RESUMEN

Nod1 is a member of the NLR/Nod/CATERPILLER family. It acts as a sensor for intracellular bacteria by recognizing specific glycopeptides derived from peptidoglycan. Nod1 activation mediates distinct cellular responses including activation of MAP kinases, IL-8 release, apoptosis and suppression of several estrogen-dependent responses in MCF-7 cells. Here we have extended these studies by identifying key regulatory steps in Nod1-dependent signaling pathways. We provide multiple lines of data showing that Nod1-dependent apoptosis is a caspase 8-mediated event and that apoptosis requires RIP2. In contrast, several lines of evidence show that Nod1-dependent JNK activation and IL-8 production did not require the presence of caspase 8 but required activation of TAK1 as well as RIP2. Thus, we have identified several key control points that lie downstream of Nod1. This work provides the basis for further studies of the biological significance and regulation of the Nod1 pathway.

Asunto(s)

Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Proteína Adaptadora de Señalización NOD1/metabolismo , Proteína Adaptadora de Señalización NOD2/metabolismo , Apoptosis/fisiología , Neoplasias de la Mama/genética , Caspasa 8/metabolismo , Línea Celular Tumoral , Proliferación Celular , Femenino , Humanos , Interleucina-8/biosíntesis , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Quinasas Quinasa Quinasa PAM/metabolismo , Mutagénesis Sitio-Dirigida , Mapeo de Interacción de Proteínas , Proteína Serina-Treonina Quinasa 2 de Interacción con Receptor/metabolismo , Transducción de Señal/fisiología , Células Tumorales Cultivadas

RESUMEN

Macrophages and dendritic cells are in the front line of host defense. When they sense host invasion, they produce cytokines that alert other innate immune cells and also abet the development of an adaptive immune response. Although lipolysaccharide (LPS), peptidoglycan, unmethylated DNA, and other microbial products were long known to be the primary targets of innate immune recognition, there was puzzlement as to how each molecule triggered a response. It is now known that the Toll-like receptors (TLRs) are the principal signaling molecules through which mammals sense infection. Each TLR recognizes a restricted subset of molecules produced by microbes, and in some circumstances, only a single type of molecule is sensed (e.g., only LPS is sensed by TLR4). TLRs direct the activation of immune cells near to and far from the site of infection, mobilizing the comparatively vast immune resources of the host to confine and defeat an invasive organism before it has become widespread. The biochemical details of TLR signaling have been analyzed through forward and reverse genetic methods, and full elucidation of the molecular interactions that transpire within the first minutes following contact between host and pathogen will soon be at hand.

Asunto(s)

Inmunidad Innata , Infecciones/inmunología , Glicoproteínas de Membrana/fisiología , Receptores de Superficie Celular/fisiología , Transducción de Señal , Proteínas Adaptadoras Transductoras de Señales , Animales , Antígenos de Diferenciación/fisiología , Humanos , Inflamación/inmunología , Lipopolisacáridos/toxicidad , Factor 88 de Diferenciación Mieloide , Receptores Inmunológicos/fisiología , Sepsis/inmunología , Receptor Toll-Like 4 , Receptores Toll-Like

RESUMEN

OBJECTIVE: This review describes efforts to define new therapeutic targets by utilizing information derived from studies of the innate immune response to infection. These targets will provide new means to design novel therapeutics for the treatment of human disease caused by dysregulation of the innate immune response. DATA SOURCES AND ANALYSIS: The results were derived from model systems that reflect the cellular changes accompanying activation of the innate immune system. These model systems include the use of a rabbit model of endotoxin shock that requires multiple exposures to lipopolysaccharide to induce severe pathophysiological changes that result in death. The cellular systems range from primary explants of myeloid lineage cells to stably transfected lines bearing key receptors of the innate immune system, the latter of which includes the Toll-like receptors and CD14. Studies in animal models include blocking monoclonal antibodies to rabbit CD14 and rabbit tumor necrosis factor-alpha. Cellular studies use measurements of cell activation as defined by activation of nuclear factor-kappaB, MAP kinase, and gene expression. In some studies, direct measurements of secreted cytokine levels were performed. Standard approaches to data analysis have been used. CONCLUSIONS: Evidence demonstrates the utility of anti-CD14 monoclonal antibody therapy in septic shock and the potential value of targeting intracellular kinases to modulate harmful cellular responses during sepsis. The key element to each of these approaches is to blunt, but not eliminate, the dysregulated inflammatory response that may occur in sepsis.

Asunto(s)

Proteínas de Drosophila , Inmunidad Innata/efectos de los fármacos , Inmunidad Innata/inmunología , Síndrome de Respuesta Inflamatoria Sistémica/inmunología , Síndrome de Respuesta Inflamatoria Sistémica/terapia , Animales , Citocinas/efectos de los fármacos , Citocinas/inmunología , Modelos Animales de Enfermedad , Expresión Génica/efectos de los fármacos , Expresión Génica/inmunología , Humanos , Inmunidad Innata/genética , Receptores de Lipopolisacáridos/efectos de los fármacos , Receptores de Lipopolisacáridos/inmunología , Glicoproteínas de Membrana/efectos de los fármacos , Glicoproteínas de Membrana/inmunología , Quinasas de Proteína Quinasa Activadas por Mitógenos/efectos de los fármacos , Quinasas de Proteína Quinasa Activadas por Mitógenos/inmunología , FN-kappa B/efectos de los fármacos , FN-kappa B/inmunología , Conejos , Receptores de Superficie Celular/efectos de los fármacos , Receptores de Superficie Celular/inmunología , Transducción de Señal/inmunología , Receptores Toll-Like , Factor de Necrosis Tumoral alfa/efectos de los fármacos , Factor de Necrosis Tumoral alfa/inmunología

RESUMEN

Cell signaling by coagulation factor Xa (Xa) contributes to pro-inflammatory responses in vivo. This study characterizes the signaling mechanism of Xa in a HeLa cell line that expresses protease-activated receptor 1 (PAR-1) but not PAR-2, -3, or -4. Xa induced NF-kappaB in HeLa cells efficiently but with delayed kinetics compared to thrombin. This delay caused no difference in gene expression patterns, as determined by high-density microarray analysis. Both proteases prominently induced the angiogenesis-promoting gene Cyr61 and connective tissue growth factor. Inhibition of PAR-1 cleavage abolished MAP kinase phosphorylation and gene induction by Xa, demonstrating that Xa signals through PAR-1 and not through a novel member of the PAR family. Activation of cell surface prothrombin with the snake venom enzyme Ecarin also produced PAR-1-dependent signaling. However, though the response to Ecarin was completely blocked by the thrombin inhibitor hirudin, the response to Xa was not. This suggests that the Xa response is not mediated by locally generated thrombin. The concentration dependence of Xa for PAR-1 activation is consistent with previously characterized Xa-mediated PAR-2 signaling, suggesting that local concentration of Xa on the cell surface, rather than sequence-specific recognition of the PAR scissile bond, determines receptor cleavage. This study demonstrates that PAR-1 cleavage by Xa can elicit the same cellular response as thrombin, but mechanistic differences in receptor recognition may be crucial for specific roles for Xa in signaling during spatial or temporal separation from thrombin generation.

Asunto(s)

Factor Xa/farmacología , Expresión Génica , Péptidos y Proteínas de Señalización Intercelular , Receptores de Trombina/genética , Transducción de Señal , Antitrombinas/farmacología , Línea Celular , Factor de Crecimiento del Tejido Conjuntivo , Proteína 61 Rica en Cisteína , Endotelio Vascular , Activación Enzimática/efectos de los fármacos , Sustancias de Crecimiento/genética , Células HeLa , Hirudinas/farmacología , Humanos , Proteínas Inmediatas-Precoces/genética , Cinética , Proteínas Quinasas Activadas por Mitógenos/metabolismo , FN-kappa B/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , Fosforilación , Receptor PAR-1 , Trombina/metabolismo , Trombina/farmacología , Venas Umbilicales

RESUMEN

Leptospira interrogans are zoonotic pathogens that have been linked to a recent increased incidence of morbidity and mortality in highly populated tropical urban centers. They are unique among invasive spirochetes in that they contain outer membrane lipopolysaccharide (LPS) as well as lipoproteins. Here we show that both these leptospiral outer membrane constituents activate macrophages through CD14 and the Toll-like receptor 2 (TLR2). Conversely, it seems that TLR4, a central component for recognition of Gram-negative LPS, is not involved in cellular responses to L. interrogans. We also show that for intact L. interrogans, it is LPS, not lipoprotein, that constitutes the predominant signaling component for macrophages through a TLR2 pathway. These data provide a basis for understanding the innate immune response caused by leptospirosis and demonstrate a new ligand specificity for TLR2.

Asunto(s)

Proteínas de Drosophila , Leptospira interrogans/inmunología , Leptospira interrogans/patogenicidad , Lipopolisacáridos/toxicidad , Activación de Macrófagos/efectos de los fármacos , Glicoproteínas de Membrana/inmunología , Receptores de Superficie Celular/inmunología , Animales , Células CHO , Línea Celular , Cricetinae , Humanos , Leptospirosis/inmunología , Receptores de Lipopolisacáridos/metabolismo , Lipopolisacáridos/inmunología , Lipoproteínas/inmunología , Activación de Macrófagos/inmunología , Glicoproteínas de Membrana/deficiencia , Glicoproteínas de Membrana/genética , Ratones , Ratones Noqueados , Receptores de Superficie Celular/deficiencia , Receptores de Superficie Celular/genética , Transducción de Señal , Receptor Toll-Like 2 , Receptor Toll-Like 4 , Receptores Toll-Like

RESUMEN

The structural features of some proteins of the innate immune system involved in mediating responses to microbial pathogens are highly conserved throughout evolution. Examples include members of the Drosophila Toll (dToll) and the mammalian Toll-like receptor (TLR) protein families. Activation of Drosophila Toll is believed to occur via an endogenous peptide rather than through direct binding of microbial products to the Toll protein. In mammals there is a growing consensus that lipopolysaccharide (LPS) initiates its biological activities through a heteromeric receptor complex containing CD14, TLR4, and at least one other protein, MD-2. LPS binds directly to CD14 but whether LPS then binds to TLR4 and/or MD-2 is not known. We have used transient transfection to express human TLRs, MD-2, or CD14 alone or in different combinations in HEK 293 cells. Interactions between LPS and these proteins were studied using a chemically modified, radioiodinated LPS containing a covalently linked, UV light-activated cross-linking group ((125)I-ASD-Re595 LPS). Here we show that LPS is cross-linked specifically to TLR4 and MD-2 only when co-expressed with CD14. These data support the contention that LPS is in close proximity to the three known proteins of its membrane receptor complex. Thus, LPS binds directly to each of the members of the tripartite LPS receptor complex.

Asunto(s)

Antígenos de Superficie/metabolismo , Proteínas de Drosophila , Receptores de Lipopolisacáridos/metabolismo , Lipopolisacáridos/metabolismo , Glicoproteínas de Membrana/metabolismo , Receptores de Superficie Celular/metabolismo , Animales , Anticuerpos Monoclonales , Antígenos de Superficie/química , Sitios de Unión , Línea Celular , Reactivos de Enlaces Cruzados , Desoxiglucosa/farmacología , Drosophila , Humanos , Receptores de Lipopolisacáridos/química , Lipopolisacáridos/química , Antígeno 96 de los Linfocitos , Glicoproteínas de Membrana/química , Ratones , Receptores de Superficie Celular/química , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Azida Sódica/farmacología , Fluoruro de Sodio/farmacología , Receptor Toll-Like 4 , Receptores Toll-Like , Transfección , Rayos Ultravioleta

RESUMEN

Herein we describe the isolation of a cDNA encoding a novel human Toll-like receptor (hTLR) that we term hTLR10. Human TLR10 contains 811 amino acid residues. Deduced amino acid sequence analysis reveals that like the other known hTLRs (hTLR1-9) it is characterized by a signal peptide followed by multiple leucine-rich repeats (LRRs), a cysteine-rich domain, a transmembrane sequence and a cytoplasmic domain homologous to that of the human interleukin-1 receptor. Phylogenetic analysis indicates that among all the hTLRs, hTLR10 is most closely related to hTLR1 and hTLR6; the overall amino acid identity is 50% and 49%, respectively. hTLR10 mRNA is most highly expressed in lymphoid tissues such as spleen, lymph node, thymus, and tonsil. Expression analysis of cell lines indicates a predominance in a variety of immune cell types. Thus, hTLR10 is preferentially expressed in tissues and cells involved in immune responses.

Asunto(s)

Receptores de Superficie Celular/genética , Secuencia de Aminoácidos , Secuencia de Bases , Línea Celular , ADN Complementario , Expresión Génica , Humanos , Ganglios Linfáticos/metabolismo , Datos de Secuencia Molecular , Tonsila Palatina/metabolismo , Receptores de Superficie Celular/clasificación , Receptores de Superficie Celular/metabolismo , Homología de Secuencia de Aminoácido , Bazo/metabolismo , Timo/metabolismo , Receptor Toll-Like 10

RESUMEN

During inflammation the balance between cell activation and cell death is determined by the tight regulation of multiple intracellular enzyme cascades. Key regulatory steps often involve protein kinases. We show that the prototypical pro-inflammatory molecule, bacterial lipopolysaccharide, activates multiple protein kinases such as p38, JNK, IKK-beta, and PKB/Akt via transforming growth factor beta-activated kinase-1 (TAK1). We also show that TAK1 plays an important role in similar activation pathways triggered by interleukin-1. Thus TAK1 must be considered as an important component of intracellular pathways in cells involved in host responses to physiological and/or environmental stress signals during inflammation.

Asunto(s)

Inflamación/enzimología , Lipopolisacáridos/farmacología , Quinasas Quinasa Quinasa PAM/fisiología , Proteínas Quinasas/metabolismo , Transducción de Señal/fisiología , Androstadienos/farmacología , Animales , Apoptosis/efectos de los fármacos , Línea Celular/efectos de los fármacos , Cromonas/farmacología , Inhibidores de Cisteína Proteinasa/farmacología , Activación Enzimática/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Flavonoides/farmacología , Humanos , Peróxido de Hidrógeno/farmacología , Soluciones Hipertónicas/farmacología , Quinasa I-kappa B , Imidazoles/farmacología , Factor I del Crecimiento Similar a la Insulina/farmacología , Interleucina-1/farmacología , Proteínas Quinasas JNK Activadas por Mitógenos , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Sistema de Señalización de MAP Quinasas/fisiología , Ratones , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Morfolinas/farmacología , FN-kappa B/metabolismo , Ácido Ocadaico/farmacología , Fosfatidilinositol 3-Quinasas/metabolismo , Prostaglandinas A/farmacología , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas c-akt , Piridinas/farmacología , Proteínas Recombinantes de Fusión/fisiología , Transducción de Señal/efectos de los fármacos , Estrés Fisiológico/enzimología , Transfección , Células Tumorales Cultivadas/efectos de los fármacos , Wortmanina , Proteínas Quinasas p38 Activadas por Mitógenos

RESUMEN

Members of the Toll-like receptor family are essential components of the innate immune system. Herein we report the molecular cloning and characterization of three novel human Toll-like receptors (hTLRs) designated hTLR7, hTLR8, and hTLR9. Human TLR7-9, like the previously described members hTLR1-6 contain an ectodomain with multiple leucine-rich repeats (LRRs) and a cytoplasmic domain homologous to that of the human interleukin-1 (IL-1) receptor. When compared with hTLR1-6, the hTLR7-9 has a higher molecular weight largely as a result of a longer ectodomain. Phylogenetic analysis shows that hTLR7-9 belong to a new sub-family of the hTLRs. Analysis of mRNA expression at the tissue levels shows differential expression patterns; hTLR7 is predominantly expressed in lung, placenta and spleen, hTLR8 is more abundant in lung, peripheral blood leukocytes, and hTLR9 is preferentially expressed in immune cell rich tissues, such as spleen, lymph node, bone marrow and peripheral blood leukocytes. The hTLR7 and hTLR8 genes are located on the sex chromosome X, hTLR9 gene is located on chromosome 3. Expression of constitutively active hTLR7-9 stimulates an NF-kappaB signaling pathway indirectly supporting the contention that these receptors are involved in cellular responses to stimuli, which activate innate immunity.

Asunto(s)

Proteínas de Drosophila , Glicoproteínas de Membrana/genética , Receptores de Superficie Celular/genética , Secuencia de Aminoácidos , Animales , Células Cultivadas , Clonación Molecular , Evolución Molecular , Exones , Regulación de la Expresión Génica , Humanos , Glicoproteínas de Membrana/química , Ratones , Datos de Secuencia Molecular , FN-kappa B/metabolismo , Especificidad de Órganos , Filogenia , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Receptores de Superficie Celular/química , Alineación de Secuencia , Homología de Secuencia de Aminoácido , Receptor Toll-Like 7 , Receptor Toll-Like 8 , Receptor Toll-Like 9 , Receptores Toll-Like

RESUMEN

The innate immune response is the first line of defence against infectious disease. The principal challenge for the host is to detect the pathogen and mount a rapid defensive response. A group of proteins that comprise the Toll or Toll-like family of receptors perform this role in vertebrate and invertebrate organisms. This reflects a remarkable conservation of function and it is therefore not surprising that studies of the mechanism by which they act has revealed new and important insights into host defence.

Asunto(s)

Proteínas de Drosophila , Inmunidad/fisiología , Glicoproteínas de Membrana/fisiología , Receptores de Superficie Celular/fisiología , Animales , Bacterias/inmunología , Humanos , Inmunidad Innata/fisiología , Transducción de Señal , Receptores Toll-Like

RESUMEN

All species require a rapid, systemic reply to pathogens in their environment. This response is known as the innate immune response and is characterized by de novo synthesis of mediators that directly or indirectly through phagocytosis remove and kill the pathogen. Innate immune responses have been preserved throughout evolution and have been studied in detail in organisms from the fruit fly Drosophila melanogaster to humans. In my laboratory, studies performed during the past 25 yr have focused on defining the molecular basis of innate immune responses to microbial pathogens. Specifically, we have used bacterial endotoxin (lipopolysaccharide) as a model stimulus to define how the innate immune system recognizes products of microbial pathogens and initiates responses to remove and/or kill such organisms. Such studies also serve as models to understand more fully the mechanisms underlying a serious human disease known as septic shock. This article discusses septic shock and its relationship to innate immunity.

Asunto(s)

Inmunidad Innata , Animales , Humanos , Lipopolisacáridos/inmunología , Choque Séptico/inmunología

RESUMEN

The p38 family of mitogen-activated protein kinases is believed to mediate a variety of leukocyte responses to pro-inflammatory stimuli. There are four members of the p38 family, and although activation of the different members has been studied in transiently transfected cells much less is known about activation of the endogenous p38s, particularly in myeloid lineage cells. To investigate activation of endogenous p38s, we have made monoclonal antibodies specific for each p38 and have used these antibodies to study p38 activation by pro-inflammatory stimuli in several human monocytic cell lines. Without stimulation endogenous p38alpha kinase activity was readily detectable, whereas that of p38beta, gamma, and delta was barely measurable. In response to inflammatory stimuli, we observed a time- and dose-dependent activation of all four p38s. The kinetics of activation of each of the p38s were similar for each stimulus used, suggesting a common upstream activation pathway. Simultaneous activation of the p38s suggests that all four may be important in inflammation.

Asunto(s)

Proteínas Quinasas Activadas por Mitógenos/metabolismo , Anticuerpos Monoclonales/farmacología , Astrocitoma , Línea Celular , Activación Enzimática , Escherichia coli , Humanos , Inflamación , Interferón gamma/farmacología , Cinética , Lipopolisacáridos/farmacología , Proteína Quinasa 12 Activada por Mitógenos , Proteínas Recombinantes/metabolismo , Acetato de Tetradecanoilforbol/farmacología , Transfección , Células Tumorales Cultivadas , Células U937 , Proteínas Quinasas p38 Activadas por Mitógenos

RESUMEN

Shigella is a diarrheal pathogen that causes disease through invasion of the large intestinal mucosa. The endotoxin of the invading bacterium may play a key role in the disease process by causing inflammation and tissue injury during infection. Earlier studies have shown that various animal species lacking functional CD14 were protected against endotoxin-mediated shock. Rabbits experimentally infected with Shigella were used to test the hypothesis that blockade of endotoxin-induced cell activation with anti-CD14 mAb would diminish inflammation and thus disease severity. Unexpectedly, we observed that the intestinal mucosa of anti-CD14-treated animals exhibited a 50-fold increase in bacterial invasion and more severe tissue injury compared with controls. Despite higher bacterial loads in treated animals, the numbers of polymorphonuclear leukocytes that were recruited to the infection site were similar to those in controls. Furthermore, the phagocytic cells of CD14-blocked animals produced IL-1 and TNF-alpha. Moreover, in vitro blockade of CD14 did not impede bactericidal activity. Thus, anti-CD14 treatment interfered with host defense mechanisms involved with removal/eradication of Shigella.

Asunto(s)

Anticuerpos Bloqueadores/farmacología , Anticuerpos Monoclonales/farmacología , Disentería Bacilar/inmunología , Disentería Bacilar/patología , Receptores de Lipopolisacáridos/inmunología , Shigella flexneri/patogenicidad , Animales , Degranulación de la Célula/inmunología , Movimiento Celular/inmunología , Citocinas/biosíntesis , Disentería Bacilar/microbiología , Mucosa Intestinal/citología , Mucosa Intestinal/inmunología , Mucosa Intestinal/microbiología , Mucosa Intestinal/patología , Lipopolisacáridos/inmunología , Macrófagos/inmunología , Macrófagos/metabolismo , Masculino , Neutrófilos/inmunología , Neutrófilos/metabolismo , Neutrófilos/patología , Conejos , Shigella flexneri/crecimiento & desarrollo , Shigella flexneri/inmunología

RESUMEN

Mammalian Toll-like receptors (TLRs) are expressed on innate immune cells and respond to the membrane components of Gram-positive or Gram-negative bacteria. When activated, they convey signals to transcription factors that orchestrate the inflammatory response. However, the intracellular signaling events following TLR activation are largely unknown. Here we show that TLR2 stimulation by Staphylococcus aureus induces a fast and transient activation of the Rho GTPases Rac1 and Cdc42 in the human monocytic cell line THP-1 and in 293 cells expressing TLR2. Dominant-negative Rac1N17, but not dominant-negative Cdc42N17, block nuclear factor-kappa B (NF-kappa B) transactivation. S. aureus stimulation causes the recruitment of active Rac1 and phosphatidylinositol-3 kinase (PI3K) to the TLR2 cytosolic domain. Tyrosine phosphorylation of TLR2 is required for assembly of a multiprotein complex that is necessary for subsequent NF-kappa B transcriptional activity. A signaling cascade composed of Rac1, PI3K and Akt targets nuclear p65 transactivation independently of I kappa B alpha degradation. Thus Rac1 controls a second, I kappa B-independent, pathway to NF-kappa B activation and is essential in innate immune cell signaling via TLR2.

Asunto(s)

Proteínas de Drosophila , Glicoproteínas de Membrana/metabolismo , FN-kappa B/metabolismo , Proteínas Serina-Treonina Quinasas , Receptores de Superficie Celular/metabolismo , Proteína de Unión al GTP rac1/metabolismo , Línea Celular , Expresión Génica , Humanos , Glicoproteínas de Membrana/genética , Modelos Biológicos , Fosfatidilinositol 3-Quinasas/metabolismo , Fosfotirosina/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas c-akt , Receptores de Superficie Celular/genética , Transducción de Señal , Staphylococcus aureus/inmunología , Receptor Toll-Like 2 , Receptores Toll-Like , Transfección , Proteína de Unión al GTP cdc42/metabolismo

Asunto(s)

Proteínas de Drosophila , Drosophila/inmunología , Glicoproteínas de Membrana/inmunología , Receptores de Superficie Celular/inmunología , Animales , Drosophila/microbiología , Humanos , Inmunidad Innata , Receptores de Lipopolisacáridos/inmunología , Lipopolisacáridos/inmunología , Macrófagos/inmunología , Glicoproteínas de Membrana/genética , Receptores de Superficie Celular/genética , Receptores Toll-Like

Asunto(s)

Antiinflamatorios/farmacología , Citocinas/genética , Inflamación/fisiopatología , Proteínas Serina-Treonina Quinasas/metabolismo , Factor de Necrosis Tumoral alfa/genética , Animales , Regulación de la Expresión Génica/inmunología , Humanos , Inflamación/genética , Péptidos y Proteínas de Señalización Intracelular , Biosíntesis de Proteínas , ARN Mensajero/genética , ARN Mensajero/metabolismo

RESUMEN

Tissue factor (TF) is the major activator of the coagulation protease cascade and contributes to lethality in sepsis. Despite several studies analyzing TF expression in animal models of endotoxemia, there remains debate about the cell types that are induced to express TF in different tissues. In this study, we performed a detailed analysis of the induction of TF mRNA and protein expression in two rabbit models of endotoxemia to better understand the cell types that may contribute to local fibrin deposition and disseminated intravascular coagulation. Northern blot analysis demonstrated that lipopolysaccharide (LPS) increased TF expression in the brain, lung, and kidney. In situ hybridization showed that TF mRNA expression was increased in cells identified morphologically as epithelial cells in the lung and as astrocytes in the brain. In the kidney, in situ hybridization experiments and immunohistochemical analysis showed that TF mRNA and protein expression was increased in renal glomeruli and induced in tubular epithelium. Dual staining for TF and vWF failed to demonstrate TF expression in endothelial cells in LPS-treated animals. These results demonstrate that TF expression is induced in many different cell types in LPS-treated rabbits, which may contribute to local fibrin deposition and tissue injury during endotoxemia.

Asunto(s)

Lipopolisacáridos/toxicidad , Tromboplastina/biosíntesis , Tromboplastina/genética , Animales , Encéfalo/metabolismo , Modelos Animales de Enfermedad , Endotoxemia/etiología , Endotoxemia/genética , Endotoxemia/metabolismo , Expresión Génica/efectos de los fármacos , Inmunohistoquímica , Hibridación in Situ , Riñón/metabolismo , Pulmón/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Conejos , Distribución Tisular

Asunto(s)

Proteínas de Drosophila , Inmunidad/efectos de los fármacos , Lipopolisacáridos/farmacología , Glicoproteínas de Membrana/genética , Receptores de Superficie Celular/genética , Animales , Resistencia a Medicamentos , Ratones , Mutación , Receptores Toll-Like

RESUMEN

Until about 10 years ago the exact mechanisms controlling cellular responses to the endotoxin - or lipopolysaccharide (LPS) - of Gram-negative bacteria were unknown. Now a considerable body of evidence supports a model where LPS or LPS-containing particles (including intact bacteria) form complexes with a serum protein known as LPS-binding protein; the LPS in this complex is subsequently transferred to another protein which binds LPS, CD14. The latter is found on the plasma membrane of most cell types of the myeloid lineage as well as in the serum in its soluble form; LPS binding to these two forms of CD14 results in the activation of cell types of myeloid and nonmyeloid lineages, respectively.

Asunto(s)

Proteínas de Fase Aguda , Endotoxinas/inmunología , Bacterias Gramnegativas/inmunología , Inmunidad Innata/inmunología , Glicoproteínas de Membrana , Animales , Antígenos Bacterianos/inmunología , Proteínas Portadoras/inmunología , Infecciones por Bacterias Gramnegativas/inmunología , Humanos , Receptores de Lipopolisacáridos/inmunología

RESUMEN

Recently there have been some substantial progresses in signal transduction of cell activation by lipopolysaccharide (LPS). After binding with LBP in the plasma, LPS is transported to the cell membrane surface of monocytes, then binds with membrane CD14 to induce cell activation. Mitogen-activated protein kinases (MAPKs) take part in the signal transduction of LPS stimulated cell activation to produce bioactive substances including tumor necrosis factor. p38 MAPK is an important regulator for TNF-alpha expression. It is possible to provide new theory and idea for the treatment of septic shock from the studies of signal transduction of cell activation by LPS.

Asunto(s)

Receptores de Lipopolisacáridos/fisiología , Lipopolisacáridos/farmacología , Proteínas Quinasas Activadas por Mitógenos/fisiología , Transducción de Señal , Animales , Humanos , Factor de Necrosis Tumoral alfa/biosíntesis , Factor de Necrosis Tumoral alfa/genética , Proteínas Quinasas p38 Activadas por Mitógenos