P-cresol and Indoxyl Sulfate Impair Osteogenic Differentiation by Triggering Mesenchymal Stem Cell Senescence.

Chronic kidney disease (CKD) patients obtained high levels of uremic toxins progressively develop several complications including bone fractures. Protein-bound uremic toxins especially p-cresol and indoxyl sulfate are hardly eliminated due to their high molecular weight. Thus, the abnormality of bone in CKD patient could be potentially resulted from the accumulation of uremic toxins. To determine whether protein-bound uremic toxins have an impact on osteogenesis, mesenchymal stem cells were treated with either p-cresol or indoxyl sulfate under in vitro osteogenic differentiation. The effects of uremic toxins on MSC-osteoblastic differentiation were investigated by evaluation of bone phenotype. The results demonstrated that p-cresol and indoxyl sulfate down-regulated the transcriptional level of collagen type I, deceased alkaline phosphatase activity, and impaired mineralization of MSC-osteoblastic cells. Furthermore, p-cresol and indoxyl sulfate gradually increased senescence-associated beta-galactosidase positive cells while upregulated the expression of p21 which participate in senescent process. Our findings clearly revealed that the presence of uremic toxins dose-dependently influenced a gradual deterioration of osteogenesis. The effects partially mediate through the activation of senescence-associated gene lead to the impairment of osteogenesis. Therefore, the management of cellular senescence triggered by uremic toxins could be considered as an alternative therapeutic approach to prevent bone abnormality in CKD patients.

Non-targeted metabolomics analyses by mass spectrometry to explore metabolic stress after six training weeks in high level swimmers.

The objective was to compare the metabolic responses of high-level national swimmers to threshold or polarised training. 22 swimmers (n = 12 males and 10 females) participated in a 28-week cross-over intervention study consisting of 2 × 6 period weeks of training. Swimmers were assigned randomly to either training group for the first period: polarised (POL) (81% in energetic zone 1: blood lactate [La]b ≤ 2 mmol.L-1; 4% in zone 2: 2 mmol.L-1 <[La]b ≤ 4 mmol.L-1; 15% in zone 3: [La]b > 4 mmol.L-1) or threshold (THR) (65%/25%/10%). Before and after each training period, urine samples were collected for non-targeted metabolomics analysis. Mixed model analysis was performed on metabolomics data including fatigue class factors and/or training and/or interaction. Ion intensities of 6-keto-decanoylcarnitine (+31%), pregnanediol-3-glucuronide (+81%), P-cresol sulphate (+18%) were higher in the threshold group (P < 0.05) indicating higher glycogenic depletion and inflammation without alteration of the neuroendocrine stress axis. 4-phenylbutanic acid sulphate was 200% higher in less fatigued swimmers (P < 0.01) linking the anti-inflammatory activity at the cell membrane level to the subjective perception of fatigue. This research suggests the importance of replenishing glycogen stores and reducing inflammation during high thresholds training loads.

Longitudinal metabolic and gut bacterial profiling of pregnant women with previous bariatric surgery.

OBJECTIVE: Due to the global increase in obesity rates and success of bariatric surgery in weight reduction, an increasing number of women now present pregnant with a previous bariatric procedure. This study investigates the extent of bariatric-associated metabolic and gut microbial alterations during pregnancy and their impact on fetal development. DESIGN: A parallel metabonomic (molecular phenotyping based on proton nuclear magnetic resonance spectroscopy) and gut bacterial (16S ribosomal RNA gene amplicon sequencing) profiling approach was used to determine maternal longitudinal phenotypes associated with malabsorptive/mixed (n=25) or restrictive (n=16) procedures, compared with women with similar early pregnancy body mass index but without bariatric surgery (n=70). Metabolic profiles of offspring at birth were also analysed. RESULTS: Previous malabsorptive, but not restrictive, procedures induced significant changes in maternal metabolic pathways involving branched-chain and aromatic amino acids with decreased circulation of leucine, isoleucine and isobutyrate, increased excretion of microbial-associated metabolites of protein putrefaction (phenylacetlyglutamine, p-cresol sulfate, indoxyl sulfate and p-hydroxyphenylacetate), and a shift in the gut microbiota. The urinary concentration of phenylacetylglutamine was significantly elevated in malabsorptive patients relative to controls (p=0.001) and was also elevated in urine of neonates born from these mothers (p=0.021). Furthermore, the maternal metabolic changes induced by malabsorptive surgery were associated with reduced maternal insulin resistance and fetal/birth weight. CONCLUSION: Metabolism is altered in pregnant women with a previous malabsorptive bariatric surgery. These alterations may be beneficial for maternal outcomes, but the effect of elevated levels of phenolic and indolic compounds on fetal and infant health should be investigated further.

Bowel Habits and the Association With Uremic Toxins in Non-Dialysis-Dependent Chronic Kidney Disease Patients.

OBJECTIVE: The aim of this study is to evaluate the association between bowel habits and microbial-derived uremic toxins p-cresyl sulfate (PCS) and indoxyl sulfate (IS) in patients with non-dialysis-dependent chronic kidney disease (NDD-CKD). DESIGN AND METHODS: This is a cross-sectional analysis including 43 nondiabetic NDD-CKD patients (58% men; 59.0 ± 13.5 years; estimated glomerular filtration rate, 21.3 ± 7.9 mL/min/1.73 m2). Bowel habit was assessed by the Bristol Stool Scale (BSS <3, characterized by hard consistency of stools and/or low frequency of evacuation and BSS ≥3, representing a more regular bowel habit) and by the Rome III criteria. PCS and IS (serum, free and total; urinary, total) were determined by high-performance liquid chromatography. Dietary intake was assessed by the 3-day food records. RESULTS: The frequency of constipation assessed by BSS and Rome III criteria was 33% (n = 14/43) and 35% (n = 15/43), respectively. The BSS <3 exhibited higher PCS, independent of renal function and dietary protein-fiber ratio (ß [95% confidence interval {CI}]: serum, total PCS = 1.54 [1.06-2.23], P = .02; serum free PCS = 1.40 [1.00-1.97], P = .05; urinary PCS = 1.78 [1.10-2.90], P < .02). According to the Rome III criteria, a tendency for a higher serum total PCS (ß [95% CI]: 1.39 [0.95-2.03 µmol/L], P = .09) and a significantly higher urinary PCS (ß [95% CI]: 1.80 [1.11-2.94 µmol/24 h], P = .02) was found in constipated participants. No effect of a compromised bowel habit (Rome III criteria or BSS) was found on IS. CONCLUSION: Constipation may lead to production of PCS in nondiabetic NDD-CKD patients.

Magnetic organic porous polymer as a solid-phase extraction adsorbent for enrichment and quantitation of gastric cancer biomarkers (P-cresol and 4-hydroxybenzoic acid) in urine samples by UPLC.

A novel magnetic organic porous polymer (denoted as Fe3O4@PC-POP) was developed for magnetic solid-phase extraction (MSPE) of two gastric cancer biomarkers (P-cresol and 4-hydroxybenzoic acid) from urine samples prior to high-performance liquid chromatographic analysis. The adsorbent was characterized by scanning electron microscope, transmission electron microscope, FTIR, powder X-ray diffraction, and other techniques. The result of dynamic light scattering shows that the particle size of the adsorbent is mainly distributed around 400 nm. Based on the design concept of the Fe3O4@PC-POP, the proposed material can effectively capture the target analytes through electrostatic and hydrophobic interaction mechanism. Furthermore, the enrichment conditions were optimized by the response surface method, and the method was utilized for the determination of P-cresol and 4-hydroxybenzoic acid in real urine samples from health and gastric cancer patients with high enrichment factors (34.8 times for P-cresol and 38.7 times for 4-hydroxybenzoic acid), low limit of detection (0.9-5.0 µg L-1), wide linear ranges (3.0-1000 µg L-1), satisfactory relative standard deviation (2.5%-8.5%), and apparent recoveries (85.3-112% for healthy people's and 86.0-112% for gastric cancer patients' urine samples). This study provides a guided principle for design of the versatile polymer with specific capturing of the target compounds from complex biological samples. Graphical abstract.

Multi-Compartment Profiling of Bacterial and Host Metabolites Identifies Intestinal Dysbiosis and Its Functional Consequences in the Critically Ill Child.

OBJECTIVES: Adverse physiology and antibiotic exposure devastate the intestinal microbiome in critical illness. Time and cost implications limit the immediate clinical potential of microbial sequencing to identify or treat intestinal dysbiosis. Here, we examined whether metabolic profiling is a feasible method of monitoring intestinal dysbiosis in critically ill children. DESIGN: Prospective multicenter cohort study. SETTING: Three U.K.-based PICUs. PATIENTS: Mechanically ventilated critically ill (n = 60) and age-matched healthy children (n = 55). INTERVENTIONS: Collection of urine and fecal samples in children admitted to the PICU. A single fecal and urine sample was collected in healthy controls. MEASUREMENTS AND MAIN RESULTS: Untargeted and targeted metabolic profiling using 1H-nuclear magnetic resonance spectroscopy and liquid chromatography-mass spectrometry or urine and fecal samples. This was integrated with analysis of fecal bacterial 16S ribosomal RNA profiles and clinical disease severity indicators. We observed separation of global urinary and fecal metabolic profiles in critically ill compared with healthy children. Urinary excretion of mammalian-microbial co-metabolites hippurate, 4-cresol sulphate, and formate were reduced in critical illness compared with healthy children. Reduced fecal excretion of short-chain fatty acids (including butyrate, propionate, and acetate) were observed in the patient cohort, demonstrating that these metabolites also distinguished between critical illness and health. Dysregulation of intestinal bile metabolism was evidenced by increased primary and reduced secondary fecal bile acid excretion. Fecal butyrate correlated with days free of intensive care at 30 days (r = 0.38; p = 0.03), while urinary formate correlated inversely with vasopressor requirement (r = -0.2; p = 0.037). CONCLUSIONS: Disruption to the functional activity of the intestinal microbiome may result in worsening organ failure in the critically ill child. Profiling of bacterial metabolites in fecal and urine samples may support identification and treatment of intestinal dysbiosis in critical illness.

Exploration of the Fecal Microbiota and Biomarker Discovery in Equine Grass Sickness.

Equine grass sickness (EGS) is a frequently fatal disease of horses, responsible for the death of 1 to 2% of the U.K. horse population annually. The etiology of this disease is currently uncharacterized, although there is evidence it is associated with Clostridium botulinum neurotoxin in the gut. Prevention is currently not possible, and ileal biopsy diagnosis is invasive. The aim of this study was to characterize the fecal microbiota and biofluid metabolic profiles of EGS horses, to further understand the mechanisms underlying this disease, and to identify metabolic biomarkers to aid in diagnosis. Urine, plasma, and feces were collected from horses with EGS, matched controls, and hospital controls. Sequencing the16S rRNA gene of the fecal bacterial population of the study horses found a severe dysbiosis in EGS horses, with an increase in Bacteroidetes and a decrease in Firmicutes bacteria. Metabolic profiling by 1H nuclear magnetic resonance spectroscopy found EGS to be associated with the lower urinary excretion of hippurate and 4-cresyl sulfate and higher excretion of O-acetyl carnitine and trimethylamine-N-oxide. The predictive ability of the complete urinary metabolic signature and using the four discriminatory urinary metabolites to classify horses by disease status was assessed using a second (test) set of horses. The urinary metabolome and a combination of the four candidate biomarkers showed promise in aiding the identification of horses with EGS. Characterization of the metabolic shifts associated with EGS offers the potential of a noninvasive test to aid premortem diagnosis.

Evaluation of pheromone-based kit: A noninvasive approach of estrus detection in buffalo.

In view of the silent nature of estrus in buffalo, a noninvasive assay kit has long been felt necessary for easy and effective estrus detection. This study was designed to detect estrus in buffalo using a kit formulated in our laboratory based on pheromone compound. Group I: Urine samples collected at estrus phase and group II: randomly collected urine samples were subjected to the test using the kit. No colour developed (i.e., positive reaction) in estrus urine after adding the kit solution. By contrast, pale and/or dark pink colour developed (i.e., negative reaction) in urine from the proestrus and diestrus buffaloes, respectively. Field evaluation of the kit in groups I and II revealed that 60.87% and 71.43% of urine samples were correctly identified as estrus and nonestrus (i.e., proestrus and diestrus), respectively. Therefore, the first of its kind estrus detection kit formulated based on urinary pheromone can as well be used as a simple device to detect estrus in buffalo.

Uremic Toxins Activates Na/K-ATPase Oxidant Amplification Loop Causing Phenotypic Changes in Adipocytes in In Vitro Models.

BACKGROUND: Oxidant stress plays a key role in the development of chronic kidney disease (CKD). Experimental CKD leads to accumulation of uremic toxins (UT) in the circulation resulting in increased ROS production, which in turn, is known to activate the Na/K-ATPase/ROS amplification loop. Studies in a murine model of obesity have shown that increased oxidative stress in plasma is due to increased ROS and cytokine production from dysfunctional adipocytes. Therefore, we hypothesized that adipocytes exposed to UTs will activate the Na/K-ATPase oxidant amplification loop causing redox imbalance and phenotypic alterations in adipocytes. We also aimed to demonstrate that the Na/K-ATPase signaling antagonist, pNaKtide, attenuates these pathophysiological consequences. METHODS: In the first set of experiments, 3T3-L1 murine pre-adipocytes were treated with varying concentrations of UTs, indoxyl sulfate (IS) (50, 100 and 250 µM) and p-cresol (50, 100 and 200 µM), with or without pNaKtide (0.7 µM) for five days in adipogenic media, followed by Oil Red O staining to study adipogenesis. RT-PCR analysis was performed to study expression of adipogenic, apoptotic and inflammatory markers, while DHE staining evaluated the superoxide levels in UT treated cells. In a second set of experiments, visceral fat was obtained from the West Virginian population. MSCs were isolated and cultured in adipogenic media for 14 days, which was treated with indoxyl sulfate (0, 25, 50 and 100 µM) with or without pNaKtide (1 µM). MSC-derived adipocytes were evaluated for morphological and molecular analysis of the above markers. RESULTS: Our results demonstrated that 3T3-L1 cells and MSCs-derived adipocytes, treated with UTs, exhibited a significant decrease in adipogenesis and apoptosis through activation of the Na/K-ATPase/ROS amplification loop. The treatment with pNaKtide in 3T3-L1 cells and MSC-derived adipocytes negated the effects of UTs and restored cellular redox in adipocytes. We noted a varying effect of pNaKtide, in adipocytes treated with UTs, on inflammatory markers, adipogenic marker and superoxide levels in 3T3-L1 cells and MSC-derived adipocytes. CONCLUSIONS: This study demonstrates for the first time that the Na/K-ATPase/ROS amplification loop activated by elevated levels of UTs has varying effect on phenotypic alterations in adipocytes in various in vitro models. Thus, we propose that, if proven in humans, inhibition of Na/K-ATPase amplification of oxidant stress in CKD patients may ultimately be a novel way to combat adipocyte dysfunction and metabolic imbalance in these patients.

New Analytical Framework for Verification of Biomarkers of Exposure to Chemicals Combining Human Biomonitoring and Water Fingerprinting.

Molecular epidemiology approaches in human biomonitoring are powerful tools that allow for verification of public exposure to chemical substances. Unfortunately, due to logistical difficulties and high cost, they tend to evaluate small study groups and as a result might not provide comprehensive large scale community-wide exposure data. Urban water fingerprinting provides a timely alternative to traditional approaches. It can revolutionize the human exposure studies as urban water represents collective community-wide exposure. Knowledge of characteristic biomarkers of exposure to specific chemicals is key to the successful application of water fingerprinting. This study aims to introduce a novel conceptual analytical framework for identification of biomarkers of public exposure to chemicals via combined human metabolism and urban water fingerprinting assay. This framework consists of the following steps: (1) in vitro HLM/S9 assay, (2) in vivo pooled urine assay, (3) in vivo wastewater fingerprinting assay, (4) analysis with HR-MSMS, (5) data processing, and (6) selection of biomarkers. The framework was applied and validated for PCMC (4-chloro-m-cresol), household derived antimicrobial agent with no known exposure and human metabolism data. Four new metabolites of PCMC (hydroxylated, sulfated/hydroxylated, sulfated PCMC, and glucuronidated PCMC) were identified using the in vitro HLM/S9 assay. But only one metabolite, sulfated PCMC, was confirmed in wastewater and in urine. Therefore, our study confirms that water fingerprinting is a promising tool for biomarker selection and that in vitro HLM/S9 studies alone, although informative, do not provide high accuracy results. Our work also confirms, for the first time, human internal exposure to PCMC.

Resistant starch alters gut microbiome and metabolomic profiles concurrent with amelioration of chronic kidney disease in rats.

Patients and animals with chronic kidney disease (CKD) exhibit profound alterations in the gut environment including shifts in microbial composition, increased fecal pH, and increased blood levels of gut microbe-derived metabolites (xenometabolites). The fermentable dietary fiber high amylose maize-resistant starch type 2 (HAMRS2) has been shown to alter the gut milieu and in CKD rat models leads to markedly improved kidney function. The aim of the present study was to identify specific cecal bacteria and cecal, blood, and urinary metabolites that associate with changes in kidney function to identify potential mechanisms involved with CKD amelioration in response to dietary resistant starch. Male Sprague-Dawley rats with adenine-induced CKD were fed a semipurified low-fiber diet or a high-fiber diet [59% (wt/wt) HAMRS2] for 3 wk (n = 9 rats/group). The cecal microbiome was characterized, and cecal contents, serum, and urine metabolites were analyzed. HAMRS2-fed rats displayed decreased cecal pH, decreased microbial diversity, and an increased Bacteroidetes-to-Firmicutes ratio. Several uremic retention solutes were altered in the cecal contents, serum, and urine, many of which had strong correlations with specific gut bacteria abundances, i.e., serum and urine indoxyl sulfate were reduced by 36% and 66%, respectively, in HAMRS2-fed rats and urine p-cresol was reduced by 47% in HAMRS2-fed rats. Outcomes from this study were coincident with improvements in kidney function indexes and amelioration of CKD outcomes previously reported for these rats, suggesting an important role for microbial-derived factors and gut microbe metabolism in regulating host kidney function.

Microbial-Derived Metabolites Reflect an Altered Intestinal Microbiota during Catch-Up Growth in Undernourished Neonatal Mice.

BACKGROUND: Protein-energy undernutrition during early development confers a lifelong increased risk of obesity-related metabolic disease. Mechanisms by which metabolic abnormalities persist despite catch-up growth are poorly understood. OBJECTIVE: We sought to determine whether abnormal metabolomic and intestinal microbiota profiles from undernourished neonatal mice remain altered during catch-up growth. METHODS: Male and female CD1 mouse pups were undernourished by timed separation from lactating dams for 4 h at 5 d of age, 8 h at 6 d of age, and 12 h/d from 7 to 15 d of age, then resumed ad libitum nursing, whereas controls fed uninterrupted. Both groups were weaned simultaneously to a standard unpurified diet. At 3 time points (0, 1, and 3 wk after ending feed deprivation), metabolites in urine, plasma, and stool were identified with the use of mass spectrometry, and fecal microbes were identified with the use of 16S metagenomic sequencing. RESULTS: Undernourished mice completely recovered deficits of 36% weight and 9% length by 3 wk of refeeding, at which time they had 1.4-fold higher plasma phenyllactate and 2.0-fold higher urinary p-cresol sulfate concentrations than did controls. Plasma serotonin concentrations in undernourished mice were 25% lower at 0 wk but 1.5-fold higher than in controls at 3 wk. Whereas most urine and plasma metabolites normalized with refeeding, 117 fecal metabolites remained altered at 3 wk, including multiple N-linked glycans. Microbiota profiles from undernourished mice also remained distinct, with lower mean proportions of Bacteroidetes (67% compared with 83%) and higher proportions of Firmicutes (26% compared with 16%). Abundances of the mucolytic organisms Akkermansia muciniphila and Mucispirillum schaedleri were altered at 0 and 1 wk. Whereas microbiota from undernourished mice at 0 wk contained 11% less community diversity (P = 0.015), refed mice at 3 wk harbored 1.2-fold greater diversity (P = 0.0006) than did controls. CONCLUSION: Microbial-derived metabolites and intestinal microbiota remain altered during catch-up growth in undernourished neonatal mice.

Resistant starch reduces colonic and urinary p-cresol in rats fed a tyrosine-supplemented diet, whereas konjac mannan does not.

The effect of resistant starch (RS) and konjac mannan (KM) to maintain and improve the large intestinal environment was compared. Wistar SPF rats were fed the following diets for 4 weeks: negative control diet (C diet), tyrosine-supplemented positive control diet (T diet), and luminacoid supplemented diets containing either high-molecular konjac mannan A (KMAT diet), low-molecular konjac mannan B (KMBT diet), high-amylose cornstarch (HAST diet), or heat-moisture-treated starch (HMTST diet). The luminacoid-fed group had an increased content of short-chain fatty acids in the cecum. HAS caused a significant decrease in p-cresol content in the cecum, whereas KM did not. Urinary p-cresol was reduced in the HAST group compared with the T group, but not the KM fed groups. Deterioration in the large intestinal environment was only improved completely in the HAST and HMTST groups, suggesting that RS is considerably more effective than KM in maintaining the large intestinal environment.

Evaluation and modeling of the impact of coexposures to VOC mixtures on urinary biomarkers.

CONTEXT: Urinary biomarkers are widely used among biomonitoring studies because of their ease of collection and nonintrusiveness. Chloroform and TEX (i.e., toluene, ethylbenzene, and m-xylene) are chemicals that are often found together because of common use. Although interactions occurring among TEX are well-known, no information exists on possible kinetic interactions between these chemicals and chloroform at the level of parent compound or urinary biomarkers. OBJECTIVE: The objective of this study was therefore to study the possible interactions between these compounds in human volunteers with special emphasis on the potential impact on urinary biomarkers. MATERIALS AND METHODS: Five male volunteers were exposed by inhalation for 6 h to single, binary, and quaternary mixtures that included chloroform. Exhaled air and blood samples were collected and analyzed for parent compound concentrations. Urinary biomarkers (o-cresol, mandelic, and m-methylhippuric acids) were quantified in urine samples. Published PBPK model for chloroform was used, and a Vmax of 3.4 mg/h/kg was optimized to provide a better fit with blood data. Adapted PBPK models from our previous study were used for parent compounds and urinary biomarkers for TEX. RESULTS: Binary exposures with chloroform resulted in no significant interactions. Experimental data for quaternary mixture exposures were well predicted by PBPK models using published description of competitive inhibition among TEX components. However, no significant interactions were observed at levels used in this study. CONCLUSION: PBPK models for urinary biomarkers proved to be a good tool in quantifying exposure to VOC.

Urinary p-cresol is elevated in young French children with autism spectrum disorder: a replication study.

The aromatic compound p-cresol (4-methylphenol) has been found elevated in the urines of Italian autistic children up to 8 years of age. The present study aims at replicating these initial findings in an ethnically distinct sample and at extending them by measuring also the three components of urinary p-cresol, namely p-cresylsulfate, p-cresylglucuronate and free p-cresol. Total urinary p-cresol, p-cresylsulfate and p-cresylglucuronate were significantly elevated in 33 French autism spectrum disorder (ASD) cases compared with 33 sex- and age-matched controls (p < 0.05). This increase was limited to ASD children aged ≤8 years (p < 0.01), and not older (p = 0.17). Urinary levels of p-cresol and p-cresylsulfate were associated with stereotypic, compulsive/repetitive behaviors (p < 0.05), although not with overall autism severity. These results confirm the elevation of urinary p-cresol in a sizable set of small autistic children and spur interest into biomarker roles for p-cresol and p-cresylsulfate in autism.

Cardiovascular disease relates to intestinal uptake of p-cresol in patients with chronic kidney disease.

BACKGROUND: Serum p-cresyl sulfate (PCS) associates with cardiovascular disease in patients with chronic kidney disease. PCS concentrations are determined by intestinal uptake of p-cresol, human metabolism to PCS and renal clearance. Whether intestinal uptake of p-cresol itself is directly associated with cardiovascular disease in patients with renal dysfunction has not been studied to date. METHODS: We performed a prospective study in patients with chronic kidney disease stage 1 - 5 (NCT00441623). Intestinal uptake of p-cresol, under steady state conditions, was estimated from 24 h urinary excretion of PCS. Primary endpoint was time to first cardiovascular event, i.e., cardiac death, myocardial infarction/ischemia, ventricular arrhythmia, cardiovascular surgery, ischemic stroke or symptomatic peripheral arterial disease. Statistical analysis was done using Kaplan-Meier estimates and Cox proportional hazard analyses. RESULTS: In a cohort of 200 patients, median 24 h urinary excretion of PCS amounted to 457.47 µmol (IQR 252.68 - 697.17). After a median follow-up of 52 months, 25 patients reached the primary endpoint (tertile 1/2/3: 5/6/14 events, log rank P 0.037). Higher urinary excretion of PCS was directly associated with cardiovascular events (univariate hazard ratio per 100 µmol increase: 1.112, P 0.002). In multivariate analysis, urinary excretion of PCS remained a predictor of cardiovascular events, independent of eGFR (hazard ratio 1.120, P 0.002). CONCLUSIONS: In patients with chronic kidney disease, intestinal uptake of p-cresol associates with cardiovascular disease independent of renal function. The intestinal generation and absorption of p-cresol may be therapeutic targets to reduce cardiovascular disease risk in patients with renal dysfunction.

Intrarenal metabolomic investigation of chronic kidney disease and its TGF-ß1 mechanism in induced-adenine rats using UPLC Q-TOF/HSMS/MS(E).

Chronic kidney disease (CKD) is becoming a worldwide public health problem. In this study, a kidney metabonomics method based on the ultra performance liquid chromatography/high-sensitivity mass spectrometry with MS(E) data collection technique was undertaken to explore the excretion pattern of low molecular mass metabolites in rat model of adenine-induced chronic renal failure (CRF). Coupled with blood biochemistry and kidney histopathology results, the significant difference in metabolic profiling between the adenine-induced CRF group and the control group by using pattern recognition analysis indicated that changes in global tissue metabolites were occurred. Some significantly changed metabolites like fatty acids, p-cresol sulfate, and indoxyl sulfate have been identified. The results showed that the most important CRF-related metabolites were polyunsaturated fatty acids, indoxyl sulfate, and p-cresyl sulfate. Indoxyl sulfate and p-cresyl sulfate (uremic toxins) were significantly increased in CRF rats. Indoxyl sulfate and p-cresyl sulfate stimulate progressive tubulointerstitial fibrosis by increasing the expression of transforming growth factor-ß1 (TGF-ß1). These biochemical changes in tissue metabolites are related to the perturbations of fatty acid metabolism and amino metabolism, which may be helpful to further understand the TGF-ß1 mechanisms of tubulointerstitial fibrosis. This work shows that the metabonomics method is a valuable tool for studying the essence of CKD.

Biomarkers of toluene exposure in rats: mercapturic acids versus traditional indicators (urinary hippuric acid and o-cresol and blood toluene).

1. Toluene (TOL) is a neurotoxic, ototoxic and reprotoxic solvent which is metabolized via the glutathione pathway, producing benzylmercapturic, o-, m- and p-toluylmercapturic acids (MAs). These metabolites could be useful as biomarkers of TOL exposure. 2. The aims of this study were (1) to provide data on MAs excretion in rat urine following TOL exposure by inhalation, (2) to compare them to data from traditional TOL biomarkers, i.e. TOL in blood (Tol-B), and urinary hippuric acid (HA) and o-cresol (oCre) and (3) to establish a relationship between these different indicators and the airborne TOL concentration (Tol-A). 3. Sprague-Dawley rats were exposed to a range of TOL concentrations. Blood and urine were collected and analyzed to determine biomarker levels. 4. Levels of the four MAs correlate strongly with Tol-A (comparable to the correlation with Tol-B). 5. MAs are thus clearly superior to oCre and HA as potential markers of exposure to TOL.

Consumption of breads containing in situ-produced arabinoxylan oligosaccharides alters gastrointestinal effects in healthy volunteers.

Arabinoxylan oligosaccharides (AXOS) are studied as food compounds with prebiotic potential. Here, the impact of consumption of breads with in situ-produced AXOS on intestinal fermentation and overall gastrointestinal characteristics was evaluated in a completely randomized, double-blind, controlled, cross-over study. Twenty-seven healthy volunteers consumed 180 g of wheat/rye bread with or without in situ-produced AXOS (WR(+) and WR(-), respectively) daily for 3 wk. Consumption of WR(+) corresponded to an AXOS intake of ~2.14 g/d. Refined wheat flour bread without AXOS (W(-)) (180 g/d) was provided during the 3-wk run-in and wash-out periods. At the end of each treatment period, participants collected urine for 48 h as well as a feces sample. Additionally, all participants completed a questionnaire about stool characteristics and gastrointestinal symptoms during the last week of each period. Urinary phenol and p-cresol excretions were significantly lower after WR(+) intake compared to WR(-). Consumption of WR(+) significantly increased fecal total SCFA concentrations compared to intake of W(-). The effect of WR(+) intake was most pronounced on butyrate, with levels 70% higher than after consumption of W(-) in the run-in or wash-out period. Consumption of WR(+) tended to selectively increase the fecal levels of bifidobacteria (P = 0.06) relative to consumption of W(-). Stool frequency increased significantly after intake of WR(+) compared to WR(-). In conclusion, consumption of breads with in situ-produced AXOS may favorably modulate intestinal fermentation and overall gastrointestinal properties in healthy humans.

Effects of a wheat bran extract containing arabinoxylan oligosaccharides on gastrointestinal health parameters in healthy adult human volunteers: a double-blind, randomised, placebo-controlled, cross-over trial.

Wheat bran extract (WBE) is a food-grade soluble fibre preparation that is highly enriched in arabinoxylan oligosaccharides. In this placebo-controlled cross-over human intervention trial, tolerance and effects on colonic protein and carbohydrate fermentation were studied. After a 1-week run-in period, sixty-three healthy adult volunteers consumed 3, 10 and 0 g WBE/d for 3 weeks in a random order, with 2 weeks' washout between each treatment period. Fasting blood samples were collected at the end of the run-in period and at the end of each treatment period for analysis of haematological and clinical chemistry parameters. Additionally, subjects collected a stool sample for analysis of microbiota, SCFA and pH. A urine sample, collected over 48 h, was used for analysis of p-cresol and phenol content. Finally, the subjects completed questionnaires scoring occurrence frequency and distress severity of eighteen gastrointestinal symptoms. Urinary p-cresol excretion was significantly decreased after WBE consumption at 10 g/d. Faecal bifidobacteria levels were significantly increased after daily intake of 10 g WBE. Additionally, WBE intake at 10 g/d increased faecal SCFA concentrations and lowered faecal pH, indicating increased colonic fermentation of WBE into desired metabolites. At 10 g/d, WBE caused a mild increase in flatulence occurrence frequency and distress severity and a tendency for a mild decrease in constipation occurrence frequency. In conclusion, WBE is well tolerated at doses up to 10 g/d in healthy adults volunteers. Intake of 10 g WBE/d exerts beneficial effects on gut health parameters.