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"Avaliação da expressão dos receptores de interleucina-8, CXCR1 e CXCR2, e da atividade proliferativa em fibroblastos de quelóide e de pele normal" ; Determination of the interleukin-8 receptors CXCR1 and CXCR2, and proliferative activity in keloids and normal skin fibroblasts

Abdo Filho, Décio
Fonte: Biblioteca Digitais de Teses e Dissertações da USP Publicador: Biblioteca Digitais de Teses e Dissertações da USP
Tipo: Tese de Doutorado Formato: application/pdf
Publicado em 05/09/2006 Português
Relevância na Pesquisa
46.27%
O quelóide é um tumor fibroso benigno que ocorre durante a cicatrização da pele em indivíduos geneticamente predispostos. A cicatrização é um processo biológico complexo e depende da interação de diferentes estruturas teciduais e de um grande número de tipos celulares residentes e infiltrativos, que produzem citocinas. A interleucina 8 (IL-8), citocina pró-inflamatória, é super-expressa pelos fibroblastos durante o desenvolvimento do tecido de granulação, acelerando o processo de cicatrização. Como o quelóide resulta de uma reparação tecidual anormal após lesão da pele, o presente estudo teve por objetivo determinar a expressão dos receptores da IL-8, CXCR1 e CXCR2, e a capacidade proliferativa, pelo ciclo celular, dos fibroblastos queloideanos cultivados e extraídos ex vivo, por citometria de fluxo. Fibroblastos de cicatriz queloideana e de pele normal foram obtidos de 21 pacientes da raça negra, com idade variando entre 10 e 40 anos, de lesões com até 2 anos de evolução. Em nosso estudo constatamos expressão reduzida dos receptores para a IL-8, CXCR1(35,7%±11,2) e CXCR2 (27,8%±11,3), em fibroblastos de cicatriz queloideana cultivados, comparando com a pele normal (44,1±16,2 e 46,3±27,1 respectivamente). Entretanto...

Expressão dos receptores das interleucinas de cadeia gama comum em linfócitos T periféricos de pacientes portadores de diabetes mellitus tipo 1 com início recente; Expression of common gamma chain cytokines receptors in periphereal T lymphocytes of recent onset type 1 diabetes patients

Crisostomo, Lindiane Gomes
Fonte: Biblioteca Digitais de Teses e Dissertações da USP Publicador: Biblioteca Digitais de Teses e Dissertações da USP
Tipo: Tese de Doutorado Formato: application/pdf
Publicado em 27/08/2010 Português
Relevância na Pesquisa
36.34%
O Diabetes Mellitus tipo 1 (DM1A) é uma doença autoimune caracterizada pela infiltração pancreática de linfócitos T e B, macrófagos e células dendríticas, levando à perda progressiva da capacidade de secreção de insulina pelas células beta pancreáticas. A homeostase das células T, ou seja, o desenvolvimento e manutenção apropriados dos números e funções das células T são essenciais para a integridade do sistema imune. Classicamente acreditava-se que as células T CD4+ poderiam se subdividir em duas populações efetoras distintas, T helper 1 e T helper 2. Recentemente, foram descritas duas novas vias de ativação de linfócitos T CD4+: a via Th17, que tem papel fundamental na autoimunidade; a via T regulatória, onde células T CD4+CD25+ high são essenciais na tolerância periférica e proteção contra autoimunidade. As Interleucinas (IL) de cadeia gama comum agem em várias etapas desta diferenciação linfocítica. A IL-21 é o membro mais recente desta família de citocinas, que inclui também: IL-2, IL-4, IL-7 , IL-9 e IL-15. A IL-21 atua através da interação com seu receptor, o IL-21R, apresentando ações pleiotrópicas e, como regra, pró-inflamatórias. Em estudos com modelos animais de diabetes autoimune verificou-se que a IL-21 e seu receptor são essenciais para o desenvolvimento da doença...

Impacto da IL-17A na predisposição ao diabetes mellitus tipo 1A; Impact of IL-17A in the predisposition to type 1 autimmune diabetes mellitus

Fores, Jéssica Pereira
Fonte: Biblioteca Digitais de Teses e Dissertações da USP Publicador: Biblioteca Digitais de Teses e Dissertações da USP
Tipo: Dissertação de Mestrado Formato: application/pdf
Publicado em 07/02/2011 Português
Relevância na Pesquisa
36.16%
Diabetes Mellitus tipo 1A (DM1A), doença autoimune clássica, decorrente da quebra de tolerância imune por fatores ambientais em indivíduos geneticamente predispostos, é caracterizada pela infiltração pancreática de linfócitos T e B, macrófagos e células dendríticas. As células T auxiliadoras 17 (Th17) são células potentes, altamente inflamatórias, que produzem a interleucina 17A (IL-17A), citocina mediadora de várias desordens imunológicas como, artrite reumatóide, esclerose múltipla, encefalite experimental autoimune, psoríase e asma, e em animais, o diabetes autoimune. No entanto, seu papel na patogênese do DM1A em humanos não está definido O objetivo de nosso estudo foi avaliar a influência da IL-17A na predisposição ao DM1A através da identificação de variantes alélicas no gene da IL-17A (por sequenciamento automático) e da determinação dos níveis séricos de IL-17A (por ELISA) e da expressão do seu receptor em linfócitos T periféricos (por citometria de fluxo). Foram analisados 103 pacientes com DM1A (idade 15,15 ± 10,38) e 102 controles normais (idade 18,29 ± 10,83). O estudo da expressão do receptor da IL-17A em linfócitos T periféricos bem como o da proteína sérica foram conduzidos em 24 pacientes com DM1A recente (duração inferior a 6 meses) e 23 controles normais. Resultados: Nos 3 exons da IL-17 A analisados...

IL-4 and IL-13 inhibit IL-1β and TNF-α induced kinin B 1 and B2 receptors through a STAT6-dependent mechanism

Souza, P. P. C.; Brechter, A. B.; Reis, R. I.; Costa, C. A. S.; Lundberg, P.; Lerner, U. H.
Fonte: Universidade Estadual Paulista Publicador: Universidade Estadual Paulista
Tipo: Artigo de Revista Científica Formato: 400-412
Português
Relevância na Pesquisa
36.33%
Background and Purpose Bone resorption induced by interleukin-1β (IL-1β) and tumour necrosis factor (TNF-α) is synergistically potentiated by kinins, partially due to enhanced kinin receptor expression. Inflammation-induced bone resorption can be impaired by IL-4 and IL-13. The aim was to investigate if expression of B1 and B2 kinin receptors can be affected by IL-4 and IL-13. Experimental Approach We examined effects in a human osteoblastic cell line (MG-63), primary human gingival fibroblasts and mouse bones by IL-4 and IL-13 on mRNA and protein expression of the B1 and B2 kinin receptors. We also examined the role of STAT6 by RNA interference and using Stat6-/- mice. Key Results IL-4 and IL-13 decreased the mRNA expression of B1 and B2 kinin receptors induced by either IL-1β or TNF-α in MG-63 cells, intact mouse calvarial bones or primary human gingival fibroblasts. The burst of intracellular calcium induced by either bradykinin (B2 agonist) or des-Arg10-Lys-bradykinin (B1 agonist) in gingival fibroblasts pretreated with IL-1β was impaired by IL-4. Similarly, the increased binding of B1 and B2 ligands induced by IL-1β was decreased by IL-4. In calvarial bones from Stat6-deficient mice, and in fibroblasts in which STAT6 was knocked down by siRNA...

Modulation of TNF-α-mediated acute lung inflammation : a role for IL-1β and soluble TNF receptors

Saperstein, Sara C. ; Finkelstein, Jacob N.
Fonte: Universidade de Rochester Publicador: Universidade de Rochester
Tipo: Tese de Doutorado
Português
Relevância na Pesquisa
36.21%
Thesis (Ph. D.)--University of Rochester. School of Medicine and Dentistry. Dept. of Environmental Medicine, 2009.; Tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) are key mediators of numerous lung inflammatory diseases. Induced by similar stimuli, in many cell types IL-1β modifies expression and solubilization of the two TNF receptors, TNFR1 and TNFR2. These proteins are cleaved from the cell surface primarily by the metalloprotease TNF-α converting enzyme (TACE) generating soluble forms still capable of binding TNF-α. Investigators postulate that the shed receptors either act as natural antagonists or as a TNF-α reservoir extending the half-life of the cognate ligand. Based on these observations, IL-1β effects on lung TNF-α-mediated inflammatory responses, as well as on TNF receptor expression and shedding, both in vitro and in vivo were investigated. Finally, the roles of the two soluble TNF receptors (sTNFR) on altering ligand-induced inflammation and cytotoxicity were analyzed. Studies utilizing an in vitro culture system, murine epithelial type II-like cells (MLE-15), and transgenic IL-1β null mice showed that the interleukin enhances acute, TNF-α-mediated pulmonary inflammatory processes. In vitro, IL-1β induced MLE-15 surface TNF receptor expression which correlated with increased mRNA levels and protein secretion of the two neutrophil chemoattractants macrophage inflammatory protein (MIP-2) and KC following TNF-α exposure. Further studies investigating the regulation of the two receptors demonstrated a counter-regulatory role for each; TNFR1 expression was necessary for interleukin-mediated TNFR2 shedding and surface expression and vice a versa. In vivo studies corroborated these findings. IL-1β KO mice instilled with TNF-α had decreased neutrophil influx measured in the lavage fluid of the lung. This correlated with a decrease in lavage sTNFR1 and MIP-2...

Reduced expression of the interleukin-2-receptor gamma chain on cord blood lymphocytes: relationship to functional immaturity of the neonatal immune response

Zola, H.; Fusco, M.; Weedon, H.; Macardle, P.; Ridings, J.; Roberton, D.
Fonte: BLACKWELL SCIENCE LTD Publicador: BLACKWELL SCIENCE LTD
Tipo: Artigo de Revista Científica
Publicado em //1996 Português
Relevância na Pesquisa
45.98%
Mutation of the interleukin-2 (IL-2) receptor gamma chain, which also serves as a component of the receptor complexes for IL-4, 7, 9 and 15, results in severe immune deficiency. We hypothesized that the immunological immaturity of healthy neonates might be associated with low levels of expression of this receptor molecule. Using monoclonal antibody and a highly sensitive immunofluorescence method, we showed that IL-2 receptor gamma chain is expressed at significantly lower levels on cord blood cells compared with adult cells. IL-2-dependent T-cell activation in vitro was reduced in cord blood cells compared with adult cells, but B-cell responses to IL-4 were not obviously impaired. The lower level of expression of the gamma chain and some other cytokine receptor chains may contribute to the immunological immaturity of the newborn, by selectively depressing particular immunological mechanisms.

Neutrophils activated by granulocyte-macrophage colony-stimulating factor express receptors for interleukin-3 which mediate class II expression

Smith, W.; Guida, L.; Qiuy, S.; Korpelainen, E.; van den Hueven, C.; Gillis, D.; Hawrylowicz, C.; Vadas, M.; Lopez, A.
Fonte: American Society of Hematology Publicador: American Society of Hematology
Tipo: Artigo de Revista Científica
Publicado em //1995 Português
Relevância na Pesquisa
46.21%
Freshly isolated peripheral blood neutrophils, unlike monocytes and eosinophils, do not bind interleukin-3 (IL-3) or respond to IL-3). We show that neutrophils cultured for 24 hours in granulocyte-macrophage colony-stimulating factor (GM-CSF) express mRNA for the IL-3 receptor (R) alpha subunit, as shown by RNase protection assays, and IL-3R alpha chain protein, as shown by cytometric analysis using two different specific monoclonal antibodies. This effect was selective for GM-CSF, because granulocyte colony-stimulating factor, tumor necrosis factor- alpha, interferon-gamma, and IL-1 failed to induce the IL-3 receptor. Saturation binding curves with 125I-IL-3 and Scatchard transformation showed the presence of about 100 high-affinity and 4,000 low-affinity receptors. Because neutrophils have been shown to express human leukocyte antigen (HLA)-DR in response to GM-CSF, we examined the possibility that IL-3 could augment HLA-DR expression on GM-CSF-treated cells. We found that neutrophils incubated with 30 ng/mL IL-3 as well as 0.1 ng/mL GM-CSF expressed a mean of 2.1-fold higher levels of HLA- DR than with GM-CSF alone (P < .005), confirming the signaling competence of the newly expressed IL-3R. This increase was seen even at maximal concentrations of GM-CSF and IL-3 can have an additive effect on mature human cells. The augmentation of HLA-DR by IL-3 was specific because it could be inhibited by a blocking anti-IL-3R antibody. Expression of class II molecules by neutrophils under these conditions may have significance for antigen presentation. These results provide further evidence for the role of GM-CSF as an amplification factor in inflammation by inducing neutrophil responsiveness to IL-3 produced by T cells or mast cells.; WB Smith...

Identification of G-protein binding sites of the human interleukin-8 receptors by functional mapping of the intracellular loops

Damaj, B.; McColl, S.; Neote, K.; Songqing, N.; Ogborn, K.; Hebert, C.; Naccache, P.
Fonte: FASEB Publicador: FASEB
Tipo: Artigo de Revista Científica
Publicado em //1996 Português
Relevância na Pesquisa
46.3%
Interleukin 8 (IL-8) is considered to be a major mediator of the inflammatory response. Recent evidence indicates that a direct physical association occurs between IL-8 receptors and the alpha subunit of guanine nucleotide regulatory protein (Gi(alpha)2) upon stimulation of human neutrophils by IL-8. In the present study, we identified by site-directed mutagenesis key residues within the three intracellular loops of the IL-8RA receptor involved in the interaction with Gi(alpha)2. We first systematically mutated, in groups of two to four, all the residues in the three intracellular loops of the IL-8 type A receptor to alanine and analyzed the mutant receptors transiently expressed in 293 cells. Four residues in the second intracellular loop (Y136, L137, I139, V140) and one residue in the third intracellular loop (M241) were shown to be crucial for mediating calcium signaling in response to IL-8. Other residues in the second and third intracellular loops were also found to affect IL-8RA-mediated signaling, but to a lesser extent. These effects were not due to lower expression or low IL-8 binding affinities to the mutated receptors. Mutagenesis of the residues in the first intracellular loop had only weak effects on the mobilization of calcium induced by IL-8. We then used a coimmunoprecipitation protocol with anti-Gi(alpha)2 antibodies to determine the involvement of the two regions defined above in Gi(alpha)2 coupling to IL-8 type A receptors. Whereas the anti-Gi(alpha)2 antibodies coimmunoprecipitated IL-8 receptors in the wild-type cells...

Diverging signal transduction pathways activated by interleukin 8 and related chemokines in human neutrophils II: IL-8 and Groa differentially stimulate calium influx through IL-8 receptors A & B

Damaj, B.; McColl, S.; Neote, K.; Hebert, C.; Naccache, P.
Fonte: AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC Publicador: AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
Tipo: Artigo de Revista Científica
Publicado em //1996 Português
Relevância na Pesquisa
46.24%
Interleukin 8 (IL-8) and Gro-α are members of the CXC branch of a family of cytokines recently designated the “chemokine” superfamily. Recent evidence indicates that, contrary to previously held beliefs, IL-8 and Gro-α may not be perceived equivalently by neutrophils. In this study, we have evaluated the effects of IL-8 and Gro-α on the rate of calcium influx in human neutrophils and in 293 cells transfected with type A or type B IL-8 receptors. Of these two chemokines, only Gro-α induced an influx of calcium in neutrophils as judged by the sensitivity of the mobilization of calcium to the extracellular calcium chelator EGTA and to the nonselective divalent cation channel inhibitor SK&F 96365, as well as by manganese quenching experiments. IL-8 was similarly without effect on the rate of Mn²⁺ influx in 293 cells transfected with IL-8 receptor A (IL-8RA) or IL-8RB. On the other hand, Gro-α induced an SK&F 96365-sensitive increase of the rate of Mn⁺² influx in IL-8RB-, but not in IL-8RA-transfected 293 cells. These results indicate not only that neutrophils respond differently to IL-8 than they do to Gro-α but, furthermore, that the consequences of the binding of IL-8 and Gro-α to IL-8RB are distinct.; Bassam B. Damaj...

Physical association of Gi₂α with Interleukin-8 receptors; Physical association of Gi(2)alpha with Interleukin-8 receptors

Damaj, B.; McColl, S.; Mahana, W.; Crouch, M.; Naccache, P.
Fonte: AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC Publicador: AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
Tipo: Artigo de Revista Científica
Publicado em //1996 Português
Relevância na Pesquisa
46.32%
Interleukin-8 (IL-8), one of the major mediators of the inflammatory response, belongs to a family of chemokines that includes NAP-2 (Graphiceutrophil-Graphicctivating Graphiceptide-2) and Gro-α and whose biological activities are directed to a great extent toward neutrophils. Two distinct receptors have been described with overlapping, but not identical, binding affinities for IL-8, NAP-2, and Gro-α. This study was designed to examine the intracellular pathways activated upon the occupation of each of the IL-8 receptors (IL-8R). The formation of a physical coupling between IL-8 receptors and the α-subunit of heterotrimeric G proteins was tested in neutrophils by examining the presence of the former in anti-Gα immune precipitates. The addition of IL-8 to a suspension of human neutrophils led to a time-dependent detection of IL-8 in anti-Gi2α (raised against amino acids 159-168 (LERIAQSDYI) of Gi2α) and anti-Gtα (raised against the COOH-terminal 10 amino acids (KENLKDCGLF) of Gtα), but not anti-Gq, immunoprecipitates. Similar results were obtained in human 293 cells stably transfected with IL-8RA or IL-8RB. The peptide derived from the COOH-terminal sequence of Gt inhibited the co-immunoprecipitation of IL-8R and Gi observed in response to the anti-Gtα and anti-Gi2α antibodies. On the other hand...

Localisation of mRNA for interleukin-1 receptor and interleukin-1 receptor antagonist in the rat ovary

Wang, L.; Brannstrom, M.; Cui, K.H.; Simula, A.; Hart, R.; Maddocks, S.; Norman, R.
Fonte: Society for Endocrinology Publicador: Society for Endocrinology
Tipo: Artigo de Revista Científica
Publicado em //1997 Português
Relevância na Pesquisa
45.98%
Interleukin-1 (IL-1) is a multifunctional cytokine with profound effects on ovarian function. The effects of IL-1 on ovarian steroidogenesis have been demonstrated in several species. IL-1 mRNA levels are increased in the thecal layer of the ovulating follicle and IL-1β has been shown to induce ovulations in vitro. In this study we have investigated the presence and distribution of the mRNAs for type I IL-1 receptor (IL-1RtI) and for the naturally occurring IL-1 receptor antagonist (IL-1ra) in ovaries of adult cycling rats, to elucidate the target cells for IL-1 action. We have demonstrated the presence of mRNA for both substances by in situ hybridisation and reverse transcription PCR. mRNA for IL-1RtI was not found in primordial follicles but was abundant in the granulosa and thecal layer in developing follicles with stronger signals in the granulosa layer. In the preovulatory and ovulatory follicles, there was a further increase in the signal for IL-1RtI mRNA in the thecal layer compared with the granulosa layer. Corpora lutea were weakly positive at all stages and atretic follicles were largely negative. No mRNA was detected in oocytes of any stage. mRNA for IL-1ra showed a similar distribution to that of IL-1RtI. The changes in distribution suggest an action of IL-1 on rat granulosa cells during follicular development and on thecal cells during ovulation.; L-J Wang...

MIF is a noncognate ligand of CXC chemokine receptors in inflammatory and atherogenic cell recruitment

Bernhagen, J.; Krohn, R.; Lue, H.; Gregory, J.; Zernecke, A.; Koenen, R.; Dewor, M.; Georgiev, I.; Schober, A.; Leng, L.; Kooistra, T.; Fingerle-Rowson, G.; Ghezzi, P.; Kleemann, R.; McColl, S.; Bucala, R.; Hickey, M.; Weber, C.
Fonte: Nature Publishing Group Publicador: Nature Publishing Group
Tipo: Artigo de Revista Científica
Publicado em //2007 Português
Relevância na Pesquisa
45.96%
The cytokine macrophage migration inhibitory factor (MIF) plays a critical role in inflammatory diseases and atherogenesis. We identify the chemokine receptors CXCR2 and CXCR4 as functional receptors for MIF. MIF triggered G(alphai)- and integrin-dependent arrest and chemotaxis of monocytes and T cells, rapid integrin activation and calcium influx through CXCR2 or CXCR4. MIF competed with cognate ligands for CXCR4 and CXCR2 binding, and directly bound to CXCR2. CXCR2 and CD74 formed a receptor complex, and monocyte arrest elicited by MIF in inflamed or atherosclerotic arteries involved both CXCR2 and CD74. In vivo, Mif deficiency impaired monocyte adhesion to the arterial wall in atherosclerosis-prone mice, and MIF-induced leukocyte recruitment required Il8rb (which encodes Cxcr2). Blockade of Mif but not of canonical ligands of Cxcr2 or Cxcr4 in mice with advanced atherosclerosis led to plaque regression and reduced monocyte and T-cell content in plaques. By activating both CXCR2 and CXCR4, MIF displays chemokine-like functions and acts as a major regulator of inflammatory cell recruitment and atherogenesis. Targeting MIF in individuals with manifest atherosclerosis can potentially be used to treat this condition.; Jürgen Bernhagen...

The role of βc subunit phosphorylation in the functioning of the GM-CSF/IL-3/IL-5 receptors.

Winnall, Wendy
Fonte: Universidade de Adelaide Publicador: Universidade de Adelaide
Tipo: Tese de Doutorado
Publicado em //2008 Português
Relevância na Pesquisa
46.13%
The cytokines GM-CSF, IL-3 and IL-5 are central regulators of haemopoietic cell functions and are pivotal in the regulation of haemopoiesis and inflammatory responses of myeloid cells. In particular, these cytokines have been shown to perform essential functions in host defence against foreign pathogens through their ability to regulate innate immune responses in myeloid cells. As key regulators of such important processes, these cytokines play an important role in human inflammatory pathologies such as rheumatoid arthritis, asthma, multiple sclerosis and psoriasis as well as a number of leukemias such as JML and CMML. GM-CSF, IL-3 and IL-5 signal through receptors containing α subunits specific to each cytokine and a common β subunit (βc). Cytokine stimulation leads to tyrosine phosphorylation of the βc and promotes specific responses such as proliferation, survival and activation of haemopoietic cells. Mouse knockout studies identified a key function of these cytokines in the activation of effector functions of myeloid cells, including production of reactive oxygen species (ROS) and phagocytosis. These earlier studies provide a link between cytokine signalling and inflammation, but the molecular mechanisms by which βc activation regulates effector cell functions...

Einflussfaktoren auf Interleukin-8-Konzentrationen in Plasma und lysiertem Vollblut beim Neugeborenen; Influencing factors on plasma and DLWB interleukin-8-concentrations in newborns

Siedler, Diana
Fonte: Universidade de Tubinga Publicador: Universidade de Tubinga
Tipo: Dissertação
Português
Relevância na Pesquisa
36.18%
Einflussfaktoren auf Interleukin-8-Konzentrationen in Plasma und lysiertem Vollblut beim Neugeborenen D. Siedler, M. Deperschmidt#, C. Henkel#, M. Eichner*, Ch. F. Poets, Th. Orlikowsky Univ.-Kinderklinik Tübingen, Abt. Neonatologie, #Univ.-Frauenklinik Tübingen, *Institut für medizinische Biometrie Tübingen Hintergrund: Trotz wirksamer Antibiotikatherapie ist die frühe neonatale bakterielle Infektion (EOBI) mit einer hohen Morbidität und Mortalität verbunden. Deshalb ist die frühzeitige Diagnose und Behandlung wichtig, um Spätfolgen zu verhindern und eine normale Entwicklung des Kindes zu gewährleisten. IL-8 ist ein Akutphase-Zytokin, welches früh bei bakteriellen Infektionen ausgeschüttet wird. Es wird neben den spezifischen IL-8-Rezeptoren auf Granulozyten auch unspezifisch an Erythrozyten gebunden. Im Plasma gemessen ist seine Halbwertszeit kurz und beträgt ein bis drei Stunden. In lysiertem Vollblut erwies es sich als Parameter, welcher der Plasmabestimmung im Hinblick auf Sensivität, Spezifität sowie dem prädiktiven Wert bei EOBI überlegen war. Ziel: Untersuchung von Einflussgrößen prä-, peri- und postnataler Faktoren, hierbei insbesondere des Erythrozytenstatus, auf IL-8-Konzentrationen in Plasma und lysiertem Vollblut...

Constitutive mutants of the GM-CSF receptor reveal multiple pathways leading to myeloid cell survival, proliferation, and granulocyte-macrophage differentiation

Brown, A.; Peters, M.; D'Andrea, R.; Gonda, T.
Fonte: Amer Soc Hematology Publicador: Amer Soc Hematology
Tipo: Artigo de Revista Científica
Publicado em //2004 Português
Relevância na Pesquisa
36.25%
Activation of the granulocyte-macrophage colony-stimulating factor (GM-CSF) family of receptors promotes the survival, proliferation, and differentiation of cells of the myeloid compartment. Several signaling pathways are activated downstream of the receptor, however it is not clear how these induce specific biologic outcomes. We have previously identified 2 classes of constitutively active mutants of the shared signaling subunit, human (h) betac, of the human GM-CSF/interleukin-3 (IL-3)/IL-5 receptors that exhibit different modes of signaling. In a factor-dependent bipotential myeloid cell line, FDB1, an activated mutant containing a substitution in the transmembrane domain (V449E) induces factor-independent proliferation and survival, while mutants in the extracellular domain induce factor-independent granulocyte-macrophage differentiation. Here we have used further mutational analysis to demonstrate that there are nonredundant functions for several regions of the cytoplasmic domain with regard to mediating proliferation, viability, and differentiation, which have not been revealed by previous studies with the wild-type GM-CSF receptor. This unique lack of redundancy has revealed an association of a conserved membrane-proximal region with viability signaling and a critical but distinct role for tyrosine 577 in the activities of each class of mutant.; Anna L. Brown...

Simultaneous antagonism of interleukin-5, granulocyte-macrophage colony-stimulting factor, and interleukin-3 stimulation of human eosinophils by targetting the common cytokine binding site of their receptors

Sun, Q.; Jones, K.; McClure, B.; Cambareri, B.; Zacharakis, B.; Iversen, P.; Stomski, F.; Woodcock, J.; Bagley, C.; D'Andrea, R.; Lopez, A.
Fonte: AMER SOC HEMATOLOGY Publicador: AMER SOC HEMATOLOGY
Tipo: Artigo de Revista Científica
Publicado em //1999 Português
Relevância na Pesquisa
46.34%
Human interleukin-5 (IL-5), granulocyte-macrophage colony-stimulating factor (GM-CSF), and IL-3 are eosinophilopoietic cytokines implicated in allergy in general and in the inflammation of the airways specifically as seen in asthma. All 3 cytokines function through cell surface receptors that comprise a ligand-specific alpha chain and a shared subunit (beta(c)). Although binding of IL-5, GM-CSF, and IL-3 to their respective receptor alpha chains is the first step in receptor activation, it is the recruitment of beta(c) that allows high-affinity binding and signal transduction to proceed. Thus, beta(c) is a valid yet untested target for antiasthma drugs with the added advantage of potentially allowing antagonism of all 3 eosinophil-acting cytokines with a single compound. We show here the first development of such an agent in the form of a monoclonal antibody (MoAb), BION-1, raised against the isolated membrane proximal domain of beta(c). BION-1 blocked eosinophil production, survival, and activation stimulated by IL-5 as well as by GM-CSF and IL-3. Studies of the mechanism of this antagonism showed that BION-1 prevented the high-affinity binding of (125)I-IL-5, (125)I-GM-CSF, and (125)I-IL-3 to purified human eosinophils and that it bound to the major cytokine binding site of beta(c). Interestingly...

Roles of the N and C terminal domains of the interleukin-3 receptor alpha chain in receptor function.

Barry, S.; Korpelainen, E.; Sun, Q.; Stomski, F.; Moretti, P.; Wakao, H.; D'Andrea, R.; Vadas, M.; Lopez, A.; Goodall, G.
Fonte: W B SAUNDERS CO Publicador: W B SAUNDERS CO
Tipo: Artigo de Revista Científica
Publicado em //1997 Português
Relevância na Pesquisa
45.94%
The interleukin-3 (IL-3), granulocyte-macrophage colony-stimulating factor, and IL-5 receptor alpha chains are each composed of three extracellular domains, a transmembrane domain and a short intracellular region. Domains 2 and 3 constitute the cytokine receptor module (CRM), typical of the cytokine receptor superfamily; however, the function of the N-terminal domain is not known. We have investigated the functions of the N-terminal and C-terminal domains of the IL-3 receptor (IL-3R) alpha chain. We find that cells transfected with the receptor beta chain (h beta c) and a truncated IL-3R alpha that is devoid of the intracellular region fail to proliferate or to activate STAT5 in response to human IL-3, despite binding the IL-3 with affinity indistinguishable from that of full-length receptor. In addition, IL-3-induced phosphorylation of h beta c was not detected. Thus, the IL-3R alpha intracellular region does not contribute detectably to stabilization of the receptor/ligand complex, but is essential for signal propagation. In contrast, a truncated IL-3R alpha with the N-terminal domain deleted interacts functionally with the beta chain; mouse cells transfected with these receptor chains proliferate in response to human IL-3 and STAT5 transcription factor is activated. High- and low-affinity binding sites are retained...

Peptide insertions in domain 4 of hbc, the shared signalling receptor subunit for GM-CSF, IL3 and IL5, induce ligand-independent activation

Jones, K.; Bagley, C.; Butcher, C.; Barry, S.; Vadas, M.; D'Andrea, R.
Fonte: Academic Press Ltd Elsevier Science Ltd Publicador: Academic Press Ltd Elsevier Science Ltd
Tipo: Artigo de Revista Científica
Publicado em //2001 Português
Relevância na Pesquisa
36.27%
A mutant form of the common beta-subunit of the GM-CSF, interleukin-3 (IL3) and IL5 receptors is activated by a 37 residue duplicated segment which includes the WSXWS motif and an adjacent, highly conserved, aliphatic/basic element. Haemopoietic expression of this mutant, hbeta(c)FIDelta, in mice leads to myeloproliferative disease. To examine the mechanism of activation of this mutant we targetted the two conserved motifs in each repeat for mutagenesis. Here we show that this mutant exhibits constitutive activity in BaF-B03 cells in the presence of mouse or human GM-CSF receptor alpha-subunit (GMRalpha) and this activity is disrupted by mutations of the conserved motifs in the first repeat. In the presence of these mutations the receptor reverts to an alternative conformation which retains responsiveness to human IL3 in a CTLL cell line co-expressing the human IL3 receptor alpha-subunit (hIL3Ralpha). Remarkably, the activated conformation is maintained in the presence of substitutions, deletions or replacement of the second repeat. This suggests that activation occurs due to insertion of extra sequence after the WSXWS motif and is not dependent on the length or specific sequence of the insertion. Thus hbeta(c) displays an ability to fold into functional receptor conformations given insertion of up to 37 residues in the membrane-proximal region. Constitutive activation most likely results from a specific conformational change which alters a dormant...

The solution structure of the cytokine-binding domain of the common -chain of the receptors for granulocyte-macrophage colony-stimulating factor, interleukin-3 and interleukin-5

Mulhern, T.; Lopez, A.; D'Andrea, R.; Gaunt, C.; Vandeleur, L.; Vadas, M.; Booker, G.; Bagley, C.
Fonte: Academic Press Ltd Publicador: Academic Press Ltd
Tipo: Artigo de Revista Científica
Publicado em //2000 Português
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46.34%
The haemopoietic cytokines, granulocyte-macrophage colony-stimulating factor, interleukin-3 and interleukin-5 bind to cell-surface receptors comprising ligand-specific alpha-chains and a shared beta-chain. The beta-chain is the critical signalling subunit of the receptor and its fourth domain not only plays a critical role in interactions with ligands, hence in receptor activation, but also contains residues whose mutation can lead to ligand-independent activation of the receptor. We have determined the NMR solution structure of the isolated human fourth domain of the beta-chain. The protein has a fibronectin type III fold with a well-defined hydrophobic core and is stabilised by an extensive network of pi-cation interactions involving Trp and Arg side-chains, including two Trp residues outside the highly conserved Trp-Ser-Xaa-Trp-Ser motif (where Xaa is any amino acid) that is found in many cytokine receptors. Most of the residues implicated in factor-independent mutants localise to the rigid core of the domain or the pi-cation stack. The loops between the B and C, and the F and G strands, that contain residues important for interactions with cytokines, lie adjacent at the membrane-distal end of the domain, consistent with their being involved cooperatively in binding cytokines. The elucidation of the structure of the cytokine-binding domain of the beta-chain provides insight into the cytokine-dependent and factor-independent activation of the receptor.

Reduced Interleukin-4 Receptor α Expression on CD8+ T Cells Correlates with Higher Quality Anti-Viral Immunity

Wijesundara, Danushka K.; Tscharke, David C.; Jackson, Ronald J.; Ranasinghe, Charani
Fonte: Public Library of Science Publicador: Public Library of Science
Tipo: Artigo de Revista Científica
Português
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With the hope of understanding how interleukin (IL)-4 and IL-13 modulated quality of anti-viral CD8(+) T cells, we evaluated the expression of receptors for these cytokines following a range of viral infections (e.g. pox viruses and influenza virus). Results clearly indicated that unlike other IL-4/IL-13 receptor subunits, IL-4 receptor α (IL-4Rα) was significantly down-regulated on anti-viral CD8(+) T cells in a cognate antigen dependent manner. The infection of gene knockout mice and wild-type (WT) mice with vaccinia virus (VV) or VV expressing IL-4 confirmed that IL-4, IL-13 and signal transducer and activator of transcription 6 (STAT6) were required to increase IL-4Rα expression on CD8(+) T cells, but not interferon (IFN)-γ. STAT6 dependent elevation of IL-4Rα expression on CD8(+) T cells was a feature of poor quality anti-viral CD8(+) T cell immunity as measured by the production of IFN-γ and tumor necrosis factor α (TNF-α) in response to VV antigen stimulation in vitro. We propose that down-regulation of IL-4Rα, but not the other IL-4/IL-13 receptor subunits, is a mechanism by which CD8(+) T cells reduce responsiveness to IL-4 and IL-13. This can improve the quality of anti-viral CD8(+) T cell immunity. Our findings have important implications in understanding anti-viral CD8(+) T cell immunity and designing effective vaccines against chronic viral infections.; This work was supported by the Australian National Health and Medical Research Council project grant award 525431 (CR) and development grant award APP1000703 (CR) and the Australian Centre for Hepatitis and HIV Virology EOI grant 2010 (CR). The funders had no role in study design...