Implementation of pure shift 1 H NMR in metabolic phenotyping for structural information recovery of biofluid metabolites with complex spin systems.

NMR spectroscopy is a mainstay of metabolic profiling approaches to investigation of physiological and pathological processes. The one-dimensional proton pulse sequences typically used in phenotyping large numbers of samples generate spectra that are rich in information but where metabolite identification is often compromised by peak overlap. Recently developed pure shift (PS) NMR spectroscopy, where all J-coupling multiplicities are removed from the spectra, has the potential to simplify the complex proton NMR spectra that arise from biosamples and hence to aid metabolite identification. Here we have evaluated two complementary approaches to spectral simplification: the HOBS (band-selective with real-time acquisition) and the PSYCHE (broadband with pseudo-2D interferogram acquisition) pulse sequences. We compare their relative sensitivities and robustness for deconvolving both urine and serum matrices. Both methods improve resolution of resonances ranging from doublets, triplets and quartets to more complex signals such as doublets of doublets and multiplets in highly overcrowded spectral regions. HOBS is the more sensitive method and takes less time to acquire in comparison with PSYCHE, but can introduce unavoidable artefacts from metabolites with strong couplings, whereas PSYCHE is more adaptable to these types of spin system, although at the expense of sensitivity. Both methods are robust and easy to implement. We also demonstrate that strong coupling artefacts contain latent connectivity information that can be used to enhance metabolite identification. Metabolite identification is a bottleneck in metabolic profiling studies. In the case of NMR, PS experiments can be included in metabolite identification workflows, providing additional capability for biomarker discovery.

Fluid intake, hydration status and body mass changes in U-15 judo athletes during a training day

Despite numerous studies related to dehydration there is still a lack of scientific literature presenting hydration status and fluid intake of judo athletes during different periods. Therefore, the aim of this study was to investigate, fluid intake, hydration status and body weight changes of young judo athletes during a typical day of training in preparation period. Twenty-two young judo athletes (age: 12 ± 0.7 y, experience: 3.5 ± 1.1) voluntarily participated in this study. Hydration status and weight were examined in the morning, before and immediately after the training. All athletes trained 90 min and they consumed fluids ad libitum during the exercise. According to morning urine specific gravity (USG) values, 81.2% of the athletes were dehydrated while only 18.8% of the athletes were euhydrated. Pre-training urine measurements showed that 63.64% of the athletes presented dehydration and 77.27% of the athletes completed the training in dehydrated condition despite fluid availability during the training. Mean body weight loss during training was -0.64 ± 0.66%. It can be concluded that young judo athletes presented high prevalence of dehydration as indicated by USG values. Most of the athletes were dehydrated during a typical training day and completed the training in more dehydrated conditions compared to pre training values despite ad libitum fluid intake. It is of great importance to evaluate hydration status of the athletes before training to refrain from common practice of fluid restriction for weight loss and adverse effects of a persistent state of fluid deficit on physical and health related state.

Estudo de estabilidade de anfetaminas e produtos de biotransformação de cocaína e tetraidrocanabinol em amostras de urina seca em papel (dried urine spots) / Stability study of amphetamines, biotransformation products of cocaine and tetrahydrocannabinol in dried urine spots

O número de pessoas utilizando substâncias ilícitas de forma recreativa aumenta a cada ano, chamando a atenção de estudiosos de diversas áreas do conhecimento. Com isso, a demanda de exames toxicológicos exigida para trabalhadores, vítimas de crimes e esportistas também tem crescido. A amostra biológica mais utilizada para análises toxicológicas continua sendo a urina, visto que sua obtenção é menos invasiva, possibilita coletar grande volume de amostra e pode-se detectar substâncias até dias após ter ocorrido a exposição ou consumo. Entretanto, estas amostras necessitam de um grande volume físico para serem armazenadas e transportadas aos laboratórios, devendo ser mantidas em temperatura baixa e controlada para conservação. Outro ponto a se considerar é a quantidade de amostra insuficientemente coletada, ou extravasamento do conteúdo, contaminando outras amostras e muitas vezes, inviabilizando a análise. Uma alternativa recente para tais problemas é utilizar a técnica chamada de dried urine spots (DUS), onde poucos microlitros de urina são colocados em um papel absorvente e secos sob temperatura ambiente, preservando de agentes degradantes os componentes presentes na urina. Assim, o objetivo deste trabalho é avaliar a estabilidade das substâncias do presente estudo em alta temperatura, temperatura ambiente e em temperaturas de 4°C e -20°C. Para este fim, foi necessário desenvolver, validar e aplicar métodos de extração e determinação de anfetaminas e produtos de biotransformação de cocaína e tetraidrocanabinol carboxílico (THCCOOH) em amostras dried urine spot, utilizando cromatografia líquida acoplada à espectrometria de massas. Os picos foram identificados por UPLC-ESI-MS/MS, com tempo total de 5 mins utilizando fase A- água, formiato de amônio e 0,1% ácido fórmico, e B- metanol: acetonitrila (6:4) + 0,1% de ácido fórmico. A extração foi feita utilizando acetonitrila: metanol: acetona (1:1:1) +ácido fórmico 0,1%. Não foi possível iniciar a validação de THCCOOH, visto uma possível complexação do analito com o papel. Para as outras substâncias, o método cromatográfico desenvolvido se mostrou eficiente e seletivo, com LOD e LOQ de 10 ng/mL para todos os analitos, sendo linear até 1000 ng/mL, atendeu as especificações de precisão e exatidão e carryover. As amostras permaneceram estáveis ao longo de 32 dias nas temperaturas estudadas, demonstrando a segurança em se utilizar a técnica de DUS para armazenamento e transporte de amostras biológicas dentro da faixa de temperatura do estudo até 32 dias

The number of people using illegal substances in a recreational way increases each year, drawing the attention of scholars from different areas of knowledge. As a result, the demand for workplaces drug tests, toxicological tests for victims of crimes and dopping has also grown. The biological sample most used for toxicological tests remains urine, since obtaining it is less invasive, it is possible to collect a large volume of sample and it is possible to detect substances up to days after exposure or consumption has occurred. However, these samples require a large physical volume to be stored and transported to the laboratories, and must be kept at a low temperature for conservation. Another point to consider is the amount of sample insufficiently collected, or leakage of the content, causing contamination of other samples and often making the analysis unfeasible. A recent alternative to such problems is to use "dried urine spots" (DUS), where few microliters of urine are placed on absorbent paper and dried at room temperature, preserving the components present in the urine from degrading agents. Thus, the objective of this work is to evaluate the stability of the substances in this study at high temperature, room temperature and at temperatures of 4°C and -20°C. For this purpose, it was necessary to develop, validate and apply methods of extraction and determination of amphetamines and biotransformation products of cocaine and carboxylic tetrahydrocannabinol (THCCOOH) in dried urine spot samples, using liquid chromatography coupled to mass spectrometry (LC-MS). The peaks were identified liquid chromatography coupled to a mass spectrometer (UPLC-ESI-MS/MS), with a total time of 5 mins using phase A- water, ammonium formate and 0.1% formic acid, and B- methanol: acetonitrile (6:4) + 0.1% formic acid. Extraction was done using acetonitrile: methanol: acetone (1:1:1) + 0.1% formic acid. It was not possible to perform the validation of THCCOOH, given a possible complexation of the analyte with the paper. To the others substances, the chromatographic method developed proved to be efficient and selective, with LOD and LOQ of 10 ng/mL for all analytes, being linear up to 1000 ng/mL, meeting the specifications of precision and accuracy and carryover. The samples remained stable for 32 days at the temperatures studied, demonstrating the safety of using the DUS technique for storage and transport of biological samples until 32 days on temperature range studied

A Method to Monitor the NAD+ Metabolome-From Mechanistic to Clinical Applications.

Nicotinamide adenine dinucleotide (NAD+) and its reduced form (NADH) are coenzymes employed in hundreds of metabolic reactions. NAD+ also serves as a substrate for enzymes such as sirtuins, poly(ADP-ribose) polymerases (PARPs) and ADP-ribosyl cyclases. Given the pivotal role of NAD(H) in health and disease, studying NAD+ metabolism has become essential to monitor genetic- and/or drug-induced perturbations related to metabolic status and diseases (such as ageing, cancer or obesity), and its possible therapies. Here, we present a strategy based on liquid chromatography-tandem mass spectrometry (LC-MS/MS), for the analysis of the NAD+ metabolome in biological samples. In this method, hydrophilic interaction chromatography (HILIC) was used to separate a total of 18 metabolites belonging to pathways leading to NAD+ biosynthesis, including precursors, intermediates and catabolites. As redox cofactors are known for their instability, a sample preparation procedure was developed to handle a variety of biological matrices: cell models, rodent tissues and biofluids, as well as human biofluids (urine, plasma, serum, whole blood). For clinical applications, quantitative LC-MS/MS for a subset of metabolites was demonstrated for the analysis of the human whole blood of nine volunteers. Using this developed workflow, our methodology allows studying NAD+ biology from mechanistic to clinical applications.

Short-term metabolic disruptions in urine of mouse models following exposure to low doses of oxygen ion radiation.

Molecular alterations as a result of exposure to low doses of high linear energy transfer (LET) radiation can have deleterious short- and long-term consequences on crew members embarking on long distance space missions. Oxygen ions (16O) are among the high LET charged particles that make up the radiation environment inside a vehicle in deep space. We used mass spectrometry-based metabolomics to characterize urinary metabolic profiles of male C57BL/6J mice exposed to a single dose of 0.1, 0.25 and 1.0 Gy of 16O (600 MeV/n) at 10 and 30 days post-exposure to delineate radiation-induced metabolic alterations. We recognized a significant down regulation of several classes of metabolites including cresols and tryptophan metabolites, ketoacids and their derivatives upon exposure to 0.1 and 0.25 Gy after 10 days. While some of these changes reverted to near normal by 30 days, some metabolites including p-Cresol sulfate, oxalosuccinic acid, and indoxylsulfate remained dysregulated at 30 days, suggesting long term prognosis on metabolism. Pathway analysis revealed a long-term dysregulation in multiple pathways including tryptophan and porphyrin metabolism. These results suggest that low doses of high-LET charged particle irradiation may have long-term implications on metabolic imbalance.

Maternal selenium deficiency in mice promotes sex-specific changes to urine flow and renal expression of mitochondrial proteins in adult offspring.

Selenium deficiency during pregnancy can impair fetal development and predispose offspring to thyroid dysfunction. Given that key selenoproteins are highly expressed in the kidney and that poor thyroid health can lead to kidney disease, it is likely that kidney function may be impaired in offspring of selenium-deficient mothers. This study utilized a mouse model of maternal selenium deficiency to investigate kidney protein glycation, mitochondrial adaptations, and urinary excretion in offspring. Female C57BL/6 mice were fed control (>190 µg selenium/kg) or low selenium (<50 µg selenium/kg) diets four weeks prior to mating, throughout gestation, and lactation. At postnatal day (PN) 170, offspring were placed in metabolic cages for 24 hr prior to tissue collection at PN180. Maternal selenium deficiency did not impact selenoprotein antioxidant activity, but increased advanced glycation end products in female kidneys. Male offspring had reduced renal Complex II and Complex IV protein levels and lower 24 hr urine flow. Although renal aquaporin 2 (Aqp2) and arginine vasopressin receptor 2 (Avpr2) mRNA were not altered by maternal selenium deficiency, a correlation between urine flow and plasma free T4 concentrations in male but not female offspring suggests that programed thyroid dysfunction may be mediating impaired urine flow. This study demonstrates that maternal selenium deficiency can lead to long-term deficits in kidney parameters that may be secondary to impaired thyroid dysfunction. Considering the significant burden of renal dysfunction as a comorbidity to metabolic diseases, improving maternal selenium intake in pregnancy may be one simple measure to prevent lifelong disease.

Beat the heat: Culex quinquefasciatus regulates its body temperature during blood feeding.

Mosquitoes are regarded as one of the most dangerous animals on earth. Because they are responsible for the spread of a wide range of both human and animal pathogens, research of the underlying mechanisms of their feeding behavior and physiology is critical. Among disease vector mosquitoes, Culex quinquefasciatus, a known carrier of West Nile virus and Western Equine Encephalitis, remains relatively understudied. As blood-sucking insects, adaptations (either at the molecular or physiological level) while feeding on warm blood are crucial to their survival, as overheating can result in death due to heat stress. Our research aims to determine how Cx. quinquefasciatus copes with the heat associated with warm blood meal ingestion and possibly uncover the adaptations this species uses to avoid thermal stress. Through the use of thermographic imaging, we analyzed the body temperature of Cx. quinquefasciatus while blood feeding. Infrared thermography has allowed us to identify a cooling strategy, evaporative cooling via the production of fluid droplets, and an overall low body temperature in comparison to the blood temperature during feeding. Understanding Cx. quinquefasciatus' adaptations and the strategies they employ to reduce their body temperature while blood feeding constitutes the first step towards discovering potential targets that could be used for their control.

Urine Metabolites Associated with the Dietary Approaches to Stop Hypertension (DASH) Diet: Results from the DASH-Sodium Trial.

SCOPE: Serum metabolomic markers of the Dietary Approaches to Stop Hypertension (DASH) diet are previously reported. In an independent study, the similarity of urine metabolomic markers are investigated. METHODS AND RESULTS: In the DASH-Sodium trial, participants are randomly assigned to the DASH diet or control diet, and received three sodium interventions (high, intermediate, low) within each randomized diet group in random order for 30 days each. Urine samples are collected at the end of each intervention period and analyzed for 938 metabolites. Two comparisons are conducted: 1) DASH-high sodium (n = 199) versus control-high sodium (n = 193), and 2) DASH-low sodium (n = 196) versus control-high sodium. Significant metabolites identified using multivariable linear regression are compared and the top 10 influential metabolites identified using partial least-squares discriminant analysis to the results from the DASH trial. Nine out of 10 predictive metabolites of the DASH-high sodium and DASH-low sodium diets are identical. Most candidate biomarkers from the DASH trial replicated. N-methylproline, chiro-inositol, stachydrine, and theobromine replicated as influential metabolites of DASH diets. CONCLUSIONS: Candidate biomarkers of the DASH diet identified in serum replicated in urine. Replicated influential metabolites are likely to be objective biomarkers of the DASH diet.

Non-targeted urine metabolomics and associations with prevalent and incident type 2 diabetes.

Better risk prediction and new molecular targets are key priorities in type 2 diabetes (T2D) research. Little is known about the role of the urine metabolome in predicting the risk of T2D. We aimed to use non-targeted urine metabolomics to discover biomarkers and improve risk prediction for T2D. Urine samples from two community cohorts of 1,424 adults were analyzed by ultra-performance liquid chromatography/mass spectrometry (UPLC-MS). In a discovery/replication design, three out of 62 annotated metabolites were associated with prevalent T2D, notably lower urine levels of 3-hydroxyundecanoyl-carnitine. In participants without diabetes at baseline, LASSO regression in the training set selected six metabolites that improved prediction of T2D beyond established risk factors risk over up to 12 years' follow-up in the test sample, from C-statistic 0.866 to 0.892. Our results in one of the largest non-targeted urinary metabolomics study to date demonstrate the role of the urine metabolome in identifying at-risk persons for T2D and suggest urine 3-hydroxyundecanoyl-carnitine as a biomarker candidate.

Specific Hypothalamic Neurons Required for Sensing Conspecific Male Cues Relevant to Inter-male Aggression.

The hypothalamus regulates innate social interactions, but how hypothalamic neurons transduce sex-related sensory signals emitted by conspecifics to trigger appropriate behaviors remains unclear. Here, we addressed this issue by identifying specific hypothalamic neurons required for sensing conspecific male cues relevant to inter-male aggression. By in vivo recording of neuronal activities in behaving mice, we showed that neurons expressing dopamine transporter (DAT+) in the ventral premammillary nucleus (PMv) of the hypothalamus responded to male urine cues in a vomeronasal organ (VNO)-dependent manner in naive males. Retrograde trans-synaptic tracing further revealed a specific group of neurons in the bed nucleus of the stria terminalis (BNST) that convey male-relevant signals from VNO to PMv. Inhibition of PMvDAT+ neurons abolished the preference for male urine cues and reduced inter-male attacks, while activation of these neurons promoted urine marking and aggression. Thus, PMvDAT+ neurons exemplify a hypothalamic node that transforms sex-related chemo-signals into recognition and behaviors.

Effects of Menstrual Cycle Phase on Fluid Regulation during Walking Exercise.

To elucidate the fluid regulation in different menstrual cycle phases during exercise. Sex hormones affect fluid regulation in different ways. Moreover, the renin angiotensin-aldosterone system is activated in the luteal phase in rest. However, there are limited studies on fluid regulation affected by such hormone excretion in the menstrual cycle during exercise, especially during a light walking exercise. A non-invasive method using urine samples to determine menstrual cycle phases was used, and the follicular and luteal phases were successfully confirmed in 10 participants (age, 21 ± 1 years; body mass index, 20.5 ± 2.1 kg/m2). The experimental exercise sessions consisted of 5-min standing and 15-min walking at 2 km/h on 15% slope (approximately 8.3°) on a treadmill. Each participant carried a backpack weighing 5% of her own weight, and performed three sessions of walking exercise. Urine aldosterone excretion was significantly higher in the luteal than in the follicular phase before and after walking (p < 0.05). Urinary excretion of aldosterone was five times higher in the luteal than in the follicular phase before and after walking exercise. Heart rates during walking, after rest, and after recovery were all significantly higher in the luteal than in the follicular phase (p < 0.05). The participants' ratings of perceived exertion during the first and third session of walking in the luteal phase was not higher than that at the follicular phase. The results of our study suggested that increased activity of the renin-angiotensin-aldosterone system in the luteal phase of the menstrual cycle might be further activated during exercise. This may increase the circulatory load, which is reflected as increased heart rate. These results suggested that premenopausal women may better take into account a possibility of an increased circulatory load in the luteal phase even when they perform light exercise.

Water transport mediated by murine urea transporters: implications for urine concentration mechanisms.

Urea transporters (UTs) facilitate urea diffusion across cell membranes and play an important role in the urinary concentration mechanisms in the kidney. Herein, we injected cRNAs encoding for c-Myc-tagged murine UT-B, UT-A2 or UT-A3 (versus water-injected control) in Lithobates oocytes and evaluated oocyte surface protein expression with biotinylation and immunoblotting, urea uptake using [14C] counts and water permeability (P f ) by video microscopy. Immunoblots of UT-injected oocyte membranes revealed bands with a molecular weight consistent with that of a UT monomer (34 kDa), and UT-injected oocytes displayed significantly increased and phloretin-sensitive urea uptake and P f when compared to day-matched control oocytes. Subtracting the water-injected urea uptake or P f values from those of UT-injected oocytes yielded UT-dependent values*. We demonstrate for the first time that UT-A2 and UT-A3 can transport water, and we confirm that UT-B is permeable to water. Moreover, the [14C] urea*/P f * ratios fell in the sequence mUT-B>mUT-A2>mUT-A3, indicating that UTs can exhibit selectivity to urea and/or water. It is likely that specific kidney regions with high levels of UTs will exhibit increased urea and/or water permeabilities, directly influencing urine concentration. Furthermore, UT-mediated water transport activity must be considered when developing UT-inhibitors as novel diuretics.This article has an associated First Person interview with the first author of the paper.

Deep Phenotyping of Urinary Leukocytes by Mass Cytometry Reveals a Leukocyte Signature for Early and Non-Invasive Prediction of Response to Treatment in Active Lupus Nephritis.

Non-invasive biomarkers are necessary for diagnosis and monitoring disease activity in lupus nephritis (LN) to circumvent risks and limitations of renal biopsies. To identify new non-invasive cellular biomarkers in the urine sediment of LN patients, which may reflect kidney inflammation and can be used to predict treatment outcome, we performed in-depth urinary immune cell profiling by mass cytometry. We established a mass cytometric workflow to comparatively analyze the cellular composition of urine and peripheral blood (PB) in 13 patients with systemic lupus erythematosus (SLE) with active, biopsy-proven proliferative LN. Clinical and laboratory data were collected at the time of sampling and 6 months after induction of therapy in order to evaluate the clinical response of each patient. Six patients with different acute inflammatory renal diseases were included as comparison group. Leukocyte phenotypes and composition differed significantly between urine and paired PB samples. In urine, neutrophils and monocytes/macrophages were identified as the most prominent cell populations comprising together about 30%-83% of nucleated cells, while T and B lymphocytes, eosinophils, and natural killer (NK) cells were detectable at frequencies of <10% each. The majority of urinary T cells showed phenotypical characteristics of activated effector memory T cells (EM) as indicated by the co-expression of CD38 and CD69 - a phenotype that was not detectable in PB. Kidney inflammation was also reflected by tissue-imprinted macrophages, which phenotypically differed from PB monocytes by an increased expression of HLA-DR and CD11c. The presence of activated urinary T cells and macrophages could be used for differential diagnosis of proliferative LN forms and other renal pathologies. Most interestingly, the amount of EM in the urine sediment could be used as a biomarker to stratify LN patients in terms of response to induction therapy. Deep immunophenotypic profiling of urinary cells in LN allowed us to identify a signature of activated T cells and macrophages, which appear to reflect leukocytic infiltrates in the kidney. This explorative study has not only confirmed but also extended the knowledge about urinary cells as a future non-invasive biomarker platform for diagnosis and precision medicine in inflammatory renal diseases.

Loading-induced antitumor capability of murine and human urine.

While urine has been considered as a useful bio-fluid for health monitoring, its dynamic changes to physical activity are not well understood. We examined urine's possible antitumor capability in response to medium-level, loading-driven physical activity. Urine was collected from mice subjected to 5-minute skeletal loading and human individuals before and after 30-minute step aerobics. Six cancer cell lines (breast, prostate, and pancreas) and a mouse model of the mammary tumor were employed to evaluate the effect of urine. Compared to urine collected prior to loading, urine collected post-activity decreased the cellular viability, proliferation, migration, and invasion of tumor cells, as well as tumor weight in the mammary fat pad. Detection of urinary volatile organic compounds and ELISA assays showed that the loading-conditioned urine reduced cholesterol and elevated dopamine and melatonin. Immunohistochemical fluorescent images presented upregulation of the rate-limiting enzymes for the production of dopamine and melatonin in the brain. Molecular analysis revealed that the antitumor effect was linked to the reduction in molecular vinculin-linked molecular force as well as the downregulation of the Lrp5-CSF1-CD105 regulatory axis. Notably, the survival rate for the high expression levels of Lrp5, CSF1, and CD105 in tumor tissues was significantly lowered in the Cancer Genome Atlas database. Collectively, this study revealed that 5- or 10-minute loading-driven physical activity was sufficient to induce the striking antitumor effect by activating the neuronal signaling and repressing cholesterol synthesis. The result supported the dual role of loading-conditioned urine as a potential tumor suppressor and a source of diagnostic biomarkers.

Urine as a Main Effector in Urological Tissue Engineering-A Double-Edged Sword.

In order to reconstruct injured urinary tract tissues, biodegradable scaffolds with autologous seeded cells are explored in this work. However, when cells are obtained via biopsy from individuals who have damaged organs due to infection, congenital disorders, or cancer, this can result in unhealthy engineered cells and donor site morbidity. Thus, neo-organ construction through an alternative cell source might be useful. Significant advancements in the isolation and utilization of urine-derived stem cells have provided opportunities for this less invasive, limitless, and versatile source of cells to be employed in urologic tissue-engineered replacement. These cells have a high potential to differentiate into urothelial and smooth muscle cells. However, urinary tract reconstruction via tissue engineering is peculiar as it takes place in a milieu of urine that imposes certain risks on the implanted cells and scaffolds as a result of the highly cytotoxic nature of urine and its detrimental effect on both growth and differentiation of these cells. Both of these projections should be tackled thoughtfully when designing a suitable approach for repairing urinary tract defects and applying the needful precautions is vital.

The Urothelium: Life in a Liquid Environment.

The urothelium, which lines the renal pelvis, ureters, urinary bladder, and proximal urethra, forms a high-resistance but adaptable barrier that surveils its mechanochemical environment and communicates changes to underlying tissues including afferent nerve fibers and the smooth muscle. The goal of this review is to summarize new insights into urothelial biology and function that have occurred in the past decade. After familiarizing the reader with key aspects of urothelial histology, we describe new insights into urothelial development and regeneration. This is followed by an extended discussion of urothelial barrier function, including information about the roles of the glycocalyx, ion and water transport, tight junctions, and the cellular and tissue shape changes and other adaptations that accompany expansion and contraction of the lower urinary tract. We also explore evidence that the urothelium can alter the water and solute composition of urine during normal physiology and in response to overdistension. We complete the review by providing an overview of our current knowledge about the urothelial environment, discussing the sensor and transducer functions of the urothelium, exploring the role of circadian rhythms in urothelial gene expression, and describing novel research tools that are likely to further advance our understanding of urothelial biology.

Effects of preventing a temperature decrease during winter on reproduction of male small-egged Kajika, Cottus pollux SE.

To investigate the effects of preventing temperature decrease on the reproductive activity of the male cold-water teleost, Cottus pollux SE, testicular development, serum 11-ketotestosterone (11-KT) levels, and physiological responses associated with nesting behavior (i.e., elevation of serum 11-KT levels and accumulation of urine in the urinary bladder) were observed from November to January. Specifically, males were exposed to three different cooling regimes (control, 16 to 6 °C; H1, 16 to 11 °C; H2, 16 to 14 °C), and the results were compared. In addition, the effects of temperature on male reproductive behavior were also clarified. At higher water temperature regimes, the rate of testicular development and serum 11-KT levels were both higher from November to mid-December than from mid-December to January. However, the results showed that high water temperature regimes in the coldest period of winter did not suppress spermatogenesis completely. Conversely, the physiological responses to nesting were affected by high water temperatures, with serum 11-KT levels increasing and urine accumulation in the urinary bladder being suppressed. Furthermore, frequencies of two behaviors associated with nesting, i.e., body undulation and face displays, were also suppressed under high water temperatures (~ 14 °C) compared with normal temperatures (~ 7 °C) during the breeding season. Based on the physiological and behavioral responses to nesting, findings showed that preventing a water temperature decrease during winter suppresses reproductive activity in Cottus pollux SE.

Genetic studies of urinary metabolites illuminate mechanisms of detoxification and excretion in humans.

The kidneys integrate information from continuous systemic processes related to the absorption, distribution, metabolism and excretion (ADME) of metabolites. To identify underlying molecular mechanisms, we performed genome-wide association studies of the urinary concentrations of 1,172 metabolites among 1,627 patients with reduced kidney function. The 240 unique metabolite-locus associations (metabolite quantitative trait loci, mQTLs) that were identified and replicated highlight novel candidate substrates for transport proteins. The identified genes are enriched in ADME-relevant tissues and cell types, and they reveal novel candidates for biotransformation and detoxification reactions. Fine mapping of mQTLs and integration with single-cell gene expression permitted the prioritization of causal genes, functional variants and target cell types. The combination of mQTLs with genetic and health information from 450,000 UK Biobank participants illuminated metabolic mediators, and hence, novel urinary biomarkers of disease risk. This comprehensive resource of genetic targets and their substrates is informative for ADME processes in humans and is relevant to basic science, clinical medicine and pharmaceutical research.

Comparison of hot water immersion at 37.8°C with or without salt for rapid weight loss in mixed martial arts athletes.

Hot water immersion, known as a hot bath, is used by MMA athletes to produce rapid weight loss (RWL) by means of passive fluid loss. This study investigated the magnitude of body mass losses using a standardized hot bath protocol with or without the addition of salt. In a crossover design, eleven male MMA athletes (28.5 ± 4.6 y; 1.83 ± 0.07 m; 82.5 ± 9.1 kg) performed a 20-min immersion at 37.8°C followed by a 40-min wrap in a warm room. This bath and wrap was performed twice per visit. During one visit, only fresh water was used (FWB), and in the other visit, magnesium sulphate (1.6% wt/vol) was added to the bath (SWB). Prior to each visit, 24 h of carbohydrate, fibre, and fluid restriction was undertaken as part of the RWL protocol. Body mass losses induced by the hot bath protocols were 1.63 ± 0.75 kg and 1.60 ± 0.80 kg for FWB and SWB, respectively, and equivalent to ~2.1% body mass. Under the conditions employed, the magnitude of body mass loss in SWB was similar to FWB. However, further research should explore bathing in a temperature that is consistent with that habitually used by fighters, and/or higher concentrations of salt.

Studying the urine microbiome in superficial bladder cancer: samples obtained by midstream voiding versus cystoscopy.

BACKGROUND: Preliminary data suggest that the urinary microbiome may play a role in bladder cancer. Information regarding the most suitable method of collecting urine specimens is needed for the large population studies needed to address this. To compare microbiome metrics resulting from 16S ribosomal RNA gene sequencing between midstream, voided specimens and those obtained at cystoscopy. METHODS: Adults, with a history of superficial urothelial cell carcinoma (non-muscle invasive bladder cancer) being followed with periodic surveillance cystoscopy had a urine sample collected by a mid-stream, voided technique and then from the bladder at cystoscopy. Urine samples underwent 16S ribosomal RNA gene sequencing on the Illumina MiSeq platform. RESULTS: 22 subjects (8 female, 14 male) were included. There was no significant difference in beta diversity (diversity between samples) in all samples between collection methods. However, analysis by sex revealed a difference between voided and cystoscopy samples from the same individual in males (p = 0.006, Adonis test) but not in females (p = 0.317, Adonis test). No differences were seen by collection method in any alpha diversity (diversity within a sample) measurement or differential abundance of taxa. CONCLUSIONS: Beta diversity of the urine microbiome did differ by collection method for males only. This suggests that the urinary microbiomes of the two collection methods are not equivalent to each other, at least in males, which is the sex that bladder cancer occurs most frequently in. Therefore, the same collection method within a given study should be used.