Systolic dysfunction by two-dimensional speckle tracking echocardiography in dogs with parvoviral enteritis.

INTRODUCTION/OBJECTIVES: Parvoviral enteritis (PVE) can cause either primary or secondary myocardial injury; the latter is associated with systemic inflammatory response syndrome and sepsis. Strain (St) and strain rate (SR) are relatively new speckle tracking echocardiographic (STE) variables used to assess myocardial function and are less influenced by preload and volume status than are conventional variables. The aim of this study was to evaluate systolic function in dogs with PVE using two-dimensional STE. ANIMALS: Forty-five client-owned dogs were included. MATERIALS AND METHODS: Dogs were classified into four groups: healthy (n = 9), PVE-mild (n = 15), PVE-severe (n = 13) and PVE-died (n = 8). Left ventricular global and segmental myocardial St and SR were assessed in radial, circumferential and longitudinal axes in the right parasternal transverse and apical 4-chamber views. In the circumferential and longitudinal axes, the value of each segment was determined separately at the endocardial and epicardial levels. RESULTS: Compared to healthy animals, all dogs with PVE showed significantly impaired St and SR values, mainly for PVE-severe and PVE-died groups. Moreover, the lowest SR value was observed in the circumferential axis at the mid-septal epicardial segment in the PVE-died group. For this variable, a cut-off value of 0.95 s-1 demonstrated 100% sensitivity and specificity for distinguishing between PVE-severe and PVE-died groups. CONCLUSIONS: In the present study, all dogs with PVE developed systolic dysfunction, which was more severe in non-survivors. Assessment of St and SR in dogs with PVE might be clinically useful for evaluating haemodynamic status and developing suitable therapeutic strategies to improve prognosis.

Clostridium difficile toxins or infection induce upregulation of adenosine receptors and IL-6 with early pro-inflammatory and late anti-inflammatory pattern.

Clostridium difficile causes intestinal inflammation, which increases adenosine. We compared the expression of adenosine receptors (AR) subtypes A1, A2A, A2B, and A3 in HCT-8, IEC-6 cells, and isolated intestinal epithelial cells, challenged or not with Clostridium difficile toxin A and B (TcdA and TcdB) or infection (CDI). In HCT-8, TcdB induced an early A2BR expression at 6 h and a late A2AR expression at 6 and 24 h. In addition, both TcdA and TcdB increased IL-6 expression at all time-points (peak at 6 h) and PSB603, an A2BR antagonist, decreased IL-6 expression and production. In isolated cecum epithelial cells, TcdA induced an early expression of A2BR at 2s and 6 h, followed by a late expression of A2AR at 6 and 24 h and of A1R at 24 h. In CDI, A2AR and A2BR expressions were increased at day 3, but not at day 7. ARs play a role in regulating inflammation during CDI by inducing an early pro-inflammatory and a late anti-inflammatory response. The timing of interventions with AR antagonist or agonists may be of relevance in treatment of CDI.

Alanyl-glutamine attenuates 5-fluorouracil-induced intestinal mucositis in apolipoprotein E-deficient mice

Apolipoprotein E (APOE=gene, apoE=protein) is a known factor regulating the inflammatory response that may have regenerative effects during tissue recovery from injury. We investigated whether apoE deficiency reduces the healing effect of alanyl-glutamine (Ala-Gln) treatment, a recognized gut-trophic nutrient, during tissue recovery after 5-FU-induced intestinal mucositis. APOE-knockout (APOE-/-) and wild-type (APOE+/+) C57BL6J male and female mice (N=86) were given either Ala-Gln (100 mM) or phosphate buffered saline (PBS) by gavage 3 days before and 5 days after a 5-fluorouracil (5-FU) challenge (450 mg/kg, via intraperitoneal injection). Mouse body weight was monitored daily. The 5-FU cytotoxic effect was evaluated by leukometry. Intestinal villus height, villus/crypt ratio, and villin expression were monitored to assess recovery of the intestinal absorptive surface area. Crypt length, mitotic, apoptotic, and necrotic crypt indexes, and quantitative real-time PCR for insulin-like growth factor-1 (IGF-1) and B-cell lymphoma 2 (Bcl-2) intestinal mRNA transcripts were used to evaluate intestinal epithelial cell turnover. 5-FU challenge caused significant weight loss and leukopenia (P<0.001) in both mouse strains, which was not improved by Ala-Gln. Villus blunting, crypt hyperplasia, and reduced villus/crypt ratio (P<0.05) were found in all 5-FU-challenged mice but not in PBS controls. Ala-Gln improved villus/crypt ratio, crypt length and mitotic index in all challenged mice, compared with PBS controls. Ala-Gln improved villus height only in APOE-/- mice. Crypt cell apoptosis and necrotic scores were increased in all mice challenged by 5-FU, compared with untreated controls. Those scores were significantly lower in Ala-Gln-treated APOE+/+ mice than in controls. Bcl-2 and IGF-1 mRNA transcripts were reduced only in the APOE-/--challenged mice. Altogether our findings suggest APOE-independent Ala-Gln regenerative effects after 5-FU challenge.

Alanyl-glutamine attenuates 5-fluorouracil-induced intestinal mucositis in apolipoprotein E-deficient mice.

Apolipoprotein E (APOE=gene, apoE=protein) is a known factor regulating the inflammatory response that may have regenerative effects during tissue recovery from injury. We investigated whether apoE deficiency reduces the healing effect of alanyl-glutamine (Ala-Gln) treatment, a recognized gut-trophic nutrient, during tissue recovery after 5-FU-induced intestinal mucositis. APOE-knockout (APOE-/-) and wild-type (APOE+/+) C57BL6J male and female mice (N=86) were given either Ala-Gln (100 mM) or phosphate buffered saline (PBS) by gavage 3 days before and 5 days after a 5-fluorouracil (5-FU) challenge (450 mg/kg, via intraperitoneal injection). Mouse body weight was monitored daily. The 5-FU cytotoxic effect was evaluated by leukometry. Intestinal villus height, villus/crypt ratio, and villin expression were monitored to assess recovery of the intestinal absorptive surface area. Crypt length, mitotic, apoptotic, and necrotic crypt indexes, and quantitative real-time PCR for insulin-like growth factor-1 (IGF-1) and B-cell lymphoma 2 (Bcl-2) intestinal mRNA transcripts were used to evaluate intestinal epithelial cell turnover. 5-FU challenge caused significant weight loss and leukopenia (P<0.001) in both mouse strains, which was not improved by Ala-Gln. Villus blunting, crypt hyperplasia, and reduced villus/crypt ratio (P<0.05) were found in all 5-FU-challenged mice but not in PBS controls. Ala-Gln improved villus/crypt ratio, crypt length and mitotic index in all challenged mice, compared with PBS controls. Ala-Gln improved villus height only in APOE-/- mice. Crypt cell apoptosis and necrotic scores were increased in all mice challenged by 5-FU, compared with untreated controls. Those scores were significantly lower in Ala-Gln-treated APOE+/+ mice than in controls. Bcl-2 and IGF-1 mRNA transcripts were reduced only in the APOE-/- -challenged mice. Altogether our findings suggest APOE-independent Ala-Gln regenerative effects after 5-FU challenge.

Is pannexin the pore associated with the P2X7 receptor?

The P2X7 receptor (P2X7R), an ATP-gated cation channel, is expressed predominantly in leukocytes. Activation of P2X7R has been implicated in the formation of a cytolytic pore (i.e., a large conductance channel) that allows the passage of molecules up to 900 Da in macrophages. At least two hypotheses have been presented to explain the conversion of a nonselective cation channel to a cytolytic pore. One hypothesis suggests that the pore is a separate molecular structure activated by P2X7R, and the second asserts that this is an intrinsic property of P2X7R (pore dilation). Based on connexin knockout and hemichannel antagonist studies, some groups have concluded that connexins and pannexins, the hemichannel-forming proteins in vertebrates, are fundamental components of the large conductance channel associated with P2X7R. Dye uptake and electrophysiology experiments were used to evaluate the efficacy and specificity of some hemichannel antagonists under conditions known to open the large conductance channel associated with P2X7R. Hemichannel antagonists and interference RNA (RNAi) targeting pannexin-1 did not affect P2X7R macroscopic currents [ATP, 1,570±189 pA; ATP+100 µM carbenoxolone (CBX), 1,498±100 pA; ATP+1 mM probenecid (Prob), 1,522±9 pA] or dye uptake in a FACS assay (ATP, 63±5%; ATP+100 µM CBX, 51.51±8.4%; ATP+1 mM Prob, 57.7±4.3%) in mouse macrophages. These findings strongly suggest that the high-permeability pore evident after prolonged P2X7R activation does not occur through connexin or pannexin hemichannels in murine macrophages. Another membrane protein may be involved in P2X7R pore formation.

Modeling the Praia dos Ingleses submarine outfall, Florianópolis, Santa Catarina, Brazil.

An outfall 1,800 m long was initially proposed by CASAN (Water and Sanitation Company of Santa Catarina) to dispose of treated domestic residual water effluent from Praia dos Ingleses beach. The final plan average flow, estimated for the summer season is 0.1543 m³/s. The characteristics on the near field of the diffuser were calculated using the NRFIELD model supplied by a series of current profiles and density, gathered at the diffuser location between March and April, 2007. Reliable information of raw sewage fecal coliform concentration and nutrients has been supplied by CASAN, based on continuous monitoring of sewage in its treatment plants. The diffuser was designed in such a way that it maximizes the initial dilution allowing the nutrients concentration to comply with legislation limits. However, results showed that the treatment plant should be designed to mainly reduce total phosphorus. A hydrodynamic model was implemented to generate current fields for the far field modeling. Information gathered in the field on variation in water level, wind, and current velocity and direction was used for boundary and/or initial conditions. Then, a Lagrangian advective-diffusive model was used to evaluate the extent of the plumes. The results of far field modeling showed a need to increase the outfall length. A new location 3,300 m from the beach has been proposed.

Determination of diflubenzuron residues in milk and cattle tissues / Determinação de resíduos de diflubenzuron em leite e tecidos bovinos

Diflubenzuron (DFB) is used to control ectoparasitic infestation by inhibiting larvae development in the manure and feces of treated animals. It is also currently been used to control tick infestations. In this study, milk and tissues from cattle treated orally with DFB for a 77-120 day period with a commercial product containing the compound were analyzed for the presence of residues. DFB residues were determined by using extraction with acetonitrile, cleanup with C18 SPE and chromatographic analysis by HPLC with UV detection (254nm). DFB was not detected in any of the analysed samples (<0.006mg kg-1 for fat, <0.014mg kg-1 for muscle, <0.015mg kg-1 for kidney, <0.016mg kg-1 for liver and <0.0006mg kg-1 for milk). In this manner, the use of this compound, according to the manufacturer's suggested doses may result in cattle milk, liver, kidneys, fat and muscles being considered safe regarding the presence of DFB residues.

O diflubenzuron (DFB) é um inibidor de desenvolvimento de insetos que inibe a síntese de quitina com atividade ovicida e larvicida e está sendo utilizado na pecuária para o controle do carrapato. Leite e tecidos provenientes de bovinos tratados por um período de 77 a 120 dias com um produto comercial contendo DFB foram analisados quanto à presença de resíduos. Os resíduos de DFB foram determinados utilizando-se extração com acetonitrila, limpeza por SPE C18 e cromatografia líquida de alta eficiência com detecção por UV (254nm). DFB não foi detectado em nenhuma das amostras analisadas (<0.006mg kg-1 para gordura, <0,014mg kg-1 para músculo, <0,015mg kg-1 para rim, <0,016mg kg-1 para fígado e <0.0006mg kg-1 para leite). Dessa forma, a utilização do princípio ativo conforme recomendado pelo fabricante e em níveis suficientes para se obter o efeito larvicida desejado deve resultar em leite, fígado, rins, gordura e músculos que podem ser considerados seguros para o consumo em termos da presença DFB.

Alterações histopatológicas da glândula mamária e qualidade do leite de cabras naturalmente infectadas com o CAEV / Histopathological changes in the mammary gland and milk quality of goats naturally infected with CAEV

Avaliou-se a influência do vírus da CAE nas características físico-químicas de amostras de leite de 54 cabras, sem predileção racial, distribuindo-as em dois grupos: cabras positivas e negativas para o teste de imunodifusão em gel de agarose. As amostras de leite foram submetidas à análise ultrassônica para obtenção de parâmetros físico-químicos - gordura, extrato seco, proteínas, lactose e densidade; realização de microbiologia - bactérias mesófilas (UCF/mL). Foram coletadas amostras de tecido mamário para exame histopatológico e imunohistoquímica. Não houve diferença significativa das características avaliadas entre os dois grupos; no microbiológico, não houve relação direta da presença de mesófilas associada à infecção pelo CAEV. Na histopatologia, observaram-se áreas com infiltração celular de monócitos, polimorfonucleares, plasmócitos, fibrose, ausência de morfologia normal do parênquima mamário, denotando processo inflamatório crônico; e foi confirmada a presença do vírus na glândula pela imunohistoquímica. Os resultados não mostraram relação direta da incidência da CAE como fator negativo no desenvolvimento do rebanho.

Aiming to evaluate the influence of CAE viruses in the chemical and physical characteristics of milk, the samples were collected from 54 goats, without racial predilection, these were divided into two groups: goats positive and negative according results of test Agarose Gel Immunodiffusion. Milk samples were ultrasonic analyzed to obtain physicochemical parameters (fat, solids, protein, lactose and density); performance microbiology (mesophilic bacteria - CFU/mL) and mammary gland samples were collected for evaluation histopathology and immunohistochemistry. The results of physical-chemical analysis showed no significant difference between the milk samples of two groups. In the microbiological analysis showed the presence of aerobic mesophilic bacteria, but this change is not associated with the presence of CAEV infection. On histopathology, there were areas with infiltration of mononuclear-leukocyte and polymorph nuclear, plasma cells, fibrosis and absence of normal morphology of the mammary tissue, indicating a chronic inflammatory process; and confirmed the presence of virus, in the gland, by immunohistochemistry. The results showed no direct relationship between incidence of CAE in the herd as a negative factor for its development, however it is known that the disease in its chronic nature, causes reduction in the productivity of the herd.

Produção de antígeno e separação da proteína p28 por microfiltragem seriada para sorodiagnóstico da artrite encefalite caprina por ensaio imunoenzimático / Production of antigen and p28 protein separation by microfiltration serial for serodiagnosis of caprine arthritis encephalitis by enzyme immunoassay

Este estudo teve como objetivo produzir um antígeno (Ag) a partir de cultura de células de membrana sinovial caprina (MSC) infectadas com o vírus de artrite encefalite caprina (CAEV), pela técnica de microfiltração seriada, substituindo a ultracentrifugação em colchão de sacarose (UCCS) para utilização em ELISA indireto (ELISA-i). Amostras de 188 soros caprinos, que previamente foram testados pelo Western blot (WB) com Ag UCCS, foram submetidas à análise pelo ELISA-i com o novo antígeno produzido, que mostrou concordância de 92% em relação ao antígeno UCCS. A sensibilidade e a especificidade do ELISA em relação ao WB foram de 95,6% e 88,5%, respectivamente. A nova técnica, criada a partir de microfiltrações, mostrou-se efetiva e de baixo custo para o diagnóstico sorológico de anticorpos para CAEV em comparação ao antígeno ultracentrifugado, e constitui uma alternativa viável para produção de antígeno purificado de lentivírus de pequenos ruminantes.

This study aimed to produce an antigen (Ag) from the culture of goat synovial membrane cells (MSC) infected by CAEV through serial microfiltering technique replacing ultra ultracentrifugation in sacarosis Mattress (UCCS) for the indirect diagnosis ELISA tests (i ELISA). Samples of 188 sera from goats previously examined by Western Blot (WB) with Ag UCCS were submitted to analysis by i ELISA with new antigen produced, demonstrating an accordance of 92% in relation to UCCS antigen. The specificity and sensitivity relating to WB were of 95,65% and 88, 5% respectively. The new technique created from the microfiltering is effective and with low cost for the serological antibodies diagnosis of CAEV comparing to the ultracentrifuged one, presenting, therefore, as a viable alternative for purified antigen of lentivirus in small ruminants.

Clinical applicability of natural product(s)-containing mouthwashes as adjunctive treatment of biofilm-induced gingivitis: a systematic review / Aplicabilidade clínica de colutórios à base de produtos naturais como tratamento adjuvante da gengivite induzida por biofilme: uma revisão sistemática

Natural products have emerged as an effective and low-cost alternative for treating various diseases of the oral cavity. This study aimed to evaluate, through a systematic literature review, if there is scientific evidence ensuring the safe and effective use of natural product(s)-containing mouthwashes as adjunctive treatment of biofilm-induced gingivitis. Searches were conducted in the databases Medline, SciELO, LILACS and Cochrane Library, by using combinations of the key words gingivitis/natural products/phytotherapy/mouthwash, in English, Portuguese and Spanish. Studies published until September 2010 were considered. Four examiners analyzed independently: study design and phase, methodological quality (Jadad scale - JE), experimental product and its concentration, dosing interval and time of usage, as well as employed statistical analysis and clinical outcome of interest. From the 503 articles found, 08 were included in the final review as phase II, controlled, randomized and blind clinical trials, scoring 4 (25%) and 5 (75%) in JE. The main natural products assessed were: Azadirachta indica, Garcinia mangostana, Lippia sidoides, Salvadora persica and Sesamum indicum whose concentration, dosing interval, time of usage and adverse effects varied according to each study. The Plaque and Gingival Index were most employed, as well as α = 5% and paired t, Student's t, Wilcoxon and Mann-Whitney tests. A total of 62.5% and 50% of the products significantly reduced supragingival biofilm and gingivitis, respectively. Mouthwashes containing the essential oil from the leaves of L. sidoides (1%) and the extract from the leaves of A. indica (25%) can be indicated as adjunctive treatment of biofilm-induced gingivitis.

Os produtos naturais têm surgido como alternativa eficaz e de baixo custo para o tratamento de várias doenças da cavidade oral. Objetivou-se avaliar, a partir de revisão sistemática da literatura, se há evidências científicas garantindo a utilização segura e eficaz de antissépticos bucais contendo produto(s) natural(is) como tratamento adjuvante da gengivite induzida por biofilme. Foram realizadas buscas nas bases de dados Medline, SciELO, LILACS e Cochrane Library, através de combinações usando as palavras-chave gengivite/produtos naturais/ fitoterápicos/bochechos, em Inglês, Português e Espanhol. Consideraram-se os estudos publicados até setembro de 2010. Quatro examinadores analisaram separadamente: desenho e fase do estudo, qualidade metodológica (escala de Jadad - EJ), produto experimental e a concentração, intervalo de administração e tempo de uso, bem como a análise estatística empregada e os resultados clínicos de interesse. Foram encontrados 503 artigos dos quais 08 foram incluídos na revisão final como sendo ensaios clínicos fase II, controlados, randomizados e cegos, marcando 4 (25%) e 5 (75%) na EJ. Os principais produtos naturais avaliados foram Azadirachta indica, Garcinia mangostana, Lippia sidoides, Salvadora persica e Sesamum indicum, cuja concentração, intervalo de administração, tempo de uso, e efeitos adversos, variaram de acordo com cada estudo. Índice de placa e Índice Gengival foram os mais utilizados, bem como α=5% e testes t-pareado, t-Student, Wilcoxon e Mann-Whitney. 62,5% e 50% dos produtos reduziram significativamente a presença de bioflme supragengival e gengivite, respectivamente. Os colutórios contendo o óleo essencial das folhas de L. sidoides (1%) e o extrato das folhas de A. indica (25%) podem ser indicados como tratamento adjuvante da gengivite induzida por biofilme.

Prevalência de anticorpos contra Leptospira spp. em bovinos, caninos, equinos, ovinos e suínos do município de Jaguapitã, estado do Paraná, Brasil / Prevalence of antibodies against Leptospira spp. in cattle, dogs, horses, sheep and swine of Jaguapitã, state of Paraná, Brazil

O objetivo deste trabalho foi determinar a prevalência de anticorpos contra Leptospira spp. em bovinos, caninos, equinos, ovinos e suínos, oriundos de 40 propriedades localizadas na área rural do Município de Jaguapitã, Estado do Paraná. Foram colhidas amostras de sangue de 370 bovinos, 161 equinos, 70 ovinos, 230 suínos e 97 caninos. O número de animais testados em cada propriedade, assim como o número de propriedades, foi determinado utilizando-se o programa Epi-info versão 6. As amostras de soros obtidas foram submetidas à prova de soroaglutinação microscópica (SAM) com 22 sorovares de Leptospira spp. Das 40 propriedades rurais pesquisadas, 38 (95,00%) tiveram pelo menos um animal sororeagente na SAM e dos 928 animais estudados, 316 (34,08%) apresentaram títulos ? 100. A prevalência observada na espécie bovina foi de 42,43%, com 87,18% das propriedades apresentando pelo menos um animal sororeagente. As prevalências de animais e propriedades reagentes para as demais espécies foram, respectivamente, 48,44% e 87,18% para equinos; 38,57% e 100% para ovinos; 18,70% e 28,00% para suínos; 11,34% e 31,25% para cães. O sorovar mais provável encontrado em bovinos foi Hardjo, em equinos Castellonis e Sentot, em ovinos, suínos e cães Icterohaemorrhagiae. Os resultados obtidos neste trabalho demonstram que as cinco espécies animais estudadas na área rural do Município de Jaguapitã tiveram contato com vários sorovares de Leptospira spp. Além disso, os resultados sorológicos sugerem uma possível transmissão do micro-organismo entre espécies animais, provavelmente em decorrência da exposição às mesmas fontes de infecção entre os animais estudados.

The objective of this work was to determine the prevalence of antibodies against Leptospira spp. in cattle, dogs, horses, sheep and swine from 40 properties located in the rural area of Jaguapitã, state of Paraná, Brazil. Blood samples were taken from 370 cattle, 97 dogs, 161 horses, 70 sheep and 230 swine. The number of animals tested on each property, as well as the number of properties was determined using the program Epi-info version 6. Samples of serum were submitted to microscopic agglutination test (MAT) with 22 Leptospira spp. serovars. From the 40 rural properties investigated, 38 (95.00%) had at least one positive animal according to SAM, and from 928 studied animals, 316 (34.08%) presented titers ? 100. The prevalence observed in the bovine species was 42.43%, with 87.18% of the properties presenting at least one positive animal. The prevalence of animals and properties reactive for the other species were, respectively: 48.44% and 87.18% for horses; 38.57% and 100% for sheep; 18.70% and 28.00% for swine; 11.34% and 31.25% for dogs. The most frequent serovar in bovines was Hardjo, in horses Castellonis and Sentot, in sheep, swine and dogs Icterohaemorrhagiae. The results obtained in this study demonstrated that the 5 studied animal species in the rural area of Jaguapitã had contact with several Leptospira spp. serovars. Moreover, serological results suggest a possible transmission of Leptospira spp. between animal species, probably because of exposure to the same sources of infection among the animals studied.

Pharmacological properties of a pore induced by raising intracellular Ca2+.

Recent studies on the P2X(7) receptor in 2BH4 cells and peritoneal macrophages have demonstrated that the raise in intracellular Ca(2+) concentration induces a pore opening similar to P2X(7) receptor pore. Herein, we have investigated whether the pore activated by the elevation of intracellular Ca(2+) concentration is associated to P2X(7) receptor. Using patch clamp in cell attached, whole cell configuration, and dye uptake, we measured the pore opening in cell types that express the P2X(7) receptor (2BH4 cells and peritoneal macrophages) and in cells that do not express this receptor (HEK-293 and IT45-RI cells). In 2BH4 cells, the stimulation with ionomycin (5-10 microM) increased intracellular free Ca(2+) concentration and induced pore formation with conductance of 421 +/- 14 pS, half-time (t(1/2)) for ethidium bromide uptake of 118 +/- 17 s, and t(1/2) for Lucifer yellow of 122 +/- 11 s. P2X(7) receptor antagonists did not block these effects. Stimulation of HEK-293 and IT45-RI cells resulted in pore formation with properties similar to those found for 2BH4 cells. Connexin hemichannel inhibitors (carbenoxolone and heptanol) also did not inhibit the pore-induced effect following the increase in intracellular Ca(2+) concentration. However, 5-(N,N-hexamethylene)-amiloride, a P2X(7) receptor pore blocker, inhibited the induced pore. Moreover, intracellular signaling modulators, such as calmodulin, phospholipase C, mitogen-activated protein kinase, and cytoskeleton components were important for the pore formation. Additionally, we confirmed the results obtained for electrophysiology by using the flow cytometry, and we discarded the possibility of cellular death induced by raising intracellular Ca(2+) at the doses used by using lactate dehydrogenase release assay. In conclusion, increased concentration in intracellular Ca(+2) induces a novel membrane pore pharmacologically different from the P2X(7) associated pore and hemigap-junction pore.

Adaptação do threshold IMT para teste de resistencia dos musculos inspiratorios / Adaptation of ThresholdIMT for endurance test on inspiratory muscles

Para avalaicao da resistencia muscular inspirtoria atraves de dispositivos do tipo threshold loading estudos anteriores empregaram equipamentos artesanais e indisponiveis comercialmente. Objetivos: verificar a aplicabilidade e reprodutibilidade de um teste de resistencia muscular inspiratoria utilizando a adaptacao de um antigo modelo do THRESHOLD IMT (nao disponivel comercialmente) com ampla gama de pressoes, avalaindo sua resistencia intrinseca ao fluxo inspiratorio, sua caracteristica limiar pressorica e sua relativa independencia do fluxo aereo na geracao de pressao durante os testes. Metodos: um antigo modelo do THRESHOLD IMT foi aberto, retirado sua mola interna e adaptado seu embolo. Apos montagem vertical, um sistema de peso externo foi instalado como gerador de carga. Valvulas foram instaladas para evitar re-inalacao de gas carbonico. Essa adaptacao foi testada aplicando-se diversas razoes de fluxosem a presenca do embolo e de carga para avaliacao da resistencia intrinseca e aplicando-se diversas razoes de fluxo quando instaladas cargas de 10 ate 130 cmH2O para avaliar a real pressao de abertura da valvula e o comportamento pressorico em altas razoes de fluxo. Resultados: a resistencia media foi de 4,8+-0,7 cmH2O/L.s-1, mas teve seu valor aumentado proporcionalmente ao fluxo. A pressao real de abertura foi inferior ao valor esperado (p=0,002), condicao encontrada em outros estudos. O comportamento pressorico apos a abertura da valvula foi similar em todas as cargas: um aumento medio de 3,8+-2,0 cmH2O para cada 1,0 L/S de fluxo gerado. Conclusao: a adaptacao proposta do THRESHOLD IMT produziu um dispositivo barato e confiavel para emprego em testes de resistencia muscular inspiratoria

Estudo preliminar da atividade antiinflamatória de Bryophillum calycinum Salisb / Preliminar study of the anti-inflammatory activity of Bryophillum calycinum Salisb

Foram avaliados a toxicidade aguda (DL50) do Extrato Bruto Aquoso Liofilizado das folhas de Bryophillum calycinum Salisb. (EBALBc) e seu efeito antiinflamatório sobre o edema de pata de rato induzido por carragenina e dextrana. Com a dose de 500 mg/kg (p.o) inibiu o edema de pata induzido por dextrana de maneira significativa (p < 0.05, ANOVA, Teste de Student Newman-Keuls) nos tempos de 60 e 90 minutos, enquanto que somente a dose oral de 1 g/kg de EBALBc inibiu o edema de pata induzido por carragenina. Os resultados indicaram efeito anti-edematogênico do extrato quando testado sobre o edema de pata induzido por dextrana e carragenina, sugerindo, entretanto, maior especificidade de ação sobre o edema induzido por dextrana. Por via oral, com as doses de 0,1 a 8 g/kg o EBALBc, não desencadeou óbito, não sendo possível determinar a DL50.

The acute toxicity (LD50) and the anti-inflammatory effect of the crude freeze-dried aqueous extract of the leaves of Bryophillumcalycinum Salisb. (EBALBc) was evaluated, on the rat paw edema induced by carrageenin and dextran. The dose of 500 mg/kg (p.o) inhibited the paw edema induced by dextran in a significative manner (p < 0.05, ANOVA, Student Newman-Keuls test) 60 and 90 minutes, after stimulus while only the oral dose of 1 g/kg of EBALBc inhibited the paw edema induced by carrageenin. The results indicated an anti-edematogenic effect of the extract when tested on the paw edema induced by dextran and carrageenin, suggesting larger specificity of action on the edema induced by dextran. The EBALBc administered orally, in the doses of the 0.1 to 8 g/kg, it did not cause death, making impossible to determine the LD50.

A novel form of cellular communication among thymic epithelial cells: intercellular calcium wave propagation.

We here describe intercellular calcium waves as a novel form of cellular communication among thymic epithelial cells. We first characterized the mechanical induction of intercellular calcium waves in different thymic epithelial cell preparations: cortical 1-4C18 and medullary 3-10 thymic epithelial cell lines and primary cultures of thymic "nurse" cells. All thymic epithelial preparations responded with intercellular calcium wave propagation after mechanical stimulation. In general, the propagation efficacy of intercellular calcium waves in these cells was high, reaching 80-100% of the cells within a given confocal microscopic field, with a mean velocity of 6-10 microm/s and mean amplitude of 1.4- to 1.7-fold the basal calcium level. As evaluated by heptanol and suramin treatment, our results suggest the participation of both gap junctions and P2 receptors in the propagation of intercellular calcium waves in thymic nurse cells and the more prominent participation of gap junctions in thymic epithelial cell lines. Finally, in cocultures, the transmission of intercellular calcium wave was not observed between the mechanically stimulated thymic epithelial cell and adherent thymocytes, suggesting that intercellular calcium wave propagation is limited to thymic epithelial cells and does not affect the neighboring thymocytes. In conclusion, these data describe for the first time intercellular calcium waves in thymic epithelial cells and the participation of both gap junctions and P2 receptors in their propagation.

Characterization of P2 receptors in thymic epithelial cells.

The presence of P2 receptors was investigated in three distinct preparations of murine thymic epithelial cells (TEC): 2BH4 murine cell line, IT45-R1 rat cell line, and a primary murine cell derived from the Nurse cell lympho-epithelial complex. In all preparations, application of ATP to the extracellular milieu triggered intracellular calcium signals indicating the presence of P2 receptor(s) in these cells. After an initial peak of calcium concentration, a plateau phase that could last more than 10 min was frequently observed. Ion replacement and channel blockage experiments indicated that the initial peak was associated with the release of calcium from intracellular stores, while the plateau phase was associated with an influx from the extracellular medium. ATP and UTP induced similar calcium signals, suggesting the presence of P2Y2 receptors in all three cell types. The murine 2BH4 cells also expressed P2X7/P2Z receptor, since under exposure to millimolar concentrations of ATP, a continuous rise in intracellular calcium concentration was observed and their plasma membranes became permeabilized to the fluorescent dyes Lucifer yellow and ethidium bromide. In addition, this permeabilization phenomenon was blocked by the P2Z-specific antagonist, oxidized ATP. RT-PCR assays confirmed the presence of mRNAs for the P2Y2 molecule in all TEC, while mRNA for the P2X7 molecule was detected only in 2BH4 cells. Our data indicate that P2Y2 purinergic receptors are widely expressed by thymic epithelial cells, whereas the expression of the P2X7 receptor appears to be more restricted, raising the possibility that its expression is related only to a particular epithelial microenvironment within/the thymus.

Oral lysine clonixinate in the acute treatment of migraine: a double-blind placebo-controlled study.

Several oral nonsteroidal anti-inflammatory drugs (NSAIDs) are effective to treat migraine attacks. Lysine clonixinate (LC) is a NSAID derived from nicotinic acid that has proven to be effective in various pain syndromes such as renal colic and muscular pain. The aim of this double-blind, placebo-controlled study was to evaluate the efficacy of oral LC compared to placebo in the acute treatment of migraine. Sixty four patients with the diagnosis of migraine, according to the IHS criteria, were studied prospectively. Patients received LC or placebo once the headache reached moderate or severe intensity for 6 consecutive attacks. With regard to the moderate attacks, LC was superior than placebo after 1, 2 and 4 hours. The consumption of other rescue medications after 4 hours was significantly higher in the placebo group. With regard to the severe attacks, there was no difference between the active drug group and the placebo group concerning headache intensity and consumption of other rescue medications. We conclude that the NSAID lysine clonixinate is effective in treating moderately severe migraine attacks. It is not superior than placebo in treating severe migraine attacks.

Pharmacologic properties of P(2Z)/P2X(7 )receptor characterized in murine dendritic cells: role on the induction of apoptosis.

In the immune system, extracellular adenosine 5'-triphosphate (ATP) mediates a variety of effects mainly through activation of a particular receptor subtype, the pore-forming P(2Z)/P2X(7) purinoceptor. This purinergic receptor has been described chiefly in cells of hemopoietic origin such as T cells, thymocytes, monocytes, macrophages, and phagocytic cells of thymic reticulum. In this study, we characterized the P(2Z)/P2X(7) purinoceptor and the ATP-mediated apoptosis in murine spleen-derived dendritic cells (DCs). Dye uptake and apoptosis were evaluated by flow cytometry. ATP-treated DCs were permeable to different low-molecular-weight fluorescent probes such as ethidium bromide, YO-PRO 1, and lucifer yellow. Such an effect was dose-dependent (EC(50): 721 micromol/L); mediated by the fully anionic agonist (ATP(4-)); and specifically stimulated by ATP, BzATP, and ATPgammaS. Additionally, an ATP-induced increase in intracellular calcium was detected by microfluorometry. Furthermore, ATP treatment induced a significant increase in apoptotic DCs (64. 46% +/- 3.8%) when compared with untreated control cells (34% +/- 5. 8%), as ascertained by the TdT-mediated dUTP nick end labeling technique. Both ATP-induced DC permeabilization and apoptosis were inhibited by oxidized ATP, a P(2Z)/P2X(7)-specific antagonist. In conclusion, we characterized the expression of the P(2Z)/P2X(7) purinoceptor in murine spleen-derived DCs and described its role on the induction of apoptosis.

Procedures to characterize and study P2Z/P2X7 purinoceptor: flow cytometry as a promising practical, reliable tool.

The expression of P2Z/P2X7 purinoceptor in different cell types is well established. This receptor is a member of the ionotropic P2X receptor family, which is composed by seven cloned receptor subtypes (P2X1 - P2X7). Interestingly, the P2Z/P2X7 has a unique feature of being linked to a non-selective pore which allows the passage of molecules up to 900 Da depending on the cell type. Early studies of P2Z/P2X7 purinoceptor were exclusively based on classical pharmacological studies but the recent tools of molecular biology have enriched the analysis of the receptor expression. The majority of assays and techniques chosen so far to study the expression of P2Z/P2X7 receptor explore directly or indirectly the effects of the opening of P2Z/P2X7 linked pore. In this review we describe the main techniques used to study the expression and functionality of P2Z/P2X7 receptor. Additionally, the increasing need and importance of a multifunctional analysis of P2Z/P2X7 expression based on flow cytometry technology is discussed, as well as the adoption of a more complete analysis of P2Z/P2X7 expression involving different techniques.