT. The enhanced energy expenditure is attributed, in part, to an increased capacity for exercise. There is no evidence that the enhanced REE is due to increased adaptive thermogenesis; instead, our results are consistent with the operation in WAT of a futile energy cycle. In contrast to FMO2 and FMO4, FMO1 is highly expressed in metabolic tissues, including liver, kidney, WAT and BAT. This and other evidence implicates FMO1 as underlying the phenotype. The identification of a novel, previously unsuspected, role for FMO1 as a regulator of energy homeostasis establishes, for the first time, a role for a mammalian FMO in endogenous metabolism. Thus, FMO1 can no longer be considered to function exclusively as a xenobiotic-metabolizing enzyme. Consequently, chronic administration of drugs that are substrates for FMO1 would be expected to affect energy homeostasis, via competition for endogenous substrates, and, thus, have important implications for the general health of patients and their response to drug therapy.

Relationships between flavin-containing mono-oxygenase 3 (FMO3) genotype and trimethylaminuria phenotype in a Japanese population.

AIM: The aim of this study was to investigate relationships between flavin-containing mono-oxygenase 3 (FMO3) genotype and phenotype (conversion of odorous trimethylamine into non-odorous trimethylamine N-oxide) in a large Japanese cohort suffering from trimethylaminuria. METHODS: Urinary excretion of trimethylamine and trimethylamine N-oxide was determined for 102 volunteers with self-reporting symptoms of trimethylaminuria. For each we determined the sequence of the entire coding region, plus 1.3 kb of flanking intronic and 2.5 kb of the upstream region of the FMO3 gene. The affect of upstream variants on transcription was determined with a reporter gene assay. RESULTS: Seventy-eight subjects were diagnosed as suffering from trimethylaminuria, based on urinary excretion of <90% of total TMA as TMA N-oxide. Of these, 13 were classified as severe, 56 as moderate and nine as mild cases, excreting <43%, 48-70% and 73-83% of trimethylamine as trimethylamine N-oxide, respectively. Twenty-seven mutations were identified in FMO3, 15 in the coding region, of which eight abolish or severely impair FMO3 activity (Pro70Leu, Cys197fsX, Thr201Lys, Arg205Cys, Met260Val, Trp388Ter, Gln470Ter and Arg500Ter), and 12 in the upstream region. The mutations segregate into 19 haplotypes, including four different combinations of upstream mutations, each of which reduces transcriptional activity in comparison with the ancestral upstream sequence of FMO3. CONCLUSIONS: Comparisons of genotype and phenotype reveal that severe trimethylaminuria is caused by loss of function mutations in FMO3. For moderate and mild cases the situation is more complex, with most resulting from factors other than FMO3 genotype. Our results have implications for the diagnosis and management of the disorder.