Tumor necrosis factor alpha increases epithelial barrier permeability by disrupting tight junctions in Caco-2 cells

The objectives of this study were to determine the effect of tumor necrosis factor alpha (TNF-α) on intestinal epithelial cell permeability and the expression of tight junction proteins. Caco-2 cells were plated onto Transwell® microporous filters and treated with TNF-α (10 or 100 ng/mL) for 0, 4, 8, 16, or 24 h. The transepithelial electrical resistance and the mucosal-to-serosal flux rates of the established paracellular marker Lucifer yellow were measured in filter-grown monolayers of Caco-2 intestinal cells. The localization and expression of the tight junction protein occludin were detected by immunofluorescence and Western blot analysis, respectively. SYBR-Green-based real-time PCR was used to measure the expression of occludin mRNA. TNF-α treatment produced concentration- and time-dependent decreases in Caco-2 transepithelial resistance and increases in transepithelial permeability to the paracellular marker Lucifer yellow. Western blot results indicated that TNF-α decreased the expression of phosphorylated occludin in detergent-insoluble fractions but did not affect the expression of non-phosphorylated occludin protein. Real-time RT-PCR data showed that TNF-α did not affect the expression of occludin mRNA. Taken together, our data demonstrate that TNF-α increases Caco-2 monolayer permeability, decreases occludin protein expression and disturbs intercellular junctions.

Rosuvastatin prevents myocardial necrosis in an experimental model of acute myocardial infarction

Dyslipidemia is related to the progression of atherosclerosis and is an important risk factor for acute coronary syndromes. Our objective was to determine the effect of rosuvastatin on myocardial necrosis in an experimental model of acute myocardial infarction (AMI). Male Wistar rats (8-10 weeks old, 250-350 g) were subjected to definitive occlusion of the left anterior descending coronary artery to cause AMI. Animals were divided into 6 groups of 8 to 11 rats per group: G1, normocholesterolemic diet; G2, normocholesterolemic diet and rosuvastatin (1 mg·kg-1·day-1) 30 days after AMI; G3, normocholesterolemic diet and rosuvastatin (1 mg·kg-1·day-1) 30 days before and after AMI; G4, hypercholesterolemic diet; G5, hypercholesterolemic diet and rosuvastatin (1 mg·kg-1·day-1) 30 days after AMI; G6, hypercholesterolemic diet and rosuvastatin (1 mg·kg-1·day-1) 30 days before and after AMI. Left ventricular function was determined by echocardiography and percent infarct area by histology. Fractional shortening of the left ventricle was normal at baseline and decreased significantly after AMI (P < 0.05 in all groups), being lower in G4 and G5 than in the other groups. No significant difference in fractional shortening was observed between G6 and the groups on the normocholesterolemic diet. Percent infarct area was significantly higher in G4 than in G3. No significant differences were observed in infarct area among the other groups. We conclude that a hypercholesterolemic diet resulted in reduced cardiac function after AMI, which was reversed with rosuvastatin when started 30 days before AMI. A normocholesterolemic diet associated with rosuvastatin before and after AMI prevented myocardial necrosis when compared with the hypercholesterolemic condition.

Protective effects of tumor necrosis factor-α blockade by adalimumab on articular cartilage and subchondral bone in a rat model of osteoarthritis

We aimed to investigate the effects of an anti-tumor necrosis factor-α antibody (ATNF) on cartilage and subchondral bone in a rat model of osteoarthritis. Twenty-four rats were randomly divided into three groups: sham-operated group (n=8); anterior cruciate ligament transection (ACLT)+normal saline (NS) group (n=8); and ACLT+ATNF group (n=8). The rats in the ACLT+ATNF group received subcutaneous injections of ATNF (20 μg/kg) for 12 weeks, while those in the ACLT+NS group received NS at the same dose for 12 weeks. All rats were euthanized at 12 weeks after surgery and specimens from the affected knees were harvested. Hematoxylin and eosin staining, Masson's trichrome staining, and Mankin score assessment were carried out to evaluate the cartilage status and cartilage matrix degradation. Matrix metalloproteinase (MMP)-13 immunohistochemistry was performed to assess the cartilage molecular metabolism. Bone histomorphometry was used to observe the subchondral trabecular microstructure. Compared with the rats in the ACLT+NS group, histological and Mankin score analyses showed that ATNF treatment reduced the severity of the cartilage lesions and led to a lower Mankin score. Immunohistochemical and histomorphometric analyses revealed that ATNF treatment reduced the ACLT-induced destruction of the subchondral trabecular microstructure, and decreased MMP-13 expression. ATNF treatment may delay degradation of the extracellular matrix via a decrease in MMP-13 expression. ATNF treatment probably protects articular cartilage by improving the structure of the subchondral bone and reducing the degradation of the cartilage matrix.

Evaluación del uso terapeutico del factor de necrosis tumoral, factor de crecimiento epidermico e insulina sobre el proceso de cicatrización en lesiones causadas por la reacción de Shwartzman en la piel de conejos diabéticos

Tesis (Maestría en Ciencias con Especialidad en Inmunología) UANL

Respuesta inflamatoria y de fase aguda en el absceso hepático amibiano mediante el estudio de leucotrienos B4, C4, E4 y D4 e interleucina1 beta, interleucina 6 y factor de necrosis tumoral alfa

Tesis (Doctorado en Ciencias con Especialidad en Química Biomédica) UANL

Factor de necrosis tumoral y granulocitos neutrófilos en la producción del síndrome de insuficiencia respiratoria del adulto causado por peroxisomicina A-1

Tesis (Doctorado en Ciencias con Especialidad en Morfología) UANL

Interacción de la peroxisomicina A1 con el factor de necrosis tumoral alfa in vitro

Tesis (Doctorado en Ciencias con Especialidad en Farmacología y Toxicología) UANL

Factors associated to avascular necrosis of the femoral head and non-union in patients younger than 65 years old with displaced femoral neck fractures treated with reduction and internal fixation.

Pocos estudios han evaluado el tratamiento de las fracturas desplazadas de cuello femoral en pacientes menores de 65 años de edad, y no han sido claramente definidos los factores de riesgo para necrosis avascular o no-unión dentro de este rango de edad. Para determinar los factores asociados a la necrosis avascular de la cabeza femoral (AVN) y no-unión en pacientes menores de 65 años de edad con fracturas desplazadas del cuello femoral tratados con reducción y fijación interna, se realizó un estudio retrospectivo de 29 fracturas desplazadas del cuello femoral en 29 pacientes consecutivos tratados en una sola institución. La influencia de la edad, la energía del trauma, tipo de reducción, y el tiempo entre la fractura y el tratamiento en desarrollo de la AVN y no-unión fueron evaluados. Los pacientes que desarrollaron NAV fueron significativamente mayores y sufrieron un trauma de más baja energía que en los casos sin AVN. Ninguna variable fue asociada con la no-unión. La regresión logística determinó que sólo la edad se asoció de forma independiente a NAV. La edad es un buen predictor para el desarrollo de NAV, con un C-estadístico de 0.861, y un mejor corte-determinado en 53,5 años. Conclusión: Los pacientes de entre 53,5 y 65 años presentan un riesgo más alto de NAV. La artroplastia primaria se debe considerar en este subgrupo.

Resultados en reducción abierta de cadera por via interna: necrosis avascular y concordancia interobservador

Introducción: la luxación típica de la cadera es una patología con alta incidencia, de ahí la necesidad de métodos efectivos para lograr una reducción de esta. Han surgido varios métodos de reducción, uno de estos métodos es la reducción abierta por vía interna. Metodología Estudió descriptivo retrospectivo, incluyo los casos operados de reducción abierta por vía interna en el Instituto de Ortopedia Infantil Roosevelt y Clínica Jorge Piñeros Corpas por uno de los tutores, desde enero de 2006 hasta junio de 2011, valorando estas caderas según los criterios de Salter de necrosis avascular (NAV), con un seguimiento mínimo de 18 meses. Se evaluó la concordancia interobservador para la clasificación de Salter en tres ortopedistas infantiles. Resultados Se evaluaron 20 caderas en 16 pacientes a los que se realizo reducción abierta de luxación típica de cadera por vía interna. El 40 % de las caderas presentaron NAV, el 75% de estas caderas presentan NAV tipo I según la clasificación de Kalamchi. El índice Kappa para la clasificación de Salter en tres ortopedistas infantiles fue 0.6. Discusión El abordaje por vía interna para la reducción de luxación de cadera típica en niños menores de 18 meses es una alternativa más para el manejo de estos pacientes, que puede producir NAV en la cadera, pero esta NAV es tipo I de Kalamchi en la mayoría de casos. La reproducibilidad de la radiografía para la evaluación de NAV, realizada por personas expertas es buena, medida con índice kappa de 0.6.

Tratamiento de Artritis reumatoide con antagonistas del factor de necrosis tumoral alfa y su asociación con el desarrollo de melanoma cutáneo: revisión sistemática de la literatura y meta-análisis.

Introducción: El tratamiento con antagonistas del factor de necrosis tumoral alfa (anti TNF) ha impactado el pronóstico y la calidad de vida de los pacientes con artritis reumatoide (AR) positivamente, sin embargo, se interroga un incremento en el riesgo de desarrollar melanoma. Objetivo: Conocer la asociación entre el uso de anti TNF y el desarrollo de melanoma maligno en pacientes con AR. Metodología: Se realizó una búsqueda sistemática en MEDLINE, EMBASE, COCHRANE LIBRARY y LILACS para ensayos clínicos, estudios observacionales, revisiones y meta-análisis en pacientes adultos con diagnóstico de AR y manejo con anti TNF (Certolizumab pegol, Adalimumab, Etanercept, Infliximab y Golimumab). Resultados: 37 estudios clínicos cumplieron los criterios de inclusión para el meta-análisis, con una población de 16567 pacientes. El análisis de heterogeneidad no fue significativo (p=1), no se encontró diferencia en el riesgo entre los grupos comparados DR -0.00 (IC 95% -0.001; -0.001). Un análisis adicional de los estudios en los que se reportó al menos 1 caso de melanoma (4222 pacientes) tampoco mostró diferencia en el riesgo DR -0.00 (IC 95% -0.004 ; -0.003). Conclusión: En la evidencia disponible a la fecha no encontramos asociación significativa entre el tratamiento con anti TNF en pacientes con diagnóstico de AR y el desarrollo de melanoma cutáneo.

Tumour necrosis factor alpha induces rapid reduction in AMPA receptor-mediated calcium entry in motor neurones by increasing cell surface expression of the GluR2 subunit: relevance to neurodegeneration

The AMPA receptor (AMPAR) subunit GluR2, which regulates excitotoxicity and the inflammatory cytokine tumour necrosis factor alpha (TNF alpha) have both been implicated in motor neurone vulnerability in Amyotrophic Lateral Sclerosis/Motor Neurone Disease. TNF alpha has been reported to increase cell surface expression of AMPAR subunits to increase synaptic strength and enhance excitotoxicity, but whether this mechanism occurs in motor neurones is unknown. We used primary cultures of mouse motor neurones and cortical neurones to examine the interaction between TNF alpha receptor activation, GluR2 availability, AMPAR-mediated calcium entry and susceptibility to excitotoxicity. Short exposure to a physiologically relevant concentration of TNFalpha (10 ng/ml, 15 min) caused a marked redistribution of both GluR1 and GluR2 to the cell surface as determined by cell surface biotinylation and immunofluorescence. Using Fura-2 AM microfluorimetry we showed that exposure to TNFalpha caused a rapid reduction in the peak amplitude of AMPA-mediated calcium entry in a PI3-kinase and p38 kinase-dependent manner, consistent with increased insertion of GluR2-containing AMPAR into the plasma membrane. This resulted in a protection of motor neurones against kainate-induced cell death. Our data therefore, suggests that TNF alpha acts primarily as a physiological regulator of synaptic activity in motor neurones rather than a pathological drive in ALS

Apical leaf necrosis as a defence mechanism against pathogen attack: effects of high nutrient availability on onset and rate of necrosis

An outdoor experiment was conducted to increase understanding of apical leaf necrosis in the presence of pathogen infection. Holcus lanatus seeds and Puccinia coronata spores were collected from two adjacent and otherwise similar habitats with differing long-term N fertilization levels. After inoculation, disease and necrosis dynamics were observed during the plant growing seasons of 2003 and 2006. In both years high nutrient availability resulted in earlier disease onset, a higher pathogen population growth rate, earlier physiological apical leaf necrosis onset and a reduced time between disease onset and apical leaf necrosis onset. Necrosis rate was shown to be independent of nutrient availability. The results showed that in these nutrient-rich habitats H. lanatus plants adopted necrosis mechanisms which wasted more nutrients. There was some indication that these necrosis mechanisms were subject to local selection pressures, but these results were not conclusive. The findings of this study are consistent with apical leaf necrosis being an evolved defence mechanism.

Apical leaf necrosis and leaf nitrogen dynamics in diseased leaves: a model study

Apical leaf necrosis is a physiological process related to nitrogen (N) dynamics in the leaf. Pathogens use leaf nutrients and can thus accelerate this physiological apical necrosis. This process differs from necrosis occurring around pathogen lesions (lesion-induced necrosis), which is a direct result of the interaction between pathogen hyphae and leaf cells. This paper primarily concentrates on apical necrosis, only incorporating lesion-induced necrosis by necessity. The relationship between pathogen dynamics and physiological apical leaf necrosis is modelled through leaf nitrogen dynamics. The specific case of Puccinia triticina infections on Triticum aestivum flag leaves is studied. In the model, conversion of indirectly available N in the form of, for example, leaf cell proteins (N-2(t)) into directly available N (N-1(t), i.e. the form of N that can directly be used by either pathogen or plant sinks) results in apical necrosis. The model reproduces observed trends of disease severity, apical necrosis and green leaf area (GLA) and leaf N dynamics of uninfected and infected leaves. Decreasing the initial amount of directly available N results in earlier necrosis onset and longer necrosis duration. Decreasing the initial amount of indirectly available N, has no effect on necrosis onset and shortens necrosis duration. The model could be used to develop hypotheses on how the disease-GLA relation affects yield loss, which can be tested experimentally. Upon incorporation into crop simulation models, the model might provide a tool to more accurately estimate crop yield and effects of disease management strategies in crops sensitive to fungal pathogens.

The global effect of follicle-stimulating hormone and tumour necrosis factor α on gene expression in cultured bovine ovarian granulosa cells

Background Oocytes mature in ovarian follicles surrounded by granulosa cells. During follicle growth, granulosa cells replicate and secrete hormones, particularly steroids close to ovulation. However, most follicles cease growing and undergo atresia or regression instead of ovulating. To investigate the effects of stimulatory (follicle-stimulating hormone; FSH) and inhibitory (tumour necrosis factor alpha; TNFα) factors on the granulosa cell transcriptome, bovine ovaries were obtained from a local abattoir and pools of granulosa cells were cultured in vitro for six days under defined serum-free conditions with treatments present on days 3–6. Initially dose–response experiments (n = 4) were performed to determine the optimal concentrations of FSH (0.33 ng/ml) and TNFα (10 ng/ml) to be used for the microarray experiments. For array experiments cells were cultured under control conditions, with FSH, with TNFα, or with FSH plus TNFα (n = 4 per group) and RNA was harvested for microarray analyses. Results Statistical analysis showed primary clustering of the arrays into two groups, control/FSH and TNFα/TNFα plus FSH. The effect of TNFα on gene expression dominated that of FSH, with substantially more genes differentially regulated, and the pathways and genes regulated by TNFα being similar to those of FSH plus TNFα treatment. TNFα treatment reduced the endocrine activity of granulosa cells with reductions in expression of FST, INHA, INBA and AMH. The top-ranked canonical pathways and GO biological terms for the TNFα treatments included antigen presentation, inflammatory response and other pathways indicative of innate immune function and fibrosis. The two most significant networks also reflect this, containing molecules which are present in the canonical pathways of hepatic fibrosis/hepatic stellate cell activation and transforming growth factor β signalling, and these were up regulated. Upstream regulator analyses also predicted TNF, interferons γ and β1 and interleukin 1β. Conclusions In vitro, the transcriptome of granulosa cells responded minimally to FSH compared with the response to TNFα. The response to TNFα indicated an active process akin to tissue remodelling as would occur upon atresia. Additionally there was reduction in endocrine function and induction of an inflammatory response to TNFα that displays features similar to immune cells.

Tumor necrosis factor alpha triggers proliferation of adult neural stem cells via IKK/NF-kappaB signaling

BACKGROUND: Brain inflammation has been recognized as a complex phenomenon with numerous related aspects. In addition to the very well-described neurodegenerative effect of inflammation, several studies suggest that inflammatory signals exert a potentially positive influence on neural stem cell proliferation, migration and differentiation. Tumor necrosis factor alpha (TNF-alpha) is one of the best-characterized mediators of inflammation. To date, conclusions about the action of TNF on neural stem or progenitor cells (NSCs, NPCs) have been conflicting. TNF seems to activate NSC proliferation and to inhibit their differentiation into NPCs. The purpose of the present study was to analyze the molecular signal transduction mechanisms induced by TNF and resulting in NSC proliferation. RESULTS: Here we describe for the first time the TNF-mediated signal transduction cascade in neural stem cells (NSCs) that results in increased proliferation. Moreover, we demonstrate IKK-alpha/beta-dependent proliferation and markedly up-regulated cyclin D1 expression after TNF treatment. The significant increase in proliferation in TNF-treated cells was indicated by increased neurosphere volume, increased bromodeoxyuridin (BrdU) incorporation and a higher total cell number. Furthermore, TNF strongly activated nuclear factor-kappa B (NF-kappaB) as measured by reporter gene assays and by an activity-specific antibody. Proliferation of control and TNF-treated NSCs was strongly inhibited by expression of the NF-kappaB super-repressor IkappaB-AA1. Pharmacological blockade of IkappaB ubiquitin ligase activity led to comparable decreases in NF-kappaB activity and proliferation. In addition, IKK-beta gene product knock-down via siRNA led to diminished NF-kappaB activity, attenuated cyclin D1 expression and finally decreased proliferation. In contrast, TGFbeta-activated kinase 1 (TAK-1) is partially dispensable for TNF-mediated and endogenous proliferation. Understanding stem cell proliferation is crucial for future regenerative and anti-tumor medicine. CONCLUSION: TNF-mediated activation of IKK-beta resulted in activation of NF-kappaB and was followed by up-regulation of the bona-fide target gene cyclin D1. Activation of the canonical NF-kappaB pathway resulted in strongly increased proliferation of NSCs.