RESUMEN
Porcine reproductive and respiratory syndrome virus (PRRSV) is a positive-stranded RNA virus that grows in macrophages and causes acute pneumonia in pigs. PRRSV causes devastating losses to the porcine industry. However, due to its high antigenic variability and poorly understood immunopathogenesis, there is currently no effective vaccine or treatment to control PRRSV infection. The common occurrence of PRRSV infection with bacterial infections as well as its inflammatory-driven pathobiology raises the question of the value of antibiotics with immunomodulating properties for the treatment of the disease it causes. The macrolide antibiotic Tulathromycin (TUL) has been found to exhibit potent anti-inflammatory and immunomodulating properties in cattle and pigs. The aim of this study was to characterize the anti-viral and immunomodulating properties of TUL in PRRSV-infected porcine macrophages. Our findings indicate that blood monocyte-derived macrophages are readily infected by PRRSV and can be used as an effective cellular model to study PRRSV pathogenesis. TUL did not change intracellular or extracellular viral titers, not did it alter viral receptors (CD163 and CD169) expression on porcine macrophages. In contrast, TUL exhibited potent immunomodulating properties, which therefore occurred in the absence of any direct antiviral effects against PRRSV. TUL had an additive effect with PRRSV on the induction of macrophage apoptosis, and inhibited virus-induced necrosis. TUL significantly attenuated PRRSV-induced macrophage pro-inflammatory signaling (CXCL-8 and mitochondrial ROS production) and prevented PRRSV inhibition of non-opsonized and opsonized phagocytic function. Together, these data demonstrate that TUL inhibits PRRSV-induced inflammatory responses in porcine macrophages and protects against the phagocytic impairment caused by the virus. Research in live pigs is warranted to assess the potential clinical benefits of this antibiotic in the context of virally induced inflammation and tissue injury.
Asunto(s)
Disacáridos/farmacología , Compuestos Heterocíclicos/farmacología , Factores Inmunológicos/farmacología , Macrófagos/virología , Síndrome Respiratorio y de la Reproducción Porcina/patología , Virus del Síndrome Respiratorio y Reproductivo Porcino/fisiología , Animales , Antígenos CD , Antígenos de Diferenciación Mielomonocítica , Apoptosis/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Línea Celular , Forma de la Célula/efectos de los fármacos , Disacáridos/uso terapéutico , Femenino , Compuestos Heterocíclicos/uso terapéutico , Factores Inmunológicos/uso terapéutico , Interleucina-10/metabolismo , Interleucina-8/metabolismo , Espacio Intracelular/metabolismo , Macrófagos/efectos de los fármacos , Masculino , Ratones , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Necrosis , Proteínas Opsoninas/metabolismo , Fagocitosis/efectos de los fármacos , Síndrome Respiratorio y de la Reproducción Porcina/tratamiento farmacológico , Virus del Síndrome Respiratorio y Reproductivo Porcino/efectos de los fármacos , Especies Reactivas de Oxígeno/metabolismo , Receptores de Superficie Celular , Receptores Virales/metabolismo , Lectina 1 Similar a Ig de Unión al Ácido Siálico/metabolismo , Porcinos , Replicación Viral/efectos de los fármacosRESUMEN
Clearance of apoptotic neutrophils is a central feature of the resolution of inflammation. Findings indicate that immuno-modulation and induction of neutrophil apoptosis by macrolide antibiotics generate anti-inflammatory benefits via mechanisms that remain obscure. Tulathromycin (TUL), a new antimicrobial agent for bovine respiratory disease, offers superior clinical efficacy for reasons not fully understood. The aim of this study was to identify the immuno-modulating effects of tulathromycin and, in this process, to establish tulathromycin as a new model for characterizing the novel anti-inflammatory properties of antibiotics. Bronchoalveolar lavage specimens were collected from Holstein calves 3 and 24 h postinfection, challenged intratracheally with live Mannheimia haemolytica (2 × 10(7) CFU), and treated with vehicle or tulathromycin (2.5 mg/kg body weight). Terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling (TUNEL) staining and enzyme-linked immunosorbent assay (ELISA) revealed that tulathromycin treatment significantly increased leukocyte apoptosis and reduced levels of proinflammatory leukotriene B(4) in M. haemolytica-challenged calves. In vitro, tulathromycin concentration dependently induced apoptosis in freshly isolated bovine neutrophils from healthy steers in a capase-3-dependent manner but failed to induce apoptosis in bovine fibroblasts, epithelial cells, and endothelial cells, as well as freshly isolated bovine blood monocytes and monocyte-derived macrophages. The proapoptotic effects of TUL were also, in part, drug specific; equimolar concentrations of penicillin G, oxytetracycline, and ceftiofur failed to cause apoptosis in bovine neutrophils. In addition, tulathromycin significantly reduced levels of phosphorylated IκBα, nuclear translocation of NF-κB p65, and mRNA levels of proinflammatory interleukin-8 in lipopolysaccharide (LPS)-stimulated bovine neutrophils. The findings illustrate novel mechanisms through which tulathromycin confers anti-inflammatory benefits.
Asunto(s)
Antibacterianos/farmacología , Antiinflamatorios/farmacología , Apoptosis/efectos de los fármacos , Caspasa 3/metabolismo , Disacáridos/farmacología , Compuestos Heterocíclicos/farmacología , Interleucina-8/genética , FN-kappa B/metabolismo , Neutrófilos/efectos de los fármacos , Animales , Western Blotting , Bovinos , Línea Celular , Células Cultivadas , Fragmentación del ADN/efectos de los fármacos , Ensayo de Inmunoadsorción Enzimática , Etiquetado Corte-Fin in Situ , Leucotrieno B4/metabolismo , Masculino , FN-kappa B/genética , Neutrófilos/citología , Neutrófilos/metabolismo , Reacción en Cadena de la Polimerasa , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , Transcripción Genética/efectos de los fármacos , Transcripción Genética/genéticaRESUMEN
The human gastrointestinal tract hosts a complex community of microorganisms that grow as biofilms on the intestinal mucosa. These bacterial communities are not well characterized, although they are known to play an important role in human health. This study aimed to develop a model for culturing biofilms (surface-adherent communities) of intestinal microbiota. The model utilizes adherent mucosal bacteria recovered from colonic biopsies to create multi-species biofilms. Culture on selective media and confocal microscopy indicated the biofilms were composed of a diverse community of bacteria. Molecular analyses confirmed that several phyla were represented in the model, and demonstrated stability of the community over 96 h when cultured in the device. This model is novel in its use of a multi-species community of mucosal bacteria grown in a biofilm mode of growth.
Asunto(s)
Bacterias/genética , Biopelículas/clasificación , Colon/microbiología , Metagenoma , Anaerobiosis , Bacterias/clasificación , Bacterias/aislamiento & purificación , Adhesión Bacteriana , Fenómenos Fisiológicos Bacterianos , Colon/química , Humanos , Mucosa Intestinal/química , Mucosa Intestinal/microbiología , Microscopía Confocal , Modelos BiológicosRESUMEN
Giardia is the most prevalent human intestinal parasitic protist in the world, and one of the most common parasite of companion animals and young livestock. Giardia is a major cause of diarrhea in children and in travelers. The host-microbial interactions that govern the outcome of infection remain incompletely understood. Findings available to date indicate that the infection causes diarrhea via a combination of intestinal malabsorption and hypersecretion. Malabsorption and maldigestion mainly result from a diffuse shortening of epithelial microvilli. This enterocytic injury is mediated by activated host T lymphocytes. Pathophysiological activation of lymphocytes is secondary to Giardia-induced disruption of epithelial tight junctions, which in turn increases intestinal permeability. Loss of epithelial barrier function is a result of Giardia-induced enterocyte apoptosis. Recent findings suggest that these effects may facilitate the development of chronic enteric disorders, including inflammatory bowel disease, irritable bowel syndrome, and allergies, via mechanisms that remain poorly understood. A newly discovered SGLT-1 glucose uptake-mediated host cytoprotective mechanism may represent an effective modulator of the epithelial apoptosis induced by this parasite, and, possibly, by other enteropathogens. A better understanding of the pathogenesis of giardiasis will shed light on new potential therapeutic targets.
Asunto(s)
Apoptosis/fisiología , Giardia/fisiología , Giardiasis/parasitología , Mucosa Intestinal/parasitología , Intestino Delgado/parasitología , Animales , Giardia/patogenicidad , Giardiasis/metabolismo , Giardiasis/veterinaria , Interacciones Huésped-Parásitos , Humanos , Absorción Intestinal , Mucosa Intestinal/metabolismo , Intestino Delgado/metabolismoRESUMEN
T lymphocyte-mediated pathogenesis is common to a variety of enteropathies, including giardiasis, cryptosporidiosis, bacterial enteritis, celiac's disease, food anaphylaxis, and Crohn's disease. In giardiasis as well as in these other disorders, a diffuse loss of microvillous brush border, combined or not with villus atrophy, is responsible for disaccharidase insufficiencies and malabsorption of electrolytes, nutrients, and water, which ultimately cause diarrheal symptoms. Other mucosal changes may include crypt hyperplasia and increased infiltration of intra-epithelial lymphocytes. Recent studies using models of giardiasis have shed new light on the immune regulation of these abnormalities. Indeed, experiments using an athymic mouse model of infection have found that these epithelial injuries were T cell-dependent. Findings from further research indicate that that the loss of brush border surface area, reduced disaccharidase activities, and increase crypt-villus ratios are mediated by CD8+ T cells, whereas both CD8+ and CD4+ small mesenteric lymph node T cells regulate the influx of intra-epithelial lymphocytes. Future investigations need to characterize the CD8+ T cell signaling cascades that ultimately lead to epithelial injury and malfunction in giardiasis and other malabsorptive disorders of the intestine.
Asunto(s)
Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Giardiasis/patología , Mucosa Intestinal/patología , Animales , Giardiasis/inmunología , Humanos , Mucosa Intestinal/inmunología , Mucosa Intestinal/parasitología , Microvellosidades/parasitología , Microvellosidades/patologíaRESUMEN
T lymphocyte-mediated pathogenesis is common to a variety of enteropathies, including giardiasis, cryptosporidiosis, bacterial enteritis, celiac's disease, food anaphylaxis, and Crohn's disease. In giardiasis as well as in these other disorders, a diffuse loss of microvillous brush border, combined or not with villus atrophy, is responsible for disaccharidase insufficiencies and malabsorption of electrolytes, nutrients, and water, which ultimately cause diarrheal symptoms. Other mucosal changes may include crypt hyperplasia and increased infiltration of intra-epithelial lymphocytes. Recent studies using models of giardiasis have shed new light on the immune regulation of these abnormalities. Indeed, experiments using an athymic mouse model of infection have found that these epithelial injuries were T cell-dependent. Findings from further research indicate that that the loss of brush border surface area, reduced disaccharidase activities, and increase crypt-villus ratios are mediated by CD8+ T cells, whereas both CD8+ and CD4+ small mesenteric lymph node T cells regulate the influx of intra-epithelial lymphocytes. Future investigations need to characterize the CD8+ T cell signaling cascades that ultimately lead to epithelial injury and malfunction in giardiasis and other malabsorptive disorders of the intestine.
Asunto(s)
Animales , Humanos , /inmunología , /inmunología , Giardiasis/patología , Mucosa Intestinal/patología , Giardiasis/inmunología , Mucosa Intestinal/inmunología , Mucosa Intestinal/parasitología , Microvellosidades/parasitología , Microvellosidades/patologíaRESUMEN
This review summarizes the main aspects and newest findings of how proteinase-activated receptor 1 (PAR-1) may modulate programmed cell death. Activation of PAR-1 has been found to induce or inhibit apoptosis in a variety of cells, depending on the dosage of its physiological agonist thrombin, or that of synthetic receptor activators. To date, cellular targets for PAR-1-mediated effects on apoptosis include neuronal, endothelial, and epithelial cells, fibroblasts, and tumor cells. The signaling pathways involved in the induction or prevention of apoptosis by PAR-1 activation are diverse, and include JAK/STAT, RhoA, myosin light chain kinase, ERK1/2, and various Bcl-2 family members. In view of the well-established involvement of microbial proteinases in host tissue malfunction, the article also elaborates on the possible significance of PAR-1 activation for the pathogenesis of infectious disease.
Asunto(s)
Apoptosis , Receptor PAR-1/metabolismo , Animales , Línea Celular Tumoral , Enfermedades Transmisibles/etiología , Humanos , Janus Quinasa 2 , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Modelos Biológicos , Cadenas Ligeras de Miosina/efectos de los fármacos , Cadenas Ligeras de Miosina/metabolismo , Proteínas Tirosina Quinasas/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Transducción de Señal , Trombina/farmacología , Proteína de Unión al GTP rhoA/metabolismoRESUMEN
The mechanisms whereby exogenous growth hormone modulates intestinal structure and function in fish were investigated. Goldfish (Carassius auratus) were fed commercial flake diet sprayed with recombinant carp growth hormone (cGH) daily for 1 month. Control animals received food sprayed with the vehicle. After 1 month of daily feedings, body mass and length were determined, and animals were sacrificed to study intestinal characteristics. Sections of foregut were removed after determination of total gut length for measurement of leucine uptake, histology, and epithelial ultrastructure. Oral administration of cGH for 1 month resulted in a 40% increase in body mass and an 8% increase in body length above controls. Gut length was 43% greater and the gut length to body length ratio was 32% greater as a result of the cGH treatment. Feeding with cGH also resulted in a significant increase in leucine uptake and increased gut mucosal thickness. Analysis of transmission electron micrographs revealed significant increases in the microvillous height and density and epithelial surface area. The findings indicate that growth hormone added to feed may increase growth in fish, in part by significantly increasing gut length, mucosal thickness, and epithelial brush border surface area, leading to enhanced epithelial absorption.
Asunto(s)
Carpa Dorada/metabolismo , Hormona del Crecimiento/farmacología , Absorción Intestinal/efectos de los fármacos , Mucosa Intestinal/ultraestructura , Intestinos/anatomía & histología , Leucina/metabolismo , Animales , Carpa Dorada/crecimiento & desarrollo , Absorción Intestinal/fisiología , Mucosa Intestinal/efectos de los fármacos , Mucosa Intestinal/crecimiento & desarrollo , Mucosa Intestinal/metabolismo , Intestinos/crecimiento & desarrollo , Intestinos/fisiologíaRESUMEN
BACKGROUND AND AIMS: The colonic epithelium plays a key role in host defence. During colitis, epithelial function is impaired, leading to elevated bacterial translocation and exacerbation of inflammation. We previously documented perturbation of epithelial function, in terms of secretion and as a barrier to bacterial translocation, that persisted long after resolution of a bout of colitis in the rat. The mechanisms underlying the epithelial dysfunction are not completely understood. METHODS: Given the ability of prostaglandin (PG) D2 to suppress colonic epithelial secretion, we investigated the potential roles of this eicosanoid and of cyclooxygenase 2 (COX-2) in mediating post-colitis epithelial secretory and barrier dysfunction. RESULTS: Six weeks after induction of colitis with trinitrobenzene sulphonic acid, there was marked elevated synthesis of PGD2 and elevated COX-2 expression. Selective COX-2 inhibition abolished the increase in PGD2 synthesis. Colonic chloride secretory responses (in vitro) were significantly diminished relative to those in controls, a defect that was reversed by pre-exposure to a selective COX-2 inhibitor (celecoxib) but not to a selective COX-1 inhibitor (SC-560). The hyporesponsiveness was mimicked by pre-exposure of normal colonic tissue to PGD2, but not to its metabolite, 15-deoxy-Delta(12-14)PGJ2. The post-colitis rats exhibited a 10-fold increase in bacterial colonisation of the colon, and >3-fold increase in bacterial translocation. Twice daily treatment for one week with a selective COX-2 inhibitor (rofecoxib) did not affect bacterial colonisation but abolished the increase in bacterial translocation. CONCLUSIONS: These studies demonstrate an important role for COX-2, possibly via generation of PGD2, in mediating the prolonged epithelial secretory and barrier dysfunction after a bout of colitis in the rat.
Asunto(s)
Colitis/metabolismo , Colon/metabolismo , Isoenzimas/metabolismo , Prostaglandina D2/metabolismo , Prostaglandina-Endoperóxido Sintasas/metabolismo , Animales , Traslocación Bacteriana/fisiología , Ciclooxigenasa 1 , Ciclooxigenasa 2 , Inhibidores de la Ciclooxigenasa 2 , Inhibidores de la Ciclooxigenasa/farmacología , Mucosa Intestinal/metabolismo , Isoenzimas/antagonistas & inhibidores , Masculino , Proteínas de la Membrana , Ratas , Ratas WistarRESUMEN
This study examines the ability of Giardia duodenalis trophozoites, isolated from a wild bird, to colonize the intestinal tracts of companion animals (kittens) and domestic ruminants (lambs). Trophozoites colonized the intestinal tracts of intraduodenally inoculated animals as demonstrated by increasing parasite burdens within the duodenum and jejunum and by fecal passage of cysts within 4 days post-inoculation. The pathogenesis of the trophozoites was further investigated in kittens. In these animals, infection significantly reduced jejunal brush border microvillous length and density, which resulted in a loss of overall epithelial brush border surface area. This injury was associated with the production of diarrhea in four of five infected kittens. These findings indicate that some bird species may carry G. duodenalis that represent a possible health threat to companion animals and livestock. Our results describe the first successful colonization of avian-derived G. duodenalis trophozoites in the small intestines of domestic kittens and lambs.
Asunto(s)
Gatos/parasitología , Giardia/aislamiento & purificación , Giardia/fisiología , Giardiasis/veterinaria , Intestinos/parasitología , Loros/parasitología , Oveja Doméstica/parasitología , Animales , Enfermedades de las Aves/parasitología , Portador Sano , Enfermedades de los Gatos/parasitología , Enfermedades de los Gatos/patología , Duodeno/parasitología , Duodeno/patología , Heces/parasitología , Giardiasis/patología , Intestinos/patología , Intestinos/ultraestructura , Yeyuno/parasitología , Yeyuno/patología , Yeyuno/ultraestructura , Microvellosidades/parasitología , Microvellosidades/patología , Microvellosidades/ultraestructura , Enfermedades de las Ovejas/parasitologíaRESUMEN
In order to improve our understanding of the host cell-parasite interactions in giardiasis, this study assessed the effects of Giardia lamblia on epithelial permeability and tight junctional ZO-1, determined whether epidermal growth factor (EGF) may affect Giardia-induced epithelial injury, and evaluated if EGF modulates epithelial colonization by live G. lamblia trophozoites. Permeability was assessed in assays of trans-epithelial fluxes of FITC-dextran, and ZO-1 integrity was characterized by confocal laser immunofluorescence microscopy in confluent epithelial cell monolayers. G. lamblia significantly increased paracellular permeability and disrupted tight-junctional ZO-1 of a novel non-transformed human small intestinal epithelial cell line (SCBN). Pre-treatment with EGF prevented the development of these abnormalities and significantly inhibited attachment of live trophozoites to the enterocytes, independently of a direct microbiocidal action. These findings demonstrate that G. lamblia may cause intestinal pathophysiology by disrupting tight junctional ZO-1 and increasing epithelial permeability. Apical administration of EGF prevents these abnormalities, and reduces epithelial colonization by the live parasites.
Asunto(s)
Giardia lamblia/crecimiento & desarrollo , Giardiasis/parasitología , Intestino Delgado/parasitología , Proteínas de la Membrana/metabolismo , Fosfoproteínas/metabolismo , Uniones Estrechas/fisiología , Animales , Permeabilidad de la Membrana Celular/fisiología , Factor de Crecimiento Epidérmico/farmacología , Células Epiteliales , Giardia lamblia/metabolismo , Giardiasis/metabolismo , Giardiasis/patología , Interacciones Huésped-Parásitos , Humanos , Mucosa Intestinal/metabolismo , Mucosa Intestinal/parasitología , Mucosa Intestinal/patología , Intestino Delgado/metabolismo , Intestino Delgado/patología , Microscopía Confocal , Proteína de la Zonula Occludens-1RESUMEN
The thrombin receptor, protease-activated receptor-1 (PAR-1), has wide tissue distribution and is involved in many physiological functions. Because thrombin is in the intestinal lumen and mucosa during inflammation, we sought to determine PAR-1 expression and function in human intestinal epithelial cells. RT-PCR showed PAR-1 mRNA expression in SCBN cells, a nontransformed duodenal epithelial cell line. Confluent SCBN monolayers mounted in Ussing chambers responded to PAR-1 activation with a Cl(-)-dependent increase in short-circuit current. The secretory effect was blocked by BaCl2 and the Ca(2+)-ATPase inhibitor thapsigargin, but not by the L-type Ca(2+) channel blocker verapamil or DIDS, the nonselective inhibitor of Ca(2+)-dependent Cl(-) transport. Responses to thrombin and PAR-1-activating peptides exhibited auto- and crossdesensitization. Fura 2-loaded SCBN cells had increased fluorescence after PAR-1 activation, indicating increased intracellular Ca(2+). RT-PCR showed that SCBN cells expressed mRNA for the cystic fibrosis transmembrane conductance regulator (CFTR) and hypotonicity-activated Cl(-) channel-2 but not for the Ca(2+)-dependent Cl(-) channel-1. PAR-1 activation failed to increase intracellular cAMP, suggesting that the CFTR channel is not involved in the Cl(-) secretory response. Our data demonstrate that PAR-1 is expressed on human intestinal epithelial cells and regulates a novel Ca(2+)-dependent Cl(-) secretory pathway. This may be of clinical significance in inflammatory intestinal diseases with elevated thrombin levels.
Asunto(s)
Calcio/fisiología , Cloruros/metabolismo , Mucosa Intestinal/metabolismo , Receptores de Trombina/fisiología , Línea Celular , AMP Cíclico/biosíntesis , Regulador de Conductancia de Transmembrana de Fibrosis Quística/biosíntesis , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Conductividad Eléctrica , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Humanos , Transporte Iónico , Oligopéptidos/farmacología , ARN Mensajero/biosíntesis , Receptor PAR-1 , Receptores de Trombina/genética , Trombina/farmacologíaRESUMEN
Bleeding and delayed healing of ulcers are well recognized clinical problems associated with the use of aspirin and other nonsteroidal antiinflammatory drugs, which have been attributed to their antiaggregatory effects on platelets. We hypothesized that antiplatelet drugs might interfere with gastric ulcer healing by suppressing the release of growth factors, such as vascular endothelial growth factor (VEGF), from platelets. Gastric ulcers were induced in rats by serosal application of acetic acid. Daily oral treatment with vehicle, aspirin, or ticlopidine (an ADP receptor antagonist) was started 3 days later and continued for 1 week. Ulcer induction resulted in a significant increase in serum levels of VEGF and a significant decrease in serum levels of endostatin (an antiangiogenic factor). Although both aspirin and ticlopidine markedly suppressed platelet aggregation, only ticlopidine impaired gastric ulcer healing and angiogenesis as well as reversing the ulcer-associated changes in serum levels of VEGF and endostatin. The effects of ticlopidine on ulcer healing and angiogenesis were mimicked by immunodepletion of circulating platelets, and ticlopidine did not influence ulcer healing when given to thrombocytopenic rats. Incubation of human umbilical vein endothelial cells with serum from ticlopidine-treated rats significantly reduced proliferation and increased apoptosis, effects reversed by an antibody directed against endostatin. Ticlopidine treatment resulted in increased platelet endostatin content and release. These results demonstrate a previously unrecognized contribution of platelets to the regulation of gastric ulcer healing. Such effects likely are mediated through the release from platelets of endostatin and possibly VEGF. As shown with ticlopidine, drugs that influence gastric ulcer healing may do so in part through altering the ability of platelets to release growth factors.
Asunto(s)
Inhibidores de la Angiogénesis/fisiología , Plaquetas/fisiología , Colágeno/fisiología , Factores de Crecimiento Endotelial/fisiología , Linfocinas/fisiología , Fragmentos de Péptidos/fisiología , Úlcera Gástrica/tratamiento farmacológico , Cicatrización de Heridas/efectos de los fármacos , Inhibidores de la Angiogénesis/sangre , Animales , Antiinflamatorios no Esteroideos/efectos adversos , Antiinflamatorios no Esteroideos/metabolismo , Antiinflamatorios no Esteroideos/uso terapéutico , Apoptosis , Aspirina/efectos adversos , Aspirina/metabolismo , Aspirina/uso terapéutico , Plaquetas/efectos de los fármacos , Plaquetas/metabolismo , División Celular/efectos de los fármacos , Células Cultivadas , Colágeno/sangre , Modelos Animales de Enfermedad , Endostatinas , Factores de Crecimiento Endotelial/sangre , Endotelio Vascular/citología , Humanos , Linfocinas/sangre , Masculino , Neovascularización Patológica , Fragmentos de Péptidos/sangre , Agregación Plaquetaria/efectos de los fármacos , Inhibidores de Agregación Plaquetaria/efectos adversos , Inhibidores de Agregación Plaquetaria/metabolismo , Inhibidores de Agregación Plaquetaria/uso terapéutico , Ratas , Ratas Wistar , Úlcera Gástrica/sangre , Úlcera Gástrica/inducido químicamente , Úlcera Gástrica/patología , Ticlopidina/efectos adversos , Ticlopidina/metabolismo , Ticlopidina/uso terapéutico , Factor A de Crecimiento Endotelial Vascular , Factores de Crecimiento Endotelial VascularRESUMEN
Twelve Giardia duodenalis-infected Holstein dairy calves were allocated into a treatment (n=6) and placebo group (n=6) according to pre-study faecal cyst counts. Calves in the treatment group received an oral dose of 5 mg/kg fenbendazole once daily for 3 days, while placebo calves received a sterile saline solution. Calves were euthanised 7 days following the initiation of treatment and intestinal were collected and prepared for trophozoite quantitation, histology, electron microscopy, and disaccharidase assays. In all calves treated with fenbendazole, intestinal trophozoites were below detection limits, while in saline-treated calves, trophozoites were observed in all intestinal segments. Histologically, no significant difference was observed between treatment groups with respect to intestinal villus height or crypt depth. However, a significant decline in the number of intraepithelial lymphocytes (IEL) was observed in fenbendazole-treated calves when compared with placebo-treated calves in the duodenum (13.9+/-1.2 vs. 17.0+/-1.1 IEL/100 enterocytes) and jejunum (21.6+/-0.8 vs. 30.7+/-1.0 IEL/100 enterocytes). In addition, measurements from TEM micrographs demonstrated a significant increase in microvillus surface area in the jejunum of fenbendazole-treated calves compared with saline-treated calves (31.2+/-10.2 vs. 22.8+/-7.6 microm(2)). This increase in microvillus surface area was also associated with an increase in jejunal maltase activity in fenbendazole-treated calves compared with calves treated with saline. These results demonstrate that fenbendazole is an effective treatment for giardiasis in calves. fenbendazole treatment eliminated Giardia trophozoites from the small intestine of calves resulting in increased microvillus surface area and greater intestinal enzyme activity. This study also demonstrates that the pathogenesis of giardiasis in calves is similar to that observed in humans and laboratory animals, and provides further evidence that Giardia is a pathogen of cattle with potential economic importance.
Asunto(s)
Antinematodos/uso terapéutico , Enfermedades de los Bovinos/tratamiento farmacológico , Industria Lechera , Fenbendazol/uso terapéutico , Giardiasis/veterinaria , Intestinos/efectos de los fármacos , Animales , Antinematodos/administración & dosificación , Bovinos , Esquema de Medicación/veterinaria , Femenino , Fenbendazol/administración & dosificación , Giardia lamblia/patogenicidad , Giardiasis/tratamiento farmacológico , Intestinos/patología , Intestinos/ultraestructura , Microscopía Electrónica/veterinaria , Relación Estructura-ActividadRESUMEN
The expression of the interleukin-8 (IL-8) gene was examined by in situ hybridization in lung tissues from calves experimentally infected with Mannheimia (Pasteurella) haemolytica and treated with tilmicosin. Interleukin-8 mRNA expression was detected in alveolar areas, particularly along interlobular septa, in the lumen, and in the epithelial cells of some bronchioles. In lesional lung tissues from animals that had received tilmicosin, we found large areas with limited inflammation. There was no staining for IL-8 mRNA in these areas. In contrast, in strongly inflamed areas, the same patterns and intensities of staining for IL-8 mRNA were detected in tilmicosin- and sham-treated animals. We conclude that tilmicosin does not affect the expression of IL-8 mRNA in tissue showing microscopic signs of inflammation. Together with previous reports, this supports the view that the pro-apoptotic properties of tilmicosin on neutrophils do not compromise the host defense mechanisms required to control the infection.
Asunto(s)
Antibacterianos/farmacología , Enfermedades de los Bovinos/tratamiento farmacológico , Interleucina-8/biosíntesis , Macrólidos , Mannheimia haemolytica , Infecciones por Pasteurella/veterinaria , Tilosina/análogos & derivados , Animales , Apoptosis/efectos de los fármacos , Bovinos/microbiología , Enfermedades de los Bovinos/microbiología , Hibridación in Situ/veterinaria , Interleucina-8/genética , Pulmón/microbiología , Neutrófilos/efectos de los fármacos , Infecciones por Pasteurella/tratamiento farmacológico , Tilosina/farmacologíaRESUMEN
The mechanisms of epithelial injury in giardiasis remain unknown. The effects of live Giardia lamblia on cellular G-actin, F-actin, alpha-actinin, and electrical resistance of human intestinal epithelial monolayers were investigated using SCBN and Caco2 cell lines grown on chamber slides or Transwell filter membranes. In separate experiments, some monolayers were also exposed to sonicated trophozoites, some to supernatant from live G. lamblia cultures, and some with or without the Ca2+ channel blocker verapamil. After 2, 24, or 48 hr of coincubation with G. lamblia, monolayers were assessed for cytoskeletal arrangement under fluorescence and confocal laser microscopy, and transepithelial electrical resistance was measured. Exposure to live G. lamblia trophozoites induced localized condensation of F-actin and loss of perijunctional alpha-actinin while G-actin remained unchanged. Confocal laser microscopy indicated that F-actin rearrangement was not affected by verapamil and was localized within the terminal web area. Coincubation of monolayers with G. lamblia lysates or with spent medium alone similarly rearranged F-actin. Verapamil alone did not alter F-actin. Electrical resistance of SCBN and Caco2 monolayers exposed to G. lamblia was significantly decreased versus controls regardless of whether live or lysed trophozoite samples were used. The results indicate that G. lamblia-induced epithelial injury is associated with F-actin and alpha-actinin rearrangements in the terminal web area via mechanisms independent of extracellular Ca2+. These alterations are associated with reduced transepithelial electrical resistance and are due at least in part to trophozoite products.
Asunto(s)
Actinina/metabolismo , Actinas/metabolismo , Colon/parasitología , Duodeno/parasitología , Giardia lamblia/fisiología , Mucosa Intestinal/fisiología , Actinina/efectos de los fármacos , Actinas/efectos de los fármacos , Animales , Células CACO-2 , Bloqueadores de los Canales de Calcio/farmacología , Línea Celular , Colon/citología , Colon/efectos de los fármacos , Colon/fisiología , Duodeno/citología , Duodeno/efectos de los fármacos , Duodeno/fisiología , Impedancia Eléctrica , Humanos , Mucosa Intestinal/parasitología , Microscopía Confocal , Microscopía Fluorescente , Verapamilo/farmacologíaRESUMEN
Pathogen virulence factors and inflammation are responsible for tissue injury associated with respiratory failure in bacterial pneumonia, as seen in the bovine lung infected with Pasteurella haemolytica. Tilmicosin is a macrolide antibiotic used for the treatment of bovine bacterial pneumonia. Recent evidence suggests that tilmicosin-induced neutrophil apoptosis may have anti-inflammatory effects. Using bovine leukocytes, we sought to define whether live P. haemolytica affected tilmicosin-induced neutrophil apoptosis, assessed the proapoptotic effects of tilmicosin in comparison with other drugs, and characterized its impact on phagocytic uptake of neutrophils by macrophages. Induction of apoptosis in the presence or absence of P. haemolytica was assessed by using an enzyme-linked immunosorbent assay for apoptotic nucleosomes. In addition, fluorescent annexin-V staining identified externalized phosphatidylserine in neutrophils treated with tilmicosin, penicillin, ceftiofur, oxytetracycline, or dexamethasone. Neutrophil membrane integrity was assessed by using propidium iodide and trypan blue exclusion. As phagocytic clearance of apoptotic neutrophils by macrophages contributes to the resolution of inflammation, phagocytosis of tilmicosin-treated neutrophils by esterase-positive cultured bovine macrophages was assessed with light microscopy and transmission electron microscopy. Unlike bovine neutrophils treated with penicillin, ceftiofur, oxytetracycline, or dexamethasone, neutrophils exposed to tilmicosin became apoptotic, regardless of the presence or absence of P. haemolytica. Tilmicosin-treated apoptotic neutrophils were phagocytosed at a significantly greater rate by bovine macrophages than were control neutrophils. In conclusion, tilmicosin-induced neutrophil apoptosis occurs regardless of the presence or absence of live P. haemolytica, exhibits at least some degree of drug specificity, and promotes phagocytic clearance of the dying inflammatory cells.
Asunto(s)
Antibacterianos/farmacología , Apoptosis , Macrólidos , Macrófagos/efectos de los fármacos , Mannheimia haemolytica/fisiología , Neutrófilos/efectos de los fármacos , Tilosina/farmacología , Animales , Transporte Biológico , Bovinos , Diferenciación Celular/efectos de los fármacos , Técnicas In Vitro , Leucocitos Mononucleares/citología , Leucocitos Mononucleares/metabolismo , Macrófagos/citología , Macrófagos/fisiología , Neutrófilos/citología , Neutrófilos/microbiología , Fagocitosis/efectos de los fármacos , Fosfatidilserinas/metabolismo , Tilosina/análogos & derivadosRESUMEN
Pathogen virulence factors and the host inflammatory response cause tissue injury associated with respiratory tract infections. The azalide azithromycin has demonstrated efficacy in the treatment of these infections. It has been demonstrated previously that induction of polymorphonuclear leucocyte (PMN) apoptosis is associated with minimization of tissue damage and inflammation in the lung. We hypothesized that, in addition to its antibacterial effects, azithromycin may promote apoptosis. The aim of the study was to determine the effects of azithromycin on PMN apoptosis, oxidative function and interleukin-8 (IL-8) production in the presence or absence of Streptococcus pneumoniae, in comparison with penicillin, erythromycin, dexamethasone or phosphate-buffered saline. Human circulating PMNs were assessed for apoptosis (by annexin V labelling and ELISA), oxidative function (by nitroblue tetrazolium reduction) and IL-8 production (by ELISA). Azithromycin significantly induced PMN apoptosis in the absence of S. pneumoniae after 1 h (10.27% +/- 1.48%, compared with 2.19% +/- 0.42% in controls) to levels similar to those after 3 h induction with tumour necrosis factor-alpha (8. 73% +/- 1.86%). This effect was abolished in the presence of S. pneumoniae. Apoptosis in PMNs exposed to the other drugs was not significantly different from that in controls. Azithromycin did not affect PMN oxidative metabolism or IL-8 production. In summary, azithromycin-induced PMN apoptosis may be detected in the absence of any effect on PMN function, and the pro-apoptotic properties of azithromycin are inhibited in the presence of S. pneumoniae.
Asunto(s)
Apoptosis/efectos de los fármacos , Azitromicina/farmacología , Interleucina-8/biosíntesis , Neutrófilos/efectos de los fármacos , Streptococcus pneumoniae/fisiología , Humanos , Neutrófilos/inmunología , Neutrófilos/metabolismo , Oxidación-ReducciónRESUMEN
Intestinal colonization with the protozoan Giardia causes diffuse brush border microvillous alterations and disaccharidase deficiencies, which in turn are responsible for intestinal malabsorption and maldigestion. The role of T cells and/or cytokines in the pathogenesis of Giardia-induced microvillous injury remains unclear. The aim of this study was to assess the role of T cells and interleukin-6 (IL-6) in the brush border pathophysiology of acute murine giardiasis in vivo. Athymic nude (nu(-)/nu(-)) CD-1 mice and isogenic immunocompetent (nu(+)/nu(+)) CD-1 mice (4 weeks old) received an axenic Giardia muris trophozoite inoculum or vehicle (control) via orogastric gavage. Weight gain and food intake were assessed daily. On day 6, segments of jejunum were assessed for parasite load, brush border ultrastructure, IL-6 content, maltase and sucrase activities, villus-crypt architecture, and intraepithelial lymphocyte (IEL) infiltration. Despite similar parasitic loads on day 6, infected immunocompetent animals, but not infected nude mice, showed a diffuse loss of brush border microvillous surface area, which was correlated with a significant reduction in maltase and sucrase activities and a decrease in jejunal IL-6 concentration. In both athymic control and infected mice, jejunal brush border surface area and disaccharidases were high, but levels of tissue IL-6 were low and comparable to the concentration measured in immunocompetent infected animals. In both immunocompetent and nude mice, infection caused a small but significant increase in the numbers of IELs. These findings suggest that the enterocyte brush border injury and malfunction seen in giardiasis is, at least in part, mediated by thymus-derived T lymphocytes and that suppressed jejunal IL-6 does not necessarily accompany microvillous shortening.