Standardization and Prevalence of the Booster Phenomenon: Evaluation Using a Two-Step Skin Test with 43 kDa Glycoprotein in Individuals from an Endemic Region of Paracoccidioidomycosis.

BACKGROUND: We estimated the occurrence rate of the booster phenomenon by using an intradermal test with 43 kDa glycoprotein in an endemic area of paracoccidioidomycosis in the central-west region of Brazil. METHODS: Individuals who had a negative result on a survey performed by using an intradermal test with 43 kDa glycoprotein in an endemic area of paracoccidioidomycosis underwent a second intradermal test after 10-15 days to determine the presence or absence of the booster phenomenon. Statistical analyses were performed using the Chi-square test, Chi-square for linear trend test, Student's t test, and binomial test; p < 0.05 was considered significant. RESULTS: For the first time, we reported the occurrence of the booster phenomenon to an intradermal reaction caused by 43 kDa glycoprotein at a rate of 5.8-8.4%, depending on the test's cutoff point. This suggests that a cutoff point should be considered for the booster phenomenon in intradermal tests with 43 kDa glycoprotein: a difference of 6-7 mm between readings according to the first and second tests, depending on the purpose of the evaluation. CONCLUSION: The results indicate that the prevalence of paracoccidioidal infection in endemic areas is underestimated, as the booster phenomenon has not been considered in epidemiological surveys for this infection.

Contribuições do paradigma de equivalência de estímulos para o estudo das atitudes / Contributions of stimulus equivalence paradigm for studying attitudes

Considerando a abordagem analítico-comportamental, atitudes podem ser consideradas respostasavaliativas emitidas de acordo com uma história prévia de aprendizagem. O paradigma deequivalência de estímulos, utilizando procedimentos que simulam o comportamento simbólico nolaboratório, tem possibilitado entender a formação de atitudes como uma rede de relações arbitráriasentre classes de estímulos e atributos avaliativos. Esta visão tem auxiliado na compreensão de comofenômenos sociais, tais como estereótipos e preconceitos, podem ser estabelecidos e modificados. Oobjetivo deste trabalho é apresentar algumas contribuições, a partir de evidências empíricasdemonstradas por pesquisas que utilizaram este paradigma nas últimas décadas, particularmente apartir da década de 90, que fortalecem a equivalência de estímulos como um modelocomportamental importante para o estudo das atitudes(AU)

Evaluation of Paracoccidioides brasiliensis infection by gp 43 intradermal test in rural settlements in Central-West Brazil.

Epidemiological studies of paracoccidioidomycosis have been based on surveys achieved with intradermal tests, and paracoccidioidin is the most common antigen used in most cases. The glycoprotein of 43-kDa (gp43) has been used in intradermal tests. It is the most antigenic component of Paracoccidioides brasiliensis, and it provides greater specificity to evaluate infection for this fungus. In this study, the prevalence of P. brasiliensis infection was estimated with intradermal tests involving gp43 for 695 people in rural Central-West Brazil. The infection rate was 45.8 % (95 % CI = 42.1-49.5), and the average age of those infected was 45.8 ± 18.2 years. The prevalence did not show gender-based differences but increased with age. The results demonstrate the importance of P. brasiliensis infection in rural settlements and the early exposure of children in the region to the fungus. Despite the high antigenicity and specificity of gp43, its usage must be standardized, so that epidemiological surveys will be comparable and more accurately reflect P. brasiliensis infection in endemic areas.

Sequence-specific binding to a subset of IscR-regulated promoters does not require IscR Fe-S cluster ligation.

IscR is an Fe-S protein that functions as a transcriptional regulator of Fe-S biogenesis and other Fe-S protein-encoding genes in Escherichia coli. In this study, we investigated the requirement for the ligation of the [2Fe-2S] cluster of IscR to regulate a subset of IscR target promoters (P(hyaA), P(ydiU), P(napF), and P(hybO)) and defined the requirements for sequence-specific binding to the IscR target site in the hyaA promoter region. In contrast to previous results with the iscR promoter, we found that the Fe-S cluster is dispensable for IscR regulation of P(hyaA), P(ydiU), P(napF), and P(hybO), since IscR mutants containing alanine substitutions of the cysteine Fe-S ligands retained IscR-dependent regulation of these promoters in vivo. In vitro assays showed that both [2Fe-2S]-IscR and an IscR mutant lacking the cluster (IscR-C92A/C98A/C104A) bound the hya site with similar affinity, explaining why the mutant protein retained its ability to repress P(hyaA) in vivo. Characterization of the oligomeric state of IscR showed that both apo-IscR and [2Fe-2S]-IscR were dimers in solution, and four protomers of either form bound to the hya site. Also, binding of either apo- or [2Fe-2S]-IscR to the hya site showed cooperativity, suggesting that both forms interact similarly with the target site. Analysis of mutations in the hya site using DNA competition assays showed that apo-IscR most likely recognizes an imperfect palindrome within the hya promoter. Furthermore, the strength of apo-IscR binding to P(sufA), P(ydiU), P(napF), and P(hybO) IscR sites correlated with the number of matches to the hya site bases shown to be important in the competition assay. Thus, our data indicated that, unexpectedly, apo-IscR is a site-specific DNA-binding protein, and the role of apo-IscR needs to be considered in developing models for how IscR globally regulates transcription.

The hemodynamic effects of intravenous digoxin-binding fab immunoglobulin in severe preeclampsia: a double-blind, randomized, clinical trial.

OBJECTIVE: An endogenous digitalis-like factor (EDLF) has been implicated in the pathophysiology of preeclampsia (PE). This hypothesis is supported by two cases of preeclampsia in which administration of digoxin immune Fab (DIF) reduced mean arterial pressure (MAP). STUDY DESIGN: To study this observation further, we performed a double-blind, placebo-controlled, randomized clinical trial to examine the effects on MAP of intravenous DIF given after delivery in 26 subjects with severe preeclampsia. Treating obstetricians were blinded to subject assignment and were allowed to use standard antihypertensive drugs during the trial. RESULTS: The primary outcome, a significant difference in blood pressure between the two groups over the 24-h period of observation after the intervention, was not supported. However, mean MAP was significantly lower in the DIF-treated subjects for the first 4 h after therapy as compared with controls (P=0.05). Six subjects (46.2%) in the placebo arm were given conventional antihypertensive medications by their obstetrician for blood pressure >160 mm Hg systolic or >110 mm Hg diastolic, compared with zero subjects in the treatment arm (P=0.01). A trend towards increased creatinine clearance was observed in DIF-treated subjects (137.6+/-42.6 versus 104.1+/-43.4, P=0.07). CONCLUSION: These results support the hypothesis that EDLF contributes to the elevated blood pressure in preeclampsia and suggests a possible role for DIF as a treatment for this condition.

Interleukin-12 response to NcSRS2 immunization of BALB/c mice against Neospora caninum.

The apicomplexan parasite Neospora caninum can cause abortions and is recognized as an important causative agent responsible for economic and reproductive losses in the cattle industry. Humoral immune response was investigated in BALB/c mice by using recombinant NcSRS2 expressed in Escherichia coli as polyhistidine-tagged fusion proteins. NcSRS2 is the major immunodominant tachyzoite surface antigen of N. caninum. Separate groups of female BALB/c mice were infected subcutaneously with (P) PBS, (N) recNcSRS2, (NIL) recNcSRS2 plus interleukin-12 or (NF) recNcSRS2, in Freunds adjuvant. Serological analysis showed that the antibodies produced by immunization recognized native protein from N. caninum tachyzoites and that, 14 days after the initial immunization, NcSRS2-specific antibodies were present in all sera tested from the groups N, NIL and NF. NcSRS2 with Freuds adjuvant led to a stronger immune response, as measured by IgG1 and IgG2a levels, than did other formulations (NF > NIL > N > P; p < 0.001), with a Th2 bias. The results corroborate the potential use of recombinant protein NcSRS2 as a vaccine aimed at reducing congenital transmission. Further studies are required to identify new adjuvants capable of improving the induction of Th1 immune response.

A role for adsorption in lead leachability from MSWI bottom ASH.

Owing to its high porosity and large surface area, bottom ash from municipal solid waste incineration (MSWI) has been previously suggested as an adsorbent for removing heavy metals from wastewater. However, since lead leachability is not clearly understood, this study reports the behavior of lead for leaching from MSWI bottom ash, classified as NH, W and H. Leaching kinetics show a sorption/desorption mechanism producing equilibrium in the leachates. Adsorption experiments reveal a similarly high saturation coverage for NH, W and H bottom ash, demonstrating a considerable adsorption capacity. Adsorption mainly reflects an ion exchange mechanism. Successive leaching experiments prove that at least two species of lead can be stripped from bottom ash: one is Pb(OH)(4)(2-) and the second, identified by XAS, is a lead complex sorbed on the surface of Fe colloïd or FeOOH polymers.

Anatomia cirúrgica do ramo externo do nervo laríngeo superior / Surgical anatomy of the external branch of the superior laryngeal nerve

Introdução: o ramo externo do nervo laríngeo superior (RELS) é um ramo motor que tem origem no nível do osso hióide e desce medialmente à artéria carótida interna para inervar o músculo cricotireóideo da laringe. Nesse trajeto, aproxima-se do pólo superior da glândula tireóide. Muitos estudos têm sido feitos com o objetivo de avaliar a necessidade e viabilidade técnica de sua identificação durante operações sobre a glândula, mas alguns com resultados adversos. Objetivo: estudar a relação anatômica entre o RELS, a tireóide e a artéria tireoidiana superior e a viabilidade técnica e o tempo despendido em sua identificação durante procedimentos cirúrgicos sobre a glândula tireóide. Pacientes e método: foram estudados 40 RELS em pacientes que se submeteram a tireoidectomia por bócios nodulares, bócio difuso tóxico e câncer de tireóide. Foi analisado o tipo de disposição do nervo em relação à glândula tireóide e artéria tireoideana superior; a viabilidade de sua identificação durante a operação; e o tempo despendido para isso. Resultados: em mais da metade dos pacientes o RELS está a menos de 1,0cm da glândula e, em 62,5% dos casos, o nervo foi devidamente identificado em menos de quatro minutos. Conclusão: o RELS está muito próximo do pólo superior do lobo tireoideano; sua identificação cuidadosa é viável; e não acrescenta tempo significativo à cirurgia, devendo ser realizada sempre, para diminuir a morbidade da operação.

Introduction: the external branch of the superior laryngeal nerve (EBSL) is a motor branch that begins near the hyoid bone, than it goes down between the internal carotid artery and the larynx, and finds the cricothyroid muscle. In the way to the muscle, the nerve crosses the superior pole of the thyroid lobe. Many studies have been performed in order to observe the technical viability of finding the nerve during the thyroid surgery, and if we have to do it. There is some disagreement about this point. Objective: to study the anatomical relationship among the EBSL, the thyroid gland and the superior thyroid artery and the technical availability and the time spent in that identification during the surgical approach on the gland. Materials and methods: 40 EBLS were studied in patients who had the thyroid operated. These people had nodular goiter, toxic diffuse goiter, follicular adenoma or thyroid cancer. The anatomical relationship among the nerve, thyroid gland and superior thyroid artery were studied; as well as the viability of the nerve identification and the time spent to do it. Results: the nerve was near the superior pole of the lobe of the thyroid gland in 52.5% of cases; it was spent less than 4 minutes to find the EBSL in 62.5% of patients. Conclusion: the nerve is frequently very close to the gland, and its identification, which is possible in a few minuts, is recommended during thyroid surgery to avoid nerve damage.

Pesticides exposure in Culturama, Brazil--knowledge, attitudes, and practices.

In this study, the knowledge, attitudes, and practices associated with pesticide use and exposure were evaluated in the agricultural community of Culturama, in the state of Mato Grosso do Sul, Brazil. A standardized questionnaire was completed by 250 farm workers aged > or =18 years old. The average age of the studied population was 43.6 years and 17.6% had never been to school. Their farms were small (approximately 30ha) and family operated and did not utilize advanced farming technology. About 92% of the interviewees had worked directly with pesticides and 59.6% reported typical intoxication symptoms. Only 44.3%, however, believe that they had been intoxicated. A significant correlation was found between hand washing after pesticide application and reporting symptoms (P=0.014). Over 90% of the farmers reported using the organophosphorus insecticide methamidophos. A great majority (>90%) considered pesticides to be harmful to human health, but less than 20% used masks, impermeable clothes, or gloves during pesticide application. These results indicate that special educational programs, legislation promoting the use of safer pesticides, and implementation of personal protective measures are necessary to decrease the pesticide exposure of farmers in Culturama.

Induction of uterine prostaglandin H synthase 2 by estradiol following fetal adrenalectomy.

In sheep, fetal cortisol stimulates the conversion of progesterone to estradiol in late gestation initiating labor. It is unclear whether an intact fetal hypothalamic-pituitary-adrenal (HPA) axis is required to induce the estradiol-triggered subsequent endocrine changes including enhanced intrauterine prostaglandin (PG) synthesis associated with the onset of labor. We have shown that maternal estradiol administration stimulates PG H synthase (PGHS)-2 expressions in pregnant ovine intra-uterine tissues. The current study was undertaken to determine whether the fetal adrenal mediates estradiol's stimulation of the intrauterine PGHS-2 in pregnant sheep. Placenta, myometrium, and endometrium were collected from two groups of ewes at 123-127 d of gestational age (dGA) after fetal adrenalectomy and vehicle treatment (ADX; n = 5); or fetal ADX and maternal estradiol administration (5 mg twice a day for 2 d, ADX+E2, n = 5). PGHS-2 mRNA and protein were analyzed by Northern and Western Blot analyses in both groups and presented as the ratios to beta actin mRNA for Northern and G protein beta subunit for Western blot analysis. Fetal plasma cortisol was measured by radioimmunoassay. Data were analyzed by Student's t test. Fetal plasma cortisol levels were low in ADX and ADX+ E2 groups (<6 ng/mL). The cervix of all ADX+E2 treated ewes was dilated at necropsy. Three out of five ADX+ E2-treated ewes delivered within 48 h. The cervix was closed in all fetal ADX ewes at necropsy. PGHS-2 mRNA and protein increased (p < 0.05) in myometrium and endometrium, but not placenta in ADX+E2-treated ewes compared with ADX group. These data provide the first in vivo evidence for estradiol upregulation of intrauterine PGHS-2 in late gestation in the absence of an intact fetal HPA axis. Thus, the fetal adrenal is not required to mediate estradiol's stimulation of uterine PGHS-2 expression associated with the onset of labor.

Ontogeny of StAR and ACTH-R genes in ovine placenta.

It has been demonstrated that the ovine placenta secretes estrogen, progesterone and cortisol, and that plasma concentrations of estrogen and cortisol increase before birth. Among the elements important for steroid production is steroidogenic acute regulatory protein (StAR) which acutely delivers cholesterol from the outer to the inner mitochondrial membrane for rapid steroidogenesis. This study was designed to determine if StAR is present in ovine placenta, and if its expression changes during fetal development. In addition, because cortisol is secreted by the placenta, we also examined the expression of adrenocorticotropic hormone receptor (ACTH-R) to determine if it was present and if the pattern of expression changed as gestation proceeded. The mRNA levels for StAR and ACTH-R were assessed by RNase protection assay (RPA) and protein levels were measured by Western blot in placentas from pregnant ewes (100-105 days of gestation, n = 8; 120 days of gestation, n = 5; 135-142 days of gestation, n = 8). The data show that the ovine placenta expresses StAR and ACTH-R. There was a significant increase in the StAR mRNA and protein between 100 and 142 days of gestation, but there were no significant age-related changes in ACTH-R mRNA and protein levels. The data suggest that the increased steroid production by the placenta in late gestation may be related to the increased expression of StAR.

Developmental aspects of ovine adrenal adrenocorticotropic hormone receptor expression.

OBJECTIVE: Maturation of adrenocortical function is important because a prepartum increase in fetal plasma glucocorticoids is required for survival after birth. Adrenal maturation may include alterations in the regulation of adrenocorticotropic hormone (ACTH) receptor expression. Therefore, we quantitated ACTH receptor expression in the ovine adrenal cortex during development and after manipulations to better understand the regulation of the adrenal receptor in vivo. METHODS: For the ontogeny study, adrenals were obtained from fetuses at different stages of development, and the cortical tissue was stored at -80C until total RNA was extracted. The ACTH binding studies were done on adrenal membranes obtained from fetuses at two different ages using I125 (Phe-2, Nle-4) ACTH as the ligand. Plasma ACTH was measured by two-site immunoradiometric assay, and cortisol was measured by radioimmunoassay. ACTH receptor mRNA was quantitated by ribonuclease protection assay. The data were analyzed by analyses of variance. RESULTS: ACTH receptor mRNA level progressively increased in fetal life; relative changes in receptor mRNA and binding were similar (3.0-fold and 2.4-fold, respectively). Physiologic increases in fetal plasma cortisol decreased adrenal ACTH receptor mRNA concentration, and there was a strong (r2=0.76, P<.002) linear relationship between fetal plasma ACTH concentration and receptor mRNA levels. Receptor mRNA stability increased in development, and message half-life was greater in adulthood than in fetal life. CONCLUSION: The data suggest that developmental increases in receptor expression are part of the maturation process in the fetal adrenal and that plasma ACTH concentration plays a major role in regulating ACTH receptor mRNA levels in vivo.

Effects of insulin and insulin-like growth factor-I on renin gene expression in the renal cortical cells of ovine fetuses.

Renin gene expression and insulin-like growth factor-I (IGFI) gene expression are both developmentally upregulated in the renal cortex of ovine fetuses and decline after birth. The infusion of IGFI into ovine fetuses in late gestation increases plasma renin activity and concentration. In order to determine whether there are direct effects of IGFI or insulin on renin gene expression in the kidneys of ovine fetuses, we treated the renal cortical cells of ovine fetuses with IGFI or insulin. The results showed that the responses of renal renin mRNA to IGFI or insulin treatment in vitro were dependent on the culture conditions. Renin mRNA levels were significantly elevated by IGFI or insulin if the cells were cultured in medium devoid of serum (starved) for 16-18 h before treatment. In contrast, no obvious changes in renal renin mRNA expression were observed in the cells cultured in the presence of serum (non starved) before treatment with IGFI or insulin. IGFI and insulin also significantly enhanced cAMP concentrations in the medium of the cells starved in vitro. The data suggest that IGFI and insulin can act directly on the renal cortical cells from ovine fetuses to stimulate renin mRNA expression. It is possible that IGFI and insulin stimulate renin mRNA expression by increasing cAMP concentration in the cells.

Importance of the renal nerves for basal and stimulated renin mRNA levels in fetal and adult ovine kidneys.

OBJECTIVE: To investigate the effect of renal denervation on renin mRNA levels in fetal and nonpregnant adult ovine renocortical tissue and in isolated juxtaglomerular cells under basal conditions and after stimulation. METHODS: The left kidney was denervated and the right kidney subjected to a sham procedure in nine ovine fetuses (136-141 days' gestation) and 20 nonpregnant ewes. After 5-7 days the denervated and intact kidneys were obtained, and renin-containing renal cortical cells were isolated and cultured overnight. Then cells were treated with isoproterenol, forskolin, or isomethylbutyl xanthine (IBMX) for 4 hours. Total RNA was isolated and renin mRNA measured by RNase protection assay. Cyclic adenosine monophosphate (cAMP) levels were measured in the incubation medium with a competitive enzyme immunoassay. RESULTS: In adults, basal renin mRNA levels were significantly lower in denervated than in sham-operated kidneys. No difference was noted between denervated and intact fetal kidneys. Renin mRNA levels were significantly higher in fetal than in adult kidney tissue, and cells from fetuses had greater increases in renin mRNA after stimulation than did cells from adults. Fetal cells also released more cAMP into the incubation medium, and there was a correlation between cAMP and renin mRNA levels. CONCLUSIONS: The data indicate the effects of renal denervation on renin mRNA expression in the kidney are age dependent and that the fetus in late gestation has a mechanism for maintaining renin mRNA levels after denervation, which is absent or nonfunctional in the adult.

Renal mRNA response to reduced perfusion pressure conserved despite denervation in mature ovine fetuses.

We hypothesized that renal denervation in mature ovine fetuses reduces renin mRNA response to 24 h of reduced renal perfusion pressure (RPP). Seven occluder (O) (132.4 +/- 1.2 days gestation) and six control (C) (131.5 +/- 1.2 days gestation) fetuses underwent left renal denervation. Postoperatively, O fetuses experienced 24 h of reduced RPP by suprarenal aortic occlusion. Femoral arterial blood pressure (FAB) and plasma active renin (pARC) and prorenin (pPRC) concentrations were obtained hourly for 6 h and at h 23 and 24. Renin mRNA was measured by RNase protection assay. We quantitated renin containing glomeruli by immunocytochemistry. Variables were compared by ANOVA. Mean O group FAB reduction from baseline was -6.60 +/- 0.41 mmHg. pARC and pPRC increased with occlusion, renal ARC and renal PRC did not increase with occlusion. No effect in renin mRNA or number of positive glomeruli was noted with denervation in the basal state; however, significant increases were noted in response to RPP irrespective of innervation status. In conclusion, 24 h or reduced RPP in mature ovine fetus increases renal renin mRNA and the immunocytochemical expression of renin. This response is conserved despite denervation.

Corticotropin-releasing hormone type I receptor messenger ribonucleic acid and protein levels in the ovine fetal pituitary: ontogeny and effect of chronic cortisol administration.

In sheep, the ACTH secretory response to CRH in vivo or in vitro changes as a function of development, with peak responses occurring several weeks before term (145 days of gestation). CRH-stimulated ACTH secretion is mediated via the G protein-coupled CRH type I (CRH R1) receptor. We used a quantitative ribonuclease protection assay and Western immunoblotting to determine messenger RNA (mRNA) and protein levels of the CRH R1 receptor in immature and mature fetuses and adults. In addition, we precociously elevated fetal plasma cortisol levels to determine whether the fetal CRH R1 receptor is sensitive to increases in plasma cortisol. CRH R1 receptor mRNA levels decreased markedly throughout gestation and into the transition to adult life (immature fetus, 1.24+/-0.17; mature fetus, 0.75+/-0.13; adult, 0.18+/-0.093 pg/microg total anterior pituitary RNA). Also, continuous cortisol infusion in immature fetuses significantly decreased CRH R1 mRNA levels by 41%. Similar decreases were noted in protein levels. Thus, the decreased ACTH response to CRH stimulation during late gestation may be related to decreased CRH R1 receptor expression. In addition, plasma cortisol levels may influence corticotroph responsiveness to CRH by decreasing CRH R1 receptor expression.

Effect of renal denervation on renin gene expression, concentration, and secretion in mature ovine fetus.

To determine the role of the renal nerves on renin secretion and expression in the mature ovine fetus, we performed bilateral renal denervation on eight fetuses of time-dated pregnant ewes (126.8 +/- 0.6 days gestation) and compared renin in them to seven fetuses that underwent sham denervation (126.7 +/- 0.6 days gestation). Fetal arterial and venous catheters were implanted, and after 5-7 days of recovery isoproterenol was infused. Plasma active renin was lower in denervated animals than in intact animals under basal conditions and at each dose of isoproterenol. Plasma prorenin levels were lower in denervated fetuses but unaffected by isoproterenol. Denervation did not change renal renin, prorenin, or renin mRNA, but it did block isoproterenol-induced increases in renin mRNA in renocortical cells in vitro. We conclude that the renal nerves are required for renin secretory mechanisms and responsiveness of renin mRNA to beta-adrenergic stimulation but not for the expression of renin in the fetal kidney. We propose that one or more of the factors that maintain renin expression in the perinatal period may be absent or may be replaced by the renal nerves in the adult.

A mortality index for postmarketing surveillance of new medications.

The rate of introduction of new pharmaceuticals is growing as a result of advances in molecular pharmacology and targeted drug development. The Fatal Toxicity Index (FTI) has been proposed as a means for monitoring drug toxicity through post-marketing surveillance. The FTI requires data regarding the general availability of a particular agent in the community which, in the US, is proprietary. The authors propose a Mortality Index as an alternative method for calculating relative lethality that does not rely on proprietary information for postmarketing surveillance. Using data from the Toxic Exposure Surveillance System (TESS) a Mortality Index was calculated from the proportion of deaths occurring among all patients who present to a health care facility with an overdose on the same agent or class of agents. The average Mortality Index for various drugs or drug classes for the years 1989 to 1997 is reported. Because the Mortality Index for desipramine appeared much greater than that for the other tricyclics, a chi-squared analysis was performed. The authors conclude, based on this analysis, that desipramine is significantly more likely to lead to death after overdosage than any other tricyclic antidepressant in the study. Also, the Mortality Index appeared to identify the impact of pediatric formulations on overdose lethality. We conclude that the Mortality Index may be a useful tool for determining the safety of agents during the postmarketing surveillance phase.