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1.
PLoS One ; 8(6): e65934, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23805193

RESUMO

BACKGROUND: Bacterial vectors have been proposed as novel vaccine strategies to induce strong cellular immunity. Attenuated strains of Brucella abortus comprise promising vector candidates since they have the potential to induce strong CD4(+) and CD8(+) T-cell mediated immune responses in the absence of excessive inflammation as observed with other Gram-negative bacteria. However, some Brucella strains interfere with the maturation of dendritic cells (DCs), which is essential for antigen-specific T-cell priming. In the present study, we investigated the interaction of human monocyte-derived DCs with the smooth attenuated B. abortus strain (S) 19, which has previously been employed successfully to vaccinate cattle. METHODOLOGY/PRINCIPAL FINDINGS: We first looked into the potential of S19 to hamper the cytokine-induced maturation of DCs; however, infected cells expressed CD25, CD40, CD80, and CD86 to a comparable extent as uninfected, cytokine-matured DCs. Furthermore, S19 activated DCs in the absence of exogeneous stimuli, enhanced the expression of HLA-ABC and HLA-DR, and was able to persist intracellularly without causing cytotoxicity. Thus, DCs provide a cellular niche for persisting brucellae in vivo as a permanent source of antigen. S19-infected DCs produced IL-12/23p40, IL-12p70, and IL-10, but not IL-23. While heat-killed bacteria also activated DCs, soluble mediators were not involved in S19-induced activation of human DCs. HEK 293 transfectants revealed cellular activation by S19 primarily through engagement of Toll-like receptor (TLR)2. CONCLUSIONS/SIGNIFICANCE: Thus, as an immunological prerequisite for vaccine efficacy, B. abortus S19 potently infects and potently activates (most likely via TLR2) human DCs to produce Th1-promoting cytokines.


Assuntos
Brucella abortus/fisiologia , Células Dendríticas/metabolismo , Interleucina-12/metabolismo , Antígeno B7-1/metabolismo , Antígenos CD40/metabolismo , Células Cultivadas , Citocinas/metabolismo , Células Dendríticas/citologia , Células Dendríticas/microbiologia , Células HEK293 , Antígenos HLA/metabolismo , Antígenos HLA-DR/metabolismo , Humanos , Interleucina-10/metabolismo , Subunidade alfa de Receptor de Interleucina-2/metabolismo , Interleucina-23/metabolismo , Interleucina-8/metabolismo , NF-kappa B/metabolismo , Receptor 2 Toll-Like/genética , Receptor 2 Toll-Like/metabolismo
2.
Infect Immun ; 81(7): 2309-17, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-23589577

RESUMO

Depletion of arginine is a recognized strategy that pathogens use to evade immune effector mechanisms. Depletion depends on microbial enzymes such as arginases, which are considered virulence factors. The effect is mostly interpreted as being a consequence of successful competition with host enzymes for the substrate. However, both arginases and arginine deiminases (ADI) have been associated with pathogen virulence. Both deplete arginine, but their reaction products differ. An ADI has been implicated in the virulence of Giardia duodenalis, an intestinal parasite that infects humans and animals, causing significant morbidity. Dendritic cells (DC) play a critical role in host defense and also in a murine G. duodenalis infection model. The functional properties of these innate immune cells depend on the milieu in which they are activated. Here, the dependence of the response of these cells on arginine was studied by using Giardia ADI and lipopolysaccharide-stimulated human monocyte-derived DC. Arginine depletion by ADI significantly increased tumor necrosis factor alpha and decreased interleukin-10 (IL-10) and IL-12p40 secretion. It also reduced the upregulation of surface CD83 and CD86 molecules, which are involved in cell-cell interactions. Arginine depletion also reduced the phosphorylation of S6 kinase in DC, suggesting the involvement of the mammalian target of rapamycin signaling pathway. The changes were due to arginine depletion and the formation of reaction products, in particular, ammonium ions. Comparison of NH(4)(+) and urea revealed distinct immunomodulatory activities of these products of deiminases and arginases, respectively. The data suggest that a better understanding of the role of arginine-depleting pathogen enzymes for immune evasion will have to take enzyme class and reaction products into consideration.


Assuntos
Amônia/metabolismo , Arginina/metabolismo , Células Dendríticas/parasitologia , Giardia lamblia/enzimologia , Hidrolases/metabolismo , Antígenos CD/imunologia , Antígeno B7-2/imunologia , Células Dendríticas/imunologia , Giardia lamblia/imunologia , Giardia lamblia/patogenicidade , Humanos , Hidrolases/genética , Imunoglobulinas/imunologia , Interleucina-10/imunologia , Subunidade p40 da Interleucina-12/imunologia , Lipopolissacarídeos/imunologia , Glicoproteínas de Membrana/imunologia , Fenótipo , Fosforilação , Proteínas Quinases S6 Ribossômicas/genética , Proteínas Quinases S6 Ribossômicas/metabolismo , Transdução de Sinais , Serina-Treonina Quinases TOR/imunologia , Fator de Necrose Tumoral alfa/imunologia , Ureia/metabolismo , Antígeno CD83
3.
Parasit Vectors ; 6(1): 317, 2013 Nov 04.
Artigo em Inglês | MEDLINE | ID: mdl-24499474

RESUMO

BACKGROUND: Effects of Giardia duodenalis on dendritic cell (DC) functions may contribute to the pathogenesis of chronic giardiasis. G. duodenalis lysate has been shown to inhibit the activation of murine DCs through the ligands of various Toll-like receptors (TLRs), including TLR2 and TLR4. Our study aimed at translating these findings to human DCs. FINDINGS: As described previously for murine DCs, also human DCs were only weakly activated by the parasite itself. LPS-stimulated DCs incubated in the presence of G. duodenalis lysate produced less IL-12/23p40 (p = 0.002), IL-12p70 (p = 0.011), and IL-23 (p = 0.004), but more IL-10 (p = 0.006) than cells incubated in the absence of the parasite. Concomitantly, the expression of CD25, CD83, CD86, and HLA-DR was reduced on G. duodenalis-incubated DCs as compared to control cells. In contrast, human DCs stimulated through TLR2 in combination with TLR1 or TLR6 and G. duodenalis lysate secreted significantly more IL-12/23p40 (p = 0.006), IL-23 (p = 0.002), and IL-10 (p = 0.014) than cells stimulated through TLR2 ligands alone. Ligands for TLR2/TLR1 or TLR2/TLR6 also induced enhanced extracellular expression of CD25, CD83, and CD86 (p < 0.05). CONCLUSIONS: In contrast to murine DCs, human DCs incubated in the presence of G. duodenalis and stimulated through TLR2 show increased activation as compared to cells incubated in the absence of the parasite. Thus, TLR2 ligands, e.g., delivered by probiotic lactobacilli, might be beneficial in human giardiasis through an adjuvant effect on the induction of cellular immune responses against G. duodenalis.


Assuntos
Antígenos CD/análise , Citocinas/metabolismo , Células Dendríticas/química , Células Dendríticas/imunologia , Giardia lamblia/imunologia , Receptor 2 Toll-Like/agonistas , Humanos , Receptor 1 Toll-Like/agonistas , Receptor 6 Toll-Like/agonistas
4.
Infect Immun ; 80(12): 4195-202, 2012 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-22988018

RESUMO

Studying the interaction of dendritic cells (DCs) with bacteria controlled by T-cell-mediated immune responses may reveal novel adjuvants for the induction of cellular immunity. Murine studies and the observation that nocardias infect predominantly immunosuppressed patients have suggested that these bacteria may possess an adjuvant potential. Moreover, adjuvants on the basis of the nocardial cell wall have been applied in clinical studies. Since the handling of adjuvants by DCs may determine the type of immune responses induced by a vaccine, the present study aimed at investigating the interaction of immature human monocyte-derived DCs with live or inactivated Nocardia farcinica in vitro and determining the cellular phenotypic changes as well as alterations in characteristic functions, such as phagocytosis, induction of T-cell proliferation, and cytokine secretion. Human DCs ingested N. farcinica and eradicated the bacterium intracellularly. DCs exposed to inactivated N. farcinica were activated, i.e., they developed a mature phenotype, downregulated their phagocytic capacity, and stimulated allogeneic T cells in mixed leukocyte reactions. Soluble factors were not involved in this process. To elucidate the potential adjuvant effect of N. farcinica on the induction of T-cell-mediated immune responses, we characterized the cytokines produced by nocardia-exposed DCs and detected substantial amounts of tumor necrosis factor alpha (TNF-α) and interleukin-12 p40 (IL-12p40). However, nocardia-treated DCs secreted only small amounts of IL-12p70, which were significantly smaller than the amounts of IL-23. Thus, N. farcinica activates DCs, but adjuvants based on this bacterium may have only a limited capacity to induce Th1 immune responses.


Assuntos
Células Dendríticas/imunologia , Interleucina-12/biossíntese , Interleucina-23/biossíntese , Nocardia/imunologia , Adjuvantes Imunológicos , Células Dendríticas/metabolismo , Humanos , Subunidade p40 da Interleucina-12/imunologia , Ativação Linfocitária , Teste de Cultura Mista de Linfócitos , Nocardia/classificação , Linfócitos T/imunologia
5.
Infect Immun ; 80(8): 2724-34, 2012 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-22615251

RESUMO

Helicobacter pylori may cause chronic gastritis, gastric cancer, or lymphoma. Myeloid antigen-presenting cells (APCs) are most likely involved in the induction and expression of the underlying inflammatory responses. To study the interaction of human APC subsets with H. pylori, we infected monocytes, monocyte-derived dendritic cells (DCs), and monocyte-derived (classically activated; M1) macrophages with H. pylori and analyzed phenotypic alterations, cytokine secretion, phagocytosis, and immunostimulation. Since we detected CD163(+) (alternatively activated; M2) macrophages in gastric biopsy specimens from H. pylori-positive patients, we also included monocyte-derived M2 macrophages in the study. Upon H. pylori infection, monocytes secreted interleukin-1ß (IL-1ß), IL-6, IL-10, and IL-12p40 (partially secreted as IL-23) but not IL-12p70. Infected DCs became activated, as shown by the enhanced expression of CD25, CD80, CD83, PDL-1, and CCR7, and secreted IL-1ß, IL-6, IL-10, IL-12p40, IL-12p70, and IL-23. However, infection led to significantly downregulated CD209 and suppressed the constitutive secretion of macrophage migration inhibitory factor (MIF). H. pylori-infected M1 macrophages upregulated CD14 and CD32, downregulated CD11b and HLA-DR, and secreted mainly IL-1ß, IL-6, IL-10, IL-12p40, and IL-23. Activation of DCs and M1 macrophages correlated with increased capacity to induce T-cell proliferation and decreased phagocytosis of dextran. M2 macrophages upregulated CD14 and CD206 and secreted IL-10 but produced less of the proinflammatory cytokines than M1 macrophages. Thus, H. pylori affects the functions of human APC subsets differently, which may influence the course and the outcome of H. pylori infection. The suppression of MIF in DCs constitutes a novel immune evasion mechanism exploited by H. pylori.


Assuntos
Células Dendríticas/microbiologia , Helicobacter pylori/fisiologia , Macrófagos/microbiologia , Monócitos/microbiologia , Células Cultivadas , Citocinas/genética , Citocinas/metabolismo , Mucosa Gástrica/citologia , Mucosa Gástrica/microbiologia , Regulação da Expressão Gênica/fisiologia , Humanos , Inflamação/imunologia , Inflamação/metabolismo , Ativação Linfocitária , Macrófagos/classificação , Fagocitose
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