TNF-alpha inhibitor adalimumab attenuates endotoxin induced cardiac damage in rats

Abstract Purpose To investigate the effects of adalimumab pretreatment on the lipopolysaccharide-mediated myocardial injury. Methods Twenty-eight Wistar rats were randomized into four groups (n=7). Control (C) group animals were injected once a day with intraperitoneal (i.p) 0.9 % saline for two days. In the Adalimumab (Ada) group, adalimumab was injected at a dose of 10 mg/kg/ day (i.p) for two days. Lipopolysaccharide (Lps) group rats were injected with a dose of 5 mg/kg (i.p) lipopolysaccharide. Lipopolysaccharide + Adalimumab (Lps+Ada) group rats received adalimumab before the administration of lipopolysaccharide. The animals were sacrificed 24 h after the last injection and blood samples were obtained for determination of biochemical cardiac injury markers and circulating levels of TNF-α and interleukin-6 (IL-6). Hearts were harvested for histological examination. Results Endotoxin exposure resulted in significant increases in serum cardiac injury markers, serum cytokines and histological myocardial injury scores in the Lps group. The levels of circulating cytokines, cardiac injury markers and histological injury scores for myocardial necrosis, perivascular cell infiltration, and inflammation were significantly reduced in Lps+Ada as compared to Lps group (p<0.05). Conclusions Adalimumab pretreatment reduces endotoxin-induced myocardial damage in rats. This beneficial effect is thought to be related to the reduction of cytokine release.

El caseinato de sodio y la caseína α inhiben la proliferación de la línea celular mieloide de ratón 32D clone 3 (32Dcl3) mediante el TNF-α / Sodium caseinate and alfa-casein inhibit proliferation of the mouse myeloid cell line 32D clone 3 (32Dcl3) via TNF-α

Resumen Introducción. Se ha demostrado que el caseinato de sodio y sus componentes (caseínas α, β y κ) inhiben la proliferación de la línea celular hematopoyética de ratón 32D clone 3 (32Dcl3) e inducen su diferenciación hacia macrófagos. Se sabe que la caseína α induce la producción de IL-1β y que esta última citocina inhibe la proliferación celular mediante la producción del factor de necrosis tumoral alfa (TNF-α), pero se desconoce si el caseinato de sodio y las caseínas inducen la producción de TNF y si este es el responsable de la inhibición de la proliferación. Objetivo. Evaluar si el caseinato de sodio y las caseínas α, β y κ inhiben la proliferación de la línea celular 32Dcl3 mediante la producción de TNF-α. Materiales y métodos. Se usaron diferentes concentraciones de caseinato de sodio y de las caseínas α, β y κ en las células 32Dcl3. Posteriormente, se evaluaron la viabilidad celular mediante una prueba con el MTT [3-(4,5-dimetiltiazol-2-ilo)-2,5-difeniltetrazol], la inducción de apoptosis con la citometría de flujo y la síntesis del TNF-α con el ELISA. Además, se hicieron pruebas de neutralización con anti-TNF-α en células 32Dcl3 tratadas con caseinato de sodio y caseína α, y se evaluó la proliferación celular. Resultados. Se encontró que el caseinato de sodio y las caseínas α, β y κ reducían la proliferación de la línea celular 32Dcl3 sin afectar la viabilidad, y que solo el caseinato y la caseína α inducían la apoptosis y la liberación al medio de TNF-α. La proliferación de células 32Dcl3 tratadas con caseinato y caseína α se restableció al usar anticuerpos anti-TNF-α. Conclusión. El TNF-α fue el principal responsable de la inhibición de la proliferación en las células 32Dcl3 tratadas con caseinato de sodio o caseína α.

Abstract Introduction: Sodium caseinate (CS) and its components (alpha-casein, beta-casein, and kappa-casein) have been shown to inhibit the proliferation of the mouse hematopoietic 32D clone 3 (32Dcl3) cell line and induce its differentiation into macrophages. It is well-known that alpha-casein induces IL-1β production and that this cytokine inhibits the proliferation via the production of tumor necrosis factor alpha (TNF-alpha), but it is not known if CS and the caseins inhibit the proliferation via TNF-alpha production. Objective: To evaluate if CS and alpha-casein, beta-casein and kappa-casein inhibit the proliferation on 32Dcl3 cell line via TNF-alpha. Materials and methods: We used different concentrations of CS, alpha-casein, beta-casein and kappa-casein in 32Dcl3 cells to evaluate cell proliferation. We assessed cell viability by MTT, induction to apoptosis by flow cytometry, and TNF-alpha synthesis by ELISA. Additionally, we performed anti-TNF-alpha neutralization assays on 32Dcl3 cells treated with CS and alpha-casein and we evaluated proliferation. Results: The results showed that CS, alpha-casein, beta-casein, and kappa-casein reduced proliferation of the 32Dcl3 cell line without affecting the viability and that only CS and alpha-casein induced apoptosis and the release of TNF-alpha. The 32Dcl3 cells treated with CS and alpha-casein reestablished their proliferation by using anti-TNF-alpha antibodies. Conclusion: TNF-alpha was the main responsible for the inhibition of proliferation in 32Dcl3 cells treated with CS or alpha-casein.

Hyperbaric Oxygen Pretreatment Improves Cognition and Reduces Hippocampal Damage Via p38 Mitogen-Activated Protein Kinase in a Rat Model

PURPOSE: To investigate the effects of hyperbaric oxygen (HBO) pretreatment on cognitive decline and neuronal damage in an Alzheimer’s disease (AD) rat model. MATERIALS AND METHODS: Rats were divided into three groups: normal saline (NS), AD, and HBO+AD. In the AD group, amyloid β peptide (Aβ)₁₋₄₀ was injected into the hippocampal CA1 region of the brain. NS rats received NS injection. In the HBO+AD group, rats received 5 days of daily HBO therapy following Aβ₁₋₄₀ injection. Learning and memory capabilities were examined using the Morris water maze task. Neuronal damage and astrocyte activation were evaluated by hematoxylin-eosin staining and immunohistochemistry, respectively. Dendritic spine density was determined by Golgi-Cox staining. Tumor necrosis factor-α, interleukin-1β, and interleukin-10 production was assessed by enzyme-linked immunosorbent assay. Neuron apoptosis was evaluated by terminal deoxynucleotidyl transferase dUTP nick end labeling. Protein expression was examined by western blotting. RESULTS: Learning and memory dysfunction was ameliorated in the HBO+AD group, as shown by significantly lower swimming distances and escape latency, compared to the AD group. Lower rates of neuronal damage, astrocyte activation, dendritic spine loss, and hippocampal neuron apoptosis were seen in the HBO+AD than in the AD group. A lower rate of hippocampal p38 mitogen-activated protein kinase (MAPK) phosphorylation was observed in the HBO+AD than in the AD group. CONCLUSION: HBO pretreatment improves cognition and reduces hippocampal damage via p38 MAPK in AD rats.

Evaluation of cytokines produced by β-hemolytic streptococcus in acute pharyngotonsillitis / Avaliação das citosinas produzidas por estreptococos β-hemolíticos em faringotonsilites agudas

INTRODUCTION: The most common pathogen in bacterial pharyngotonsillitis is group A β-hemolytic streptococcus, although groups B, C, F,and G have also been associated with pharyngotonsillitis.OBJECTIVE: To assess the levels of the cytokines TNF-α, IL-6,IL-4, and IL-10 in bacterial pharyngotonsillitis caused by group A and non-A (groups B, C, F and G) β-hemolytic streptococcus.METHODS: The study was conducted at a pediatric emergency care unit. The sample comprised children (5-9 years old) with acute bacterial pharyngotonsillitis diagnosed between December of 2011 and May of 2012. The research involved collection of blood samples from the patients, enzyme-linked immunosorbent assay detection of TNF-α, IL-6,IL-4, and IL-10, and collection of two oropharyngeal swabs for bacterial isolation. Additionally, the medical history of the study participants was also collected.RESULTS: In the studied group (mean age: 5.93 years), higher pharyngotonsillitis incidence was observed in the female gender (64.76%). Higher incidence of tonsillar exudates was observed with groups A and C. No statistically significant differences in cytokine levels were observed among groups. However, the group A and the control group showed a difference in the IL-6 level (p = 0.0016).CONCLUSIONS: The Groups A and C showed higher cytokine levels than the Groups B and control, suggesting similar immunological patterns.

INTRODUÇÃO: O patógeno mais comumente associado à faringotonsilite bacteriana é o estreptococo β-hemolítico do grupo A, a despeito dos grupos B, C, F e G terem também sido associados com a faringotonsilite.OBJETIVO: Determinar os níveis das citosinas TNF-α, IL-6, IL-4, e IL-10 na faringotonsilite bacteriana causada pelos estreptococos β-hemolíticos do grupo A e não-A (grupos B, C, F e G).MÉTODO: O estudo foi conduzido em uma emergência pediátrica. A amostra estudada compreendeu crianças (entre 5 e 9 anos) com faringotonsilite aguda bacteriana diagnosticada entre dezembro de 2011 e maio de 2012. A pesquisa envolveu a coleta de amostras sanguíneas dos pacientes, a detecção, através do ELISA, de TNF-α, IL-6, IL-4, and IL-10, além da coleta de dois swabs orofaríngeos para isolamento bacteriano. Adicionalmente foi coletada a história médica dos participantes do estudo.RESULTADOS: No grupo estudado (idade média: 5,93 anos), a maior incidência de faringotonsilite foi observada no gênero feminino (64,76%). Foram detectadas maiores incidências de exsudatos tonsilares nos grupos A e C. Não foram observadas diferenças estatisticamente significantes dos níveis de citosinas entre os grupos. Porém os grupos A e o controle mostraram diferença nos níveis de IL-6 (p = 0.0016).CONCLUSÕES: Os grupos A e C mostraram maiores níveis de citosinas que os grupos B e o controle, sugerindo mecanismos imunológicos similares.

Avaliação de consumo alimentar, medidas antropométricas e tempo de menopausa de mulheres na pós-menopausa / Food consumption in postmenopausal women and its relation with anthropometric measurements and time since menopause

OBJETIVO: Avaliar o hábito alimentar e nutricional de mulheres na pós-menopausa e compará-los com o perfil antropométrico, faixa etária e tempo de menopausa. MÉTODOS: No período de junho a agosto de 2011, 148 mulheres na pós-menopausa residentes no Estado de São Paulo (região Sudeste do Brasil) foram avaliadas com um questionário estruturado contendo dados socioeconômicos, clínicos, antropométricos e alimentares. Avaliou-se nível de atividade física, variáveis bioquímicas, Índice de Massa Corporal (IMC), circunferência abdominal (CA) e consumo alimentar (energia, proteínas, carboidratos e gorduras, fibra, colesterol, vitaminas A e C, minerais, cálcio e ferro) de acordo com a faixa etária e o tempo de pós-menopausa (TPM). RESULTADOS: A média de IMC foi 29,0±5,6 kg/m2 e da CA, 95,7±12,9 cm. O consumo médio calórico diário atingiu 1.406,3±476,5 kcal. A ingestão e a adequação calórica foram significantemente mais apropriadas entre as mulheres eutróficas e com CA<88 cm. O mesmo ocorreu quanto ao consumo de proteínas (p<0,001 e p=0,006, respectivamente). Na análise por faixa etária ou TPM não houve diferenças significantes, exceto a média do consumo proteico, maior no grupo com 5 anos ou menos de menopausa (p=0,048). CONCLUSÃO: O perfil antropométrico de mulheres na pós-menopausa mostrou predominância de sobrepeso ou obesidade. O consumo alimentar apresentou-se adequado quanto às calorias e percentuais de macronutrientes, entre as eutróficas e com CA<88 cm. .

PURPOSE: To evaluate eating in postmenopausal women and its relation to anthropometry, age and time since menopause in São Bernardo do Campo residents. METHODS: During the period from June to August of 2011, 148 postmenopausal women residents in state of São Paulo (Southeast region of Brazil) were evaluated using a structured questionnaire containing socioeconomic, clinical, anthropometric and food data. The level of physical activity, biochemical variables, Body Mass Index (BMI), abdominal circumference (AC) and dietary intake (energy, protein, carbohydrates and fats, fiber, cholesterol, vitamins A and C, minerals, calcium and iron) were analyzed according to age and time after menopause. RESULTS: Mean BMI was 29.0≤5.6 kg/m2 and abdominal circumference was 95.7±12.9 cm. The average daily caloric consumption was 1,406.3±476.5 kcal. The calorie intake was significantly more appropriate in normal-weight women and women with AC<88 cm. The same was observed for protein intake (p<0.001 and p=0.006, respectively). No association was observed with age or duration of the postmenopausal period, except for average protein consumption that was higher in the group with five years or less of menopause (p=0.048). CONCLUSION: The anthropometry of postmenopausal women showed a predominance of overweight and obesity. Dietary intake was adequate in relation to the percentage of calories and macronutrients and calories among most normal-weight women and women with AC<88 cm. .

Human amnion mesenchymal cells inhibit lipopolysaccharide-induced TNF-α and IL-1ß production in THP-1 cells

BACKGROUND: Human amnion mesenchymal cells (hAMCs), isolated from the amniotic membrane of human placenta, are a unique population of mesenchymal stem cells. Recent studies demonstrated that hAMCs could inhibit the activities and functions of several immune cells. However, their effect on inflammatory macrophages is largely unknown. This study investigated the effect of hAMCs on expression of inflammatory cytokines and mitogen-activated protein kinases (MAPKs)/NF-kB pathway in human THP-1 macrophages induced by lipopolysaccharide (LPS). RESULTS: The levels of TNF-α and IL-1ß secreted by LPS- stimulated THP-1 cells were increased significantly compared with those in the control group. After co-culture with different numbers of hAMCs, the levels of TNF-α and IL-1ß in LPS-stimulated THP-1 cells were significantly reduced compared with the LPS group. The mRNA expression of TNF-α and IL-1ß were also markedly inhibited. Moreover, treating LPS-stimulated THP-1 cells with hAMCs supernatants could also suppress TNF-α and IL-1ß production in THP-1 cells. Important signaling pathways involved in the production of TNF-α and IL-1ß were affected by hAMCs co-culture: hAMCs remarkably suppressed NF-kB activation and down-regulated the phosphorylation of ERK and JNK in LPS- stimulated THP-1 cells. CONCLUSIONS: Human amnion mesenchymal cells inhibited the production of TNF-α and IL-1ß secreted by LPS-stimulated THP-1 cells, partly through the suppression of NF-kB activation and ERK and JNK phosphorylation.

Instituição de longa permanência para idosos: um lugar de cuidado para quem não tem opção? / Long-stay institutions for the elderly: a place of care for those who have no choice? / Institución de larga estadía para las personas mayores: ¿un lugar de atención para aquellos que no tienen otra opción?

Pesquisa descritiva, qualitativa, com abordagem compreensiva, que teve por objetivo compreender o significado da instituição de longa permanência para idosos institucionalizados. Os dados foram coletados com 13 idosos institucionalizados, no período de 5 de abril a 25 de maio de 2013 por meio da entrevista narrativa, e submetidos a análise de conteúdo, na modalidade de análise temática. Os resultados indicam que ser idoso institucionalizado significa ter suas necessidades de cuidado atendidas, no que concerne a suas necessidades básicas; ao acesso a serviços e recursos de saúde, e a ter um lugar onde possam envelhecer e morrer. O estudo permitiu concluir que a instituição aparece como um lugar ambíguo para os idosos, pois ao mesmo tempo em que os acolhe, abriga e atende suas necessidades, é um ambiente que inviabiliza a vida independente e autônoma.

This is a descriptive, qualitative research, with comprehensive approach, which aimed to understand the meaning that the longterm institution has to institutionalized elderly. Data were collected with 13 institutionalized elderly in the period from April 5 to May 25, 2013, through narrative interview, and subjected to content analysis, in the form of thematic analysis. The results indicated that being elderly institutionalized means having their care needs met, with respect to their basic needs; access to health services and resources, and to have a place where they can grow old and die. The study concluded that the institution appears as an ambiguous place for the elderly because, even embracing and housing them and meeting their needs, is an environment that prevents the independent and autonomous life.

Investigación descriptiva, cualitativa con enfoque comprehensivo, que tuve como objetivo comprender el significado de la institución a largo plazo tiene para ancianos institucionalizados. Los datos fueron recolectados con 13 ancianos institucionalizados en el periodo comprendido entre el 5 abril-25 mayo de 2013, a través de entrevista narrativa, y tratados mediante análisis de contenido, en la modalidad de análisis temático. Los resultados indican que estar en edad avanzada y estar institucionalizado significa tener sus atendidas sus necesidades básicas; el acceso a los servicios de salud y los recursos, y tener un lugar donde pueden envejecer y morir. El estudio llegó a la conclusión de que la institución se presenta como un lugar ambiguo para las personas mayores, ya que si bien les acoja, albergue y atiendan sus necesidades, es un ambiente que impide la vida independiente y autónoma.

Metotrexate como inductor en la producción de citocinas proinflamatorias en células epiteliales / Methotrexate as inducer of proinflammatory cytokines by epithelial cells

El metotrexate (MTX) es uno de los medicamentos frecuentemente utilizados en el cáncer infantil señalándose además, como agente citotóxico de la mucosa bucal, que puede desencadenar el proceso inflamatorio e incremento de la vascularidad en los tejidos epiteliales durante las fases iniciales de la mucositis oral. El presente trabajo tiene como objetivo determinar la producción de citocinas proinflamatorias IL-1b, IL-6 y TNF-a en cultivos de células epiteliales tratadas con MTX. Se realizaron cultivos de células epiteliales de laringe humana obtenidas de la línea celular Hep-2, con diferentes dosis de MTX en distintos tiempos de incubación, y a su vez se analizó la citotoxicidad del fármaco mediante el ensayo colorimétrico, el cual se basa en la reducción metabólica del bromuro de 3-(4,5- dimetiltiazol-2-ilo)-2,5-difeniltetrazol (MTT), y la producción de citocinas proinflamatorias mediante el ensayo inmuno enzimático indirecto (ELISA). En cuanto a los resultados se observó, que los cultivos de células Hep-2 presentaron aumento en la producción de las citocinas proinflamatorias a las 72 horas al utilizar las dosis de 0.32µM MTX. Estos resultados sugieren que la dosis y el tiempo de exposición del MTX alteran la fisiología de las células epiteliales humanas, lo cual podrían desempeñar un papel importante durante las fases de iniciación y de desarrollo de la mucositis oral.

Methotrexate (MTX), a drug commonly used in childhood cancer, has also been indicated as a cytotoxic agent of the oral mucosa, which can trigger the inflammatory process and increase the vascularity of epithelial tissues during the early stages of oral mucositis. The aim of this study was to determine the production of proinflammatory cytokines IL-1b, IL-6 y TNF-a in epithelial cell cultures treated with MTX. Epithelial cells of human larynx, obtained from the cell line Hep-2, were cultured with different doses of MTX during different incubation times. The drug cytotoxicity was analyzed by means of the colorimetric test, which is based on the metabolic reduction of the bromide of 3-(4, 5-dimetiltiazol-2-ilo)-2,5-difeniltetrazol (MTT); and the proinflammatory cytokines production by the test enzyme-linked immunosorbent assay (ELISA). Cultures of HEp-2 cells showed increased production of proinflammatory cytokines at 72 hours with 0.32µM of MTX. These results suggest that depending on the dose and exposure time, MTX alters the physiology of human epithelial cells, which may play an important role during the phases of initiation and development of oral mucositis.

Wound healing potential of flowers extracts of Woodfordia fruticosa Kurz.

Wound healing or repair is the body’s natural process of regenerating dermal and epidermal tissue. Woodfordia fruticosa Kurz (Family: Lythraceae) is used traditionally in wound healing by the tribals of Chhattisgarh district. However, there is a paucity of scientific data in support. In this study, we evaluated antimicrobial activity of petroleum ether, chloroform, ethanolic and aqueous extracts against a diverse range of gram +ve and gram -ve bacteria along with pathogenic fungi. The wound healing activity of ethanolic extract was also evaluated at dose levels of 250 and 500 mg/kg body wt in rats by excision, incision and dead space wound healing models along with histopathology of wound area of skin. The ethanolic extract showed potent wound healing activity, as evident from the increase in the wound contraction and breaking strength in dose-dependent manner. Treatment with ethanolic extract (250 and 500 mg/kg body wt) showed significant dose-dependently decrease in epithelization period and scar area. Hydroxyproline, hexuronic acid and hexosamine contents, the important constituents of extracellular matrix of healing were also correlated with the observed healing pattern. During early wound healing phase, pro-inflammatory cytokines TNF-α, IL-6 and anti-inflammatory cytokine IL-10 levels were found to be upregulated by the ethanolic extract treatment. The ethanolic extract exhibited a strong and broad spectrum antimicrobial activity, as compared to other extracts. It showed very low Minimum inhibitory concentration (MIC) values and inhibited the growth of E. coli, Staphylococcus aureus and Candida albicans in concentration of 2.5 µg/disc. Thus, the results of the present study demonstrated the strong wound healing potential and antimicrobial activities of W. fruticosa, flowers, supporting the folklore use of the plant by the tribal people of Chhattisgarh district.

Correlation between the response to Mycobacterium tuberculosis antigens and the tuberculin skin test in patients with rheumatoid arthritis in Colombia / Correlación de la respuesta a antígenos de Mycobacterium tuberculosis y la prueba cutánea de tuberculina en pacientes con artritis reumatoide

Introduction. Rheumatoid arthritis patients under treatment with anti-TNF-α are at a high risk of developing active tuberculosis, and therefore, screening for latent tuberculosis infection is recommended before anti-TNF-α therapy. Objective. To compare the tuberculin test and IFNγ production induced by culture filtrate proteins(CFPs) and Mycobacterium tuberculosis-specific CFP-10 antigens in rheumatoid arthritis patients. Materials and methods. An analytic transversal study was conducted in rheumatoid arthritis patients treated at Hospital Universitario San Vicente Fundación between January and December 2007. IFNγ production in response to CFPs and CFP-10 was measured in the supernatants of whole blood cultures and evaluated for correlations with tuberculin reactivity. The degree of concordance between both tests was also established. Results. Forty-five patients were included, of which 14 (31.1%) had a tuberculin reaction of ≥10 mm of induration, 9 (20%) produced IFNγ in response to CFP-10, and 7 were positive for both tests. The correlation between tests was r=0.53 (IC 95%:0.28-0.72), and the global concordance between tests was80%, with a Kappa coefficient of 0.48 (IC95%:0.20-0.76). Conclusions. Only two tuberculin (-)/CFP-10+ "anergic" patients were observed. By contrast, six tuberculin +/CFP-10(-) "tuberculin false-positive" patients were observed. These data suggest that the tuberculin test is not an appropriate tool for determining the need for tuberculosis prophylaxis.

Introducción. Los pacientes con artritis reumatoide bajo tratamiento con anti-TNFα están en alto riesgo de desarrollar tuberculosis activa, por lo cual se recomienda hacer la tamización para infección latente de tuberculosis, antes de iniciar el tratamiento. Objetivo. Comparar la prueba de tuberculina y la producción de IFNγ inducida por antígenos CFP (Culture Filtrate Protein) y antígenos específicos de Mycobacterium tuberculosis (CFP-10) para el diagnóstico de infección latente de tuberculosis en pacientes con artritis reumatoide. Materiales y métodos. Se llevó a cabo un estudio transversal analítico en pacientes con artritis reumatoide atendidos en el Hospital Universitario San Vicente Fundación, entre enero y diciembre de 2007, a los cuales se les determinó la producción de IFNγ en respuesta a CFP y CFP-10 en sobrenadantes de cultivos de sangre total, y se correlacionó con la reacción en la prueba de tuberculina. Además, se estableció el grado de concordancia entre ambas pruebas. Resultados. Se incluyeron 45 pacientes, de los cuales, 14 (31,1 %) tuvieron un diámetro de induración ≥10 mm (tuberculina positiva), nueve (20 %) produjeron IFNγ en respuesta a CFP-10, y siete fueron positivos para ambas pruebas. La correlación entre las pruebas fue de r=0,53 (IC95%: 0,28-0,72) y la concordancia global entre pruebas fue de 80 %, con un coeficiente kappa de 0,48 (IC95%: 0,20-0,76). Conclusiones. Solo se observaron dos pacientes con tuberculina positiva y CFP-10 positivo "anérgicos" y se encontraron seis pacientes con tuberculina positiva y CFP-10 negativa "falsos positivos para tuberculina", lo cual sugiere que la prueba de la tuberculina no es la más adecuada para indicar profilaxis para tuberculosis.

Expressão de mediadores neurotróficos e pró-inflamatórios na endometriose de reto e sigmoide / Expression of neurotrophic and inflammatory mediators in rectosigmoid endometriosis

OBJETIVO: Avaliar a expressão de mediadores neurotróficos (NGF, NPY E VIP) e pró-inflamatórios (TNF-α) em fragmentos de reto e sigmoide comprometidos por endometriose. MÉTODOS: Foram selecionadas 24 pacientes submetidas ao tratamento cirúrgico de endometriose de reto e sigmoide com técnica de ressecção segmentar, seguido de anastomose mecânica término-terminal, com grampeador circular, no período de janeiro de 2005 a dezembro de 2007. Neste estudo incluímos mulheres no menacme que se submeteram a tratamento cirúrgico por endometriose profunda infiltrativa com acometimento do reto e sigmoide, atingindo o nível da camada muscular, submucosa ou mucosa. Para o grupo de estudo foram utilizados 24 fragmentos de reto e sigmoide com endometriose confirmada histologicamente, sendo um fragmento de cada uma das 24 pacientes selecionadas. Para o grupo controle, utilizou-se um fragmento da margem distal da ressecção, denominado anel de anastomose, de cada uma das 24 pacientes selecionadas e incluídas no estudo. As amostras foram agrupadas em blocos de Tissue Micro Array (TMA) e submetidas à reação imunoistoquímica para avaliar a expressão do fator de necrose tumoral alfa (TNF-α), do fator de crescimento neural (NGF), do neuropeptídeo Y (NPY) e do peptídeo intestinal vasoativo P (VIP), e posterior análise semiquantitativa da imunomarcação por meio da leitura da densidade ótica relativa (DO). RESULTADOS: Observou-se maior densidade ótica relativa da imunomarcação para TNF-α e NGF no grupo de estudo (amostras com endometriose intestinal), DO= 0,01, respectivamente, para as duas proteínas (p<0,05), em relação aos controles sem endometriose. Não houve diferença estatística na densidade ótica da imunomarcação do NPY e VIP. CONCLUSÃO: Identificou-se o aumento da imunomarcação dos anticorpos TNF-α e NGF em fragmentos de reto e sigmoide comprometidos por endometriose em relação aos controles livres da doença. Não identificamos diferença estatística na imunomarcação das proteínas NPY e VIP.

PURPOSE: To evaluate the expression of neurotrophic (NGF, NPY and VIP) and pro-inflammatory (TNF-α) mediators in the rectum and sigmoid fragments compromised by endometriosis. METHODS: Twenty-four patients were selected to undergo surgical treatment of endometriosis of the rectum and sigmoid colon with a segmental resection technique, followed by end-to-end anastomosis with a circular stapler from January 2005 to December 2007. The study included premenopausal women who underwent surgical treatment for deep endometriosis infiltrating the rectum with involvement of the rectum and sigmoid, reaching the level of the muscle layer, submucosa or mucosa. Twenty-four rectum and sigmoid fragments with histologically confirmed endometriosis, one from each of the 24 selected patients, were used for the study group. For the control group, we used a fragment of the distal resection margin called anastomosis ring from each of the 24 patients enrolled in the study. Samples were grouped into Tissue Micro Array (TMA) blocks and subjected to immunohistochemistry to evaluate the expression of tumor necrosis factor alpha (TNF-α), nerve growth factor (NGF), neuropeptide Y (NPY) and P vasoactive intestinal peptide (VIP), followed by semiquantitative analysis of immunostaining by reading the relative optical density (OD). RESULTS: There was higher optical density relative to TNF-α immunostaining and NGF in the study group (samples with intestinal endometriosis), DO=0.01, for the two proteins, respectively (p<0.05), compared to controls without endometriosis. There was no statistically significant difference in the optical density of immunostaining of NPY and VIP. CONCLUSION: We identified increased immunostaining of TNF-α antibodies and fragments of NGF in the rectum and sigmoid compromised by endometriosis compared to disease-free controls. We did not identify any statistical difference in immunostaining of NPY and VIP proteins.

Production of TNF-α, nitric oxide and hydrogen peroxide by macrophages from mice with paracoccidioidomycosis that were fed a linseed oil-enriched diet

Omega-3 polyunsaturated fatty acids (n-3 PUFA) can modulate the immune system and their primary effect is on macrophage function. Paracoccidioidomycosis (PCM) is an endemic systemic mycosis in Latin America that is caused by the dimorphic fungus Paracoccidioides brasiliensis (Pb). Macrophages are the main defence against this pathogen and have microbicidal activity that is dependent on interferon-Γ and tumour necrosis factor (TNF)-α. These cytokines stimulate the synthesis of nitric oxide (NO) and hydrogen peroxide (H2O2), leading to the death of the fungus. To study the effect of n-3 PUFA on the host immune response during experimental PCM, macrophages that were obtained from animals infected with Pb18 and fed a diet enriched by linseed (LIN) oil were cultured and challenged with the fungus in vitro. The macrophage function was analysed based on the concentrations of TNF-α, NO and H2O2. LIN oil seems to influence the production of TNF-α during the development of disease. A diet enriched with LIN oil influences the microbicidal activity of the macrophages by inducing the production of cytokines and metabolites such as NO and H2O2, predominantly in the chronic phase of infection.

Distinct cytokine profiles of circulating mononuclear cells stimulated with Staphylococcus aureus enterotoxin A in vitro during early and late episodes of chronic osteomyelitis

We investigated the cytokine profile of peripheral mononuclear cells from chronic osteomyelitis (OST) patients following in vitro stimulation with staphylococcal enterotoxin A (SEA). We demonstrate that stimulation with SEA induced prominent lymphocyte proliferation and high levels of tumour necrosis factor (TNF)-α, interleukin (IL)-4 and IL-10 secretion in both OST and non-infected individuals (NI). Even though stimulation with SEA had no impact on IL-6 production in either patient group, the baseline level of IL-6 production by cells from OST patients was always significantly less than that produced by cells from NI. After classifying the osteomyelitic episodes based on the time after the last reactivation event as "early" (1-4 months) or "late" osteomyelitis (5-12 months), we found that increased levels of TNF-α and IL-4 in combination with decreased levels of IL-6 were observed in the early episodes. By contrast, increased levels of IL-10, IL-2 and IL-6 were hallmarks of late episodes. Our data demonstrate that early osteomyelitic episodes are accompanied by an increased frequency of "high producers" of TNF-α and IL-4, whereas late events are characterised by increased frequencies of "high producers" of IL-10, IL-6 and IL-2. These findings demonstrate the distinct cytokine profiles in chronic osteomyelitis, with a distinct regulation of IL-6 production during early and late episodes.

Recognition of enteroinvasive Escherichia coli and Shigella flexneri by dendritic cells: distinct dendritic cell activation states

The innate and adaptive immune responses of dendritic cells (DCs) to enteroinvasive Escherichia coli (EIEC) infection were compared with DC responses to Shigella flexneri infection. EIEC triggered DCs to produce interleukin (IL)-10, IL-12 and tumour necrosis factor (TNF)-α, whereas S. flexneri induced only the production of TNF-α. Unlike S. flexneri, EIEC strongly increased the expression of toll like receptor (TLR)-4 and TLR-5 in DCs and diminished the expression of co-stimulatory molecules that may cooperate to inhibit CD4+ T-lymphocyte proliferation. The inflammation elicited by EIEC seems to be related to innate immunity both because of the aforementioned results and because only EIEC were able to stimulate DC transmigration across polarised Caco-2 cell monolayers, a mechanism likely to be associated with the secretion of CC chemokine ligands (CCL)20 and TNF-α. Understanding intestinal DC biology is critical to unravelling the infection strategies of EIEC and may aid in the design of treatments for infectious diseases.

Inmunosupresión en donantes renales: Expresión de citoquinas proinflamatorias / Immunosuppression in kidney donors with rapamycin and tacrolimus: Proinflammatory cytokine expression

La lesión por isquemia y reperfusión (IRI) es uno de los principales problemas en el trasplante. Nuestro objetivo fue evaluar el efecto del pre - acondicionamiento al donante con rapamicina y tacrolimus para prevenir la lesión por IRI. Las ratas Wistar donantes, 12 horas antes de la nefrectomía, recibieron fármacos inmunosupresores. La muestra se dividió en cuatro grupos experimentales: un grupo con intervención simulada (sham), un grupo control sin tratamiento, otro tratado con rapamicina (2 mg/kg) y el restante tratado con tacrolimus (0.3 mg/kg). Se retiró el riñón izquierdo y después de tres horas de isquemia fría, se lo trasplantó. Veinticuatro horas después, el órgano trasplantado se recuperó para el análisis histológico y la evaluación de la expresión de citoquinas. El tratamiento de pre-acondicionamiento con rapamicina o con tacrolimus redujo significativamente el nitrógeno ureico en sangre y los niveles de creatinina en comparación con el control (BUN: p < 0.001; creatinina: p < 0.001). La necrosis tubular aguda fue significativamente menor en las ratas donantes tratadas con inmunosupresores en comparación con el grupo control (p < 0.001). Finalmente, las citoquinas inflamatorias, como TNF-α, IL-6 y rIL-21, mostraron niveles más bajos en el injerto de los animales que recibieron tratamiento. Este estudio experimental exploratorio muestra que el pre-acondicionamiento en donantes con rapamicina y tacrolimus en dos grupos distintos mejora los resultados clínicos y anatomopatológicos en receptores, con una reducción in situ de citoquinas pro-inflamatorias relacionadas con la diferenciación Th17, y de este modo crea un ambiente favorable para la diferenciación de células T regulatorias (Tregs).

The ischemia-reperfusion injury (IRI) remains a major problem in transplantation. The objective of this study was to evaluate the effects of preconditioning a donor group with rapamycin and another donor group with tacrolimus to prevent IRI. Twelve hours before nephrectomy, donor Wistar rats received immunosuppressive drugs. The sample was divided into four experimental groups: a sham group, an untreated control group, a group treated with rapamycin (2 mg/kg) and a group treated with tacrolimus (0.3 mg/kg). Left kidneys were removed and, after three hours of cold ischemia, grafts were transplanted. Twenty-four hours later, the transplanted organs were recovered for histological analysis and evaluation of cytokine expression. The pre-conditioning treatment with rapamycin or tacrolimus significantly reduced donor blood urea nitrogen and creatinine levels compared with control group (BUN: p < 0.001 vs. control and creatinine: p < 0.001 vs. control). Acute tubular necrosis was significantly lower in donors treated with immunosuppressant drugs compared with the control group (p < 0.001). Finally, inflammatory cytokines such as TNF-α, IL-6 and rIL-21 showed lower levels in the graft of pre-treated animals. This exploratory experimental study shows that preconditioning donors with rapamycin and tacrolimus in different groups improves clinical outcome and pathology in recipients and reduces in situ pro-inflammatory cytokines associated with Th17 differentiation, creating a favorable environment for the differentiation of regulatory T cells (Tregs).

High-density lipoprotein prevents SAA-induced production of TNF-α in THP-1 monocytic cells and peripheral blood mononuclear cells

In this study, we evaluated whether human serum and lipoproteins, especially high-density lipoprotein (HDL), affected serum amyloid A (SAA)-induced cytokine release. We verified the effects of SAA on THP-1 cells in serum-free medium compared to medium containing human serum or lipoprotein-deficient serum. SAA-induced tumour necrosis factor-alpha (TNF-α) production was higher in the medium containing lipoprotein-deficient serum than in the medium containing normal human serum. The addition of HDL inhibited the SAA-induced TNF-α release in a dose-dependent manner. This inhibitory effect was specific for HDL and was not affected by low-density lipoprotein or very low-density lipoprotein. In human peripheral blood mononuclear cells, the inhibitory effect of HDL on TNF-α production induced by SAA was less pronounced. However, this effect was significant when HDL was added to lipoprotein-deficient medium. In addition, a similar inhibitory effect was observed for interleukin-1 beta release. These findings confirm the important role of HDL and support our previous hypothesis that HDL inhibits the effects of SAA during SAA transport in the bloodstream. Moreover, the HDL-induced reduction in the proinflammatory activity of SAA emphasizes the involvement of SAA in diseases, such as atherosclerosis, that are characterized by low levels of HDL.

Schistosoma mansoni antigens alter the cytokine response in vitro during cutaneous leishmaniasis

Schistosoma mansoni infection or associated products are able to down-modulate the type 1 CD4+ T cell inflammatory response characteristic of autoimmune diseases. In this study, we evaluated how S. mansoni antigens altered the immune response that was induced by the soluble Leishmania antigen (SLA) from cutaneous leishmaniasis (CL) patients. Cytokines were measured from the supernatants of peripheral blood mononuclear cell cultures stimulated with SLA. This was performed using the sandwich enzyme linked immunosorbent assay technique in the presence or absence of S. mansoni recombinant antigens Sm29, SmTSP-2 and PIII. The addition of S. mansoni antigens to the cultures resulted in the reduction of interferon gamma (IFN-γ) levels in 37-50 percent of patients. Although to a lesser extent, the antigens were also able to decrease the production of tumour necrosis factor-alpha (TNF-α). We compared patients that either had or did not have reduction in IFN-γ and TNF-α production in cultures stimulated with SLA in the presence of S. mansoni antigens. We found that there was no significant difference in the levels of interleukin (IL)-10 and IL-5 in response to S. mansoni antigens between the groups. The antigens used in this study down-modulated the in vitro proinflammatory response induced by SLA in a group of CL patients through a currently undefined mechanism.

Effect of TNF-α production inhibitors on the production of pro-inflammatory cytokines by peripheral blood mononuclear cells from HTLV-1-infected individuals

Human T lymphotropic virus type 1 (HTLV-1) is the causal agent of myelopathy/tropical spastic paraparesis (HAM/TSP), a disease mediated by the immune response. HTLV-1 induces a spontaneous proliferation and production of pro-inflammatory cytokines by T cells, and increasing interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α) levels are potentially involved in tissue damage in diseases related to HTLV-1. This exaggerated immune response is also due to an inability of the natural regulatory mechanisms to down-modulate the immune response in this group of patients. TNF-α inhibitors reduce inflammation and have been shown to improve chronic inflammatory diseases in clinical trials. The aim of this study was to evaluate the ability of pentoxifylline, forskolin, rolipram, and thalidomide to decrease in vitro production of TNF-α and IFN-γ in cells of HTLV-1-infected subjects. Participants of the study included 19 patients with HAM/TSP (mean age, 53 ± 11; male:female ratio, 1:1) and 18 HTLV-1 carriers (mean age, 47 ± 11; male:female ratio, 1:2.6). Cytokines were determined by ELISA in supernatants of mononuclear cell cultures. Pentoxifylline inhibited TNF-α and IFN-γ synthesis with the minimum dose used (50 µM). The results with forskolin were similar to those observed with pentoxifylline. The doses of rolipram used were 0.01-1 µM and the best inhibition of TNF-α production was achieved with 1 µM and for IFN-γ production it was 0.01 µM. The minimum dose of thalidomide used (1 µM) inhibited TNF-α production but thalidomide did not inhibit IFN-γ production even when the maximum dose (50 µM) was used. All drugs had an in vitro inhibitory effect on TNF-α production and, with the exception of thalidomide, all of them also decreased IFN-γ production.

Tnf-a production and apoptosis in hepatocytes after listeria monocytogenes and salmonella typhimurium invasion / Produção de tnf-a e apoptose em hepatócitos após invasão por listeria monocytogenes e salmonella typhimurium

Invasion of hepatocytes by Listeria monocytogenes (LM) and Salmonella Typhimurium (ST) can stimulate tumor necrosis factor alpha (TNF-α) release and induce apoptosis. In this study, we compared the behavior of hepatocytes invaded by three L. monocytogenes serotypes (LM-4a, LM-4b and LM-1/2a) and by ST to understand which bacterium is more effective in the infectious process. We quantified TNF-α release by ELISA, apoptosis rates by annexin V (early apoptosis) and TUNEL (late apoptosis) techniques. The cell morphology was studied too. TNF-α release rate was highest in ST-invaded hepatocytes. ST and LM-1/2a induced the highest apoptosis production rates evaluated by TUNEL. LM-4b produced the highest apoptosis rate measured by annexin. Invaded hepatocytes presented various morphological alterations. Overall, LM-4b and LM-1/2a proved to be the most efficient at cell invasion, although ST adapted faster to the environment and induced earlier hepatocyte TNF-α release.

A invasão de hepatócitos por Listeria monocytogenes (LM) e Salmonella Typhimurium (ST) pode estimular a liberação do Fator de Necrose Tumoral (TNF-α) e induzir a apoptose celular. Neste estudo comparamos o comportamento de hepatócitos invadidos por três sorotipos de L. monocytogenes (LM-4a, LM-4b e LM-1/2a) e por ST para entender qual bacteria é mais efetiva no processo infeccioso. Nós quantificamos a liberação de TNF-α pelos hepatócitos por ELISA e as taxas de apoptose pelas técnicas de anexina V (apoptose precoce) e TUNEL (apoptose tardia). A morfologia das células foi estudada também. A taxa de liberação de TNF-α foi mais alta em hepatócitos invadidos por ST. ST e LM-1/2a induziram as maiores taxas de apoptose pelo método TUNEL, enquanto LM-4b produziu as maiores taxas de apoptose por anexina V. Os hepatócitos invadidos apresentaram várias alterações morfológicas. Na análise do conjunto de dados, os sorotipos LM-4b e LM-1/2a provaram ser os mais eficientes na invasão celular, enquanto que ST adaptou-se mais rápido ao meio e induziu a liberação precoce de TNF-α pelos hepatócitos.

Ehrlichia canis (Jaboticabal strain) induces the expression of TNF-α in leukocytes and splenocytes of experimentally infected dogs / Amostra Ehrlichia canis (Jaboticabal) induz a expressão de TNF-α em leucócitos e esplenócitos de cães experimentalmente infectados

Canine ehrlichiosis is caused by the bacterium Ehrlichia canis and is characterized by a systemic febrile disease of unknown pathogenesis. This study evaluated the expression of cytokines TNF-α, IL-10, IFN-γ, in splenic cells and blood leukocytes during the acute phase of ehrlichiosis and after treatment with doxycycline hyclate in dogs experimentally infected with the E. canis Jaboticabal strain. The study results showed a significant expression of TNF-α 18 days post-inoculation, reducing by approximately 70 percent after treatment. There was a unique peak of expression of IL-10 and IFN-γ 18 and 30 days post-inoculation, respectively. This study suggests that TNF-α plays a role in the pathogenesis of the acute phase of canine ehrlichiosis and that treatment with doxycycline hyclate reduces the systemic effects of this cytokine, possibly by reducing or eliminating parasitemia.

A erliquiose canina é causada pela bactéria Ehrlichia canis, que desencadeia no hospedeiro uma doença febril e sistêmica, de patogênese pouco conhecida. O presente estudo avaliou a expressão das citocinas TNF-α, IL-10, IFN-γ, em células esplênicas e em leucócitos sanguíneos, durante a fase aguda da erliquiose e após o tratamento com hiclato de doxiciclina, em cães experimentalmente infectados com a amostra E. canis Jaboticabal. Os resultados mostraram expressão significativa de TNF-α 18 dias após a inoculação, reduzindo aproximadante 70 por cento após o tratamento. Houve um único pico de expressão de IL-10 e de IFN-γ entre 18 e 30 dias após a inoculação, respectivamente. Este estudo sugere que o TNF-α participa da patogenia da fase aguda da erliquiose canina, e que o tratamento com hiclato de doxiciclina reduz os efeitos sistêmicos dessa citocina, possivelmente por reduzir ou eliminar a parasitemia.