Ácido fólico: validação do método espectrofotométrico

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

SEDIMENT BIOTURBATION POTENTIAL of UCA RAPAX and UCA URUGUAYENSIS AS A RESULT of THEIR FEEDING ACTIVITY

Bioturbation of mangrove sediments by Uca uruguayensis (Nobili, 1901) and U. rapax (Smith, 1870) was compared based on the grain-size composition and organic content in surface sediment around the burrow and feeding pellets in two mangrove zones of the Sao Vicente Estuary, state of São Paulo, Brazil. For each species, 25 burrows with active crabs were selected. All pellets within a 15-cm radius of each burrow were carefully collected; samples of substrate were taken; and the crab occupant was excavated, sexed, and measured for carapace width (CW). The number of spoon-tipped setae on the second maxilliped of each species was estimated; U. uruguayensis showed more of these setae than U. rapax. For both species, the sediment post-processed by feeding activity (feeding pellets) showed a similar increase of coarser fractions and a smaller organic content. However. U. uruguayensis was more efficient in removing organic matter (88.1%) from the sediment than U. rapax (37.5%). These results suggest that different numbers of spoon-tipped setae on the second maxillipeds of the fiddler crabs do not affect the potential for grain-size selection, but result in differing abilities to remove organic matter from the sediment.

Comparison of the quantification of caffeine in human plasma by gas chromatography and ELISA

In the present study we evaluated the precision of the ELISA method to quantify caffeine in human plasma and compared the results with those obtained by gas chromatography. A total of 58 samples were analyzed by gas chromatography using a nitrogen-phosphorus detector and routine techniques. For the ELISA test, the samples were diluted to obtain a concentration corresponding to 50% of the absorbance of the standard curve. To determine whether the proximity between the I50 of the standard curve and that of the sample would bring about a more precise result, the samples were divided into three blocks according to the criterion of difference, in modulus, of the I50 of the standard curve and of the I50 of the sample. The samples were classified into three groups. The first was composed of 20 samples with I50 up to 1.5 ng/ml, the second consisted of 21 samples with I50 ranging from 1.51 to 3 ng/ml, and the third of 17 samples with I50 ranging from 3.01 to 13 ng/ml. The determination coefficient (R² = 0.999) showed that the data obtained by gas chromatography represented a reliable basis. The results obtained by ELISA were also reliable, with an estimated Pearson correlation coefficient of 0.82 between the two methods. This coefficient for the different groups (0.88, 0.79 and 0.49 for groups 1, 2 and 3, respectively) showed greater reliability for the test with dilutions closer to I50.

Lymphocyte transformation assay for C neoformans antigen is not reliable for detecting cellular impairment in patients with Neurocryptococcosis

Background: Cryptococcus neoformans causes meningitis and disseminated infection in healthy individuals, but more commonly in hosts with defective immune responses. Cell-mediated immunity is an important component of the immune response to a great variety of infections, including yeast infections. We aimed to evaluate a specific lymphocyte transformation assay to Cryptococcus neoformans in order to identify immunodeficiency associated to neurocryptococcosis (NCC) as primary cause of the mycosis.Methods: Healthy volunteers, poultry growers, and HIV-seronegative patients with neurocryptococcosis were tested for cellular immune response. Cryptococcal meningitis was diagnosed by India ink staining of cerebrospinal fluid and cryptococcal antigen test (Immunomycol-Inc, SP, Brazil). Isolated peripheral blood mononuclear cells were stimulated with C. neoformans antigen, C. albicans antigen, and pokeweed mitogen. The amount of H-3-thymidine incorporated was assessed, and the results were expressed as stimulation index (SI) and log SI, sensitivity, specificity, and cut-off value (receiver operating characteristics curve). We applied unpaired Student t tests to compare data and considered significant differences for p<0.05.Results: The lymphotoxin alpha showed a low capacity with all the stimuli for classifying patients as responders and non-responders. Lymphotoxin alpha stimulated by heated-killed antigen from patients with neurocryptococcosis was not affected by TCD4+ cell count, and the intensity of response did not correlate with the clinical evolution of neurocryptococcosis.Conclusion: Response to lymphocyte transformation assay should be analyzed based on a normal range and using more than one stimulator. The use of a cut-off value to classify patients with neurocryptococcosis is inadequate. Statistical analysis should be based on the log transformation of SI. A more purified antigen for evaluating specific response to C. neoformans is needed.

Relevância do estado de hidratação na interpretação de parâmetros nutricionais em diálise peritoneal

OBJETIVO: Identificar determinantes do estado de hidratação de pacientes em diálise peritoneal crônica, bem como investigar os efeitos da sobrecarga líquida sobre o estado nutricional. MÉTODOS: Foi feito estudo transversal, realizado em 2006, avaliando 27 pacientes em diálise peritoneal crônica, acompanhados no Hospital das Clínicas da Faculdade de Medicina de Botucatu (SP), quanto a parâmetros clínicos, dialíticos, laboratoriais, antropométricos e de bioimpedância elétrica. Para avaliar a influência de parâmetros sobre o estado de hidratação empregou-se modelo de regressão linear múltipla. A amostra foi estratificada quanto ao estado de hidratação pela relação entre água extracelular e água corporal total (0,47 para homens e 0,52 para mulheres), parâmetros obtidos por meio de bioimpedância elétrica. Comparações foram realizadas por análise de covariância, Mann-Whitney, Qui-quadrado ou teste exato de Fisher. Considerou-se significância estatística quando p≤0,05. RESULTADOS: Pacientes com maior volume urinário e em modalidade dialítica automatizada apresentaram melhor estado de hidratação. Pacientes com maior sobrecarga líquida, comparados àqueles com menor sobrecarga, apresentaram menor ângulo de fase (M=4,2, DP=0,9 vs M=5,7, DP=0,7º; p=0,006), menor albumina (M=3,06, DP=0,46 vs M=3,55, DP=0,52g/dL; p=0,05) e maior % prega cutânea tricipital (M=75,3, DP=36,9 vs M=92,1, DP=56,9%; p=0,058), sem outras evidências antropométricas. CONCLUSÃO: Pode-se sugerir que os níveis reduzidos de albumina e ângulo de fase nos pacientes com maior sobrecarga líquida não estiveram relacionados a pior estado nutricional. Para o diagnóstico nutricional em vigência de sobrecarga líquida, deve-se considerar o conjunto de variáveis obtidas por diversos métodos, buscando relacioná-las e interpretá-las de maneira abrangente, possibilitando um diagnóstico nutricional fidedigno.

Cell blocks allow reliable evaluation of expression of basal (CK5/6) and luminal (CK8/18) cytokeratins and smooth muscle actin (SMA) in breast carcinoma

Objective:Gene expression studies have revealed several molecular subtypes of breast carcinoma with distinct clinical and biological behaviours. DNA microarray studies correlated with immunohistochemical profiling of breast carcinomas using cytokeratin (CK) markers, Her2/neu, oestrogen receptor (ER), and basal myoepithelial cell markers have identified five breast tumour subtypes: (i) luminal A (ER+; Her2/neu-), (ii) luminal B (ER+; Her2/neu+), (iii) Her2 overexpression (ER-; Her2/neu+), (iv) basal-like (ER-; Her2/neu-, CK5/6 and 14+), and (v) negative for all markers. Luminal carcinomas express cytokeratins in a luminal pattern (CK8/18), and the basal-like type expresses CK5/6 and CK14 or basal epithelial cell markers. CK5/6, CK8/18, and smooth muscle actin (SMA) expression were assessed in cell blocks and compared with expression in surgical specimens.Methods:Sixty-two cases of breast carcinoma diagnosed by fine needle aspiration cytology with cell blocks and available surgical specimens were included. Cell blocks containing at least 10 high-power fields each with at least 10 tumour cells and surgical specimens were immunostained for CK5/6, CK8/18 and SMA.Results:Percentage sensitivity, specificity, positive predictive value, negative predictive value and accuracy were, respectively, 77, 100, 100, 92 and 94 for CK5/6; 98, 66, 96, 80 and 95 for CK8/18; and 92, 96, 85, 98 and 95 for SMA.Conclusion:The identification of CK5/6, CK8/18 and SMA by immunohistochemistry in cell blocks can be a reliable method that yields results close to those obtained in surgical specimens, and can contribute to the classification of breast carcinomas with luminal and basal expression patterns, providing helpful information in the choice of treatment and in the evaluation of prognostic and predictive factors.

Motile sperm organelle morphology examination (MSOME): intervariation study of normal sperm and sperm with large nuclear vacuoles

Background: Although the motile sperm organelle morphology examination (MSOME) was developed only as a selection criterion, its application as a method for classifying sperm morphology may represent an improvement in evaluation of semen quality, with potential clinical repercussions. The present study aimed to evaluate individual variations in the motile sperm organelle morphology examination (MSOME) analysis after a time interval.Methods: Two semen samples were obtained from 240 men from an unselected group of couples undergoing infertility investigation and treatment. Mean time interval between the two semen evaluations was 119 +/- 102 days. No clinical or surgical treatment was realized between the two observations. Spermatozoa were analyzed at greater than or equal to 8400 x magnification by inverted microscope equipped with DIC/Nomarski differential interference contrast optics. At least 200 motile spermatozoa per semen sample were evaluated and percentages of normal spermatozoa and spermatozoa with large nuclear vacuoles (LNV/one or more vacuoles occupying >50% of the sperm nuclear area) were determined. A spermatozoon was classified as morphologically normal when it exhibited a normal nucleus (smooth, symmetric and oval nucleus, width 3.28 +/- 0.20 mu m, length 4.75 +/- 0.20 mu m/absence of vacuoles occupying >4% of nuclear area) as well as acrosome, post-acrosomal lamina, neck and tail, besides not presenting cytoplasm around the head. One examiner, blinded to subject identity, performed the entire study.Results: Mean percentages of morphologically normal and LNV spermatozoa were identical in the two MSOME analyses (1.6 +/- 2.2% vs. 1.6 +/- 2.1% P = 0.83 and 25.2 +/- 19.2% vs. 26.1 +/- 19.0% P = 0.31, respectively). Regression analysis between the two samples revealed significant positive correlation for morphologically normal and for LNV spermatozoa (r = 0.57 95% CI: 0.47-0.65 P < 0.0001 and r = 0.50 95% CI: 0.38-0.58 P < 0.0001, respectively).Conclusions: The significant positive correlation and absence of differences between two sperm samples evaluated after a time interval with respect to normal morphology and LNV spermatozoa indicated that MSOME seems reliable (at least for these two specific sperm forms) for analyzing semen. The present result supports the future use of MSOME as a routine method for semen analysis.

A new ovarian response prediction index (ORPI): implications for individualised controlled ovarian stimulation

Background: The objective was to present a new ovarian response prediction index (ORPI), which was based on anti-Mullerian hormone (AMH) levels, antral follicle count (AFC) and age, and to verify whether it could be a reliable predictor of the ovarian stimulation response.Methods: A total of 101 patients enrolled in the ICSI programme were included. The ORPI values were calculated by multiplying the AMH level (ng/ml) by the number of antral follicles (2-9 mm), and the result was divided by the age (years) of the patient (ORPI=(AMH x AFC)/Patient age).Results: The regression analysis demonstrated significant (P<0.0001) positive correlations between the ORPI and the total number of oocytes and of MII oocytes collected. The logistic regression revealed that the ORPI values were significantly associated with the likelihood of pregnancy (odds ratio (OR): 1.86; P=0.006) and collecting greater than or equal to 4 oocytes (OR: 49.25; P<0.0001), greater than or equal to 4 MII oocytes (OR: 6.26; P<0.0001) and greater than or equal to 15 oocytes (OR: 6.10; P<0.0001). Regarding the probability of collecting greater than or equal to 4 oocytes according to the ORPI value, the ROC curve showed an area under the curve (AUC) of 0.91 and an efficacy of 88% at a cut-off of 0.2. In relation to the probability of collecting greater than or equal to 4 MII oocytes according to the ORPI value, the ROC curve had an AUC of 0.84 and an efficacy of 81% at a cut-off of 0.3. The ROC curve for the probability of collecting greater than or equal to 15 oocytes resulted in an AUC of 0.89 and an efficacy of 82% at a cut-off of 0.9. Finally, regarding the probability of pregnancy occurrence according to the ORPI value, the ROC curve showed an AUC of 0.74 and an efficacy of 62% at a cut-off of 0.3.Conclusions: The ORPI exhibited an excellent ability to predict a low ovarian response and a good ability to predict a collection of greater than or equal to 4 MII oocytes, an excessive ovarian response and the occurrence of pregnancy in infertile women. The ORPI might be used to improve the cost-benefit ratio of ovarian stimulation regimens by guiding the selection of medications and by modulating the doses and regimens according to the actual needs of the patients.

A rapid, environmentally friendly, and reliable method for pesticide analysis in high-fat samples

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

Is urine interleukin-8 level a reliable laboratory test for diagnosing late onset sepsis in premature infants?

The present study is aimed to determine serum and urine interleukin-8 (IL-8) levels in premature infants with late onset sepsis (LOS) and to evaluate if urine IL-8 is a useful test for LOS diagnosis. Fifty-six premature infants admitted to the NICU over 1 year had serum and urine IL-8 determined by ELISA. They were divided into three groups: I definite sepsis, II probable sepsis and III non-infected. Results were expressed as mean or median. Differences between groups were assessed by ANOVA, Kruskal-Wallis ANOVA and Dunns Method. Sensitivity, specificity and positive and negative predictive values were calculated and a receiver operator characteristic curve was constructed to determine serum and urine IL-8 accuracy. There were no differences between groups for birth weight, and gestational and post-natal age. Median serum and urine IL-8 levels were significantly higher in GI and GII: 929 x 906 x 625pg/ml; P=0.024, and 249 x 189 x 42pg/mgCr; P< 0.001. Optimal cut-off point was 625pg/ml for serum IL-8 with 69 sensitivity and 75pg/mgCr for urine IL-8 with 92 sensitivity. IL-8 can be determined in urine from premature infants with LOS and is an accurate and feasible diagnosis method.

Is the ultrasound-estimated bladder weight a reliable method for evaluating bladder outlet obstruction?

OBJECTIVETo evaluate the correlation between ultrasound-estimated bladder weight (UEBW) in patients with different degrees of bladder outlet obstruction (BOO).METHODSWe evaluated 50 consecutive non-neurogenic male patients with lower urinary tract symptoms (LUTS) referred to urodynamic study (UDS). All patients self-answered the International Prostate Score Symptoms (IPSS) questionnaire. After the UDS, the bladder was filled with 150 mL to determine UEBW.Patients with a bladder capacity under 150 mL, a previous history of prostate surgery or pelvic irradiation, an IPSS score <8, a bladder stone or urinary tract infection were excluded.After a pressure-flow study, the Schafer linear passive urethral resistance relation nomogram was plotted to determine the grade of obstruction: Grades I-II/VI were defined as mild obstruction, Grades III-IV/VI as moderate obstruction, and Grades V-VI/VI as severe obstruction.RESULTSThe UEBW was 51.7 +/- 26.9, 54.1 +/- 30.0 and 54.8 +/- 28.2 in patients with mild, moderate and severe BOO, respectively (P = 0.130). The UEBW allowed us to define four groups: (i) UEBW < 35 g; (ii) 35 g <= UEBW < 50 g; (iii) 50 g <= UEBW < 70 g; and (4) UEBW >= 70 g.We did not find any differences in age, prostate weight, IPSS, PVR, cystometric bladder capacity, presence of detrusor overactive and degree of obstruction in the aforementioned groups.CONCLUSIONDespite the fact that some studies have emphasized the value of UEBW as an efficient non-invasive method for evaluating lower urinary tract obstruction, our study suggests that UEBW does not present any individual correlation with LUTS or objective measurements of BOO.

Central angiotensin II induces sodium bicarbonate intake in the rat

The aim of this work was to test mineral preference in hydrated rats that received a pulse intracerebroventricular (icv(p)) injection of ANG II at a dipsogenic dose (50 ng). The icv(p) ANG II induced a four-fold higher ingestion of 0.15 M NaHCO(3) than of other mineral solutions at palatable concentrations (0.15 M NaCl, 0.05 mM CaCl(2) and 0.01 M KCl) in a five-bottle test with water available in a fifth bottle; water intake was not consistently high in this test. Contrary to what is predicted by the mineralocorticoid/angiotensin II synergy hypothesis, the 0.15 M NaCl intake in the five-bottle test was not enhanced by icvp ANG H preceded by deoxycorticosterone (DOCA) treatment (2.5 mg/day for 3 days); neither was the NaHCO(3) intake. This result contrasted with the vigorous ingestion of both isotonic sodium solutions, but mostly of NaCl, rather than of other fluids, by sodium-depleted (furosemide 10 mg sc + 24 h removal ambient sodium) rats in a sodium appetite test. The results suggest that mineralocorticoid combined to icv(p) ANG II does not simulate the sodium preference shown during sodium appetite. The results also show that a dipsogenic dose of central ANG II induces a reliable ingestion of isotonic sodium bicarbonate in the rat. (C) 2007 Elsevier Ltd. All rights reserved.

Toward reliable estimates of seed removal by small mammals and birds in the Neotropics

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

Molecular identification of Pichia guilliermondii, Debaryomyces hansenii and Candida palmioleophila

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