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.

Diagnoses after newly recorded abdominal pain in primary care: observational cohort study.

BACKGROUND: Non-acute abdominal pain in primary care is diagnostically challenging. AIM: To quantify the 1-year cumulative incidence of 35 non-malignant diagnoses and nine cancers in adults after newly recorded abdominal pain in primary care. DESIGN AND SETTING: Observational cohort study of 125 793 Clinical Practice Research Datalink GOLD records. METHOD: Participants, aged ≥40 years, had newly recorded abdominal pain between 1 January 2009 and 31 December 2013. Age- and sex-stratified 1-year cumulative incidence by diagnosis is reported. RESULTS: Most (>70%) participants had no pre-specified diagnoses after newly recorded abdominal pain. Non-malignant diagnoses were most common: upper gastrointestinal problems (gastro-oesophageal reflux disease, hiatus hernia, gastritis, oesophagitis, and gastric/duodenal ulcer) in males and urinary tract infection in females. The incidence of upper gastrointestinal problems plateaued at age ≥60 years (aged 40-59 years: males 4.9%, 95% confidence interval [CI] = 4.6 to 5.1, females 4.0%, 95% CI = 3.8 to 4.2; aged 60-69 years: males 5.8%, 95% CI = 5.4 to 6.2, females 5.4%, 95% CI = 5.1 to 5.8). Urinary tract infection incidence increased with age (aged 40-59 years: females 5.1%, 95% CI = 4.8 to 5.3, males 1.1%, 95% CI = 1.0 to 1.2; aged ≥70 years: females 8.0%, 95% CI = 7.6 to 8.4, males 3.3%, 95% CI = 3.0 to 3.6%). Diverticular disease incidence rose with age, plateauing at 4.2% (95% CI = 3.9 to 4.6) in males aged ≥60 years, increasing to 6.1% (95% CI = 5.8 to 6.4) in females aged ≥70 years. Irritable bowel syndrome incidence was higher in females (aged 40-59 years: 2.9%, 95% CI = 2.7 to 3.1) than males (aged 40-59 years: 2.1%, 95% CI = 1.9 to 2.3), decreasing with age to 1.3% (95% CI = 1.2 to 1.5) in females and 0.6% (95% CI = 0.5 to 0.8) in males aged ≥70 years. CONCLUSION: Although abdominal pain commonly remains unexplained, non-malignant diagnosis are more likely than cancer.

Intra-abdominal cancer risk with abdominal pain: a prospective cohort primary care study.

BACKGROUND: Quantifying cancer risk in primary care patients with abdominal pain informs diagnostic strategies. AIM: To quantify oesophagogastric, colorectal, liver, pancreatic, ovarian, uterine, kidney, and bladder cancer risks associated with newly reported abdominal pain with or without other symptoms, signs, or abnormal blood tests (that is, features) indicative of possible cancer. DESIGN AND SETTING: This was an observational prospective cohort study using Clinical Practice Research Datalink records with English cancer registry linkage. METHOD: The authors studied 125 793 patients aged ≥40 years with newly reported abdominal pain in primary care between 1 January 2009 and 31 December 2013. The 1-year cumulative incidence of cancer, and the composite 1-year cumulative incidence of cancers with shared additional features, stratified by age and sex are reported. RESULTS: With abdominal pain, overall risk was greater in men and increased with age, reaching 3.4% (95% confidence interval [CI] = 3.0 to 3.7, predominantly colorectal cancer 1.9%, 95% CI = 1.6 to 2.1) in men ≥70 years, compared with their expected incidence of 0.88% (95% CI = 0.87 to 0.89). Additional features increased cancer risk; for example, for men, colorectal or pancreatic cancer risk with abdominal pain plus diarrhoea at 60-69 years of age was 3.1% (95% CI = 1.9 to 4.9) predominantly colorectal cancer (2.2%, 95% CI = 1.2 to 3.8). CONCLUSION: Abdominal pain increases intra-abdominal cancer risk nearly fourfold in men aged ≥70 years, exceeding the 3% threshold warranting investigation. This threshold is surpassed for the >60 years age group only with additional features. These results will help direct appropriate referral and testing strategies for patients based on their demographic profile and reporting features. The authors suggest non-invasive strategies first, such as faecal immunochemical testing, with safety-netting in a shared decision-making framework.

Recognizing sinonasal cancer in primary care: a matched case-control study using electronic records.

BACKGROUND: Cancers of the nasopharynx, nasal cavity, and accessory sinuses ("sinonasal") are rare in England, with around 750 patients diagnosed annually. There are no specific National Institute for Health and Care Excellence (NICE) referral guidelines for these cancers and no primary care research published. OBJECTIVE: To identify and quantify clinical features of sinonasal cancer in UK primary care patients. METHODS: This matched case-control study used UK Clinical Practice Research Datalink (CPRD) data. Patients were aged ≥40 years with a diagnosis of sinonasal cancer between January 1, 2000 and December 31, 2009 and had consulted their GP in the year before diagnosis. Clinical features of sinonasal cancer were analysed using conditional logistic regression. Positive predictive values (PPVs) for single and combined features were calculated. RESULTS: In total, 155 cases and 697 controls were studied. Nine symptoms and one abnormal investigation were significantly associated with the cancer: nasal mass; odds ratio, 95 (95% confidence interval 7.0, 1315, P = 0.001); head and neck lumps, 68 (12, 387, P < 0.001); epistaxis, 17 (3.9, 70, P < 0.001); rhinorrhoea, 14 (4.6, 44, P < 0.001); visual disturbance, 12 (2.2, 67, P = 0.004); sinusitis, 7.3 (2.2, 25, P = 0.001); sore throat, 6.0 (2.0, 18, P = 0.001); otalgia, 5.4 (1.6, 18, P = 0.007); headache, 3.6 (1.4, 9.5, P = 0.01); raised white cell count, 8.5 (2.8, 27, P < 0.001). Combined PPVs for epistaxis/rhinorrhoea, epistaxis/sinusitis, and rhinorrhoea/sinusitis were 0.62%. CONCLUSION: This is the first primary care study identifying epistaxis, sinusitis, and rhinorrhoea as part of the clinical prodrome of sinonasal cancer. Although no PPVs meet the 3% NICE referral threshold, these results may help clinicians identify who warrants safety-netting and possible specialist referral, potentially reducing the number of advanced-stage diagnoses of sinonasal cancer.

Sinonasal cancer occurs in the back of the nose or in the sinuses. It is rare in the United Kingdom, with most cases being diagnosed at an advanced stage. Delayed presentation and non-specific symptoms often lead to diagnosis at a later stage, with consequently poorer survival outcomes. Currently, there is no research describing the symptoms presented by these patients to their general practitioner (GP), nor referral guidelines for primary healthcare professionals. The aim of this study was to detect the symptoms of patients aged ≥40 years, diagnosed with sinonasal cancer in primary care. Three symptoms in the year before their diagnosis were linked with sinonasal cancer: nosebleeds, runny nose, and sinusitis. These symptoms may help GPs to identify possible sinonasal cancer patients earlier, though each symptom was low-risk on its own.

Quantifying the impact of pre-existing conditions on the stage of oesophagogastric cancer at diagnosis: a primary care cohort study using electronic medical records.

BACKGROUND: Pre-existing conditions interfere with cancer diagnosis by offering diagnostic alternatives, competing for clinical attention or through patient surveillance. OBJECTIVE: To investigate associations between oesophagogastric cancer stage and pre-existing conditions. METHODS: Retrospective cohort study using Clinical Practice Research Datalink (CPRD) data, with English cancer registry linkage. Participants aged ≥40 years had consulted primary care in the year before their incident diagnosis of oesophagogastric cancer in 01/01/2010-31/12/2015. CPRD records pre-diagnosis were searched for codes denoting clinical features of oesophagogastric cancer and for pre-existing conditions, including those providing plausible diagnostic alternatives for those features. Logistic regression analysed associations between stage and multimorbidity (≥2 conditions; reference category: no multimorbidity) and having 'diagnostic alternative(s)', controlling for age, sex, deprivation and cancer site. RESULTS: Of 2444 participants provided, 695 (28%) were excluded for missing stage, leaving 1749 for analysis (1265/1749, 72.3% had advanced-stage disease). Multimorbidity was associated with stage [odds ratio 0.63, 95% confidence interval (CI) 0.47-0.85, P = 0.002], with moderate evidence of an interaction term with sex (1.76, 1.08-2.86, P = 0.024). There was no association between alternative explanations and stage (odds ratio 1.18, 95% CI 0.87-1.60, P = 0.278). CONCLUSIONS: In men, multimorbidity is associated with a reduced chance of advanced-stage oesophagogastric cancer, to levels seen collectively for women.

Diagnosing cancer is complicated by existing medical conditions. Diagnosis may be delayed if conditions explain cancer symptoms, or dominate appointments. Diagnosis may be quicker if conditions increase doctor­patient contact. We studied the association between existing illness and stage (early or advanced) of diagnosis with cancer of the stomach or gullet. We studied the primary-care records of patients aged ≥40 years, diagnosed in 01/01/2010­31/12/2015, and got stage from English cancer registry data. We searched the primary-care records for cancer symptoms (e.g. difficulty swallowing), and for 27 conditions that were common or explained cancer symptoms (e.g. difficulty swallowing following a stroke). We analysed cancer stage, looking at age, sex, multimorbidity (two or more conditions) and explanations for symptoms. We studied 1749 patients, of whom 1265 (72.3%) had advanced-stage cancer. The chance of advanced stage was similar in women with (71%, 95% CI 66­75%) or without (69%, 62­76%) multimorbidity. It was lower for men with (70%, 67­74%) than without (79%, 75­83%) multimorbidity. Stage of cancer was not affected by having explanations for cancer symptoms. In summary, for men, multimorbidity is associated with a reduced chance of advanced-stage cancer of the stomach or gullet to levels seen collectively for women.

Microcytosis as a risk marker of cancer in primary care: a cohort study using electronic patient records.

BACKGROUND: Microcytosis (smaller than normal red blood cells) has previously been identified as a possible early risk marker for some cancers. However, the role of microcytosis across all cancers has not been fully investigated. AIM: To examine cancer incidence in a cohort of patients with microcytosis, with and without accompanying anaemia. DESIGN AND SETTING: Cohort study of patients aged ≥40 years using UK primary care electronic patient records. METHOD: The 1-year cancer incidence was compared between cohorts of patients with a mean red cell volume of <85 femtolitres (fL) (low) or 85-101 fL (normal). Further analyses examined sex, age group, cancer site, and haemoglobin values. RESULTS: Of 12 289 patients with microcytosis, 497 had a new cancer diagnosis within 1 year (4.0%, 95% confidence interval [CI] = 3.7 to 4.4), compared with 1465 of 73 150 without microcytosis (2.0%, CI = 1.9 to 2.1). In males, 298 out of 4800 with microcytosis were diagnosed with cancer (6.2%, CI = 5.5 to 6.9), compared with 940 out of 34 653 without (2.7%, CI = 2.5 to 2.9). In females with microcytosis, 199 out of 7489 were diagnosed with cancer (2.7%, CI = 2.3 to 3.1), compared with 525 out of 38 497 without (1.4%, CI = 1.3 to 1.5). In patients with microcytosis but normal haemoglobin, 86 out of 2637 males (3.3%, CI = 2.6 to 4.0) and 101 out of 5055 females (2.0%, CI = 1.6 to 2.4) were diagnosed with cancer. CONCLUSION: Microcytosis is a predictor of underlying cancer even if haemoglobin is normal. Although a benign explanation is more likely, clinicians in primary care should consider simple testing for cancer on encountering unexplained microcytosis, particularly in males.

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.

Recognising laryngeal cancer in primary care: a large case-control study using electronic records.

BACKGROUND: Over 1700 people are diagnosed with laryngeal cancer annually in England. Current National Institute for Health and Care Excellence (NICE) guidelines on referral for suspected laryngeal cancer were based on clinical consensus, in the absence of primary care studies. AIM: To identify and quantify the primary care features of laryngeal cancer. DESIGN AND SETTING: Matched case-control study of patients aged ≥40 years using data from the UK's Clinical Practice Research Datalink. METHOD: Clinical features of laryngeal cancer with which patients had presented to their GP in the year before diagnosis were identified and their association with cancer was assessed using conditional logistic regression. Positive predictive values (PPVs) for each clinical feature were calculated for the consulting population aged >60 years. RESULTS: In total, 806 patients diagnosed with laryngeal cancer between 2000 and 2009 were studied, together with 3559 age-, sex-, and practice-matched controls. Ten features were significantly associated with laryngeal cancer: hoarseness odds ratio [OR] 904 (95% confidence interval [CI] = 277 to 2945); sore throat, first attendance OR 6.2 (95% CI = 3.7 to 10); sore throat, re-attendance OR 7.7 (95% CI = 2.6 to 23); dysphagia OR 6.5 (95% CI = 2.7 to 16); otalgia OR 5.0 (95% CI = 1.9 to 13); dyspnoea, re-attendance OR 4.7 (95% CI = 1.9 to 12); mouth symptoms OR 4.7 (95% CI = 1.8 to 12); recurrent chest infection OR 4.5 (95% CI = 2.4 to 8.5); insomnia OR 2.7 (95% CI = 1.3 to 5.6); and raised inflammatory markers OR 2.5 (95% CI = 1.5 to 4.1). All P-values were <0.01. Hoarseness had the highest individual PPV of 2.7%. Symptom combinations currently not included in NICE guidance were sore throat plus either dysphagia, dyspnoea, or otalgia, for which PPVs were >5%. CONCLUSION: These results expand current NICE guidance by identifying new symptom combinations that are associated with laryngeal cancer; they may help GPs to select more appropriate patients for referral.

Selection of men for investigation of possible testicular cancer in primary care: a large case-control study using electronic patient records.

BACKGROUND: Testicular cancer incidence has risen over the last two decades and is expected to continue to rise. There are no primary care studies on the clinical features of testicular cancer, with recent National Institute for Health and Care Excellence (NICE) guidance based solely upon clinical consensus. AIM: To identify clinical features of testicular cancer and to quantify their risk in primary care patients, with the aim of improving the selection of patients for investigation. DESIGN AND SETTING: A matched case-control study in males aged ≥17 years, using Clinical Practice Research Datalink records. METHOD: Putative clinical features of testicular cancer were identified and analysed using conditional logistic regression. Positive predictive values (PPVs) were calculated for those aged <50 years. RESULTS: In all, 1398 cases were available, diagnosed between 2000 and 2012, with 4956 age-, sex-, and practice-matched controls. Nine features were independently associated with testicular cancer, the top three being testicular swelling (odds ratio [OR] 280, 95% confidence interval [CI] = 110 to 690), testicular lump (OR 270, 95% CI = 100 to 740), and scrotal swelling (OR 170, 95% CI = 35 to 800). The highest PPV for 17-49-year-olds was testicular lump, at 2.5% (95% CI = 1.1 to 5.6). Combining testicular lump with testicular swelling or testicular pain produced PPVs of 17% and 10%, respectively. CONCLUSION: Testicular enlargement carries a risk of cancer of 2.5% - close to the current 3% threshold in UK referral guidance. Contrary to traditional teaching, painful testicular enlargement may signify cancer. Some initial hydrocele diagnoses appear to be wrong, with missed cancers, suggesting an ultrasound may be useful when a hydrocele diagnosis is uncertain. These results support the existing NICE guidelines, and help to characterise when an ultrasound should be considered in symptomatic men.

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.

Clinical relevance of thrombocytosis in primary care: a prospective cohort study of cancer incidence using English electronic medical records and cancer registry data.

BACKGROUND: Thrombocytosis (raised platelet count) is an emerging risk marker of cancer, but the association has not been fully explored in a primary care context. AIM: To examine the incidence of cancer in a cohort of patients with thrombocytosis, to determine how clinically useful this risk marker could be in predicting an underlying malignancy. DESIGN AND SETTING: A prospective cohort study using Clinical Practice Research Datalink data from 2000 to 2013. METHOD: The 1-year incidence of cancer was compared between two cohorts: 40 000 patients aged ≥40 years with a platelet count of >400 × 109/L (thrombocytosis) and 10 000 matched patients with a normal platelet count. Sub-analyses examined the risk with change in platelet count, sex, age, and different cancer sites. RESULTS: A total of 1098 out of 9435 males with thrombocytosis were diagnosed with cancer (11.6%; 95% confidence interval [CI] = 11.0 to 12.3), compared with 106 of 2599 males without thrombocytosis (4.1%; 95% CI = 3.4 to 4.9). A total of 1355 out of 21 826 females with thrombocytosis developed cancer (6.2%; 95% CI = 5.9 to 6.5), compared with 119 of 5370 females without (2.2%; 95% CI = 1.8 to 2.6). The risk of cancer increased to 18.1% (95% CI = 15.9 to 20.5) for males and 10.1% (95% CI = 9.0 to 11.3) for females, when a second raised platelet count was recorded within 6 months. Lung and colorectal cancer were more commonly diagnosed with thrombocytosis. One-third of patients with thrombocytosis and lung or colorectal cancer had no other symptoms indicative of malignancy. CONCLUSION: Thrombocytosis is a risk marker of cancer in adults; 11.6% and 6.2% cancer incidence in males and females, respectively, is worthy of further investigation for underlying malignancy. These figures well exceed the National Institute for Health and Care Excellence-mandated risk threshold of 3% risk to warrant referral for suspected cancer.

Clinical features of bowel disease in patients aged <50 years in primary care: a large case-control study.

BACKGROUND: Incidences of colorectal cancer (CRC) and inflammatory bowel disease (IBD) are increasing in those aged <50 years. AIM: To identify and quantify clinical features in primary care of CRC/IBD in those aged <50 years. This study considered the two conditions together and aimed to determine which younger patients, presenting in primary care with symptoms, would benefit from investigation for potentially serious colorectal disease. DESIGN AND SETTING: Matched case-control study using primary care records from the Clinical Practice Research Datalink, UK. METHOD: Incident cases (aged <50 years) of CRC (n = 1661) and IBD (n = 9578) diagnosed between 2000 and 2013 were each matched with up to three controls (n = 3979 CRC; n = 22 947 IBD). Odds ratios (OR) and positive predictive values (PPV) were estimated for features of CRC/IBD in the year before diagnosis. RESULTS: Ten features were independently associated with CRC/IBD (all P<0.001): rectal bleeding, change in bowel habit, diarrhoea, raised inflammatory markers, thrombocytosis, abdominal pain, low mean cell volume (MCV), low haemoglobin, raised white cell count, and raised hepatic enzymes. PPVs were >3% for rectal bleeding with diarrhoea, thrombocytosis, low MCV, low haemoglobin or raised inflammatory markers; for change in bowel habit with low MCV, thrombocytosis or low haemoglobin; and for diarrhoea with thrombocytosis. CONCLUSION: This study quantified the risk of serious bowel disease in symptomatic patients aged <50 years in primary care. Rectal bleeding and change in bowel habit are strongly predictive of CRC/IBD when combined with abnormal haematology. The present findings help prioritise patients for colonoscopy where the diagnosis is not immediately apparent.

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.