RESUMO
Cut-off values for anti-dsDNA, anti-nucleosome and anti-C1q antibodies tests and for complementmediated hemolytic activity (CH50) were explored to identify patients with high risk of developing severe lupus nephritis (LN). Forty-one patients with confirmed systemic lupus erythematosus (SLE) were identified; their levels for the three antibodies and complement had been measured on a same serum sample. These patients were classified based on the presence of renal involvem ent; sixteen had active proliferative LN. With the cut-off values accepted in the laboratory for SLE diagnosis (anti-dsDNA > 100 UI/ml, anti-nucleosome > 50 U/ml or CH50 < 190 UCH50%) no significant differences were found between patients with and without LN. Anti-C1q > 40 U/ml showed a statistically significant association with LN and had 80% of specificity. Cut-off values for LN identified by Receiver Operating Characteristic curves (ROC) were higher for anti-dsDNA (> 455 IU/ml) and antinucleosome (>107 U/ml), lower for CH50 (< 150 UCH50%) and, for anti-C1q (> 41 U/ml) coincided with the cut-off values accepted for SLE. Anti-C1q > 134 U/ml had a 92% of specificity, 56% of sensibility and was associated with a fifteen-fold increased risk of LN. The simultaneous presence of anti-nucleosome > 107 U/ml and anti-C1q > 134 U/ml was associated with a 27-fold higher probability for LN. According to these results, the cut-off values used to detect SLE activity could be inadequate to identify patients at high risk of severe LN.
Assuntos
Testes Imunológicos/normas , Nefrite Lúpica/sangue , Nefrite Lúpica/diagnóstico , Adolescente , Adulto , Anticorpos Antinucleares/sangue , Biomarcadores/sangue , Criança , Pré-Escolar , Complemento C1q/imunologia , Ensaio de Atividade Hemolítica de Complemento/métodos , Ensaio de Atividade Hemolítica de Complemento/normas , Feminino , Humanos , Testes Imunológicos/métodos , Lactente , Lúpus Eritematoso Sistêmico/sangue , Lúpus Eritematoso Sistêmico/diagnóstico , Lúpus Eritematoso Sistêmico/imunologia , Nefrite Lúpica/imunologia , Masculino , Pessoa de Meia-Idade , Nucleossomos/imunologia , Padrões de Referência , Estudos Retrospectivos , Medição de Risco/métodos , Fatores de Risco , Sensibilidade e Especificidade , Índice de Gravidade de Doença , Adulto JovemRESUMO
Cut-off values for anti-dsDNA, anti-nucleosome and anti-C1q antibodies tests and for complement-mediated hemolytic activity (CH50) were explored to identify patients with high risk of developing severe lupus nephritis (LN). Forty-one patients with confirmed systemic lupus erythematosus (SLE) were identified; their levels for the three antibodies and complement had been measured on a same serum sample. These patients were classified based on the presence of renal involvem ent; sixteen had active proliferative LN. With the cut-off values accepted in the laboratory for SLE diagnosis (anti-dsDNA > 100 UI/ml, anti-nucleosome > 50 U/ ml or CH50 < 190 UCH50%) no significant differences were found between patients with and without LN. Anti-C1q > 40 U/ml showed a statistically significant association with LN and had 80% of specificity. Cut-off values for LN identified by Receiver Operating Characteristic curves (ROC) were higher for anti-dsDNA (> 455 IU/ml) and anti-nucleosome (>107 U/ml), lower for CH50 (< 150 UCH50%) and, for anti-C1q (> 41 U/ml) coincided with the cut-off values accepted for SLE. Anti-C1q > 134 U/ml had a 92% of specificity, 56% of sensibility and was associated with a fifteen-fold increased risk of LN. The simultaneous presence of anti-nucleosome > 107 U/ml and anti-C1q > 134 U/ml was associated with a 27-fold higher probability for LN. According to these results, the cut-off values used to detect SLE activity could be inadequate to identify patients at high risk of severe LN.
Se exploraron valores de corte para los ensayos de anti-ADNdc, anti-nucleosoma, anti-C1q y complemento hemolítico total (CH50) capaces de identificar los casos con mayor riesgo de nefritis lúpica (NL) grave. Se seleccionaron 41 pacientes ≥ 16 años con lupus eritematoso sistémico (LES) confirmado que tenían titulados los niveles de los tres anticuerpos y CH50, en una misma muestra de suero. Fueron clasificados según presencia de compromiso renal; 16 presentaron formas proliferativas de NL activa. Con los valores de corte aceptados por el laboratorio para el diagnóstico de LES (anti-ADNdc > 100 UI/ml, anti-nucleosoma > 50 U/ml o un CH50 < 190 UCH50%) no se encontraron diferencias significativas entre casos con y sin NL. Un anti-C1q > 40 U/ml tuvo una especificidad del 80% y mostró una asociación estadísticamente significativa con NL. Al aplicar curvas Receiver Operating Characteristic (ROC) para NL, se identificaron valores de corte más altos para anti-ADNdc (> 455 IU/ml) y anti-nucleosoma (> 107 U/ml), más bajo para CH50 (< 150 UCH50%) y para el anti-C1q (> 41 U/ml) coincidió con el aceptado para diagnóstico de LES. Un anti-C1q > 134 U/ml presentó una sensibilidad del 56%, una especificidad del 92% y se asoció con quince veces más riesgo de NL. La presencia simultánea de anti-C1q > 134 U/ml y anti-nucleosoma > 107 U/ml se asoció 27 veces más riesgo de NL. De acuerdo a estos resultados los valores de corte empleados para actividad en pacientes con LES podrían resultar inadecuados para identificar pacientes con mayor riesgo de NL grave.
Assuntos
Humanos , Masculino , Feminino , Lactente , Pré-Escolar , Criança , Adolescente , Adulto , Pessoa de Meia-Idade , Adulto Jovem , Testes Imunológicos/normas , Nefrite Lúpica/sangue , Padrões de Referência , Índice de Gravidade de Doença , Testes Imunológicos/métodos , Nefrite Lúpica/diagnóstico , Nucleossomos/imunologia , Biomarcadores/sangue , Complemento C1q/imunologia , Ensaio de Atividade Hemolítica de Complemento/métodos , Ensaio de Atividade Hemolítica de Complemento/normas , Anticorpos Antinucleares/sangue , Estudos Retrospectivos , Fatores de Risco , Sensibilidade e Especificidade , Medição de Risco/métodos , Lúpus Eritematoso Sistêmico/imunologia , Lúpus Eritematoso Sistêmico/sangueRESUMO
Abstract In the study presented here, a new series of 2-furyl(4-{4-[(substituted)sulfonyl]benzyl}-1-piperazinyl)methanone derivatives was targeted. The synthesis was initiated by the treatment of different secondary amines (1a-h) with 4-bromomethylbenzenesulfonyl chloride (2) to obtain various 1-{[4-(bromomethyl)phenyl]sulfonyl}amines (3a-h). 2-Furyl(1-piperazinyl)methanone (2-furoyl-1-piperazine; 4) was then dissolved in acetonitrile, with the addition of K2CO3, and the mixture was refluxed for activation. This activated molecule was further treated with equi-molar amounts of 3a-h to form targeted 2-furyl(4-{4-[(substituted)sulfonyl]benzyl}-1-piperazinyl)methanone derivatives (5a-h) in the same reaction set up. The structure confirmation of all the synthesized compounds was carried out by EI-MS, IR and 1H-NMR spectral analysis. The compounds showed good enzyme inhibitory activity. Compound 5h showed excellent inhibitory effect against acetyl- and butyrylcholinesterase with respective IC50 values of 2.91±0.001 and 4.35±0.004 µM, compared to eserine, a reference standard with IC50 values of 0.04±0.0001 and 0.85±0.001 µM, respectively, against these enzymes. All synthesized molecules were active against almost all Gram-positive and Gram-negative bacterial strains tested. The cytotoxicity of the molecules was also checked to determine their utility as possible therapeutic agents.
Assuntos
Simulação por Computador/estatística & dados numéricos , Anti-Infecciosos/análise , Piperazinas/análise , Ensaio de Atividade Hemolítica de Complemento , Colinesterases/farmacologiaRESUMO
abstract A series of N-substituted 2-{[5-(1H-indol-3-ylmethyl)-1,3,4-oxadiazol-2-yl]sulfanyl}acetamides (8a-w) was synthesized in three steps. The first step involved the sequential conversion of 2-(1H-indol-3-yl)acetic acid (1) to ester (2) followed by hydrazide (3) formation and finally cyclization in the presence of CS2 and alcoholic KOH yielded 5-(1H-indole-3-yl-methyl)-1,3,4-oxadiazole-2-thiol (4). In the second step, aryl/aralkyl amines (5a-w) were reacted with 2-bromoacetyl bromide (6) in basic medium to yield 2-bromo-N-substituted acetamides (7a-w). In the third step, these electrophiles (7a-w) were reacted with 4 to afford the target compounds (8a-w). Structural elucidation of all the synthesized derivatives was done by 1H-NMR, IR and EI-MS spectral techniques. Moreover, they were screened for antibacterial and hemolytic activity. Enzyme inhibition activity was well supported by molecular docking results, for example, compound 8q exhibited better inhibitory potential against α-glucosidase, while 8g and 8b exhibited comparatively better inhibition against butyrylcholinesterase and lipoxygenase, respectively. Similarly, compounds 8b and 8c showed very good antibacterial activity against Salmonella typhi, which was very close to that of ciprofloxacin, a standard antibiotic used in this study. 8c and 8l also showed very good antibacterial activity against Staphylococcus aureus as well. Almost all compounds showed very slight hemolytic activity, where 8p exhibited the least. Therefore, the molecules synthesized may have utility as suitable therapeutic agents.
resumo Uma série de acetamidas 2-{[5-(1H-indol-3-ilmetil)-1,3,4-oxadiazol-2-il]sulfanila} N-substituídas (8a-w) foi sintetizada em três fases. A primeira etapa envolveu a conversão sequencial de ácido 2-(1H-indol-3-il)acético (1) a éster (2), seguido por hidrazida (3) e, finalmente, a e ciclização na presença de CS2 e KOH alcoólico produziu 5-(1H-indol-3-il- metil)-1,3,4-oxadiazole-2-tiol (4). Na segunda etapa, aminas arílicas/aralquílicas(5a-w) reagiram com brometo de 2-bromoacetila (6), em meio básico, para se obter acetamidas 2-bromo-N-substituídas (7a-w). Na terceira etapa, estes eletrófilos (7a- w) reagiram com 4, para se obter os compostos alvo (8a-w). A elucidação estrutural de todos os derivados sintetizados foi realizada por 1H-NMR, IR e técnicas de espectrometria de EI-MS. Além disso, eles foram submetidos a triagem de atividade antibacteriana e hemolítica. Análise da inibição enzimática foi bem apoiada pelos resultados de docking molecular. Por exemplo, o composto 8q exibiu melhor potencial inibitório contra α-glicosidase, e os compostos 8g e 8b exibiram, comparativamente, melhor inibição contra butirilcolinesterase (BChE) elipoxigenase (LOX), respectivamente. Do mesmo modo os compostos 8b e 8c mostraram excelente potencial antibacteriano contra SalmonellaTyphi, semelhante ao do ciprofloxacino, antibiótico padrão usado neste estudo. Os compostos 8c e 8l também mostraram excelente potencial antibacteriano contra Staphylococcus aureus . Quase todos os compostos mostraram pequena atividade hemolítica, sendo que o composto 8p apresentou menor atividade. Assim, as moléculas sintetizadas podem ter a sua utilidade como agentes terapêuticos adequados.
Assuntos
Ácido Hidroxi-Indolacético/análise , Acetamidas/análise , Butirilcolinesterase/análise , Ensaio de Atividade Hemolítica de Complemento/classificação , Lipoxigenases/farmacocinética , Glicosídeo Hidrolases/farmacocinéticaRESUMO
Blood levels of regulators of the complement system in preterm babies were reported in few studies only. The aim of this study was to set up a complement profile in premature and term babies focusing on the development of blood levels of MBL, key regulatory proteins and on classical pathway activity, which may allow an estimation of potential susceptibility to infection. Complement activity (CH50), levels of mannan-binding lectin (MBL), complement regulators (factors H and I, C1 inhibitor, properdin) and C3a as marker of complement activation were assessed in three groups of healthy newborns: (1) prematures (≤34 weeks); (2) late prematures (>34-<37 weeks) and (3) term neonates (≥37 weeks). CH50 increased with gestational age with lower titres in cord blood than in day 5 post-delivery venous blood. MBL concentrations were not significantly different among groups. Quantitative and functional C1 inhibitor were below adult normal range in prematures <34 weeks and lower in cord blood as compared to day 5. Factor I, factor H and properdin remained below adult values in all groups. Low C3a levels excluded that low complement titres were due to activation-induced consumption. These results demonstrate the relative immaturity of the complement system and its regulation, especially in premature infants.
Assuntos
Proteína Inibidora do Complemento C1/metabolismo , Complemento C3a/metabolismo , Nascimento Prematuro/imunologia , Adulto , Ativação do Complemento , Proteína Inibidora do Complemento C1/genética , Complemento C3a/genética , Fator H do Complemento/genética , Fator H do Complemento/metabolismo , Ensaio de Atividade Hemolítica de Complemento , Feminino , Fibrinogênio/genética , Fibrinogênio/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Idade Gestacional , Humanos , Recém-Nascido , Lectina de Ligação a Manose/genética , Lectina de Ligação a Manose/metabolismo , Gravidez , Properdina/genética , Properdina/metabolismoRESUMO
Background: Previous works had shown that scorpion venom induced neurotransmitter elevation and an inflammatory response associated with various anatomo-pathological modifications. The most dangerous scorpions species in Algeria responsible for these effects are Androctonus australis hector (Aah) and Androctonus amoreuxi (Aam). Results: Comparison of the physiopathological effects induced by the two venoms showed differences in the kinetic of cytokine release and in lung injury. The lung edema was only observed in response to Aah venom and it was correlated with cell infiltration. In order to better understand the involved mechanism in inflammatory response, we used two antagonists, atropine (non-selective muscarinic antagonist) and propranolol (ß adrenergic antagonist), which lead to a decrease of cell infiltration but has no effect on edema forming. Conclusion: These results suggest another pathway in the development of lung injury following envenomation with Aam or Aah venom.(AU)
Assuntos
Animais , Masculino , Feminino , Pele/metabolismo , Bufo rana , Hemólise/fisiologia , Anfíbios/fisiologia , Ensaio de Atividade Hemolítica de Complemento , Técnica de Placa Hemolítica/métodos , OsmorregulaçãoRESUMO
The impact of agrichemicals on aquatic vertebrate species has been a matter of increasing concern to researchers and environmentalist. In the present study, we evaluated the effects of a sublethal concentration of atrazine (10% of the LC(50-96 h)), a world-wide used herbicide, on the innate immune system of silver catfish (Rhamdia quelen). A significant reduction on phagocytic index, bacteria agglutination and bactericidal activity of the serum, serum lysozyme and total serum peroxidase activity was observed in fish exposed to atrazine for 24 h. After 10 days exposure to atrazine, only bactericidal activity of the serum, bacteria agglutination and total serum peroxidase activity were significantly reduced. Atrazine had no effect on the natural complement hemolytic activity. Our results demonstrate that atrazine decreases the innate immune response of fingerlings, which might increase its susceptibility to opportunistic pathogens.
Assuntos
Atrazina/toxicidade , Peixes-Gato/imunologia , Peixes-Gato/microbiologia , Doenças dos Peixes/imunologia , Herbicidas/toxicidade , Imunidade Inata , Infecções por Actinomycetales/imunologia , Infecções por Actinomycetales/veterinária , Aeromonas hydrophila/fisiologia , Animais , Aquicultura , Ensaio de Atividade Hemolítica de Complemento/veterinária , Relação Dose-Resposta a Droga , Feminino , Infecções por Bactérias Gram-Negativas/imunologia , Infecções por Bactérias Gram-Negativas/veterinária , Masculino , Micrococcus luteus/fisiologia , Muramidase/sangue , Peroxidase/sangue , Poluentes Químicos da Água/toxicidadeRESUMO
En el ensayo de microlinfocitotoxicidad, el suero de conejo como fuente de complemento desempeña un rol esencial en la clasificación del complejo mayor de histocompatibilidad humano, tanto para la tipificación antigénica como en el cross-match linfocitario de la pareja donante-receptor antes de realizar los trasplantes. En este trabajo, se obtuvieron 3 lotes experimentales de dicho hemoderivado con óptimos indicadores de rendimiento, actividad biológica y estabilidad. El exanguíneo de los animales se realizó por la técnica de yugulación. El suero fue separado por centrifugación en condiciones ambientales y de temperatura controladas. Posteriormente, el producto se sometió a filtración esterilizante y se tomaron muestras para los ensayos de calidad. El estudio mostró que el producto conserva la actividad biológica al menos durante 18 meses y que los valores de citotoxicidad se encuentran dentro de los límites de aceptación para su empleo en los ensayos de histocompatibilidad pretrasplante
Microlinfocitotoxicity assay is one of the serological methods used to classify human major histocompatibility complex. In this technique the rabbit´s serum as complement source, plays an essential role in antigens determination and in lymphocytes cross match assay as necessary stage before the selection of the best donor-receiver pair to realize the organ transplant. Three experimental lots of normal rabbit serum were developed with excellent indicators of efficiency, biological activity and stability. Rabbit's blood was obtained through jugulating technique and serum was separated by centrifugation in controlled environment and temperature´s conditions. The sterilization process was performed by filtration and the samples were taken for quality´s control assays. Stability studies showed that the product kept the biological activity at least during 18 months after it was processed and cytotoxicity's values were adequate for its employment in histocompatibility pre-transplant assays
Assuntos
Coelhos , Ensaio de Atividade Hemolítica de Complemento/métodos , Antígenos de Histocompatibilidade , Transplante de Órgãos , Teste de Histocompatibilidade/métodosRESUMO
Liposomes have been employed as potential drug carriers. However, after their in vivo administration, they can be destabilized by proteins of complement system, contributing to the clearance of vesicles from blood circulation. Antioxidant flavonoids such as quercetin have been reported to be beneficial to human health, but their low water solubility and bioavailability limit their enteric administration. Therefore, the development of appropriate flavonoid-carriers could be of great importance to drug therapy. The aim of the present study was to evaluate the activation of human complement system proteins by liposomes composed of soya phosphatidylcholine (SPC) and cholesterol (CHOL) or cholesteryl ethyl ether (CHOL-OET) loaded with quercetin or not. The consumption of complement, via classical (CP) and alternative (AP) pathways, by different vesicles was evaluated using a hemolytic assay and quantitative determination of iC3b and natural antibodies deposited on empty liposomal surfaces by ELISA. The main results showed that empty liposomes composed of large amounts of CHOL consumed more complement components than the others for both CP and AP. Furthermore, replacement of CHOL with CHOL-OET reduced complement consumption via both CP and AP. Incorporation of quercetin did not change CP and AP consumption. Deposition of iC3b, IgG and IgM in vesicles composed of SPC:CHOL-OET at a molar ratio of 1.5:1 was lower compared to the others. Taken together, these observations suggest that liposomes composed of SPC:CHOL-OET at a molar ratio of 1.5:1 are the most appropriate among the vesicles studied herein to be used as a drug carrier system in further investigations.
Assuntos
Ativação do Complemento , Portadores de Fármacos/química , Lipossomos/química , Quercetina/química , Animais , Ensaio de Atividade Hemolítica de Complemento , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Lipossomos/sangue , Masculino , Quercetina/sangue , Coelhos , Valores de Referência , Ovinos , Propriedades de SuperfícieRESUMO
Opportunistic fungi are those that normally would not cause diseases in otherwise healthy people, but are able to cause problems under some circumstances, and for this they need to possess a certain virulence potential. The objective of this study was to identify samples of filamentous fungi isolated from poultry barns in Cascavel, Paraná, and also to evaluate their virulence potential by assessing proteinase production, hemolytic activity, urease production, and growth rate at 37 ºC. We have evaluated the following samples: Acremonium hyalinulum (1 sample), Aspergillus sp. (12), Beauveria bassiana (1), Curvularia brachyspora (1), Paecilomyces variotti (1), and Penicillium sp. (2). Out of the 18 samples analyzed, 44.4 percent showed proteolytic activity using albumin as the substrate versus 66.7 percent when using casein; 66.7 percent showed hemolytic activity, 83.3 percent were positive for urea, and 88.9 percent grew at a temperature of 37 ºC. The results demonstrated that the majority of the isolates expressed virulence factors. Therefore, these isolates have the potential to harm human hosts, such as those working at poultry barns, especially predisposed or susceptible individuals.
Fungos oportunistas são aqueles que normalmente não causariam doenças em pessoas saudáveis, mas eles são capazes de causar problemas sob certas circunstâncias e, para isso, eles necessitam possuir algum potencial de virulência. O objetivo deste trabalho foi identificar amostras de fungos filamentosos isolados de granjas de aves em Cascavel, Paraná, e também avaliar o seu potencial de virulência, verificando a produção de proteinase, atividade hemolítica, produção de urease e crescimento a 37 ºC. Foram avaliados Acremonium hyalinulum (01), Aspergillus sp (12), Beauveria bassiana (01), Curvularia brachyspora (01), Paecylomices variotti (01) e Penicillium sp (02). Das 18 amostras, 44,4 por cento apresentaram atividade proteolítica usando como substrato a albumina e 66,7 por cento com caseína; 66,7 por cento demonstraram atividade hemolítica, 83,3 por cento foram uréia positivas e 88,9 por cento cresceram em temperatura de 37 ºC. Os resultados demonstram que a maioria dos isolados expressaram fatores de virulência e, portanto, têm potencial para causar danos a hospedeiros humanos como os trabalhadores dos aviários, sobretudo em indivíduos predispostos ou suscetíveis.
Assuntos
Animais , Aves Domésticas/análise , Brasil , Fatores de Virulência/análise , Fungos/virologia , Ensaio de Atividade Hemolítica de Complemento , Fungos/classificação , Peptídeo Hidrolases/química , Urease/química , VirulênciaRESUMO
Using agrichemicals to control unwanted species has become a necessary and common worldwide practice to improve crop production. Although most currently used agrichemicals are considered relatively safe, continuous usage contributes for soil and water contamination and collateral toxic effects on aquatic species. Few studies correlated the presence of agrichemicals on fish blood cells and natural immune system. Thus, in this study, silver catfish (Rhamdia quelen) were exposed to sublethal concentrations (10% of the LC(50-96 h)) of a glyphosate based herbicide and hematological and natural immune system parameters were evaluated. Silver catfish fingerlings exposed to glyphosate for 96 h had a significant reduction on blood erythrocytes, thrombocytes, lymphocytes and total leukocytes in contrast to a significant increase in the number of immature circulating cells. The effect of glyphosate on natural immune system was evaluated after 24h or 10 days exposure by measuring the phagocytic index of coelomic cells, and lysozyme, total peroxidase, bacteria agglutination, bactericidal activity and natural complement hemolytic activity in the serum of fingerlings. A significant reduction on phagocytic index, serum bacteria agglutination and total peroxidase was observed only after 24h exposure to glyphosate. In contrast, fingerlings exposed to glyphosate for 10 days had a significant lower serum bacteria agglutination and lysozyme activity. Glyphosate had no effect on serum bactericidal and complement natural hemolytic activity after 24h or 10 days exposure. Nonetheless, the information obtained in this study indicates that glyphosate contaminated water contributes to alter blood cells parameters and to reduce the activity of natural immune components important to mediate fish resistance to infecting microorganisms.
Assuntos
Peixes-Gato , Doenças dos Peixes/induzido quimicamente , Glicina/análogos & derivados , Herbicidas/toxicidade , Poluentes Químicos da Água/toxicidade , Aeromonas , Animais , Ensaio de Atividade Hemolítica de Complemento/veterinária , Relação Dose-Resposta a Droga , Feminino , Doenças dos Peixes/sangue , Doenças dos Peixes/imunologia , Glicina/administração & dosagem , Glicina/toxicidade , Herbicidas/administração & dosagem , Masculino , Muramidase/sangue , Peroxidase/sangue , Fatores de Tempo , GlifosatoRESUMO
The dimorphic fungus Paracoccidioides brasiliensis is the etiological agent of paracoccidioidomycosis, a human granulomatous disease. Recently the first case of natural disease in dogs was reported. The complement system is an important effector component of humoral immunity against infectious agents. Therefore, the aim of this study was to evaluate the activation of the dog alternative complement pathway by P. brasiliensis. Initially, the ability of erythrocytes of guinea pig, rabbit, sheep, chicken and swine to activate the dog alternative pathway was evaluated. The guinea pig erythrocytes showed the greatest capacity to activate dog alternative pathway. The alternative (AH50) hemolytic activity was evaluated in 27 serum samples from healthy dogs and the mean values were 87.2 AH50/ml. No significant differences were observed in relation to sex and age. The alternative pathway activation by P. brasiliensis was higher in serum samples from adult dogs when compared to puppies and aged dogs (p < 0.05). This is the first report of dog alternative complement pathway activation by P. brasiliensis and suggests that it may play a protective role in canine paracoccidioidomycosis.
O fungo dimórfico Paracoccidioides brasiliensis é o agente etiológico da paracoccidioidomicose, uma doença granulomatosa humana. Recentemente, foi relatado o primeiro caso da doença natural em cães. O sistema complemento é um importante componente efetor da imunidade humoral contra agentes infecciosos. Portanto, o objetivo deste trabalho foi avaliar a ativação da via alternativa do complemento canina pelo P. brasiliensis. Inicialmente, foi avaliada a capacidade de eritrócitos de cobaia, coelho, carneiro, galinha e suíno ativarem a via alternativa do complemento canino. Os eritrócitos de cobaia apresentaram maior capacidade de ativar a via alternative do complemento canino. A atividade hemolítica da via alternativa (AH50) foi avaliada em 27 amostras de soro de cães saldáveis e os valores médios observados foram de 87,2 AH50/ml. Não foi observada diferença significativa ao sexo e idade. A ativação da via alternativa pelo P. brasiliensis foi maior nas amostras de soro de cães adultos quando comparada aos cães filhotes e idosos (p < 0.05). Este é o primeiro relato da ativação da via alternative do complemento canino pelo fungo P. brasiliensis e sugere que pode ter um papel protetor na paracoccidioidomicose canina.
Assuntos
Animais , Cães , Formação de Anticorpos , Ensaio de Atividade Hemolítica de Complemento , Eritrócitos , Paracoccidioidomicose , Cães , Métodos , Técnicas e Procedimentos DiagnósticosRESUMO
Malnutrition modifies the hostfis resistance to infection, altering many physiological processes, including hematopoiesis and immunological functions. In this study, we evaluated the total complement system and its C3 component factor in animals subjected to a protein malnutrition model with lipopolysaccharide (LPS) stimulation. The cellularity of blood, bone marrow and spleen, as well as the production of C3 and CH50, were evaluated. Two-month old male Swiss mice were submitted to protein malnutrition with a low-protein diet containing 4% protein. A control group received a control diet containing 20% protein. When the experimental group had reached a loss of about 20% in their original body mass, they were intravenously inoculated with LPS. Cells in the blood, bone marrow and spleen were counted and the circulating levels of C3 and CH50 were evaluated in these animals. Malnourished animals presented anemia, leucopenia, and a severe reduction in bone marrow and spleen cellularity. The survival rate of the malnourished animals was lower, as well as the production of C3 and CH50, if compared to the control animals. These findings suggest that malnourished mice present a deficient response to LPS. The decrease in the complement synthesis may be partially responsible for the immunodeficiency observed. These data lead us to conclude that the nutritional status interferes in the immune response activation.
La desnutrición proteico-energética modifica la resistencia a la infección, alterando numerosos procesos fisiológicos, incluyendo la hematopoyesis y la función inmunológica. En este estudio medimos las concentraciones séricas del factor C3 y del Sistema Complemento Total (CH50) en ratos con desnutrición proteico-energética estimulados con lipopolisacárido (LPS). Fue evaluada la celularidad periférica, medular y esplénica. Ratos Swiss, machos, adultos, con dos meses de edad fueron sometidos a desnutrición proteica con una dieta de 4% de proteína .El grupo control consumía una dieta con 20% de proteínas. Cuando los animales desnutridos perdieron 20% de su peso inicial, fueron inoculados por vía endovenosa con LPS. Fueron determinadas las concentraciones de células sanguíneas, de la médula ósea, del bazo y los valores de C3 y CH50 circulantes en los animales estimulados. Los animales desnutridos presentaron anemia y leucopenia además de una reducción significativa de celularidad de la médula ósea y del bazo. La sobrevivencia de este grupo era menor y también eran más bajas las concentraciones del factor C3 del complemento y del complemento total, en relación a los animales del grupo control. Los resultados sugieren que animales desnutridos muestran una respuesta deficiente a LPS. La síntesis menor de complemento puede ser en parte responsable por la inmunodeficiencia observada. Estos resultados nos conducen a inferir que la desnutrición proteico-energética interfiere en la activación de la respuesta inmune.
A desnutrição proteico-energética modifica a resistência à infecção, modi? cando diversos processos fisiológicos, incluindo a hematopoiese e as funções imunológicas. Neste estudo, avaliamos as concentrações séricas do fator C3 e do Sistema Complemento total (CH50) em um modelo no qual camundongos submetidos à desnutrição proteico-energética são estimulados com lipopolissacarídeo (LPS), e avaliamos a celularidade periférica, medular e esplênica. Camundongos Swiss, machos, adultos, com dois meses de idade foram submetidos ao processo de desnutrição proteica com uma dieta contendo 4% de proteína em comparação aos animais controles com uma dieta contendo 20% de proteína. Quando os animais do grupo desnutrido alcançaram aproximadamente 20% de perda de peso, em relação ao inicial, foram inoculados endovenosamente com LPS. As células do sangue, da medula óssea e do baço foram quantificadas, bem como as concentrações circulantes de C3 e CH50 em animais estimulados com LPS. Os animais desnutridos apresentaram anemia e leucopenia, além de redução significativa da celularidade da medula óssea e do baço. Os animais desnutridos apresentaram menor taxa de sobrevivência, bem como das concentrações do fator C3 do complemento e do complemento total em relação aos animais controles. Estes resultados sugerem que animais desnutridos apresentam uma resposta deficiente aos LPS. A síntese menor do complemento pode ser em parte responsável pela imunodeficiência observada. Estes resultados conduzem-nos a inferir que a desnutrição proteico-energética interfere na ativação da resposta imune.
Assuntos
Animais , /biossíntese , Desnutrição Proteico-Calórica , Lipopolissacarídeos , Muridae/sangue , Ensaio de Atividade Hemolítica de Complemento , Interpretação Estatística de DadosRESUMO
This work evaluated a crude hydroalcoholic extract (ExT) from the pulp of the tamarind (Tamarindus indica) fruit as a source of compounds active on the complement system (CS) in vitro. ExT, previously characterized by other authors, had time and concentration dependent effects on the lytic activity of the CS. The activity of 0.8 mg/mL of the extract on the classical/lectin pathways (CP/LP) increased after 30 min of pre-incubation, while that of the alternative pathway (AP) decreased after 15 min at 1mg/mL. Since the CS is a mediator of inflammation, studies were also made in vivo, taking advantage of a model of hypercholesterolemia in hamsters to investigate the role of CS in the phase preceding the inflammatory process of atherosclerosis. Hamsters submitted to a diet rich in cholesterol showed increased lytic activity of the CP/LP and AP after 45 days. The activity levels of C2 and factor B components on respectively, classical/lectin and alternative pathways of the CS also increased. Early events cooperating to trigger the process of atherosclerotic lesions are not completely understood, and these alterations of complement may participate in these events. When treatment with a diet rich in cholesterol was associated to the furnishing of ExT, evaluation of complement components and complement lytic activity showed values similar to those of the controls, showing that treatment with ExT blocked the increase of complement activity caused by the cholesterol-rich diet. By itself, ExT had no effect on the complement system in vivo. ExT activity on the CS may be of interest for therapy and research purposes.
Assuntos
Complemento C2/imunologia , Fator B do Complemento/imunologia , Via Alternativa do Complemento/efeitos dos fármacos , Via Clássica do Complemento/efeitos dos fármacos , Hiperlipidemias/imunologia , Extratos Vegetais/farmacologia , Tamarindus/química , Alanina Transaminase/sangue , Animais , Aspartato Aminotransferases/sangue , Colesterol/sangue , Complemento C2/metabolismo , Fator B do Complemento/metabolismo , Ensaio de Atividade Hemolítica de Complemento , Cricetinae , Frutas/química , Hiperlipidemias/tratamento farmacológico , Masculino , Mesocricetus , Estatísticas não Paramétricas , Triglicerídeos/sangueRESUMO
The scorpion Tityus serrulatus is considered one of the most dangerous species in Brazil. Its venom evokes an inflammatory response, although the exact mechanism of this effect is still unknown. The aim of the present study was to investigate the effect of Tityus serrulatus venom (TsV) on the complement system (CS) and on leukocyte recruitment. Complement consumption by TsV was evaluated using in vitro hemolytic assays, immunoelectrophoresis and two-dimensional immunoelectrophoresis of complement components (factor B and C3). In order to evaluate neutrophil migration induced in normal human serum (NHS) in the presence of TsV, in vitro chemotaxis assays were performed using the Boyden chamber model. In vitro TsV induced a concentration- and time-dependent reduction in hemolytic activity of the classical/lectin and alternative complement pathways, with samples of 43.0 microg and 43.4 microg, respectively, inhibiting 50% of the lytic activity. Alterations in C3 and factor B electrophoretic mobility after incubation of NHS with TsV, were identical to those obtained with zymosan (positive control). Incubation of NHS with TsV induced neutrophil chemotaxis similar to that observed with zymosan-activated serum. Our results show that TsV activates the CS, leading to factor B and C3 cleavage, to reduction of serum lytic activity and generation of complement chemotactic factors. Therefore, CS may play an important role in the inflammatory response observed upon scorpion envenomation.
Assuntos
Ativação do Complemento/efeitos dos fármacos , Leucócitos/efeitos dos fármacos , Venenos de Escorpião/farmacologia , Animais , Quimiotaxia de Leucócito/efeitos dos fármacos , Complemento C3/metabolismo , Fator B do Complemento/metabolismo , Ensaio de Atividade Hemolítica de Complemento , Via Alternativa do Complemento/efeitos dos fármacos , Via Clássica do Complemento/efeitos dos fármacos , Eritrócitos/efeitos dos fármacos , Feminino , Humanos , Imunoeletroforese Bidimensional , Técnicas In Vitro , Lectinas , Masculino , Coelhos , OvinosRESUMO
OBJETIVO: Avaliar a atividade funcional das vias clássica e alternativa do sistema complemento e os níveis de C3, C4 e fator B durante o primeiro episódio de infecção meningocócica e durante a convalescença. PACIENTES E MÉTODOS: Dez crianças brasileiras com idades entre 8 meses e 8 anos, admitidas de 1991 a 1993, com diagnóstico clínico-laboratorial de meningite meningocócica, foram estudadas durante infecção aguda (até 7 dias do diagnóstico) e no período de convalescença (entre 1 e 6 meses após). C3, C4 e fator B foram quantificados por nefelometria e a atividade lítica das vias clássica e alternativa foi avaliada por método cinético e expressa como tempo necessário para lisar 50% de uma suspensão de eritrócitos (T1/2, expresso em segundos). Baixos valores de T1/2 das vias clássica e alternativa se correlacionam com elevadas atividades de via clássica e via alternativa, respectivamente. RESULTADOS: Observaram-se diferenças significativas entre a atividade lítica da via alternativa durante a infecção e no período de convalescença (282 e 238 segundos, respectivamente, P= .01). Nenhuma diferença foi detectada nos outros parâmetros analisados. CONCLUSÕES: Na presença de meningite meningocócica a via alternativa é preferencialmente ativada, provavelmente devido à maior capacidade da endotoxina meningocócica para ativar esta via, in vivo.
Assuntos
Humanos , Lactente , Pré-Escolar , Criança , Complemento C3 , Complemento C4 , Fator B do Complemento/análise , Via Alternativa do Complemento/imunologia , Via Clássica do Complemento/imunologia , Meningite Meningocócica/imunologia , Doença Aguda , Brasil , Ensaio de Atividade Hemolítica de Complemento , Convalescença , Meningite Meningocócica/sangue , Nefelometria e Turbidimetria , Valores de ReferênciaRESUMO
A total of 376 chickens from different ecotypes were immunized with the non-pathogenic multi-determinant antigen sheep red blood cells (SRBC). The ecotypes included indigenous chickens from various locations in Tanzania (n=102), India (n=86) and Bolivia (n=89). In addition, eight German Dahlem Red (GDR) chicken lines with different major genes (dwarf, naked neck and frizzled) of tropical interest were also immunized with SRBC. Immune competence of the breeds was assessed by measuring complement haemolytic activity, both from the classical calcium-dependent complement pathway (CPW) and alternative calcium-independent complement pathway (APW), alongside IgTotal, IgG and IgM antibody responses to SRBC at 7 days post immunization. Large variations in complement activity and antibody responses to SRBC were observed within and between the indigenous breeds. Many indigenous chickens, especially from Bolivia, showed decreased complement activity (APW) following immunization with SRBC. Breeds from India showed the highest CPW activity and humoral (especially IgM) responses to SRBC, suggesting high immune competence. In contrast, Bolivian chickens were characterized by low CPW activity, low APW activity and low antibody levels to SRBC suggesting an overall low immune competence. In the GDR chickens, characterized by high CPW activity and high IgG antibody responses to SRBC, the major genes for naked neck, frizzling and dwarfism had no significant effect on the antibody responses and complement activity to SRBC.
Assuntos
Galinhas/genética , Galinhas/imunologia , Proteínas do Sistema Complemento/análise , Imunidade Inata/genética , Animais , Formação de Anticorpos , Bolívia , Ensaio de Atividade Hemolítica de Complemento/veterinária , Via Alternativa do Complemento , Via Clássica do Complemento , Proteínas do Sistema Complemento/genética , Eritrócitos , Feminino , Testes de Hemaglutinação/veterinária , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Índia , Ovinos , Tanzânia , Clima TropicalRESUMO
Low molecular weight hemorrhagins were purified from crude Bothrops atrox snake venom by gel filtration followed by ionic strength chromatography. The protein fractions obtained, designated HI-1 to HI-8, contained proteins with molecular masses lower than 30 kDa. HI-5, the most representative among of these fractions, exhibited, in vitro, proteolytic and C inactivating properties, as analyzed by proteolysis of a protein substrate, and C system consumptive activities as assayed by reduction of the hemolytic C activity in normal human serum and by cleavage of partially purified component C3. HI-5 hemorrhagin injected i.m. into C-sufficient BALB/c mice induced a local inflammation characterized by edema, accumulation of polymorphonuclear leucocytes (PMN) and hemorrhage. In contrast, when injected into BALB/c mice previously C-depleted, the number of PMN per tissue section, but not hemorrhage, was significantly reduced (129.668 +/- 31.341 cells per microscopic field) as compared with the control C-sufficient mice (812.168 +/- 111.194 cells per microscopic field). The observations were confirmed by using C5-deficient mice instead of C-depleted mice. The average number of PMN per tissue section in C5-defficient A/J mice was 72.666 +/- 19.416 cells per microscopic field. These data indicate that the C system is involved in PMN accumulation, but not in the hemorrhage, at the local induced lesions by low molecular mass B. atrox hemorrhagins. HI-5 apparently is not contaminated with other direct or indirect inflammation mediators, PMN accumulation and hemorrhage, however, an independent phenomenon, could be mediated by the same hemorrhagin proteinase domain.
Assuntos
Bothrops , Proteínas do Sistema Complemento/fisiologia , Venenos de Crotalídeos/enzimologia , Inflamação/induzido quimicamente , Metaloendopeptidases/toxicidade , Neutrófilos/metabolismo , Animais , Complemento C5/imunologia , Ensaio de Atividade Hemolítica de Complemento , Proteínas do Sistema Complemento/deficiência , Venenos de Crotalídeos/toxicidade , Edema/induzido quimicamente , Hemorragia/induzido quimicamente , Humanos , Inflamação/patologia , Injeções Intramusculares , Metaloendopeptidases/isolamento & purificação , Camundongos , Camundongos Endogâmicos A , Camundongos Endogâmicos BALB C , Peso Molecular , Músculo Esquelético/patologia , Neutrófilos/efeitos dos fármacos , Neutrófilos/patologia , Soro/metabolismo , Especificidade da EspécieRESUMO
Complement-depleted and -non-depleted BALB/c mice were inoculated with Leishmania (Leishmania) amazonensis promastigotes into the hind footpad to study the role of the complement system in cutaneous leishmaniasis. Total serum complement activity was measured by hemolytic assay and C3 fragment deposit at the inoculation site was determined by direct immunofluorescence in the early period of infection, i.e., at 3, 24, 48 h and 7 days post-infection. The inflammatory reaction and the parasite burden were evaluated in the skin lesion at 7 and 30 days post-infection. Total serum complement activity decreased in the early phase of infection, from 3 to 24 h, in non-depleted mice compared to non-infected and non-depleted mice. C3 fragment deposit at the site of parasite inoculation was present throughout the period of infection in non-depleted mice. In contrast, no C3 fragment deposit was observed at the inoculation site in complement-depleted mice. Complement-depleted mice showed a significant decrease in the inflammatory response and a significant increase in the number of parasites (70.0 +/- 5.3 vs 5.3 +/- 1.5) at 7 days of infection (P<0.05). A higher number of parasites were also present at 30 days of infection at the inoculation site of complement-depleted mice (78.5 +/- 24.9 vs 6.3 +/- 5.7). These experiments indicate that complement has an important role at the beginning of experimental cutaneous leishmaniasis caused by L. (L.) amazonensis by controlling the number of parasites in the lesion.
Assuntos
Proteínas do Sistema Complemento/fisiologia , Leishmania , Leishmaniose Cutânea/imunologia , Animais , Ativação do Complemento , Complemento C3/fisiologia , Ensaio de Atividade Hemolítica de Complemento , Leishmaniose Cutânea/parasitologia , Depleção Linfocítica , Masculino , Camundongos , Camundongos Endogâmicos BALB CRESUMO
Complement-depleted and -non-depleted BALB/c mice were inoculated with Leishmania (Leishmania) amazonensis promastigotes into the hind footpad to study the role of the complement system in cutaneous leishmaniasis. Total serum complement activity was measured by hemolytic assay and C3 fragment deposit at the inoculation site was determined by direct immunofluorescence in the early period of infection, i.e., at 3, 24, 48 h and 7 days post-infection. The inflammatory reaction and the parasite burden were evaluated in the skin lesion at 7 and 30 days post-infection. Total serum complement activity decreased in the early phase of infection, from 3 to 24 h, in non-depleted mice compared to non-infected and non-depleted mice. C3 fragment deposit at the site of parasite inoculation was present throughout the period of infection in non-depleted mice. In contrast, no C3 fragment deposit was observed at the inoculation site in complement-depleted mice. Complement-depleted mice showed a significant decrease in the inflammatory response and a significant increase in the number of parasites (70.0 ± 5.3 vs 5.3 ± 1.5) at 7 days of infection (P < 0.05). A higher number of parasites were also present at 30 days of infection at the inoculation site of complement-depleted mice (78.5 ± 24.9 vs 6.3 ± 5.7). These experiments indicate that complement has an important role at the beginning of experimental cutaneous leishmaniasis caused by L. (L.) amazonensis by controlling the number of parasites in the lesion.