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1.
Ann Biol Clin (Paris) ; 72(2): 252-4, 2014.
Artigo em Francês | MEDLINE | ID: mdl-24736149

RESUMO

On the way for the accreditation which should lead us until 2020, we wish to share some reflections stemming from the daily practice concerning the compulsory quality approach for everyone. Several themes as training and skills evaluation, external quality controls, risk management and action plans have a great relevance and are a matter of public concern. Their consideration contributes not only to the reassurance of processes but also to knowledge improvement. In the following paragraphs we will present an overview of these themes which are all key elements for project management.


Assuntos
Acreditação/normas , Competência Clínica/normas , Avaliação de Desempenho Profissional , Laboratórios Hospitalares/normas , Melhoria de Qualidade , Humanos , Conhecimento , Controle de Qualidade
2.
Int J Pharm ; 403(1-2): 29-36, 2011 Jan 17.
Artigo em Inglês | MEDLINE | ID: mdl-20951782

RESUMO

Phase and habit selection is a very important step in the early stages of pharmaceutical development of new APIs. In this paper, we show how observation, diffraction and thermal analysis are complementary methods of solid habit and phase characterization. At the end of phase screening of an API several habits and phases can be discriminated by microscopy, XRPD or Raman spectroscopy. Using thermal methods here allows us to separate the 12 phases discriminated by XRPD into: anhydrous, monohydrate, organic monosolvate and heterosolvate phases.


Assuntos
Preparações Farmacêuticas/química , Varredura Diferencial de Calorimetria , Química Farmacêutica , Cristalização , Microscopia Eletrônica de Varredura , Transição de Fase , Soluções , Propriedades de Superfície , Termogravimetria
3.
Cell Microbiol ; 10(4): 908-20, 2008 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-18005238

RESUMO

Toxoplasma gondii is an obligate intracellular parasite that causes severe disease in humans. It is able to infect all nucleated mammalian cells leading to lifelong persistence of the parasite in the host. Here, we studied the effect of T. gondii infection on host cell proliferation and explored the molecular mechanisms involved in host cell cycle progression. We found that T. gondii induced G1/S transition in host cells in the presence of UHRF1, followed by G2 arrest after cyclin B1 downregulation which is probably the major cause of the arrest. Other molecules at the G2/M checkpoint including p53, p21 and Cdk1 were normally regulated. Interestingly, while parasite proliferation was normal in cells that were in the G2 phase, it was suppressed in G1-arrested cells induced by UHRF1-siRNA, indicating the importance of the G2 phase via UHRF1-induced G1/S transition for T. gondii growth.


Assuntos
Proteínas Estimuladoras de Ligação a CCAAT/metabolismo , Ciclo Celular/fisiologia , Proliferação de Células , Fase G2/fisiologia , Toxoplasma/fisiologia , Animais , Western Blotting , Proteínas Estimuladoras de Ligação a CCAAT/genética , Proteína Quinase CDC2/genética , Proteína Quinase CDC2/metabolismo , Linhagem Celular , Ciclina B/genética , Ciclina B/metabolismo , Ciclina B1 , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Citometria de Fluxo , Interações Hospedeiro-Parasita , Humanos , Imuno-Histoquímica , Imunoprecipitação , RNA Interferente Pequeno/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Toxoplasma/crescimento & desenvolvimento , Transfecção , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Ubiquitina-Proteína Ligases
4.
Int J Parasitol ; 38(2): 249-58, 2008 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-17822706

RESUMO

IFN-gamma production is a hallmark of acute infection with the protozoan parasite Toxoplasma gondii. The tryptophan-catabolising enzyme indoleamine 2,3-dioxygenase (IDO), as well as inducible nitric oxide synthase (NOS2) are induced by IFN-gamma and can play extremely diverse roles in immune regulation, defence against pathogens and physiological homeostasis. We investigated the regulation of these two central enzymes in the placenta during acute infection of pregnant female mice. Using IFN-gamma receptor knockout (IFNgammaR-/-) mice, we showed that IDO is not constitutively expressed in term placentas. In contrast, NOS2 expression was observed, largely dependent on IFN-gamma signalling. Upon infection with the avirulent PRU strain of T. gondii, IDO mRNA expression was induced in an IFNgammaR-dependent manner. Surprisingly, NOS2 mRNA was severely suppressed. Importantly, we showed in crossing experiments of heterozygote (IFNgammaR+/-) mothers with IFNgammaR-/- males and vice versa that IDO expression largely depends on the presence of IFN-gamma receptors on foetal cells, and to a lesser extent on maternal cells. Immunohistochemical analysis localised foetal IDO production to invasive trophoblasts within the maternal part of the placenta. The placental vascular endothelium only stained positive when the mothers possessed functional IFN-gamma receptors. In contrast, placental NOS2 expression, but also its suppression following infection, seems to be largely dependent on IFN-gamma signalling in maternal cells. Neither factor appears to regulate placental T. gondii growth, as we observed no difference in parasite numbers between (+/-) and (-/-) foetuses. Taken together, our results demonstrate the crucial role of the foetus in placental IDO, but not NOS2, production following T. gondii infection.


Assuntos
Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Placenta/enzimologia , Complicações Parasitárias na Gravidez/enzimologia , Toxoplasma/fisiologia , Toxoplasmose/enzimologia , Animais , Feminino , Feto/imunologia , Feto/metabolismo , Feto/parasitologia , Genes de Protozoários , Imuno-Histoquímica , Indolamina-Pirrol 2,3,-Dioxigenase/análise , Indolamina-Pirrol 2,3,-Dioxigenase/genética , Interferon gama/genética , Interferon gama/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos , Camundongos Knockout , Óxido Nítrico Sintase Tipo II/análise , Óxido Nítrico Sintase Tipo II/genética , Parasitemia , Placenta/imunologia , Placenta/parasitologia , Gravidez , Complicações Parasitárias na Gravidez/imunologia , RNA Mensageiro/análise , Receptores de Interferon/análise , Receptores de Interferon/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Toxoplasma/genética , Toxoplasmose/imunologia , Receptor de Interferon gama
5.
Int J Parasitol ; 35(14): 1569-76, 2005 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-16185692

RESUMO

Materno-foetal transmission causes one of the most severe forms of infection with the protozoan parasite Toxoplasma gondii. Several studies have shown T. gondii in placental trophoblast cells, which form the barrier between maternal blood circulation and foetal tissue. Parasite multiplication in trophoblast cells is thus a critical step leading to infection of the foetus. Here, we show that multiplication of T. gondii tachyzoites was slow in BeWo trophoblast cells, compared with MRC-5 fibroblast cells. However, unlike MRC-5 cells, even combined stimulation with interferon-gamma and tumor necrosis factor-alpha did not reduce T. gondii replication in BeWo cells. This was associated with a lack of indoleamine-2,3-dioxygenase induction by these cytokines. Neither low availability of iron salts, nor an immunosuppressive action of cyclooxygenase-2 could be attributed to the low T. gondii multiplication rate in BeWo cells. However, treatment with the nitric oxide synthesis inhibitor N(G)-methyl-l-arginine and addition of ornithine enhanced the proliferation rate of the intracellular pathogen. Despite detection of inducible nitric oxide synthase-II mRNA in BeWo cells, nitric oxide production could not be detected during cell culture. Thus, inhibition of arginase activity by nitric oxide synthesis may be partially responsible for the lower multiplication rate in BeWo cells.


Assuntos
Óxido Nítrico/metabolismo , Poliaminas/metabolismo , Complicações Parasitárias na Gravidez/metabolismo , Toxoplasma/fisiologia , Toxoplasmose/transmissão , Trofoblastos/parasitologia , Animais , Arginase/metabolismo , Linhagem Celular , Inibidores de Ciclo-Oxigenase 2/farmacologia , Inibidores Enzimáticos , Feminino , Fibroblastos/parasitologia , Interações Hospedeiro-Parasita , Humanos , Indometacina/farmacologia , Transmissão Vertical de Doenças Infecciosas , Molécula 1 de Adesão Intercelular/metabolismo , Interferon gama/farmacologia , Óxido Nítrico Sintase/antagonistas & inibidores , Ornitina/farmacologia , Parasitologia/métodos , Gravidez , Reprodução , Toxoplasmose/metabolismo , Trofoblastos/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , ômega-N-Metilarginina/farmacologia
6.
Immunol Cell Biol ; 83(5): 483-9, 2005 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-16174097

RESUMO

Materno-foetal transmission causes one of the most serious forms of infection with the intracellular protozoan parasite Toxoplasma gondii. In the placenta, trophoblast cells constitute the barrier between maternal circulation and foetal tissue. We looked at the factors that determine the extent of cell adhesion to human BeWo trophoblast cells during T. gondii infection. BeWo monolayers stimulated with the supernatant of T. gondii-infected PBMC showed a large increase in THP-1 cell adhesion and upregulation of the intercellular adhesion molecule (ICAM)-1. Neutralization of cytokines by corresponding antibodies demonstrated that anti-IFN-gamma, but not anti-TNF-alpha or anti-IL-1beta, led to a significant reduction of THP-1 adhesion to a BeWo monolayer. Treatment of BeWo cells with single cytokines failed to induce upregulation of adhesion. In contrast, simultaneous treatment with IFN-gamma and either TNF-alpha or IL-1beta mimicked strongly the effect of infected cell supernatant. The results suggest that IFN-gamma plays a pivotal role in the cell adhesion process through upregulation of ICAM-1 and in the process of congenital transmission of T. gondii.


Assuntos
Molécula 1 de Adesão Intercelular/fisiologia , Monócitos/fisiologia , Toxoplasma/fisiologia , Toxoplasmose/metabolismo , Trofoblastos/fisiologia , Animais , Adesão Celular/fisiologia , Moléculas de Adesão Celular/biossíntese , Moléculas de Adesão Celular/genética , Linhagem Celular , Citocinas/fisiologia , Feminino , Regulação da Expressão Gênica/fisiologia , Humanos , Transmissão Vertical de Doenças Infecciosas , Gravidez , Toxoplasmose/transmissão
7.
Biochem Pharmacol ; 70(4): 570-9, 2005 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-15964557

RESUMO

Antigen-induced cell death is essential for function, growth and differentiation of T-lymphocytes through legation of the T cell receptor. Since TCR-induced cell death occurs at late G1 checkpoint of the cell cycle and considering that ICBP90 is critical for G1/S transition, we studied the ICBP90 regulation through the TCR pathway in Jurkat cells. ICBP90 expression was strongly decreased after TCR triggering concomitantly to cyclin D3 and topoisomerase IIalpha expression decreases. Cell stimulation with PMA and/or calcium ionophore A23187 down-regulated ICBP90 expression. The decrease of ICBP90 protein and mRNA expressions was accompanied with cell growth arrest. A luciferase reporter assay demonstrated that activation of TCR pathways inhibit ICBP90 gene promoter activity. Three consensus E2F binding sites (called from E2F-a to E2F-c) were identified in the ICBP90 gene promoter and were subjected to mutations. The E2F-a, located in a highly active promoter fragment, shows a strong positive functional activity in proliferating cells. E2F-a and E2F-c binding sites are involved in the TCR-induced down-regulation of ICBP90 gene transcription. Altogether, our data demonstrate that TCR signaling pathways regulate ICBP90 gene expression through pRb/E2F complex. We propose that ICBP90 down-regulation is a key event in G1 arrest preceding T cell death.


Assuntos
Proteínas Estimuladoras de Ligação a CCAAT/genética , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ligação a DNA/metabolismo , Regulação para Baixo/fisiologia , Receptores de Antígenos de Linfócitos T/fisiologia , Fatores de Transcrição/metabolismo , Sequência de Bases , Sítios de Ligação , Proteínas Estimuladoras de Ligação a CCAAT/metabolismo , Calcimicina/farmacologia , Cálcio/metabolismo , Ciclina D3 , Ciclinas/metabolismo , Primers do DNA , Fatores de Transcrição E2F , Humanos , Células Jurkat , Regiões Promotoras Genéticas , Proteína do Retinoblastoma/metabolismo , Fase S , Transdução de Sinais , Acetato de Tetradecanoilforbol/farmacologia , Ubiquitina-Proteína Ligases
8.
J Steroid Biochem Mol Biol ; 96(2): 179-85, 2005 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-15939587

RESUMO

The hormonal form of Vitamin D, 1,25-dihydroxyvitamin D3, is well known for its immunosuppressive, anti-proliferative and pro-apoptotic activities. In the present work, we studied the effect of 1,25-dihydroxyvitamin D3 on Toxoplasma gondii-infected mice. We observed that 1,25-dihydroxyvitamin D3 reduces the survival rate of infected mice by up to 37% at day 10 post-infection compared to untreated infected mice (P < 0.0001). IFN-gamma and IL-12p40 levels were significantly reduced by 1,25-dihydroxyvitamin D3 in infected mice sera indicating an inhibition of Th-1-type cytokines. CD4+ T lymphocyte and splenocyte counts were also reduced following 1,25-dihydroxyvitamin D3 treatment and a marked induction of apoptosis, accompanied with down-regulation of the anti-apoptotic proteins Bcl-2 and Bcl-X(L), was observed. The above results indicate that 1,25-dihydroxyvitamin D3 induces splenocyte apoptosis and enhances host susceptibility to toxoplasmosis.


Assuntos
Calcitriol/farmacologia , Suscetibilidade a Doenças , Baço/citologia , Toxoplasmose/fisiopatologia , Animais , Divisão Celular/efeitos dos fármacos , Imunidade Inata/efeitos dos fármacos , Imunossupressores/farmacologia , Camundongos , Camundongos Endogâmicos BALB C , Baço/efeitos dos fármacos , Baço/fisiologia , Taxa de Sobrevida , Toxoplasmose/mortalidade
9.
J Immunol ; 174(11): 7393-7, 2005 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-15905587

RESUMO

Focal adhesion kinase (FAK) is a nonreceptor protein tyrosine kinase involved in signaling downstream of integrins, linking bacterial detection, cell entry, and initiation of proinflammatory response through MAPKs and NF-kappaB activation. In this study, using protein I/II from Streptococcus mutans as a model activator of FAK, we investigated the potential link between FAK and TLR pathways. Using macrophages from TLR- or MyD88-deficient mice, we report that MyD88 plays a major role in FAK-dependent protein I/II-induced cytokine release. However, response to protein I/II stimulation was independent of TLR4, TLR2, and TLR6. The data suggest that there is a cross talk between FAK and MyD88 signaling pathways. Moreover, MyD88-dependent, LPS-induced IL-6 secretion by human and murine fibroblasts required the presence of FAK, confirming that MyD88 and FAK pathways are interlinked.


Assuntos
Antígenos de Diferenciação/fisiologia , Proteínas Tirosina Quinases/fisiologia , Receptor Cross-Talk/imunologia , Receptores Imunológicos/fisiologia , Transdução de Sinais/imunologia , Proteínas Adaptadoras de Transdução de Sinal , Animais , Antígenos de Diferenciação/genética , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/fisiologia , Células Cultivadas , Fibroblastos/enzimologia , Fibroblastos/imunologia , Fibroblastos/metabolismo , Quinase 1 de Adesão Focal , Proteína-Tirosina Quinases de Adesão Focal , Humanos , Integrina alfa5beta1/metabolismo , Interleucina-6/metabolismo , Ligantes , Lipopolissacarídeos/farmacologia , Macrófagos/enzimologia , Macrófagos/imunologia , Macrófagos/metabolismo , Glicoproteínas de Membrana/metabolismo , Glicoproteínas de Membrana/fisiologia , Camundongos , Camundongos Knockout , Fator 88 de Diferenciação Mieloide , Óxido Nítrico/biossíntese , Fosforilação , Proteínas Tirosina Quinases/metabolismo , Receptores de Superfície Celular/fisiologia , Receptores Imunológicos/deficiência , Receptores Imunológicos/genética , Streptococcus mutans/imunologia , Membrana Sinovial/enzimologia , Membrana Sinovial/imunologia , Membrana Sinovial/metabolismo , Receptor 2 Toll-Like , Receptor 4 Toll-Like , Receptor 6 Toll-Like , Fator de Necrose Tumoral alfa/biossíntese , Fator de Necrose Tumoral alfa/metabolismo
10.
Cell Microbiol ; 6(6): 593-8, 2004 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-15104599

RESUMO

Proinflammatory cytokines such as tumour necrosis factor (TNF)-alpha, interleukin (IL)-1 beta and IL-18 are key mediators of joint inflammation during rheumatoid arthritis (RA). This chronic inflammation may result from a non-specific innate immune response that could be triggered by a wide variety of microorganisms, because numerous bacterial fragments have been identified in the joints of RA patients. As we have demonstrated previously that protein I/II, a pathogen-associated molecular pattern (PAMP) from oral streptococci, triggers IL-6 and IL-8 gene expression and release from either THP-1 cells or fibroblast-like synoviocytes (FLSs), we next explored the capacity of protein I/II to induce the synthesis and release of IL-18 in THP-1 cells and in FLSs isolated from either RA or osteoarthritis (OA) patients. We demonstrate that protein I/II induced IL-18 mRNA in both THP-1 cells and FLSs but, in contrast to THP-1 cells, gene expression was not associated with the synthesis of the corresponding protein in FLSs. Furthermore, our studies revealed that FLSs did not express the biologically inactive precursor, pro-IL-18, in response to protein I/II. Using actinomycin D, we also showed that IL-18 mRNA is unstable in FLSs. Taken together, these data indicate that lack of IL-18 release from activated FLSs results from a defect in translation of IL-18 mRNA into pro-IL-18 because of rapid degradation of IL-18 mRNA.


Assuntos
Proteínas de Bactérias/fisiologia , Fibroblastos/imunologia , Interleucina-18/metabolismo , Streptococcus mutans/patogenicidade , Membrana Sinovial/imunologia , Artrite Reumatoide/imunologia , Artrite Reumatoide/patologia , Linhagem Celular , Células Cultivadas , Dactinomicina/farmacologia , Regulação da Expressão Gênica , Humanos , Interleucina-18/biossíntese , Interleucina-18/genética , Inibidores da Síntese de Ácido Nucleico/farmacologia , Osteoartrite/imunologia , Osteoartrite/patologia , Biossíntese de Proteínas , Estabilidade de RNA , RNA Mensageiro/análise , RNA Mensageiro/isolamento & purificação , Membrana Sinovial/citologia , Transcrição Gênica
11.
Infect Immun ; 72(3): 1397-401, 2004 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-14977944

RESUMO

Protective immunity in mice infected with Toxoplasma gondii is mainly mediated by NK cells, CD4 and CD8 T cells, and type 1 cytokines, such as gamma interferon (IFN-gamma). To clarify the roles of NK cells and IFN-gamma in protection against primary congenital toxoplasmosis, we used recombination activating gene 2 knockout (RAG-2(-/-)) mice, which lack T and B lymphocytes, in comparison with the wild-type BALB/c model. RAG-2(-/-) mice had a significantly lower risk of fetal toxoplasmosis than BALB/c mice (25 versus 63.9%; P = 0.003). This protection was associated with an increased number of maternal NK cells, IFN-gamma secretion by spleen cells, and decreased parasitemia. In the RAG-2(-/-) mice, NK cell depletion increased both the rate of fetal infection, to 56.5% (P = 0.02), and the blood parasite burden. Conversely, in the BALB/c mice, this treatment did not modify maternofetal transmission or the blood parasite burden. Neutralization of IFN-gamma in both infected RAG-2(-/-) and BALB/c mice decreased congenital Toxoplasma transmission, contrasting with an exacerbation of maternal infection. These data suggest that a partially protective immunity against congenital toxoplasmosis is achieved due to the increased number of NK cells in RAG-2(-/-) mice. However, it seems that IFN-gamma enhances, directly or indirectly, the transplacental transmission.


Assuntos
Interferon gama/biossíntese , Células Matadoras Naturais/imunologia , Toxoplasmose Animal/imunologia , Toxoplasmose Animal/transmissão , Animais , Contagem de Células , Proteínas de Ligação a DNA/deficiência , Proteínas de Ligação a DNA/genética , Feminino , Transmissão Vertical de Doenças Infecciosas , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Testes de Neutralização , Gravidez , Complicações Parasitárias na Gravidez/imunologia , Toxoplasmose Animal/complicações , Toxoplasmose Congênita/etiologia , Toxoplasmose Congênita/imunologia
12.
Carcinogenesis ; 25(8): 1477-84, 2004 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-14742316

RESUMO

Several studies reported linkage between bacterial infections and carcinogenesis. Streptococcus bovis was traditionally considered as a lower grade pathogen frequently involved in bacteremia and endocarditis. This bacterium became important in human health as it was shown that 25-80% of patients who presented a S.bovis bacteremia had also a colorectal tumor. Moreover, in previous experiments, we demonstrated that S.bovis or S.bovis wall extracted antigens (WEA) were able to promote carcinogenesis in rats. The aim of the present study was: (i) to identify the S.bovis proteins responsible for in vitro pro-inflammatory properties; (ii) to purify them; (iii) to examine their ability to stimulate in vitro IL-8 and COX-2 expression by human colon cancer cells; and (iv) to assess in vivo their pro-carcinogenic potential in a rat model of colon carcinogenesis. The purified S300 fraction, as determined by proteomic analysis, contained 72 protein spots in two-dimensional gel electrophoresis representing 12 different proteins able to trigger human epithelial colonic Caco-2 cells and rat colonic mucosa to release CXC chemokines (human IL-8 or rat CINC/GRO) and prostaglandins E2, correlated with an in vitro over-expression of COX-2. Moreover, these proteins were highly effective in the promotion of pre-neoplastic lesions in azoxymethane-treated rats. In the presence of these proteins, Caco-2 cells exhibited enhanced phosphorylation of the three classes of MAP kinases. Our results show a relationship between the pro-inflammatory potential of S.bovis proteins and their pro-carcinogenic properties, confirming the linkage between inflammation and colon carcinogenesis. These data support the hypothesis that colonic bacteria can contribute to cancer development particularly in chronic infection/inflammation diseases where bacterial components may interfere with cell function.


Assuntos
Carcinógenos , Streptococcus bovis/metabolismo , Animais , Western Blotting , Células CACO-2 , Diferenciação Celular , Linhagem Celular Tumoral , Neoplasias do Colo/metabolismo , Ciclo-Oxigenase 2 , Dinoprostona/metabolismo , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Eletroforese em Gel Bidimensional , Inibidores Enzimáticos/farmacologia , Humanos , Concentração de Íons de Hidrogênio , Inflamação , Interleucina-8/metabolismo , Isoenzimas/metabolismo , Espectrometria de Massas , Proteínas de Membrana , Mucosa/patologia , Fosforilação , Prostaglandina-Endoperóxido Sintases/metabolismo , Proteoma , Ratos , Frações Subcelulares , Fatores de Tempo
13.
Ann N Y Acad Sci ; 1030: 355-60, 2004 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-15659817

RESUMO

Protein kinase 2 (casein kinase 2 [CK2]) is a protein serine/threonine kinase involved in cell proliferation with an expression that is dysregulated in tumors. ICBP90, a transcription factor exhibiting antiapoptotic properties, has several putative CK2 phosphorylation sites. The aim of the present study was to investigate whether ICBP90 could behave as a CK2 substrate. We observed that ICBP90 was more efficiently phosphorylated by the free CK2a subunit than by the heterotetrameric CK2 (alpha(2), beta(2)). Our results suggest that CK2 is an important regulator of the transcriptional activity of ICBP90 and therefore of the antiapoptotic properties of ICBP90. We propose that the "ICBP90 family" members may be substrates for CK2.


Assuntos
Proteínas Estimuladoras de Ligação a CCAAT/metabolismo , Caseína Quinase II/metabolismo , Animais , Células COS , Humanos , Mapeamento de Peptídeos , Fosforilação , Especificidade por Substrato , Ubiquitina-Proteína Ligases
14.
Infect Immun ; 71(11): 6615-9, 2003 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-14573684

RESUMO

We evaluated the effect of vaccination with the SAG1 protein of Toxoplasma gondii against congenital toxoplasmosis in mice with different genetic backgrounds. In BALB/c mice (H-2(d)), vaccination reduced the number of infected fetuses by 50% and was associated with a mixed type 1 and type 2 immunity. In CBA/J mice (H-2(k)), vaccination increased the number of infected fetuses by 50% and was associated with a predominant type 2 response. Our results indicate that the effect of vaccination with SAG1 is controlled by the genetic background of the mouse.


Assuntos
Antígenos de Protozoários/imunologia , Proteínas de Protozoários/imunologia , Vacinas Protozoárias/imunologia , Toxoplasma/imunologia , Toxoplasmose Congênita/prevenção & controle , Animais , Anticorpos Antiprotozoários/sangue , Citocinas/biossíntese , Feminino , Imunização , Transmissão Vertical de Doenças Infecciosas , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos CBA , Especificidade da Espécie , Vacinação
15.
Infect Immun ; 71(9): 5169-77, 2003 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-12933861

RESUMO

Streptococcus mutans possesses different cell wall molecules, such as protein of the I/II family, the serotype f polysaccharide rhamnose glucose polymer (RGP), and lipoteichoic acid (LTA), which act as adhesins and modulins, allowing S. mutans to colonize teeth and cause dental caries and pulpitis. We tested several isogenic mutants of S. mutans defective in protein I/II and/or RGP, as well as purified modulins such as protein I/II, RGP, and LTA, for their binding and activation abilities on monocytic, dental pulp (DP), and periodontal ligament (PDL) cells. Our results demonstrate that both protein I/II and RGP play important roles in streptococcal adherence to human monocytic and fibroblastic cells, whereas LTA is only a minor adhesin. In the activation process, the cytokine response elicited is polarized toward a Th1 response which seems principally due to protein I/II and RGP. Even if protein I/II seems to be more efficient in its purified form in triggering cells to release interleukin-8 (IL-8), RGP is the most efficient cytokine-stimulating component in intact bacteria, while LTA plays only a minor role. In cell activation, we showed, by using either cytochalasin D or coated ligands, that internalization of either S. mutans, S. mutans isogenic mutants, or purified ligands is not necessary to trigger cells to release IL-8. We also showed that, besides the implication of monocytes in pulpal inflammation, fibroblast-like cells such as DP and PDL cells are also actively implicated in local inflammation and in the generation of a Th1 response after stimulation with S. mutans cells or antigens.


Assuntos
Genes Bacterianos , Interleucina-6/biossíntese , Interleucina-8/biossíntese , Glicoproteínas de Membrana , Streptococcus mutans/genética , Streptococcus mutans/imunologia , Fator de Necrose Tumoral alfa/biossíntese , Adesinas Bacterianas/imunologia , Aderência Bacteriana , Proteínas de Bactérias/genética , Proteínas de Bactérias/imunologia , Linhagem Celular , Cárie Dentária/imunologia , Cárie Dentária/microbiologia , Polpa Dentária/imunologia , Polpa Dentária/microbiologia , Humanos , Hidroliases/genética , Hidroliases/imunologia , Monócitos/imunologia , Monócitos/microbiologia , Mutagênese Insercional , Ligamento Periodontal/imunologia , Ligamento Periodontal/microbiologia , Infecções Estreptocócicas/imunologia , Infecções Estreptocócicas/microbiologia , Streptococcus mutans/patogenicidade
16.
J Biol Chem ; 278(30): 27721-8, 2003 Jul 25.
Artigo em Inglês | MEDLINE | ID: mdl-12761229

RESUMO

Protein I/II, a pathogen-associated molecular pattern from oral streptococci, is a potent inducer of interleukin-6 (IL-6) and IL-8 synthesis and release from fibroblast-like synoviocytes (FLSs), cells that are critically involved in joint inflammation. This synthesis implicates ERK 1/2 and JNKs as well as AP-1-binding activity and nuclear translocation of NF-kappaB. The mechanisms by which protein I/II activates MAPKs remain, however, elusive. Because focal adhesion kinase (FAK) was proposed to play a role in signaling to MAPKs, we examined its ability to contribute to the MAPKs-dependent synthesis of IL-6 and IL-8 in response to protein I/II. We used FAK-/- fibroblasts as well as FAK+/+ fibroblasts and FLSs transfected with FRNK, a dominant negative form of FAK. The results demonstrate that IL-6 and IL-8 release in response to protein I/II was strongly inhibited in both protein I/II-stimulated FAK-/- and FRNK-transfected cells. Cytochalasin D, which inhibits protein I/II-induced phosphorylation of FAK (Tyr-397), had no effect either on activation of ERK 1/2 and JNKs or on IL-6 and IL-8 release. Taken together, these results indicate that IL-6 and IL-8 release by protein I/II-activated FLSs is regulated by FAK independently of Tyr-397 phosphorylation.


Assuntos
Fibroblastos/metabolismo , Interleucina-6/biossíntese , Interleucina-8/biossíntese , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Proteínas Tirosina Quinases/fisiologia , Membrana Sinovial/citologia , Transporte Ativo do Núcleo Celular , Androstadienos/farmacologia , Anticorpos Monoclonais/metabolismo , Western Blotting , Células Cultivadas , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/farmacologia , Quinase 1 de Adesão Focal , Proteína-Tirosina Quinases de Adesão Focal , Genes Dominantes , Humanos , Inflamação , Integrina alfa5beta1/metabolismo , Interleucina-8/metabolismo , Proteína Quinase 3 Ativada por Mitógeno , NF-kappa B/metabolismo , Inibidores da Síntese de Ácido Nucleico/farmacologia , Fosforilação , Inibidores da Síntese de Proteínas/farmacologia , Proteínas Tirosina Quinases/metabolismo , RNA Mensageiro/metabolismo , Transdução de Sinais , Fatores de Tempo , Tirosina/metabolismo , Wortmanina
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