Investigation of urinary volatile organic metabolites as potential cancer biomarkers by solid-phase microextraction in combination with gas chromatography-mass spectrometry

BACKGROUND: Non-invasive diagnostic strategies aimed at identifying biomarkers of cancer are of great interest for early cancer detection. Urine is potentially a rich source of volatile organic metabolites (VOMs) that can be used as potential cancer biomarkers. Our aim was to develop a generally reliable, rapid, sensitive, and robust analytical method for screening large numbers of urine samples, resulting in a broad spectrum of native VOMs, as a tool to evaluate the potential of these metabolites in the early diagnosis of cancer. METHODS: To investigate urinary volatile metabolites as potential cancer biomarkers, urine samples from 33 cancer patients (oncological group: 14 leukaemia, 12 colorectal and 7 lymphoma) and 21 healthy (control group, cancer-free) individuals were qualitatively and quantitatively analysed. Dynamic solid-phase microextraction in headspace mode (dHS-SPME) using a carboxenpolydimethylsiloxane (CAR/PDMS) sorbent in combination with GC-qMS-based metabolomics was applied to isolate and identify the volatile metabolites. This method provides a potential non-invasive method for early cancer diagnosis as a first approach. To fulfil this objective, three important dHS-SPME experimental parameters that influence extraction efficiency (fibre coating, extraction time and temperature of sampling) were optimised using a univariate optimisation design. The highest extraction efficiency was obtained when sampling was performed at 501C for 60min using samples with high ionic strengths (17% sodium chloride, wv 1) and under agitation. RESULTS: A total of 82 volatile metabolites belonging to distinct chemical classes were identified in the control and oncological groups. Benzene derivatives, terpenoids and phenols were the most common classes for the oncological group, whereas ketones and sulphur compounds were the main classes that were isolated from the urine headspace of healthy subjects. The results demonstrate that compound concentrations were dramatically different between cancer patients and healthy volunteers. The positive rates of 16 patients among the 82 identified were found to be statistically different (Po0.05). A significant increase in the peak area of 2-methyl3-phenyl-2-propenal, p-cymene, anisole, 4-methyl-phenol and 1,2-dihydro-1,1,6-trimethyl-naphthalene in cancer patients was observed. On average, statistically significant lower abundances of dimethyl disulphide were found in cancer patients. CONCLUSIONS: Gas chromatographic peak areas were submitted to multivariate analysis (principal component analysis and supervised linear discriminant analysis) to visualise clusters within cases and to detect the volatile metabolites that are able to differentiate cancer patients from healthy individuals. Very good discrimination within cancer groups and between cancer and control groups was achieved.

Solid phase microextraction, mass spectrometry and metabolomic approaches for detection of potential urinary cancer biomarkers: a powerful strategy for breast cancer diagnosis

A sensitive assay to identify volatile organic metabolites (VOMs) as biomarkers that can accurately diagnose the onset of breast cancer using non-invasively collected clinical specimens is ideal for early detection. Therefore the aim of this study was to establish the urinary metabolomic profile of breast cancer patients and healthy individuals (control group) and to explore the VOMs as potential biomarkers in breast cancer diagnosis at early stage. Solid-phase microextraction (SPME) using CAR/PDMS sorbent combined with gas chromatography–mass spectrometry was applied to obtain metabolomic information patterns of 26 breast cancer patients and 21 healthy individuals (controls). A total of seventy-nine VOMs, belonging to distinct chemical classes, were detected and identified in control and breast cancer groups. Ketones and sulfur compounds were the chemical classes with highest contribution for both groups. Results showed that excretion values of 6 VOMs among the total of 79 detected were found to be statistically different (p < 0.05). A significant increase in the peak area of (−)-4-carene, 3-heptanone, 1,2,4-trimethylbenzene, 2-methoxythiophene and phenol, in VOMs of cancer patients relatively to controls was observed. Statiscally significant lower abundances of dimethyl disulfide were found in cancer patients. Bioanalytical data were submitted to multivariate statistics [principal component analysis (PCA)], in order to visualize clusters of cases and to detect the VOMs that are able to differentiate cancer patients from healthy individuals. Very good discrimination within breast cancer and control groups was achieved. Nevertheless, a deep study using a larger number of patients must be carried out to confirm the results.

Avaliação da incontinência urinária de esforço em mulheres na pós-menopausa com e sem queixa de perda urinária através da aplicação do pad-test de 1 hora

Stress urinary incontinence (SUI) is defined as "involuntary loss of urine" due to several processes that alter the ability of the bladder to hold urine properly, regarded as a social and hygienic problem that adversely affects quality of life. In postmenopausal women, IU is associated with atrophy and weakness of the pelvic floor muscles. The objective this study was investigate, using the onehour pad test, stress urinary leakage (SUI), evaluate and compare their results in postmenopausal and premenopausal women. The survey was characterized as a cross-sectional study. The study consisted of 60 postmenopausal women were divided into GIU - consisting of 34 volunteers complaining of involuntary loss of urine during stress - and GSIU - consisting of 26 volunteers without complaints of loss of urine during stress, and 15 women, during the premenopausal (GPM), and ovulatory with normal menstrual cycle. All volunteers were evaluated clinically, subjected to one-hour pad test, after the biochemical evaluation of blood and sex hormones. Statistical analysis was performed by descriptive analysis, ANOVA, Turkey´s post-test and Pearson correlation. The results showed that 100% of postmenopausal patients had involuntary loss of urine during one hour pad test (GIU: 4.0 g; GSIU: 4.5 g). GPM remained continent after an hour pad test (GPM: 0.4 g). In addition, Pearson showed a strong correlation between urine loss with time since menopause (r = 0.8, p <0.01) and body mass index - BMI (r = 0.7; p = 0.01). These data suggest that the one-hour pad test is a useful test to assess and quantify urinary leakage, including those volunteers who had no previous complaint of SUI

Incontinência urinária após gonadectomia em fêmeas caninas

A incontinência urinária adquirida é uma condição debilitante e, muitas vezes, incurável que acomete fêmeas castradas e raramente fêmeas inteiras ou machos. A manifestação clínica pode ocorrer em qualquer momento após a gonadectomia e resulta em graves problemas no manejo do paciente. Os mecanismos que desencadeiam a incontinência após ovariectomia envolvem decréscimo na pressão de fechamento uretral, alterações hormonais, aumento na deposição de colágeno na musculatura lisa da bexiga, diminuição na contratilidade do músculo detrusor e redução na resposta aos estímulos elétricos e ao carbachol. O diagnóstico é realizado pelo histórico do animal, pelo exame físico, pelos exames laboratoriais, pelo perfil de pressão uretral, pela ultrassonografia e pelas radiografias abdominais. O tratamento clínico envolve utilização de fármacos -adrenérgicos, estrógenos, análogos de GnRH e agentes antidepressivos. As técnicas cirúrgicas recomendadas correspondem à uretropexia, cistouretropexia, aplicação de colágeno na uretra e colpossuspensão. Melhor compreensão da etiologia, da fisiopatologia, dos métodos de diagnóstico e tratamentos é fundamental em razão do pouco conhecimento e da identificação dessa condição no Brasil.

Study of caffeine urine and saliva of horses subjected to urinary acidification

The study of caffeine in racing horses has been of growing concern in veterinary sports medicine since the Association of Racing Commissioners International (ARCI) stated that it has no valid therapeutic use in racehorses. We examined the kinetic alterations in the urinary excretion and salivary secretion of caffeine in seven horses subjected to urinary acidification using ascorbic acid because this procedure can simulate the acidosis that follows anaerobic exercise. They participated in two treatment groups: the control group (SG) received 500 ml of saline and then 2.0 mg kg(-1) caffeine i.v. 30 min later; and the acidified group (AG) was subjected to urinary acidification with ascorbic acid at a dose of 0.5 g kg(-1) i.v. and then 2.0. mg kg(-1) caffeine i.v. 30 min later. Samples were collected 30 min before caffeine administration, immediately before caffeine administration (time zero) and at 0.25, 0.5, 1, 2, 4, 6, 8, 12, 24, 48 and 72 h afterwards. The samples were assayed by gas chromatography. The mean urinary pH for SG was 8.2, but for AG it was as low as 5.9 at 4 h, extending acidosis for up to 8 h. The kinetic curves for the two groups were similar for urinary excretion and salivary secretion. Differences occurred only in peak excretion and peak secretion in SG obtained at 1 h and 30 min, respectively, and in AG at 2 h and 1 h, respectively. This could be explained, in part, to the diuresis in AG compared with SG, resulting in less concentrated urine in the former group. The large difference between the pK(a) of caffeine and the pH of the medium may be responsible for the similar pharmacokinetics observed for the two groups. Copyright (C) 2004 John Wiley Sons, Ltd.

Kinetics of cellular component in inflammatory response induced by different stimuli in the swim bladder of pacu, Piaractus mesopotamicus Holmberg 1887 (Characidae)

The present study assessed the kinetics of cell accumulation at the site of inflammation induced by thioglycolate, Escherichia coli lipopolysaccharide (LPS) and heat-inactivated Aeromonas hydrophila, in the pacu, Piaractus mesopotamicus (Characidae), swim bladder. A quantitative, as well as qualitative, assessment was done of all the cells present in the exudate at 6, 24, and 48 h (n = 8) after inoculation of inflammatory agents. The results show that the thioglycolate was the irritant to induce higher total inflammatory cell accumulation when compared to the control group, 6 h after insult (P < 0.05). Inoculation of heat-inactivated Aeromonas hydrophila induced progressive accumulation of total inflammatory cells, with cell number peaking after 24 h and being significantly higher than observed in the other groups (P < 0.05). Injection of LPS also induced greater cell accumulation when compared to the control group (P < 0.05), although in lower numbers than those induced by the other two irritants. All irritants injected induced significantly greater accumulation of lymphocytes and thrombocytes when compared to the control group (P < 0.05).

Phenotypical characterization and adhesin identification in Escherichia coli strains isolated from dogs with urinary tract infections

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Avaliação da dor oncológica: estudo comparativo com pacientes internados em um hospital de referência em Natal/RN

Quasi-experimental study, with prospective data, comparative with quantitative approach, performed in a reference hospital, aiming to identify the effectiveness of the Numerical Rating Scale (NRS) and McGill Pain Questionnaire, used simultaneously, to evaluate a group of patients with oncologic pain (Experimental Group); to identify the effectiveness of the Numerical Rating Scale (NRS) to evaluate a group of patients with oncologic pain (Control Group); to identify the resolution of pain according to prescribed medication, considering the result of the rating scales, and to compare it between the two groups of patients in the study. The population consisted of 100 patients, with both the experimental and control groups being composed of 50 people, with data collected from February to April 2010. The results show that in the experimental group, 32% of the patients were aged 60 to 69, 80% were female; 30% had a primary tumor in the breast, 58% had metastasis, and on 70% the disease was localized. In the first pain evaluation, 26% identified it as light; 46%, moderate; and 28%, severe; with an average of 5.50. In the second pain evaluation, 2% reported no pain; 70%, light; 26%, moderate. and 2%, severe, with an average of 3.30. On those with moderate pain, 60% used non-opioid medicine, 25% under severe pain were medicated with non-opioids and 41.67% with weak opioids. Regarding the Pain Management Index (PMI), 44.0% were rated as "-1". In the control group, 28% were aged 40 to 49, and 54% were male; 20% had primary tumor in the breast and genital-urinary system, consecutively; 56% presented metastasis; on 64% the disease was localized. In the first pain evaluation, 14% considered it light; 42%, moderate; and 44%, severe; with an average of 6.26. In the second pain evaluation, 18% did not signal pain; on 38% pain was light; 40%, moderate; and 4%, severe; with an average of 3.0. Regarding medicine therapy, 71.43% with moderate pain used non-opioids, 22.73% with severe pain used non-opioids and 27.27% weak opioids. Considering PMI, 42% were rated "-1"; and 42%, rated "0". We conclude that, despite the importance of pain as the 5th vital sign, it is still under-identified and under-treated by professionals. Nevertheless, studied oncologic patients had a tendency to report pain more easily when evaluated with the NRS instrument than with the combined use of NRS and MPQ. We believe, however, that the combination of these two instruments represents a more effective evaluation of pain, as it allows comprehension of its quantitative and qualitative aspects. We recommend, however, the replication of this study on a larger population, for a longer span of time, and consequently generating more evaluations, so this can confirm or deny the hypothesis that NRS and MPQ can, together, better evaluate pain on the oncologic patient

Dose-dependent effect of Resveratrol on bladder cancer cells: Chemoprevention and oxidative stress

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Urinary Mutagenicity in Chemical Laboratory Workers Exposed to Solvents

Solvents represent an important group of environmental pollutants to which people are exposed daily in the workplace. The physico chemical properties of solvents may result in disturbances to cellular structures, including damage to DNA. However, the effects of mixtures of solvents are not well known. Mutations caused by environmental agents are related to cancer development and other degenerative diseases. The work in a research laboratory that uses several types of solvents is equally predisposed to these hazards. In this study, we evaluated the mutagenicity of urine from 29 subjects exposed occupationally to solvents in a chemistry research laboratory and 29 subjects without occupational exposure (controls). Urine samples were collected in polyethylene containers at the end of the work shift. For the concentration and extraction of urine samples the XAD-2 resin was used with acetone as an eluting agent. Several strains of Salmonella typhimurium (TA100, TA98, TA97a, TA1535, YG1024) should be used to assess mutagenic susceptibilities among workers exposed to organic solvents. Different doses of extract (1.5; 3.0; 6.0 and 12.0 m/ equivalents of urine per plate) were tested on S. typhimurium strains TA100 and YG 1024, with and without metabolic activation. The mutagenic activity, measured in Salmonella typhimurium YGI1024 with S9 mix, was significantly greater in urine from workers than from controls (p <= 0.05). These results indicate the relevance of using biomarkers to assess the risk of occupational exposure to organic solvents.

Efeitos da ligadura do ducto pancreático e da secção ductal com livre drenagem de secreções para o peritônio sobre as funções endócrina e exócrina do pâncreas: estudo clínico e laboratorial em coelhos

As ressecções pancreáticas seguidas de anastomoses acompanham-se de altas taxas de morbidade, que incluem: surtos de pancreatite e, principalmente, fístulas digestivas. Nos transplantes pancreáticos a drenagem da parte exócrina do enxerto para o intestino ou para a bexiga, além das peculiaridades técnicas, também não é isenta de diversas complicações. Visando evitar ou atenuar tais conseqüências e simplificar a técnica cirúrgica, têm sido usadas outras abordagens para o tratamento ductal do coto/enxerto pancreático, tais como: drenagem livre de secreções para o peritônio, com o ducto pancreático aberto, ligadura ductal e oclusão do ducto com polímeros sintéticos. OBJETIVO: O presente estudo visa avaliar clínica e laboratorialmente as funções endócrina e exócrina do pâncreas de coelhos com o ducto aberto e ligado. MÉTODOS: Foram realizadas 150 operações, divididas em 3 grupos: N - manipulação/controle (n=50), A - grupo aberto (n=50), e L - ligado (n=50). Os momentos de observação foram pré-operatório, dia 0 (dia da operação) e pós-operatório (observação e sacrifício): 7 dias, 14 dias, 28 dias, 90 dias e 180 dias. Os parâmetros analisados foram: estado geral, atividade, controles do peso corporal, ingestão hídrica, ingestão alimentar e dosagens da amilase sangüínea, glicemia, glicose urinária e insulina plasmática. RESULTADOS: Todos os grupos tiveram evolução clínica similar, com bom estado geral, ganho ponderal progressivo e valores normais da ingestão hídrica e ingestão alimentar. Exceto uma significativa elevação da amilase sangüínea nas primeiras 24-48h para os 2 grupos com interferência ductal, não houve qualquer alteração dos níveis basais da glicemia, glicosúria e insulinemia entre os 3 grupos experimentais, em todos os momentos de observação. CONCLUSÃO: À exceção da amilase sangüínea, cujos níveis basais foram significativamente elevados no 1º e 2º dias de pós-operatório, as duas modalidades técnicas de abordagem da secreção exócrina do pâncreas, utilizadas em coelhos, não determinaram quaisquer alterações clínicas, bem como dos níveis basais de glicose sangüínea, glicose urinária e insulina plasmática, durante 6 meses de seguimento.

Metabolic assessment in patients with urinary lithiasis

INTRODUCTION: Metabolic investigation in patients with urinary lithiasis is very important for preventing recurrence of disease. The objective of this work was to diagnose and to determine the prevalence of metabolic disorders, to assess the quality of the water consumed and volume of diuresis as potential risk factors for this pathology. PATIENTS and METHODS: We studied 182 patients older than 12 years. We included patients with history and/or imaging tests confirming at least 2 stones, with creatinine clearance > 60 mL/min and negative urine culture. The protocol consisted in the collection of 2, 24-hour urine samples, for dosing Ca, P, uric acid, Na, K, Mg, Ox and Ci, glycemia and serum levels of Ca, P, Uric acid, Na, K, Cl, Mg, U and Cr, urinary pH and urinary acidification test. RESULTS: 158 patients fulfilled the inclusion criteria. Among these, 151 (95.5%) presented metabolic changes, with 94 (62.2%) presenting isolated metabolic change and 57 (37.8%) had mixed changes. The main disorders detected were hypercalciuria (74%), hypocitraturia (37.3%), hyperoxaluria (24.1%), hypomagnesuria (21%), hyperuricosuria (20.2%), primary hyperparathyroidism (1.8%) secondary hyperparathyroidism (0.6%) and renal tubular acidosis (0.6). CONCLUSION: Metabolic change was diagnosed in 95.5% of patients. These results warrant the metabolic study and follow-up in patients with recurrent lithiasis in order to decrease the recurrence rate through specific treatments, modification in alimentary and behavioral habits.

Serum and urinary measurements of prostatic acid phosphatase (PAP) and prostatic specific antigen (PSA) in dogs

Realizaram-se mensurações sérica e urinária de fosfatase ácida prostática (PAP) e antígeno prostático específico (PSA) de 20 cães. Os testes de PAP e PSA foram feitos em um equipamento automatizado, com o uso de kits comerciais para humanos. A média de PAP sérico foi de 0,7U/l e urinário 0,U/l. As médias do PSA sérico e urinário foram 0,005ng/dL e 0,004ng/dl, respectivamente. A determinação do dois biomarcadores in vivo é uma nova opção de diagnóstico na medicina veterinária e os valores obtidos devem ser correlacionados com a lesão morfológica da próstata.

Hematologic, serum biochemistry and urinary values for captive Crab-eating Fox (Cerdocyon thous) in São Paulo state, Brazil

A importância do estudo dos valores hematológicos, de bioquímica sérica e urinários de Cachorro-do-mato (Cerdocyon thous) baseia-se na necessidade de cuidados e manutenção da sanidade destas populações. Este estudo visou investigar os parâmetros fisiológicos hematológicos, de bioquímica sérica e urinários dos Cachorros-do-mato de cativeiro, comparando as possíveis diferenças sexuais e de faixa etária. Foram colhidas amostras de sangue de 52 animais, pertencentes a diversos Zoológicos do Estado de São Paulo, Brasil. Foram colhidos cerca de 7mL de sangue, que foram utilizados para se obter os valores hematológicos e o perfil de bioquímica sérica. Também foram colhidos 5mL de urina para realização da urinálise. Não se encontraram diferenças entre os valores obtidos para machos e fêmeas tanto na hematologia, quanto na bioquímica sérica. Alguns parâmetros hematológicos e de bioquímica sérica foram afetados pela idade, mostrando diferenças significativas. Os resultados da urinálise foram demonstrados apenas em forma descritiva. Os principais valores encontrados foram, hemácias 4,35+0,73 x 106 células /µL, leucócitos totais 7,72+3,66 x 103 células /µL (predomínio de neutrófilos segmentados), plaquetas 227,06+111,58 x 103 células /µL, ureia 43,06+14,28mg/dL e creatinina 1,03+0,24mg/dL. Os valores hematológicos, de bioquímica sérica e urinários obtidos neste estudo podem ser utilizados como valores fisiológicos de Cachorros-do-mato de cativeiro. Pode-se concluir que as espécies silvestres necessitam de seus próprios valores de referência com necessidade de se diferenciar animais em cativeiro de animais de vida livre.

Complications and risk factors in transrectal ultrasound-guided prostate biopsies

CONTEXTO E OBJETIVO: A biópsia da próstata não é um procedimento isento de riscos. Existe preocupação com respeito às complicações e quais seriam os melhores antibióticos usados antes do procedimento. O objetivo foi determinar a taxa de complicações e os possíveis fatores de risco para complicação na biópsia da próstata. TIPO DE ESTUDO E LOCAL: Estudo prospectivo clínico, realizado no Hospital das Clínicas de Botucatu. MÉTODOS: Foram realizadas biópsias em 174 pacientes que apresentavam anormalidade ao exame digital da próstata ou antígeno prostático específico maior que 4 ng/ml ou ambos. Todos os pacientes realizaram enema e antibioticoprofilaxia previamente ao exame. As complicações foram anotadas após o término do procedimento e em consultas posteriores. Algumas condições foram investigadas como possíveis fatores de risco para biópsias de próstata: idade, câncer da próstata, diabetes melito, hipertensão arterial sistêmica, antecedentes de prostatite, uso de ácido acetilsalicílico, volume prostático, número de biópsias e uso de sonda vesical. RESULTADOS: As complicações hemorrágicas foram mais comuns (75,3%) enquanto que as infecciosas ocorreram em 19% dos casos. O tipo mais freqüente foi a hematúria, ocorrendo em 56% dos pacientes. A infecção do trato urinário ocorreu em 16 pacientes (9,2%). Sepse foi observada em três pacientes (1,7%). Não houve óbitos. em 20% dos pacientes não foram observadas complicações após o exame. A presença da sonda vesical foi fator de risco para complicações infecciosas (p < 0,05). O número maior de amostras nas biópsias foi relacionado à hematúria, sangramento retal e complicações infecciosas (p < 0,05). As demais condições investigadas não se relacionaram com complicações pós-biópsia da próstata. CONCLUSÕES: As complicações pós-biópsia da próstata foram em sua maioria autolimitadas. A taxa de complicações graves foi baixa, sendo a biópsia de próstata guiada pelo ultra-som segura e eficaz. A retirada de um maior número de fragmentos na biópsia relaciona-se com hematúria, sangramento retal e complicações infecciosas. A sonda vesical foi um fator de risco para complicações infecciosas.