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Background: 14-3-3 proteins comprise a family of eukaryotic multifunctional proteins involved in several cellular processes. The Pb14-3-3 of Paracoccidioides brasiliensis seems to play an important role in the Paracoccidioides-host interaction. Paracoccidioides brasiliensis is an etiological agent of paracoccidioidomycosis, which is a systemic mycosis that is endemic in Latin America. In the initial steps of the infection, Paracoccidioides spp. synthetizes adhesins that allow it to adhere and invade host cells. Therefore, the aim of this work was to perform a functional analysis of Pb14-3-3 using Saccharomyces cerevisiae as a model., Results: The functional analysis of Pb14-3-3 was performed in S. cerevisiae, and it was found that Pb14-3-3 partially complemented S. cerevisiae proteins Bmh1p and Bmh2p, which are recognized as two yeast 14-3-3 homologues. When we evaluated the adhesion profile of S. cerevisiae transformants, Pb14-3-3 acted as an adhesin in S. cerevisiae; however, Bmh1p did not show this function. The influence of Pb14-3-3 in S. cerevisiae ergosterol pathway was also evaluated and our results showed that Pb14-3-3 up-regulates genes involved in ergosterol biosynthesis., Conclusions: Our data showed that Pb14-3-3 was able to partially complement Bmh1p and Bmh2p proteins in S. cerevisiae; however, we suggest that Pb14-3-3 has a differential role as an adhesin. In addition, Pb-14-3-3 may be involved in Paracoccidioides spp. ergosterol biosynthesis which makes it an interest as a therapeutic target.

Leptospirosis is a zoonotic disease of global importance, and has a worldwide distribution. Equine leptospirosis is commonly manifested by recurrent uveitis, reproductive disorders, as abortions, embryonic absorption, stillbirth and the birth of weak foals. The aim of this study was to verify the presence of Leptospira sp or its DNA in genital tract of mares with reproductive problems. A total of 38 mares with reproductive problems were studied. All the mares were sampled for blood (for serology), urine (for culturing and qPCR), vaginal fluid-VF and endometrial biopsy-EB (for culturing, qPCR and indirect immunofluorescence). PCRs products were sequenced for secY gene. Seventeen (44.7%) serum samples were reactive, predominantly against serogroups Australis (76.4%) and Pomona (23.6%). No positive culture was obtained, but DNA was detected by qPCR on urine samples (26.3%), VF (44.7%) and EB (18.4%) collected 2 months or longer following diagnosis of early fetal death and endometritis. Leptospira cell aggregations were visible by indirect immunofluorescence on 57.1% (4/7) EBs and 17.6% (3/17) VFs. A total of 18 amplicons showed interpretable sequences. Out of those 18 amplicons, 15 presented 100% of identity with the species L. interrogans (sv Bratislava and Pomona), while three were L. borgpertersenii. This study suggests the presence of leptospires in the uterus of mares with reproductive problems. Moreover, serology was shown not to be indicated for the diagnosis of presumptive Leptospira infection in early gestation. The most common agent of the genital infection in those mares was L. interrogans, most probably sg Australis., (Copyright © 2015 Elsevier B.V. All rights reserved.)

The fungal strain Paracoccidioides brasiliensis remains viable inside of epithelial cells and can induce apoptosis in this population. However, until now, the molecules that participate in this process remained unknown. Thus, this study evaluated the contribution of two P. brasiliensis molecules, the 14-3-3 and glycoprotein of 43 kDa proteins, which had been previously described as extracellular matrix adhesins and apoptosis inductors in human pneumocytes. Accordingly, epithelial cells were treated with these molecules for different periods of time and the expression of the apoptosis regulating-proteins Bak, Bax, Bcl-2, p53 and caspases were evaluated by terminal deoxynucleotidyl transferase dUTP nick end labelling, flow cytometry and real-time polymerase chain reaction analysis. Our results demonstrated that treatment with these molecules induces apoptosis signalling in pulmonary epithelial cells, showing the same pattern of programmed cell-death as that observed during infection with P. brasiliensis. Thus, we could conclude that P. brasiliensis uses these molecules as virulence factors that participate not only in the fungal adhesion process to host cells, but also in other important cellular mechanisms such as apoptosis.

Members of the Paracoccidioides genus are the etiologic agents of paracoccidioidomycosis (PCM). This genus is composed of two species: Paracoccidioides brasiliensis and Paracoccidioides lutzii. The correct molecular taxonomic classification of these fungi has created new opportunities for studying and understanding their relationships with their hosts. Paracoccidioides spp. have features that permit their growth under adverse conditions, enable them to adhere to and invade host tissues and may contribute to disease development. Cell wall proteins called adhesins facilitate adhesion and are capable of mediating fungi-host interactions during infection. This study aimed to evaluate the adhesion profile of two species of the genus Paracoccidioides, to analyze the expression of adhesin-encoding genes by real-time PCR and to relate these results to the virulence of the species, as assessed using a survival curve in mice and in Galleria mellonella after blocking the adhesins. A high level of heterogeneity was observed in adhesion and adhesin expression, showing that the 14-3-3 and enolase molecules are the most highly expressed adhesins during pathogen-host interaction. Additionally, a survival curve revealed a correlation between the adhesion rate and survival, with P. brasiliensis showing higher adhesion and adhesin expression levels and greater virulence when compared with P. lutzii. After blocking 14-3-3 and enolase adhesins, we observed modifications in the virulence of these two species, revealing the importance of these molecules during the pathogenesis of members of the Paracoccidioides genus. These results revealed new insights into the host-pathogen interaction of this genus and may enhance our understanding of different isolates that could be useful for the treatment of this mycosis.

In the present study, we explored the possibility of using the acetylcholinesterase (AChE) as a biomarker after deltamethrin (pyrethroid insecticide) exposure with three strains of the cladoceran Daphnia magna. Four calculated time-weighted deltamethrin concentrations (20.1, 40.3, 80.6 and 161.3 ng L(-1)) were compared against control acetylcholinesterase activity. Our results showed that after 48 h of deltamethrin exposure, all treatments induced a significant decrease of AChE activities whatever the three considered strains. However, diverse responses were registered in terms of lowest observed effect concentrations (LOEC: 80.6 ng L(-1) for strain 1 and 20.1 ng L(-1) for strains 2 and 3) revealing differences in sensitivity among the three tested strains of D. magna. Our results suggest that after deltamethrin exposure, the AChE activity responses can be also used as a biomarker of susceptibility (i.e., variation of strain specific response). Moreover, our results show that strain 1 is the less sensitive in terms of IC50-48 h of AChE, whereas it became the most sensitive when considering the EC50-48 h estimated in the standard ecotoxicity test., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Paracoccidioides species are dimorphic fungi and are the etiologic agents of paracoccidioidomycosis, which is a serious disease that involves multiple organs. The many tissues colonized by this fungus suggest a variety of surface molecules involved in adhesion. A surprising finding is that most enzymes in the glycolytic pathway, tricarboxylic acid (TCA) cycle and glyoxylate cycle in Paracoccidioides spp. have adhesive properties that aid in interacting with the host extracellular matrix and thus act as 'moonlighting' proteins. Moonlighting proteins have multiple functions, which adds a dimension to cellular complexity and benefit cells in several ways. This phenomenon occurs in both eukaryotes and prokaryotes. For example, moonlighting proteins from the glycolytic pathway or TCA cycle can play a role in bacterial pathogenesis by either acting as proteins secreted in a conventional pathway and/or as cell surface components that facilitate adhesion or adherence. This review outlines the multifunctionality exhibited by many Paracoccidioides spp. enzymes, including aconitase, aldolase, glyceraldehyde-3-phosphate dehydrogenase, isocitrate lyase, malate synthase, triose phosphate isomerase, fumarase, and enolase. We discuss the roles that moonlighting activities play in the virulence characteristics of this fungus and several other human pathogens during their interactions with the host.

Transcutaneous electrical nerve stimulation (TENS) is a non-pharmacological therapy for the treatment of pain. The present work investigated the effect of cannabidiol, naloxone and diazepam in combination with 10 Hz and 150 Hz TENS. Male Wistar rats were submitted to the tail-flick test (baseline), and each rodent received an acute administration (intraperitoneal) of naloxone (3.0mg/kg), diazepam (1.5mg/kg) or cannabidiol (0.75 mg/kg, 1.5mg/kg, 3.0mg/kg, 4.5mg/kg, 6.0mg/kg and 12.0mg/kg); 10 min after the acute administration, 10 Hz or 150 Hz TENS or a sham procedure was performed for 30 min. Subsequently, tail-flick measures were recorded over a 90-min period, at 5-min intervals. 10 Hz TENS increased the nociceptive threshold during the 90-min period. This antinociceptive effect was reversed by naloxone pre-treatment, was not altered by diazepam pre-treatment and was abolished by cannabidiol pre-treatment (1.5mg/kg). Moreover, 150 Hz TENS increased tail-flick latencies by 35 min post-treatment, which was partially inhibited by naloxone pre-treatment and totally inhibited by cannabidiol (1.5mg/kg). These data suggest the involvement of the endogenous opioid system and the cannabinoid-mediated neuromodulation of the antinociception induced by transcutaneous electrostimulation at 10 Hz and 150 Hz TENS., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Background: Paracoccidioides spp is a fungi genus and the agent of paracoccidioidomycosis. The strategies of infection used by these pathogens involve the expression of proteins related to adaptation to the host, particularly regarding the uptake of micronutrients. This study analyzed the adhesion of Paracoccidioides lutzii during conditions of copper (Cu) and iron (Fe) deprivation, while also evaluating the proteins expressed in conditions of Cu depletion in the presence of four extracellular matrix (ECM) components (laminin, fibronectin and types I and IV collagen)., Results: We cultured the P. lutzii in a chemically defined media without Cu and Fe. The fungus was then placed in contact with different ECM components and adhesion was evaluated. A significant increase in binding to all ECM components was observed when the fungus was cultured without Cu; which might be related to some adhesins expression. A proteomic assay was developed and revealed 39 proteins expressed that are involved in processes such as virulence, protein synthesis, metabolism, energy, transcription, transport, stress response and the cell cycle when the fungus was interacting with the ECM components. The up-regulated expression of two important adhesins, enolase and 14-3-3, was observed at the fungal cell wall during the interaction with the ECM components, indicating the role of these proteins in the Paracoccidioides-host interaction., Conclusions: This study is important for determining prospective proteins that may be involved in the interaction of Paracoccidioides with a host. Understanding the adaptive response to different growth conditions, elucidating the processes of adhesion and cell invasion, and identifying the proteins that are differentially expressed during the fungus-host interaction may help elucidate mechanisms used for survival and growth of Paracoccidioides in various human tissues.

Although infection by Corynebacterium diphtheriae is a model of extracellular mucosal pathogenesis, different clones have been also associated with invasive infections such as sepsis, endocarditis, septic arthritis and osteomyelitis. The mechanisms that promote C. diphtheriae infection and haematogenic dissemination need further investigation. In this study we evaluated the association and invasion mechanisms with human umbilical vein endothelial cells (HUVECs) and experimental arthritis in mice of endocarditis-associated strains and control non-invasive strains. C. diphtheriae strains were able to adhere to and invade HUVECs at different levels. The endocarditis-associated strains displayed an aggregative adherence pattern and a higher number of internalized viable cells in HUVECs. Transmission electron microscopy (TEM) analysis revealed intracellular bacteria free in the cytoplasm and/or contained in a host-membrane-confined compartment as single micro-organisms. Data showed bacterial internalization dependent on microfilament and microtubule stability and involvement of protein phosphorylation in the HUVEC signalling pathway. A high number of affected joints and high arthritis index in addition to the histopathological features indicated a strain-dependent ability of C. diphtheriae to cause severe polyarthritis. A correlation between the arthritis index and increased systemic levels of IL-6 and TNF-α was observed for endocarditis-associated strains. In conclusion, higher incidence of potential mechanisms by which C. diphtheriae may access the bloodstream through the endothelial barrier and stimulate the production of pro-inflammatory cytokines such as IL-6 and TNF-α, in addition to the ability to affect the joints and induce arthritis through haematogenic spread are thought to be related to the pathogenesis of endocarditis-associated strains.

The present study addressed the effects of sleep deprivation (SD) on AMPA receptor (AMPAR) binding in brain regions associated with learning and memory, and investigated whether treatment with drugs acting on AMPAR could prevent passive avoidance deficits in sleep deprived animals. [(3)H]AMPA binding and GluR1 in situ hybridization signals were quantified in different brain regions of male Wistar rats either immediately after 96 h of sleep deprivation or after 24h of sleep recovery following 96 h of sleep deprivation. Another group of animals were sleep deprived and then treated with either the AMPAR potentiator, aniracetam (25, 50 and 100mg/kg, acute administration) or the AMPAR antagonist GYKI-52466 (5 and 10mg/kg, acute and chronic administration) before passive avoidance training. Task performance was evaluated 2h and 24h after training. A significant reduction in [(3)H]AMPA binding was found in the hippocampal formation of SD animals, while no alterations were observed in GluR1 mRNA levels. The highest dose of aniracetam (100mg/kg) reverted SD-induced impairment of passive avoidance performance in both retention tests, whereas GYKI-52466 treatment had no effect. Pharmacological enhancement of AMPAR function may revert hippocampal-dependent learning impairments produced after SD. We argue that such effects might be associated with reduced AMPAR binding in the hippocampus of sleep deprived animals., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Background: The ability to apply standard and interoperable solutions for implementing and managing medical registries as well as aggregate, reproduce, and access data sets from legacy formats and platforms to advanced standard formats and operating systems are crucial for both clinical healthcare and biomedical research settings., Purpose: Our study describes a reproducible, highly scalable, standard framework for a device registry implementation addressing both local data quality components and global linking problems., Methods and Results: We developed a device registry framework involving the following steps: (1) Data standards definition and representation of the research workflow, (2) Development of electronic case report forms using REDCap (Research Electronic Data Capture), (3) Data collection according to the clinical research workflow and, (4) Data augmentation by enriching the registry database with local electronic health records, governmental database and linked open data collections, (5) Data quality control and (6) Data dissemination through the registry Web site. Our registry adopted all applicable standardized data elements proposed by American College Cardiology / American Heart Association Clinical Data Standards, as well as variables derived from cardiac devices randomized trials and Clinical Data Interchange Standards Consortium. Local interoperability was performed between REDCap and data derived from Electronic Health Record system. The original data set was also augmented by incorporating the reimbursed values paid by the Brazilian government during a hospitalization for pacemaker implantation. By linking our registry to the open data collection repository Linked Clinical Trials (LinkedCT) we found 130 clinical trials which are potentially correlated with our pacemaker registry., Conclusion: This study demonstrates how standard and reproducible solutions can be applied in the implementation of medical registries to constitute a re-usable framework. Such approach has the potential to facilitate data integration between healthcare and research settings, also being a useful framework to be used in other biomedical registries.

Paracoccidoides brasiliensis adhesion to lung epithelial cells is considered an essential event for the establishment of infection and different proteins participate in this process. One of these proteins is a 30 kDa adhesin, pI 4.9 that was described as a laminin ligand in previous studies, and it was more highly expressed in more virulent P. brasiliensis isolates. This protein may contribute to the virulence of this important fungal pathogen. Using Edman degradation and mass spectrometry analysis, this 30 kDa adhesin was identified as a 14-3-3 protein. These proteins are a conserved group of small acidic proteins involved in a variety of processes in eukaryotic organisms. However, the exact function of these proteins in some processes remains unknown. Thus, the goal of the present study was to characterize the role of this protein during the interaction between the fungus and its host. To achieve this goal, we cloned, expressed the 14-3-3 protein in a heterologous system and determined its subcellular localization in in vitro and in vivo infection models. Immunocytochemical analysis revealed the ubiquitous distribution of this protein in the yeast form of P. brasiliensis, with some concentration in the cytoplasm. Additionally, this 14-3-3 protein was also present in P. brasiliensis cells at the sites of infection in C57BL/6 mice intratracheally infected with P. brasiliensis yeast cells for 72 h (acute infections) and 30 days (chronic infection). An apparent increase in the levels of the 14-3-3 protein in the cell wall of the fungus was also noted during the interaction between P. brasiliensis and A549 cells, suggesting that this protein may be involved in host-parasite interactions, since inhibition assays with the protein and this antibody decreased P. brasiliensis adhesion to A549 epithelial cells. Our data may lead to a better understanding of P. brasiliensis interactions with host tissues and paracoccidioidomycosis pathogenesis.

Paracoccidioidomycosis is a systemic mycosis caused by the dimorphic fungus Paracoccidioides brasiliensis. Understanding the interactions between P. brasiliensis and the host tissue depends on the study of the different steps of the process of colonization, especially adhesion, in which the pathogen recognizes ligands on the surface of host cells. This study aimed to verify the role of enolase in the host cell-fungus interaction and the ability of enolase to bind to extracellular matrix components, to determine its subcellular localization, and to study the P. brasiliensis enolase amino acid sequence. The data revealed that fibronectin is the major ligand of enolase. Evaluation of the location of enolase at an ultrastructural level revealed that it is distributed in various cellular compartments, but at a high level in the cell wall. The analysis of the amino acid sequence revealed an internal plasminogen-binding motif ((254)FYKADEKKY(262)), which is conserved in most organisms and described as an important interaction site of the enolase with the host cell surface. This suggests that enolase performs additional functions related to the glycolytic pathway and also plays a role of adhesion in P. brasiliensis. Therefore, this study increases the knowledge about the characteristics of enolase and its influence on the binding process of P. brasiliensis., (© 2012 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.)

The postictal state is generally followed by antinociception. It is known that connections between the dorsal raphe nucleus, the periaqueductal gray matter, and the locus coeruleus, an important noradrenergic brainstem nucleus, are involved in the descending control of ascending nociceptive pathways. The aim of the present study was to determine whether noradrenergic mechanisms in the locus coeruleus are involved in postictal antinociception. Yohimbine (an α(2)-receptor antagonist) or propranolol (a β-receptor antagonist) was microinjected unilaterally into the locus coeruleus, followed by intraperitoneal administration of pentylenetetrazole (PTZ), a noncompetitive antagonist that blocks GABA-mediated Cl(-) influx. Although the administration of both yohimbine and propranolol to the locus coeruleus/subcoeruleus area resulted in a significant decrease in tonic or tonic-clonic seizure-induced antinociception, the effect of yohimbine restricted to the locus coeruleus was more distinct compared with that of propranolol, possibly because of the presynaptic localization of α(2)-noradrenergic receptors in locus coeruleus neurons. These effects were related to the modulation of noradrenergic activity in the locus coeruleus. Interestingly, microinjections of noradrenaline into the locus coeruleus also decrease the postictal antinociception. The present results suggest that the mechanism underlying postictal antinociception involves both α(2)- and β-noradrenergic receptors in the locus coeruleus, although the action of noradrenaline on these receptors causes a paradoxical effect, depending on the nature of the local neurotransmission., (Copyright © 2011 Elsevier Inc. All rights reserved.)

In the present study, we investigated the possible additive effects of epilepsy and aging on the expression of m1 muscarinic acetylcholine receptors (AChR) in the rat hippocampus. Young (3 months) and Aged (20 months) male, Wistar rats were treated with pilocarpine to induce status epilepticus (SE). Immunohistochemical procedure for m1 AChR detection was performed 2 months after pilocarpine-induced SE. In the CA1 pyramidal region m1 AChR staining was significantly decreased in aged epileptic animals when compared to young epileptic and aged control rats, indicating that the aging effect is worsened by the epileptic condition. However, the Nissl-stained cell analysis indicated that the number of pyramidal CA1 neurons was similarly reduced in both epileptic groups, young and aged animals. Therefore, our data suggest that the progressive reduction of m1 AChR expression in CA1 pyramidal cells of aged epileptic rats might bear relevance to the associated progressive cognitive impairment.

Gene expression is disrupted by sepsis. Genetic markers can only reveal a patient's genotype, and they are not affected by environmental biological processes. These processes are expressed by proteins. This study was aimed to advance the insight into the molecular foundations of sepsis. It employed proteomic techniques to identify and analyze differential serum protein expressions taken from a patient throughout the stages of sepsis (sepsis, severe sepsis and septic shock). Serum samples were collected at each stage of sepsis and submitted to one-dimensional electrophoresis, on gradient strips of immobilized pH, followed by two-dimensional 12.5% polyacrylamide gel electrophoresis. The gels obtained were stained, scanned and analyzed by the ImageMasterPlatinum program. Proteins that were differentially expressed in the gels were excised, digested with trypsin and identified through mass spectrometry. Fourteen differentially expressed proteins were identified throughout the stages of sepsis, as well as a protein that was not expressed in all stages, suggesting the potential existence of a biomarker. The differentially expressed proteins identified were: serum amyloid A, apolipoprotein A-1 (2 isoforms), zinc finger protein 222, human albumin, PRO 2619, immunoglobulin kappa light chain VLJ region, monoclonal immunoglobulin M cold agglutinin, 7 proteinase inhibitors - alpha-1 antitrypsin. The findings of this pilot study demonstrate the involvement of the complement and coagulation pathways, of the lipid metabolism and of genetic information in sepsis. The vast majority of proteins identified are involved in the immune system and the proteinase inhibitor proteins are predominant.

The regulation of the gene encoding the extracellular exo-beta-1,3-glucanase (tag83) produced by the mycoparasite Trichoderma asperellum was studied. Enzyme activity was detected in all carbon sources, but the highest levels were found when starch and purified cell walls from Rhizoctonia solani were used. These results are supported by the appearance of one strong band with enzyme activity in non-denaturing PAGE. Experiments using RT-PCR showed that exo-beta-1,3-glucanase induction in T. asperellum occurred at the transcriptional level. We used RT-PCR and real-time PCR analysis to examine the expression of tag83 gene during in vivo assay of T. asperellum against R. solani. We showed that the expression of tag83 is significantly increased by the presence of R. solani.

Hospital wastewater is considered a complex mixture populated with pathogenic microorganisms. The genetic constitution of these microorganisms can be changed through the direct and indirect effects of hospital wastewater constituents, leading to the appearance of antibiotic multi-resistant bacteria. To avoid environmental contamination hospital wastewaters must be treated. The objective of this study was to evaluate the efficiency of hospital wastewater treated by a combined process of biological degradation (septic tank) and the Fenton reaction. Thus, after septic tank biodegradation, batch Fenton reaction experiments were performed in a laboratory-scale reactor and the effectiveness of this sequential treatment was evaluated by a physico-chemical/microbiological time-course analysis of COD, BOD(5), and thermotolerant and total coliforms. The results showed that after 120min of Fenton treatment BOD(5) and COD values decreased by 90.6% and 91.0%, respectively. The BOD(5)/COD ratio changed from 0.46 to 0.48 after 120min of treatment. Bacterial removal efficiency reached 100%, while biotests carried out with Scenedesmus subspicatus and Daphnia magna showed a significant decrease in the ecotoxicity of hospital wastewater after the sequential treatment. The use of this combined system would ensure that neither multi-resistant bacteria nor ecotoxic substances are released to the environment through hospital wastewater discharge.

Cadmium is a nonessential, highly toxic heavy metal that shows ionic properties similar to calcium. These ionic similarities imply that the cadmium ion, Cd2+, is a calcium ion, Ca2+, receptor-agonist, affecting the same biochemical pathways involved in Ca2+ homeostasis. In the yeast Saccharomyces cerevisiae, the PMC1 and PMR1 genes encode vacuolar and Golgi Ca2+-ATPases, respectively. The PMR1 protein product Pmr1p is involved in both Ca2+ and Mn2+ homeostasis. This study investigated the importance of Pmc1p and Pmr1p for Cd2+ cellular detoxification. Using the standard techniques of yeast molecular research and a multielemental procedure named particle-induced X-ray emission, Pmr1p was identified as a protein that directly participates in the detoxification of Cd2+, possibly through the secretory pathway. The results allow us to posit a model of Cd2+ detoxification where Pmr1p has a central role in cell survival in a Cd2+-rich environment.

Background and Objectives: Weaning patients from mechanical ventilation is still a challenge in Intensive Care Units (ICU) and is related to complications and mortality. Daily rounds at the bedside, which are part of good care, can identify patients able to undergo to spontaneous breathing trials. The authors suggest one mnemonic technique with the term "ESTRATEGIA" (strategy) considering some key aspects, as a checklist, which can be applied by any person of the multidisciplinary team, during the bedside rounds, in order to shorten the weaning time., Contents: To introduce the word Strategy as a mnemonic method based on the studies related to weaning from mechanical ventilation and its applicability as a checklist in any intensive care unit by the multidisciplinary team where each letter reminds some key aspects related to the subject., Conclusions: The applicability of a mnemonic mechanism as a checklist for weaning patients from mechanical ventilation easily practiced during the daily round to identify those who are able to undergo to spontaneous breathing trials.

Background and Objectives: Anemia of critical illness is a multifactorial condition caused by blood loss, frequent phlebotomies and inadequate production of red blood cells (RBC). Controversy surrounds the most appropriate hemoglobin concentration "trigger" for transfusion of RBC. We aimed to evaluate transfusion practices in Brazilian ICUs., Methods: A prospective study throughout a 2-week period in 19 Brazilian ICUs. Hemoglobin (Hb) level, transfusion rate, organ dysfunction assessment and 28-day mortality were evaluated. Primary indication for transfusion and pretransfusion hemoglobin level were collected for each transfusion., Results: Two hundred thirty-one patients with an ICU length of stay longer than 48h were included. An Hb level lower than 10 g/dL was found in 33% on admission in the ICU. A total of 348 RBC units were transfused in 86 patients (36.5%). The mean pretransfusion hemoglobin level was 7.7 ± 1.1 g/dL. Transfused-patients had significantly higher SOFA score (7.9 ± 4.6 vs 5.6 ± 3.8, p < 0.05, respectively), days on mechanical ventilation (10.7 ± 8.2 vs 7.2 ± 6.4, p < 0.05) and days on vasoactive drugs (6.7 ± 6.4 vs 4.2 ± 4.0, p < 0.05) than non-transfused patients despite similar APACHE II scores (15.2 ± 8.1 vs 14.2 ± 8.1, NS). Transfused patients had higher mortality rate (43.5%) than non-transfused patients (36.3%) (RR 0.60-1.15, NS). Only one patient (0.28%) had febrile non-hemolytic transfusion and urticarial reactions., Conclusions: Anemia is common in critically ill patients.It seems from the present study that transfusion practices in Brazil have had a more restrictive approach with a lower limit "transfusion trigger".

The present study was carried out in Wistar rats, using the jaw-opening reflex and dental pulp stimulation, to investigate noradrenaline- and serotonin-mediated antinociceptive circuits. The effects of microinjections of bradykinin into the principal sensory trigeminal nucleus (PSTN) before and after neurochemical lesions of the locus coeruleus noradrenergic neurons were studied. Neuroanatomical experiments showed evidence for reciprocal neuronal pathways connecting the locus coeruleus (LC) to trigeminal sensory nuclei and linking monoaminergic nuclei of the pain inhibitory system to spinal trigeminal nucleus (STN). Fast blue (FB) injections in the locus coeruleus/subcoeruleus region retrogradely labeled neurons in the contralateral PSTN and LC. Microinjections of FB into the STN showed neurons labeled in both ipsilateral and contralateral LC, as well as in the ipsilateral Barrington's nucleus and subcoeruleus area. Retrograde tract-tracing with FB also showed that the mesencephalic trigeminal nucleus sends neural pathways towards the ipsilateral PSTN, with outputs from cranial and caudal aspects of the brainstem. In addition, neurons from the lateral and dorsolateral columns of periaqueductal gray matter also send outputs to the ipsilateral PSTN. Microinjections of FB in the interpolar and caudal divisions of the STN labeled neurons in the caudal subdivision of STN. Microinjections in the STN interpolar and caudal divisions also retrogradely labeled serotonin- and noradrenaline-containing nucleus of the brainstem pain inhibitory system. Finally, the gigantocellularis complex (nucleus reticularis gigantocellularis/paragigantocellularis), nucleus raphe magnus and nucleus raphe pallidus also projected to the caudal divisions of the STN. Microinjections of bradykinin in the PSTN caused a statistically significant long-lasting antinociception, antagonized by the damage of locus coeruleus-noradrenergic neuronal fibres with (N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine) (DSP4), a neurotoxin that specifically depleted noradrenaline from locus coeruleus terminal fields. These data suggest that serotonin- and noradrenaline-containing nuclei of the endogenous pain inhibitory system exert a key-role in the antinociceptive mechanisms of bradykinin and the locus coeruleus is crucially involved in this effect.

Background and Objectives: The main cardiovascular function is to maintain the adequate perfusion e oxygen delivery to the cells. Physiologically, this is controlled by the cellular metabolic rate. The critically ill patients are in high danger of tissue hipoperfusion and this is directly related to cellular injury and organ dysfunction. Therefore, the tissue perfusion monitoring makes part and is indissociated of hemodynamic evaluation of the critically ill patient and is indicated to all this patients. The objective was to define recommendations about clinical utility of different tolls to bedside perfusion monitoring., Methods: Modified Delphi methodology was used to create and quantify the consensus between the participants. AMIB indicated a coordinator who invited more six experts in the area of monitoring and hemodynamic support to constitute the Consensus Advisory Board. Twenty five physician and two nurses selected from different regions of the country completed the expert panel, which reviewed the pertinent bibliography listed at the MedLine in the period from 1996 to 2004., Results: Recommendations were done about the utility of clinical monitoring of tissue perfusion, temperature gradient and transcutaneous oxygen monitoring, serum lactate, base excess, SvO² and ScvO², gastric and sublingual capnometry, CO² venous-arterial gradient and Orthogonal Polarization Spectral (OPS)., Conclusions: The homodynamic compensation of a critically ill patient isn’t complete unless the tissue perfusion is corrected. Many different methods of monitoring is available and are useful in clinical practice, however, none has accuracy and effectiveness characteristics to be used independently of clinical context.

Background and Objectives: Shock occurs when the circulatory system cannot maintain adequate cellular perfusion. If this condition is not reverted irreversible cellular injury establishes. Shock treatment has as its initial priority the fast and vigorous correction of mean arterial pressure and cardiac output to maintain life and avoid or lessen organic dysfunctions. Fluid challenge and vasoactive drugs are necessary to warrant an adequate tissue perfusion and maintenance of function of different organs and systems, always guided by cardiovascular monitorization. The recommendations built in this consensus are aimed to guide hemodynamic support needed to maintain adequate tisular perfusion., Methods: Modified Delphi methodology was used to create and quantify the consensus between the participants. AMIB indicated a coordinator who invited more six experts in the area of monitoring and hemodynamic support to constitute the Consensus Advisory Board. Twenty five physician and two nurses selected from different regions of the country completed the expert panel, which reviewed the pertinent bibliography listed at the MEDLINE in the period from 1996 to 2004., Results: Recommendations were made answering 17 questions about hemodynamic support with focus on fluid challenge, red blood cell transfusions, vasoactive drugs and perioperative hemodynamic optimization., Conclusions: Hemodynamic monitoring by itself does not reduce the mortality of critically ill patients, however, we believe that the correct interpretation of the data obtained by the hemodynamic monitoring and the use of hemodynamic support protocols based on well defined tissue perfusion goals can improve the outcome of these patients.

In this work, cadmium phytotoxicity and quantitative sensitivity relationships between different hierarchical endpoints in plants cultivated in a contaminated soil were studied. Thus, germination rate, biomass growth and antioxidative enzyme activity (i.e. superoxide dismutase, peroxidase, catalase and glutathione reductase) in three terrestrial plants (Avena sativa L., Brassica campestris L. cv. Chinensis, Lactuca sativa L. cv. hanson) were analyzed. Plant growth tests were carried out according to an International Standard Organization method and the results were analyzed by ANOVA followed by Williams' test. The concentration of Cd2+ that had the smallest observed significant negative effect (LOEC) on plant biomass was 6.25, 12.5 and 50 mg Cd/kg dry soil for lettuce, oat and Chinese cabbage, respectively. Activity of all enzymes studied increased significantly compared to enzyme activity in plant controls. For lettuce, LOEC values (mg Cd/kg dry soil) for enzymic activity ranged from 0.05 (glutathione reductase) to 0.39 (catalase). For oat, LOEC values (mg Cd/kg dry soil) ranged from 0.19 (for superoxide dismutase and glutathione reductase) to 0.39 (for catalase and peroxidase). For Chinese cabbage, LOEC values (mg Cd/kg dry soil) ranged from 0.19 (peroxidase, catalase and glutathione reductase) to 0.39 (superoxide dismutase). Classical (i.e. germination and biomass) and biochemical (i.e. enzyme activity) endpoints were compared to establish a sensitivity ranking, which was: enzyme activity>biomass>germination rate. For cadmium-soil contamination, the determination of quantitative sensitivity relationships (QSR) between classical and antioxidative enzyme biomarkers showed that the most sensitive plant species have, generally, the lowest QSR values.

Background and Objectives: Sepsis represents the major cause of death in the ICUs all over the world. Many studies have shown an increasing incidence over time and only a slight reduce in mortality. Many new treatment strategies are arising and we should define the incidence and features of sepsis in Brazil., Methods: Prospective cohort study in sixty-five hospitals all over Brazil. The patients who were admitted or who developed sepsis during the month of September, 2003 were enrolled. They were followed until the 28th day and/or until their discharge. The diagnoses were made in accordance to the criteria proposed by ACCP/SCCM. It were evaluated demographic features, APACHE II score, SOFA (Sepsis-related Organ Failure Assessment) score, mortality, sources of infections, microbiology, morbidities and length of stay (LOS)., Results: Seventy-five ICUs from all regions of Brazil took part in the study.3128 patients were identified and 521 (16.7%) filled the criteria of sepsis, severe sepsis or septic shock. Mean age was 61.7 (IQR 39-79), 293 (55.7%) were males, and the overall 28-day mortality rate was 46.6%. Average APACHE II score was 20 and SOFA score on the first day was 7 (IQR 4-10). SOFA score in the mortality group was higher on day 1 (8, IQR 5-11), and had increased on day 3 (9, IQR 6-12). The mortality rate for sepsis, severe sepsis and septic shock was 16.7%, 34.4% and 65.3%, respectively. The average LOS was 15 days (IQR 5-22). The two main sources of infection were the respiratory tract (69%) and the abdomen one (23.1%). Gram-negative bacilli were more prevalent (40.1%). Gram-positive cocci were identified in 32.8% and fungi infections in 5%. Mechanical ventilation was observed in 82.1% of the patients, Swan-Ganz catheter in 18.8%, vasopressors in 66.2% and hemotransfusion in 44.7%., Conclusions: It was observed a high mortality of sepsis in the ICUs in Brazil. The high frequency of septic shock demonstrated a group at high risk of death. In order to have a better use of the resources and reduce in mortality during the next 5 years, it is very important to identify our specific features related to this syndrome.

Background and Objectives: Honeybees first used in the honey production came from Europe. They were gentle but their productivity was very low. In 1956 it was brought from Africa some species of honeybees that were more productible but also extremely aggressive. There was an accidental release of those bees, that proliferated by hybridizing with the European honeybees generating a new specimen of bees: the Africanized honeybees. The objective of this article is to suggest a pattern of treatment for this poisoning syndrome, because of the crescent number of these attacks and a few data about it., Case Report: Male, 19 years old, soldier that in the course of his military training was attacked by a swarm of Africanized honeybees., Conclusions: As the Africanized honeybees are very aggressive, they attack their victims with lots of stings releasing a large quantity of venom. The reactions to the stings can vary from a local inflammatory reaction in non sensibilized people, hipersensibility reaction and anaphylactic shock in sensibilized people and poisoning syndrome when there is a big amount of inoculated venom. The medical attendance to this kind of accident must be as fast as possible. It must be done an adequate clinical support and a quick mechanical remotion of the stingers. The hemodynamic stabilization is a very important point.

Background and Objectives: Cardiac output and preload as absolute data do not offer helpful information about the hemodynamic of critically ill patients. However, monitoring the response of these variables to volume challenge or inotropic drugs is a very useful tool in the critical care setting, particularly for patients with signs of tissue hypoperfusion. Although PAC remains the " gold standard" to measure cardiac output and preload, new and alternative technologies were developed to evaluate these hemodynamic variables., Methods: Modified Delphi methodology was used to create and quantify the consensus between the participants. AMIB indicated a coordinator who invited more six experts in the area of monitoring and hemodynamic support to constitute the Consensus Advisory Board. Twenty three physician and two nurses selected from different regions of the country completed the expert panel, which reviewed the pertinent bibliography listed at the MEDLINE in the period from 1996 to 2004., Results: Recommendations regarding the use of arterial pulse pressure variation during mechanical ventilation, continuous arterial pulse contour and lithium dilution cardiac output measurements, esophageal Doppler waveform, thoracic electrical bioimpedance, echocardiography and partial CO2 rebreathing for monitoring cardiac output and preload were created., Conclusions: The new and less invasive techniques for the measurement of cardiac output, preload or fluid responsiveness are accurate and may be an alternative to PAC in critically ill patients.

Background and Objectives: Monitoring of vital functions is one of the most important tools in the management of critically ill patients. Nowadays is possible to detect and analyze a great deal of physiologic data using a lot of invasive and non-invasive methods. The intensivist must be able to select and carry out the most appropriate monitoring technique according to the patient requirements and taking into account the benefit/risk ratio. Despite the fast development of non invasive monitoring techniques, invasive hemodynamic monitoring using Pulmonary Artery Catheter still is one of the basic procedures in Critical Care. The aim was to define recommendations about clinical utility of basic hemodynamic monitoring methods and the Use of Pulmonary Artery Catheter., Methods: Modified Delphi methodology was used to create and quantify the consensus between the participants. AMIB indicated a coordinator who invited more six experts in the area of monitoring and hemodynamic support to constitute the Consensus Advisory Board. Twenty-five physicians and nurses selected from different regions of the country completed the expert panel, which reviewed the pertinent bibliography listed at the MEDLINE in the period from 1996 to 2004., Results: Recommendations were made based on 55 questions about the use of central venous pressure, invasive arterial pressure, pulmonary artery catheter and its indications in different settings., Conclusions: Evaluation of central venous pressure and invasive arterial pressure, besides variables obtained by the PAC allow the understanding of cardiovascular physiology that is of great value to the care of critically ill patients. However, the correct use of these tools is fundamental to achieve the benefits due to its use.

Simaba ferruginea (Simaroubaceae) is a Brazilian medicinal plant used in traditional medicine to treat several ailments, including gastric ulcers, fever, diarrhea, and dolorous and inflammatory processes. This study examines the chemical composition and antiulcerogenic effects of rhizomes from this plant. Bioassay-guided fractionation led to the isolation of two bioactive indole alkaloids called canthin-6-one (1) and 4-methoxycanthin-6-one (2). The alkaloid fraction and both alkaloids demonstrated potent antiulcerogenic effects when evaluated in gastric lesion-induced animals, as well as significant antinociceptive activity in mice. These results confirm and justify the popular use of S. ferruginea against gastric ulcers and dolorous processes.

Biomass growth, micronucleus induction, and antioxidative stress enzymes (superoxide dismutase, peroxidase, glutathione reductase, and catalase) were investigated simultaneously in the Vicia faba plant exposed to cadmium in solution. The biomass lowest-observed-effect concentration (LOEC) value was 2,000 microM Cd2+. In the shoots, enzymic activities increased without concentration-response relationships. In the roots, after an initial increase, activities of all enzymes showed negative concentration-response relationships. A significant increase in micronucleus induction was observed at 20 microM Cd2+. Regarding sensitivity, our results showed that biomass endpoint was less sensitive than micronucleus induction, which was less sensitive than antioxidative enzyme activities. The increase of antioxidant stress enzyme activities in response to cadmium exposure may be taken as evidence for an enhanced detoxification capacity of V. faba plants toward reactive oxygen species (and derivatives) that might be generated in the stressed plants. Concomitant micronucleus induction may be also interpreted as a consequence of oxidative stress, upholding the view that cadmium-induced DNA damage is, to some extent, via generation of reactive (intermediate) oxygen species.

Background: Few data are available on the risk of aseptic meningitis following vaccination with the Leningrad-Zagreb (L-Z) strain of mumps vaccine. In 1997 the mumps vaccine was introduced into the state of Rio Grande do Sul in Brazil through mass vaccination with mumps-measles-rubella (MMR), targeting children aged 1-11 years. Five municipalities used exclusively MMR vaccine containing the L-Z strain of mumps. An outbreak of aseptic meningitis was observed shortly after the mass campaign., Methods: To estimate the risk of aseptic meningitis associated with this strain, we analysed vaccination and meningitis case surveillance data from the selected municipalities. A case of vaccine-associated aseptic meningitis was defined as one with a pleocytosis of 10-1,500 leukocytes/ml and occurring within 15-35 days after vaccine receipt., Results: We estimated a risk of 2.9 cases per 10,000 doses of L-Z administered, equivalent to 1 case per 3,390 doses administered. The overall risk of aseptic meningitis following the campaign was increased 12.2-fold (95% CI: 6.0-24.7) compared with the same period in 1995-1996. Following the mass campaign, the incidence of mumps declined 93% during 1998-2000., Conclusions: Vaccination with the L-Z strain of mumps vaccine as part of a mass campaign was associated with a significantly increased risk of aseptic meningitis. Decisions about type of mumps vaccine and mumps vaccination strategies must consider vaccine safety issues in addition to other criteria.

Purpose: We studied the surface expression of intracellular adhesion molecule 1 (ICAM-1) on peripheral and intra-ocular T lymphocytes in patients with active uveitis., Methods: Two-colour flow-cytometric analysis was performed on cells isolated from aqueous humour and peripheral blood of 23 patients with active uveitis and 16 control patients who were to undergo cataract extraction, in order to determine the percentage of cells expressing CD4, CD8 and ICAM-1 (CD54) molecules., Results: In the aqueous humour of patients with uveitis, we found an increase in the percentage of CD4+ and CD8+ lymphocytes, co-expressing the ICAM-1 molecule as compared to control patients (p < 0.0001). In peripheral blood, these uveitis patients exhibited a significant decrease in the percentage of CD4+ICAM-1+ (p = 0.0106) and CD8+ICAM-1+ (p = 0.0014) as compared to control subjects. The comparative study of cells from the aqueous humour and peripheral blood showed that the percentage of CD4+ICAM-1+ cells was higher in the aqueous humour (p < 0.0001). Comparison of the aqueous humour and peripheral blood for CD8+ICAM-1+ cells revealed no significant differences. In addition, we found a high negative correlation for the CD8+ICAM-1+ subset between the aqueous humour and peripheral blood., Conclusions: Our results suggest a greater local participation for CD4+ICAM-1+ cells but not for CD8+ICAM-1+ cells in the pathogenesis of uveitis., (Copyright 2002 S. Karger AG, Basel)