RESUMO
Streptococcus suis (S. suis) is an important porcine pathogen, causing severe disease like meningitis and septicemia primarily in piglets. Previous work showed that the IgM-degrading enzyme of S. suis (Ide Ssuis ) specifically cleaves soluble porcine IgM and is involved in complement evasion. The objective of this study was to investigate Ide Ssuis cleavage of the IgM B cell receptor and subsequent changes in B cell receptor mediated signaling. Flow cytometry analysis revealed cleavage of the IgM B cell receptor by recombinant (r) Ide Ssuis _homologue as well as Ide Ssuis derived from culture supernatants of S. suis serotype 2 on porcine PBMCs and mandibular lymph node cells. Point-mutated rIde Ssuis _homologue_C195S did not cleave the IgM B cell receptor. After receptor cleavage by rIde Ssuis _homologue, it took at least 20 h for mandibular lymph node cells to restore the IgM B cell receptor to levels comparable to cells previously treated with rIde Ssuis _homologue_C195S. B cell receptor mediated signaling after specific stimulation via the F(ab')2 portion was significantly inhibited by rIde Ssuis _homologue receptor cleavage in IgM+ B cells, but not in IgG+ B cells. Within IgM+ cells, CD21+ B2 cells and CD21- B1-like cells were equally impaired in their signaling capacity upon rIde Ssuis _homologue B cell receptor cleavage. In comparison, intracellular B cell receptor independent stimulation with tyrosine phosphatase inhibitor pervanadate increased signaling in all investigated B cell types. In conclusion, this study demonstrates Ide Ssuis cleavage efficacy on the IgM B cell receptor and its consequences for B cell signaling.
Assuntos
Streptococcus suis , Animais , Suínos , Linfócitos B , Transdução de Sinais , Receptores de Antígenos de Linfócitos B , Imunoglobulina MRESUMO
Streptococcus suis (S. suis) is a common swine pathogen but also poses a threat to human health in causing meningitis and severe cases of streptococcal toxic shock-like syndrome (STSLS). Therefore, it is crucial to understand how S. suis interacts with the host immune system during bacteremia. As S. suis has the ability to introduce d-alanine into its lipoteichoic acids (LTAs), we investigated the working hypothesis that cell wall modification by LTA d-alanylation influences the interaction of S. suis with porcine blood immune cells. We created an isogenic mutant of S. suis strain 10 by in-frame deletion of the d-alanine d-alanyl carrier ligase (DltA). d-alanylation of LTAs was associated with reduced phagocytosis of S. suis by porcine granulocytes, reduced deposition of complement factor C3 on the bacterial surface, increased hydrophobicity of streptococci, and increased resistance to cationic antimicrobial peptides (CAMPs). At the same time, survival of S. suis was not significantly increased by LTA d-alanylation in whole blood of conventional piglets with specific IgG. However, we found a distinct cytokine pattern as IL-1ß but not tumor necrosis factor (TNF)-α levels were significantly reduced in blood infected with the ΔdltA mutant. In contrast to TNF-α, activation and secretion of IL-1ß are inflammasome-dependent, suggesting a possible influence of LTA d-alanylation on inflammasome regulation. Especially in the absence of specific antibodies, the association of S. suis with porcine monocytes was reduced by d-alanylation of its LTAs. This dltA-dependent phenotype was also observed with a non-encapsulated dltA double mutant indicating that it is independent of capsular polysaccharides. High antibody levels caused high levels of S. suis-monocyte-association followed by inflammatory cell death and strong production of both IL-1ß and TNF-α, while the influence of LTA d-alanylation of the streptococci became less visible. In summary, the results of this study expand previous findings on d-alanylation of LTAs in S. suis and suggest that this pathogen specifically modulates association with blood leukocytes through this modification of its surface.
RESUMO
A vaccine protecting against different Streptococcus suis serotypes is highly needed in porcine practice to improve animal welfare and reduce the use of antibiotics. We hypothesized that immunogens prominently recognized by convalescence sera but significantly less so by sera of susceptible piglets are putative protective antigens. Accordingly, we investigated immunogenicity and protective efficacy of a multicomponent vaccine including six main conserved immunogens, namely SSU0934, SSU1869, SSU0757, SSU1950, SSU1664 and SSU0187. Flow cytometry confirmed surface expression of all six immunogens in S. suis serotypes 2, 9 and 14. Although prime-booster vaccination after weaning resulted in significantly higher specific IgG levels against all six immunogens compared to the placebo-treated group, no significant differences between bacterial survival in blood from either vaccinated or control animals were recorded for serotype 2, 9 and 14 strains. Furthermore, vaccinated piglets were not protected against morbidity elicited through intranasal challenge with S. suis serotype 14. As ~50% of animals in both groups did not develop disease, we investigated putative other correlates of protection. Induction of reactive oxygen species (ROS) in blood granulocytes was not associated with vaccination but correlated with protection as all piglets with >5% ROS survived the challenge. Based on these findings we discuss that the main immunogens of S. suis might actually not be a priori good candidates for protective antigens. On the contrary, expression of immunogens that evoke antibodies that do not mediate killing of this pathogen might constitute an evolutionary advantage conserved in many different S. suis strains.
Assuntos
Imunogenicidade da Vacina , Infecções Estreptocócicas/veterinária , Vacinas Estreptocócicas/imunologia , Streptococcus suis/imunologia , Doenças dos Suínos/prevenção & controle , Animais , Infecções Estreptocócicas/microbiologia , Infecções Estreptocócicas/prevenção & controle , Vacinas Estreptocócicas/administração & dosagem , Sus scrofa , Suínos , Doenças dos Suínos/microbiologia , Resultado do TratamentoRESUMO
Weaning piglets are susceptible to the invasive Streptococcus (S.) suis infection, which can result in septicemia. The aim of this study was to investigate the cytokine profile induced upon S. suis infection of blood, to determine the cellular sources of those cytokines, and to study the potential effects of the induced cytokines on bacterial killing. We measured TNF-α, IL-6, IFN-γ, IL-17A and IL-10 after an experimental intravenous infection with S. suis serotype 2 in vivo, and analyzed whole blood, peripheral blood mononuclear cells (PBMC) and separated leukocytes to identify the cytokine-producing cell type(s). In addition, we used a reconstituted whole blood assay to investigate the effect of TNF-α on bacterial killing in the presence of different S. suis-specific IgG levels. An increase in IL-6 and IL-10, but not in IFN-γ or IL-17A, was observed in two of three piglets with pronounced bacteremia 16 to 20 h after infection, but not in piglets with controlled bacteremia. Our results confirmed previous findings that S. suis induces TNF-α and IL-6 and could demonstrate that TNF-α is produced by monocytes in vitro. We further found that IL-10 induction resulted in reduced secretion of TNF-α and IL-6. Rapid induction of TNF-α was, however, not crucial for in vitro bacterial killing, not even in the absence of specific IgG.
RESUMO
Bacteremia is a hallmark of invasive Streptococcus suis infections of pigs, often leading to septicemia, meningitis, or arthritis. An important defense mechanism of neutrophils is the generation of reactive oxygen species (ROS). In this study, we report high levels of ROS production by blood granulocytes after intravenous infection of a pig with high levels of S. suis-specific antibodies and comparatively low levels of bacteremia. This prompted us to investigate the working hypothesis that the immunoglobulin-mediated oxidative burst contributes to the killing of S. suis in porcine blood. Several S. suis strains representing serotypes 2, 7, and 9 proved to be highly susceptible to the oxidative burst intermediate hydrogen peroxide, already at concentrations of 0.001%. The induction of ROS in granulocytes in ex vivo-infected reconstituted blood showed an association with pathogen-specific antibody levels. Importantly, inhibition of ROS production by the NADPH oxidase inhibitor apocynin led to significantly increased bacterial survival in the presence of high specific antibody levels. The oxidative burst rate of granulocytes partially depended on complement activation, as shown by specific inhibition. Furthermore, treatment of IgG-depleted serum with a specific IgM protease or heat to inactivate complement resulted in >3-fold decreased oxidative burst activity and increased bacterial survival in reconstituted porcine blood in accordance with an IgM-complement-oxidative burst axis. In conclusion, this study highlights an important control mechanism of S. suis bacteremia in the natural host: the induction of ROS in blood granulocytes via specific immunoglobulins such as IgM.
Assuntos
Granulócitos/fisiologia , Explosão Respiratória/fisiologia , Streptococcus suis/imunologia , Doenças dos Suínos/microbiologia , Acetofenonas/farmacologia , Animais , Inibidores Enzimáticos/farmacologia , Granulócitos/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Streptococcus suis/efeitos dos fármacos , Suínos , Doenças dos Suínos/imunologiaRESUMO
Vaccination of weaning piglets with the recombinant IgM degrading enzyme of Streptococcus suis (S. suis), rIde Ssuis, elicits protection against disease caused by serotype (cps) 2 infection. In Europe, S. suis cps9 is at least as important as cps2 in causing severe herd problems associated with meningitis, septicemia and arthritis. The objective of this study was to determine humoral and cellular immunogenicities of rIde Ssuis suckling piglet vaccination and to investigate protection against a virulent cps9 strain. Vaccination in the 2nd and 4th week of life with rIde Ssuis and an oil-in-water adjuvant induced seroconversion against Ide Ssuis in 13 of 20 vaccinated piglets. In the 5th week, survival of the S. suis cps9 strain was significantly reduced in the blood of prime-booster vaccinated piglets. After a 2nd booster vaccination Ide Ssuis -reactive T helper (Th) cells partially producing TNF-α, IL-17A or IFN-É£ were detectable in rIde Ssuis -vaccinated but not in placebo-treated piglets and frequencies of Ide Ssuis -reactive Th cells correlated with α-Ide Ssuis-IgG levels. An intravenous challenge, conducted with a cps9 strain of sequence type (ST) 94, led to 89% mortality in placebo-treated piglets due to septicemia and meningitis. In contrast, all rIde Ssuis prime-booster-booster vaccinated littermates survived the challenge despite signs of disease such as fever and lameness. In conclusion, the described rIde Ssuis vaccination induces humoral and detectable Ide Ssuis -reactive Th cell responses and leads to protection against a highly virulent cps9 strain.
RESUMO
Streptococcus suis (S. suis) causes meningitis, arthritis and endocarditis in piglets. The aim of this study was to characterize the IgM degrading enzyme of S. suis (IdeSsuis) and to investigate the role of IgM cleavage in evasion of the classical complement pathway and pathogenesis. Targeted mutagenesis of a cysteine in the putative active center of IdeSsuis abrogated IgM cleavage completely. In contrast to wt rIdeSsuis, point mutated rIdeSsuis_C195S did not reduce complement-mediated hemolysis indicating that complement inhibition by rIdeSsuis depends on the IgM proteolytic activity. A S. suis mutant expressing IdeSsuis_C195S did not reduce IgM labeling, whereas the wt and complemented mutant showed less IgM F(ab')2 and IgM Fc antigen on the surface. IgM cleavage increased survival of S. suis in porcine blood ex vivo and mediated complement evasion as demonstrated by blood survival and C3 deposition assays including the comparative addition of rIdeSsuis and rIdeSsuis_C195S. However, experimental infection of piglets disclosed no significant differences in virulence between S. suis wt and isogenic mutants without IgM cleavage activity. This work revealed for the first time in vivo labeling of S. suis with IgM in the cerebrospinal fluid of piglets with meningitis. In conclusion, this study classifies IdeSsuis as a cysteine protease and emphasizes the role of IgM cleavage for bacterial survival in porcine blood and complement evasion though IgM cleavage is not crucial for the pathogenesis of serotype 2 meningitis.
Assuntos
Proteínas do Sistema Complemento/imunologia , Cisteína Proteases/imunologia , Evasão da Resposta Imune , Imunoglobulina M/metabolismo , Streptococcus suis/enzimologia , Streptococcus suis/imunologia , Animais , Proteínas de Bactérias/genética , Proteínas de Bactérias/imunologia , Sítios de Ligação de Anticorpos , Cisteína Proteases/genética , Interações Hospedeiro-Patógeno/imunologia , Imunoglobulina M/imunologia , Meningite/líquido cefalorraquidiano , Meningite/microbiologia , Mutagênese , Proteólise , Sorogrupo , Infecções Estreptocócicas/sangue , Infecções Estreptocócicas/imunologia , Suínos , Doenças dos Suínos/microbiologiaRESUMO
There is growing interest in studying host-pathogen interactions in human-relevant large animal models such as the pig. Despite the progress in developing immunological reagents for porcine T cell research, there is an urgent need to directly assess pathogen-specific T cells-an extremely rare population of cells, but of upmost importance in orchestrating the host immune response to a given pathogen. Here, we established that the activation marker CD154 (CD40L), known from human and mouse studies, identifies also porcine antigen-reactive CD4+ T lymphocytes. CD154 expression was upregulated early after antigen encounter and CD4+CD154+ antigen-reactive T cells coexpressed cytokines. Antigen-induced expansion and autologous restimulation enabled a time- and dose-resolved analysis of CD154 regulation and a significantly increased resolution in phenotypic profiling of antigen-responsive cells. CD154 expression identified T cells responding to staphylococcal Enterotoxin B superantigen stimulation as well as T cells responding to the fungus Candida albicans and T cells specific for a highly prevalent intestinal parasite, the nematode Ascaris suum during acute and trickle infection. Antigen-reactive T cells were further detected after immunization of pigs with a single recombinant bacterial antigen of Streptococcus suis only. Thus, our study offers new ways to study antigen-specific T lymphocytes in the pig and their contribution to host-pathogen interactions.
RESUMO
IL-9-secreting Th9 cells have been considered to play a pivotal role in the pathogenesis of atopic diseases. To what extent IL-9-producing cells are induced or regulated by sensitization with naturally occurring allergens is not yet clear. Naturally occurring allergens are capable of inducing IL-6 production in dendritic cells (DCs). Whether allergen-induced IL-6 supports a Th9 subtype by increasing IL-9 production, as observed in in vitro studies, or rather favors Th17 differentiation is not finally resolved. Therefore, in the present study we have investigated the impact of IL-6 on the Th9/Th17 balance depending on the predominant cytokine milieu and, additionally, in vivo using a DC-driven murine asthma model. In vitro, IL-6 increases Th9 cells under strong IL-4 and TGF-ß activation, whereas under moderate IL-4 and TGF-ß activation the presence of IL-6 shifts naive CD4(+) cells to Th17 cells. To induce allergic airway inflammation, OVA-pulsed DCs from IL-6-deficient or wild-type donors were adoptively transferred into BALB/c mice. Recipients receiving IL-6-producing wild-type DCs showed a significant decrease of Th9- and IL-4-producing Th2 cells but an increase of Th17 cells in lung tissue in comparison with recipients sensitized with IL-6-deficient DCs. Our data suggest that the IL-6-mediated reduction of Th2-related IL-4 leads to a decline of the Th9 immune response and allows Th17 differentiation.
Assuntos
Alérgenos/imunologia , Asma/imunologia , Linfócitos T CD4-Positivos/imunologia , Inflamação/imunologia , Interleucina-17/imunologia , Interleucina-6/imunologia , Interleucina-9/imunologia , Animais , Modelos Animais de Doenças , Feminino , Interleucina-6/biossíntese , Camundongos , Camundongos Endogâmicos BALB CRESUMO
Syndecan-4 (SDC4), expressed on dendritic cells (DCs) and activated T cells, plays a crucial role in DC motility and has been shown as a potential target for activated T-cell-driven diseases. In the present study, we investigate the role of SDC4 in the development of T-helper 2 cell-mediated allergic asthma. Using SDC4-deficient mice or an anti-SDC4 antibody we show that the absence or blocking of SDC4 signalling in ovalbumin-sensitized mice results in a reduced asthma phenotype compared with control animals. Most importantly, even established asthma is significantly decreased using the anti-SDC4 antibody. The disturbed SDC4 signalling leads to an impaired motility and directional migration of antigen-presenting DCs and therefore, to a modified sensitization leading to diminished airway inflammation. Our results demonstrate that SDC4 plays an important role in asthma induction and indicate SDC4 as possible target for therapeutic intervention in this disease.
Assuntos
Asma/imunologia , Movimento Celular/imunologia , Células Dendríticas/imunologia , Pulmão/imunologia , Sindecana-4/imunologia , Células Th2/imunologia , Adjuvantes Imunológicos , Hidróxido de Alumínio , Animais , Asma/patologia , Asma/fisiopatologia , Movimento Celular/genética , Citocinas/imunologia , Células Dendríticas/metabolismo , Modelos Animais de Doenças , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Imunoglobulina E/imunologia , Imunoglobulina G/imunologia , Pulmão/patologia , Pulmão/fisiopatologia , Hidróxido de Magnésio , Camundongos , Camundongos Knockout , Ovalbumina , Pletismografia , Hipersensibilidade Respiratória/imunologia , Hipersensibilidade Respiratória/patologia , Hipersensibilidade Respiratória/fisiopatologia , Sindecana-4/genéticaRESUMO
The T helper cell subsets Th1, Th2, Th17, and Treg play an important role in immune cell homeostasis, in host defense, and in immunological disorders. Recently, much attention has been paid to Th17 cells which seem to play an important role in the early phase of the adoptive immune response and autoimmune disease. When generating Th17 cells under in vitro conditions the amount of IL-17A producing cells hardly exceeds 20% while the nature of the remaining T cells is poorly characterized. As engagement of the aryl hydrocarbon receptor (AHR) has also been postulated to modulate the differentiation of T helper cells into Th17 cells with regard to the IL-17A expression we ask how far do Th17 polarizing conditions in combination with ligand induced AHR activation have an effect on the production of other T helper cell cytokines. We found that a high proportion of T helper cells cultured under Th17 polarizing conditions are IL-8 and IL-9 single producing cells and that AHR activation results in an upregulation of IL-8 and a downregulation of IL-9 production. Thus, we have identified IL-8 and IL-9 producing T helper cells which are subject to regulation by the engagement of the AHR.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Linfócitos T CD4-Positivos/metabolismo , Interleucina-8/metabolismo , Interleucina-9/metabolismo , Receptores de Hidrocarboneto Arílico/metabolismo , Células Th17/citologia , Doenças Autoimunes/metabolismo , Técnicas de Cultura de Células , Diferenciação Celular , Citocinas/metabolismo , Homeostase , Humanos , LigantesRESUMO
In dogs a CD4(+)CD8(+) double positive T cell subpopulation exists that has not been phenotypically defined yet. We demonstrate that canine CD4(+)CD8(+) T cells are mature CD1a(-) and TCRαß(+) T cells. To analyse the activation potential of CD4(+)CD8(+) T cells, PBMC from dogs vaccinated against canine distemper virus (CDV) were re-stimulated with CDV. Upon antigen-specific stimulation, the CD4(+)CD8(+) T cell fraction increases and consists nearly exclusively of proliferated cells. Similarly, other features of activated effector/memory T cells such as up-regulation of CD25 and MHC-II as well as down-regulation of CD62L (L-selectin) were observed in CD4(+)CD8(+) T cells after stimulation. Canine CD4(+)CD8(+) T cells are less abundant, but more heterogeneous than porcine ones, comprising a small proportion expressing the ß chain of CD8 in addition to the CD8α chain, like human CD4(+)CD8(+) T cells. In summary, this analysis provides the basis for functional characterisation of the in vivo relevance of CD4(+)CD8(+) T cells in T-cell mediated immunity.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Cães/imunologia , Animais , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD8-Positivos/citologia , Vírus da Cinomose Canina/imunologia , Cães/sangue , Feminino , Citometria de Fluxo/veterinária , Antígenos de Histocompatibilidade Classe II/sangue , Imunofenotipagem/veterinária , Subunidade alfa de Receptor de Interleucina-2/sangue , Selectina L/sangue , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/imunologia , Ativação Linfocitária/imunologia , MasculinoRESUMO
BACKGROUND: Epidemiological studies suggest an association between exposure to volatile organic compounds (VOCs) and adverse allergic and respiratory symptoms. However, whether VOCs exhibit a causal role as adjuvants in asthma development remains unclear. METHODS: To investigate the effect of VOC exposure on the development of allergic airway inflammation Balb/c mice were exposed to VOCs emitted by new polyvinylchloride (PVC) flooring, sensitized with ovalbumin (OVA) and characterized in acute and chronic murine asthma models. Furthermore, prevalent evaporated VOCs were analyzed and mice were exposed to selected single VOCs. RESULTS: Exposure of mice to PVC flooring increased eosinophilic lung inflammation and OVA-specific IgE serum levels compared to un-exposed control mice. The increased inflammation was associated with elevated levels of Th2-cytokines. Long-term exposure to PVC flooring exacerbated chronic airway inflammation. VOCs with the highest concentrations emitted by new PVC flooring were N-methyl-2-pyrrolidone (NMP) and 2,2,4-trimethyl-1,3-pentanediol diisobutyrate (TXIB). Exposure to NMP or TXIB also increased the allergic immune response in OVA-sensitized mice. In vitro or in vivo exposure to NMP or TXIB reduced IL-12 production in maturing dendritic cells (DCs) and enhanced airway inflammation after adoptive DC transfer into Balb/c mice. At higher concentrations both VOCs induced oxidative stress demonstrated by increased isoprostane and glutathione-S-transferase-pi1 protein levels in the lung of non-sensitized mice. Treatment of PVC flooring-exposed mice with N-acetylcysteine prevented the VOC-induced increase of airway inflammation. CONCLUSIONS: Our results demonstrate that exposure to VOCs may increase the allergic immune response by interfering with DC function and by inducing oxidative stress and has therefore to be considerate as risk factor for the development of allergic diseases.
Assuntos
Asma/imunologia , Glicóis/efeitos adversos , Pneumonia/induzido quimicamente , Pirrolidinonas/efeitos adversos , Compostos Orgânicos Voláteis/efeitos adversos , Acetilcisteína/uso terapêutico , Poluição do Ar em Ambientes Fechados/efeitos adversos , Animais , Células Dendríticas/imunologia , Modelos Animais de Doenças , Feminino , Pisos e Cobertura de Pisos , Interleucina-12/biossíntese , Pulmão/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Ovalbumina/imunologia , Estresse Oxidativo/efeitos dos fármacos , Cloreto de Polivinila/efeitos adversos , Células Th2/imunologiaRESUMO
RATIONALE: Epidemiological studies have shown that indoor molds are associated with increased prevalence and exacerbation of respiratory symptoms and asthma. Mycotoxins, secondary metabolites of molds, may contribute to these effects. OBJECTIVES: To investigate the adjuvant activity of mycotoxins on allergic airway inflammation. METHODS: Balb/c mice were exposed via the airways to gliotoxin and via the intestine to patulin, sensitized with ovalbumin (OVA), and then analyzed in acute and chronic murine asthma models. In addition, the effect of mycotoxin exposure on dendritic cell (DC) function was investigated using murine bone marrow-derived DCs. MEASUREMENTS AND MAIN RESULTS: Exposure of mice to both mycotoxins enhanced dose-dependently airway hyperreactivity, eosinophilic lung inflammation, and OVA-specific IgE serum levels compared with mice that received only the antigen. These findings correlated with increased Th2 cytokine levels and decreased IFN-gamma production. Long-term mycotoxin exposure exacerbated chronic airway inflammation and airway remodeling. In vitro or in vivo mycotoxin exposure inhibited IL-12 production in maturing DCs and enhanced airway inflammation after adoptive DC transfer into Balb/c mice. Mycotoxin exposure enhanced OVA-induced lung lipid peroxidation and moderately increased isoprostane levels in naive mice. Treatment of mycotoxin-exposed DCs with the antioxidants N-acetylcysteine or glutathione ethyl ester restored IL-12 secretion and pretreatment of exposed mice with N-acetylcysteine prevented the mycotoxin-induced increase of airway inflammation and AHR. CONCLUSIONS: Our results demonstrate that gliotoxin and patulin increase the allergic immune response in mice by modulating the Th1/Th2 balance via direct effects on IL-12 secretion in DCs and by inducing oxidative stress.
Assuntos
Asma/imunologia , Gliotoxina/toxicidade , Imunossupressores/toxicidade , Patulina/toxicidade , Venenos/toxicidade , Acetilcisteína/farmacologia , Actinas/metabolismo , Remodelação das Vias Aéreas/efeitos dos fármacos , Remodelação das Vias Aéreas/imunologia , Animais , Líquido da Lavagem Broncoalveolar/citologia , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Modelos Animais de Doenças , Eosinófilos/metabolismo , Sequestradores de Radicais Livres/farmacologia , Glutationa/análogos & derivados , Glutationa/efeitos dos fármacos , Glutationa/metabolismo , Glutationa/farmacologia , Imunoglobulina E/sangue , Interferon gama/metabolismo , Interleucina-12/biossíntese , Interleucina-12/imunologia , Interleucinas/metabolismo , Isoprostanos/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Pulmão/metabolismo , Linfócitos/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , NF-kappa B/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Pletismografia Total , Antígeno Nuclear de Célula em Proliferação/metabolismo , Células Th2/imunologia , Células Th2/metabolismo , Translocação Genética/efeitos dos fármacos , Proteínas Quinases p38 Ativadas por Mitógeno/efeitos dos fármacosRESUMO
IL-23 is a key cytokine in promotion of chronic inflammation. Here, we address if its pro-inflammatory potential can be harnessed to protect against chronic cryptococcosis. Mice were infected with Cryptococcus neoformans and treated with recombinant IL-23. Administration of IL-23 led to prolonged survival and reduced fungal burden but was inferior to IL-12 treatment. Independent of endogenous IL-23/IL-12, IL-23 treatment induced an altered cytokine profile accompanied by marked changes in composition of the inflammatory infiltrate characterized by T cell and dendritic cell recruitment. Although IL-23 induced hallmarks of the T(h)17 pathway, also non-T cells produced IL-17A and IL-22. IL-23 treatment of T-cell-deficient mice resulted in increased IL-17A and IL-22 production and modulation of the cellular response at the site of infection with elevated expression of CD86 on macrophages. Our data show that IL-23 treatment induces innate and adaptive tissue inflammation with limited impact on resistance to chronic cryptococcosis.
Assuntos
Imunidade Adaptativa/efeitos dos fármacos , Antifúngicos/administração & dosagem , Criptococose/tratamento farmacológico , Cryptococcus neoformans/patogenicidade , Imunidade Inata/efeitos dos fármacos , Interleucina-23/administração & dosagem , Animais , Células Cultivadas , Criptococose/imunologia , Criptococose/microbiologia , Citocinas/metabolismo , Proteínas de Ligação a DNA/deficiência , Proteínas de Ligação a DNA/genética , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Células Dendríticas/microbiologia , Modelos Animais de Doenças , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Mediadores da Inflamação/metabolismo , Interleucina-12/administração & dosagem , Interleucina-12/deficiência , Interleucina-12/genética , Subunidade p40 da Interleucina-12/deficiência , Subunidade p40 da Interleucina-12/genética , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/microbiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Recombinantes/administração & dosagem , Linfócitos T Auxiliares-Indutores/efeitos dos fármacos , Linfócitos T Auxiliares-Indutores/imunologia , Linfócitos T Auxiliares-Indutores/microbiologia , Fatores de TempoRESUMO
We have recently shown that inactivated parapoxvirus ovis (iPPVO) effectively stimulates canine blood phagocytes. However, a potential link between innate and adaptive immunity induced by iPPVO remained open. The objective of this study was to define the effects of repeated iPPVO treatment of dogs to evaluate (i) iPPVO-specific antibody production, and (ii) modulation of iPPVO-induced oxidative burst by anti-iPPVO antibodies. Serum analysis of dogs treated repeatedly with iPPVO (Zylexis) showed transient production of non-neutralising iPPVO-specific IgG. There was a correlation between iPPVO-specific IgG levels and enhanced oxidative burst rates in vitro upon transfer of immune sera. Even four years after Zylexis treatment considerably stronger oxidative burst rates in response to iPPVO were observed in monocytes and PMN, whereas only moderate burst rates were detected in monocytes, but not in PMN, from dogs treated with a placebo. Depletion of serum IgG by protein A-sepharose or by parapoxvirus ovis coupled to sepharose abolished the increase of oxidative burst responses and resulted in burst rates similar to blood leukocytes from control dogs. However, uptake of viral particles was found to be independent of iPPVO-specific IgG and restricted to cells with dendritic and monocytic morphology. These data demonstrate that non-neutralising iPPVO-specific IgG is produced during treatment with Zylexis. Moreover, for the first time the interaction of iPPVO with antibodies is shown to enhance oxidative burst.
Assuntos
Antígenos Virais/imunologia , Leucócitos Mononucleares/imunologia , Neutrófilos/imunologia , Parapoxvirus/imunologia , Infecções por Poxviridae/veterinária , Explosão Respiratória/imunologia , Vacinas Virais/imunologia , Animais , Anticorpos Antivirais/sangue , Bovinos , Linhagem Celular , Células Cultivadas , Cães , Soros Imunes/imunologia , Imunidade Inata/imunologia , Imunização Passiva , Imunoglobulina G/sangue , Infecções por Poxviridae/imunologia , Infecções por Poxviridae/virologiaRESUMO
Early activation of the IL-12/IFN-gamma axis has been shown following Salmonella enterica serovar Enteritidis (S. Enteritidis) infection. We were interested to study whether IL-22 and IL-17A production is initiated early in response to S. Enteritidis. We demonstrate here that IL-22 was strongly elevated in the peritoneal lavage fluid and in serum already 1 day post-intraperitoneal infection (d.p.i.) of mice; not only IL-22 but also IL-17A was produced ex vivo by activated peritoneal exudate cells (PEC). Peritoneal gammadelta T cells were identified as cellular source of IL-17A. The early IL-22 production was completely IL-23-dependent. In contrast, IL-17A production was only partially IL-23-dependent. To investigate the local production of upstream cytokines important for induction of IL-22, IL-17A and IFN-gamma during salmonellosis, the production of IL-23 and IL-12 was studied. Elevated p19 and p40 mRNA levels were found in PEC at 1 d.p.i., whereas p35 mRNA levels were not changed. Besides, the T(h)17-promoting cytokines IL-6, IL-1beta and transforming growth factor-beta were produced in response to S. Enteritidis. However, IL-6 was not required for IL-22 or IL-17A production by PEC. By ex vivo analysis of PEC at 1 d.p.i., we show that the major producers of early IL-12/23p40 in the peritoneal cavity were dendritic cells (DC), whereas macrophages notably contributed to IL-6 production. Taken together, these data suggest that DC initiate early IL-22 production at the site of infection which may contribute to resistance against salmonellosis. Furthermore, we provide evidence that production of IL-22 and IL-17A is differentially regulated during infection.
Assuntos
Células Dendríticas/imunologia , Macrófagos/imunologia , Infecções por Salmonella/imunologia , Linfócitos T/imunologia , Animais , Células Dendríticas/metabolismo , Imunidade Inata , Interferon gama/imunologia , Interferon gama/metabolismo , Interleucina-12/genética , Interleucina-12/imunologia , Interleucina-12/metabolismo , Interleucina-17/imunologia , Interleucina-17/metabolismo , Interleucina-23/genética , Interleucina-23/imunologia , Interleucina-23/metabolismo , Interleucina-6/genética , Interleucina-6/imunologia , Interleucina-6/metabolismo , Interleucinas/imunologia , Interleucinas/metabolismo , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores de Antígenos de Linfócitos T gama-delta/imunologia , Infecções por Salmonella/microbiologia , Salmonella enteritidis/imunologia , Linfócitos T/metabolismo , Interleucina 22RESUMO
Inactivated parapoxvirus ovis (iPPVO) shows strong immunomodulatory activities in several species and is used in veterinary medicine as an immunostimulatory biological for the prevention and/or treatment of infectious diseases. In this study the immunostimulatory capacity of iPPVO on the innate immune system was investigated in vitro by the evaluation of induction of the oxidative burst and modulation of phagocytosis by canine blood leukocytes (polymorphonuclear cells and monocytes) of dogs. In addition, the activation of canine T lymphocytes was also studied. After stimulation with iPPVO the phagocytosis of FITC-labeled Listeria monocytogenes was increased in canine blood monocytes and neutrophils. Enhanced burst rates by canine monocytes stimulated with iPPVO were observed and the MHC-II expression on canine CD14+ monocytes was elevated following stimulation with iPPVO compared to the stabiliser control. Canine CD4+ T cells were activated for oligoclonal proliferation in response to iPPVO. This study shows that iPPVO is able to stimulate both phagocytotic and T-cell-dependent immune mechanisms in canine blood leukocytes.
Assuntos
Ativação Linfocitária/fisiologia , Parapoxvirus/fisiologia , Fagócitos/fisiologia , Linfócitos T/fisiologia , Animais , Proliferação de Células , Células Cultivadas , Cães , Genes MHC da Classe II/genética , Genes MHC da Classe II/fisiologia , Fagocitose/fisiologia , Explosão Respiratória , Regulação para Cima , Inativação de VírusRESUMO
IL-12 is essential for protective T cell-mediated immunity against Salmonella infection. To characterize the role of the related cytokine IL-23, wild-type (WT) C57BL/6 and p19(-/-) mice were infected systemically with an attenuated strain of Salmonella enterica serovar Enteritidis (S. Enteritidis). IL-23-deficient mice controlled infection with S. Enteritidis similarly as WT mice. Similar IFN-gamma production as compared with WT mice, but defective IL-17A and IL-22 production was found in the absence of IL-23. Nevertheless, although IL-23 is required for T cell-dependent cytokine responses, IL-23 is dispensable for protection against S. Enteritidis when IL-12 is present. To analyze the role of IL-23 in the absence of IL-12, low doses of S. Enteritidis were administered to p35(-/-) mice (lacking IL-12), p35/19(-/-) mice (lacking IL-12 and IL-23), p35/40(-/-) mice (lacking IL-12, IL-23, and homodimeric IL-12p40), or p35/IL-17A(-/-) mice (lacking IL-12 and IL-17A). We found survival of p35(-/-) and p35/IL-17A(-/-) mice, whereas p35/19(-/-) and p35/40(-/-) mice died within 3-6 wk and developed liver necrosis. This indicates that IL-23, but not homodimeric IL-12p40, is required for protection, which, surprisingly, is independent of IL-17A. Moreover, protection was associated with IL-22, but not IL-17F or IL-21 expression or with neutrophil recruitment. Finally, anti-IL-22 treatment of S. Enteritidis-infected p35(-/-) mice resulted in liver necrosis, indicating a central role of IL-22 in hepatocyte protection during salmonellosis. In conclusion, IL-23-dependent IL-22, but not IL-17 production is associated with protection against systemic infection with S. Enteritidis in the absence of IL-12.
Assuntos
Subunidade p40 da Interleucina-12/imunologia , Interleucina-17/imunologia , Interleucina-23/imunologia , Interleucinas/imunologia , Infecções por Salmonella/imunologia , Salmonella enteritidis/imunologia , Animais , Feminino , Hepatócitos/imunologia , Interferon gama/genética , Interferon gama/imunologia , Subunidade p40 da Interleucina-12/genética , Interleucina-17/genética , Interleucina-23/genética , Interleucinas/genética , Fígado/imunologia , Camundongos , Camundongos Knockout , Necrose/genética , Necrose/imunologia , Neutrófilos/imunologia , Infecções por Salmonella/genética , Interleucina 22RESUMO
In this review, we summarize our investigations concerning the differential importance of CD14 and LBP in toll-like receptor 4 (TLR4)/myeloid differentiation protein-2 (MD-2)-mediated signaling by smooth and rough-form lipopolysaccharide (LPS) chemotypes and include the results obtained in studies with murine and human TLR4-transgenic mice. Furthermore, we present more recent data on the mechanisms involved in the induction of LPS hypersensitivity by bacterial and viral infections and on the reactivity of the hypersensitive host to non-LPS microbial ligands and endogenous mediators. Finally, the effects of pre-existing hypersensitivity on the course and outcome of a super-infection with Salmonella typhimurium or Listeria monocytogenes are summarized.