Impact of Bacille Calmette-Guérin revaccination on serum IgE levels in a randomized controlled trial

Abstract INTRODUCTION: Bacille Calmette-Guérin (BCG) downmodulates allergen-specific IgE levels and prevents other atopic responses in experimental models but fails to protect against respiratory allergies. Human responsiveness to BCG is variable and may interfere with protection. METHODS: Multivariate models were evaluated to test the possible effect of responsiveness (assessed by IFN-γ production) to BCG revaccination on the modulation of total and allergen-specific serum IgE levels in healthy volunteers participating in a randomized controlled trial. RESULTS: Serum total or Derp-specific IgE levels did not change regardless of the increase in IFN-γ levels. CONCLUSIONS: BCG responsiveness does not affect protection against atopy.

Comparative study of lymphocytes from individuals that were vaccinated and unvaccinated against the pandemic 2009-2011 H1N1 influenza virus in Southern Brazil

ABSTRACTINTRODUCTION:While no single factor is sufficient to guarantee the success of influenza vaccine programs, knowledge of the levels of immunity in local populations is critical. Here, we analyzed influenza immunity in a population from Southern Brazil, a region with weather conditions that are distinct from those in the rest of country, where influenza infections are endemic, and where greater than 50% of the population is vaccinated annually.METHODS:Peripheral blood mononuclear cells were isolated from 40 individuals. Of these, 20 had received the H1N1 vaccine, while the remaining 20 were unvaccinated against the disease. Cells were stimulated in vitro with the trivalent post-pandemic influenza vaccine or with conserved major histocompatibility complex I (MHC I) peptides derived from hemagglutinin and neuraminidase. Cell viability was then analyzed by [3-(4,5-dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide)]-based colorimetric assay (MTT), and culture supernatants were assayed for helper T type 1 (Th1) and Th2-specific cytokine levels.RESULTS:Peripheral blood lymphocytes from vaccinated, but not unvaccinated, individuals exhibited significant proliferation in vitro in the presence of a cognate influenza antigen. After culturing with vaccine antigens, cells from vaccinated individuals produced similar levels of interleukin (IL)-10 and interferon (IFN)-γ, while those from unvaccinated individuals produced higher levels of IFN-γ than of IL-10.CONCLUSIONS:Our data indicate that peripheral blood lymphocytes from vaccinated individuals are stimulated upon encountering a cognate antigen, but did not support the hypothesis that cross-reactive responses related to previous infections can ameliorate the immune response. Moreover, monitoring IL-10 production in vaccinated individuals could comprise a valuable tool for predicting disease evolution.

Producción in vitro de interferón gamma en ratones vacunados con ´Mycobacterium habana´ frente a antígenos de Mycobacterium tuberculosis / In vitro interferon gamma production in mice vaccinated with ´Mycobacterium habana´ against Mycobacterium tuberculosis antigens

Introducción: el desarrollo de nuevas vacunas antituberculosas requiere de la caracterización de la respuesta de inmunidad celular, inducida por el nuevo candidato vacunal frente a los antígenos principales de Mycobacterium tuberculosis. Objetivo: determinar el potencial inmunogénico de ´Mycobacterium habana´ TMC-5135, cuando se usa como vacuna subcutánea en ratones Balb/c. Métodos: en este estudio se inocularon subcutáneamente ratones Balb/c con la cepa viva ´Mycobacterium habana´ TMC-5135 y se determinó la producción in vitro de IFN gamma en cultivos celulares de pulmón, bazo y ganglios inguinales estimulados con antígenos solubles totales y el antígeno 85b. Como grupo control se vacunaron ratones con BCG subcepa Phipps. Resultados: particularmente en los ganglios linfáticos inguinales, ambos antígenos indujeron mayor producción de IFN gamma en los ratones vacunados con ´Mycobacterium habana´que con BCG. Conclusiones: los resultados justifican la realización de nuevas investigaciones usando ´Mycobacterium habana´ TMC-5135 como candidato vacunal para prevenir la tuberculosis.

Introduction: development of new antituberculosis vaccines requires the characterization of the cell-mediated immune responses induced by mycobacterial antigens. Objective: to determine the immunogenic potential of ´Mycobacterium habana´ TMC-5135 when using subcutaneous vaccine in Balb/c mice. Methods: in this study, Balb/c mice were inoculated subcutaneously with live ´Mycobacterium habana´ TMC-5135. The production of IFN gamma in cell suspensions obtained from the lungs, the spleen and the lymph nodes after stimulation with mycobacterial antigens Ag85b or culture filtrate antigens (CFA) was recorded. Results: the production of IFN gamma after stimulation with CFA and Ag85b was higher in mice vaccinated with ´M. habana´ than in animals immunized with BCG. Conclusions: these results encourage new research on ´M. habana´ as vaccinal candidate against tuberculosis.

Interferon-gamma and interleukin-10 production by mononuclear cells from patients with advanced head and neck cancer

OBJECTIVE: This study aims to evaluate the production of interferon-gamma and interleukin-10 by stimulated peripheral blood mononuclear cells isolated from patients with supraglottic laryngeal cancer before and after surgical treatment. METHODS: Fourteen patients with advanced supraglottic laryngeal cancer were studied. Cultures of peripheral blood mononuclear cells isolated during the preoperative and late postoperative periods were stimulated with concanavalin A and Bacille Calmette-Guerin, and the supernatant concentrations of interferon-gamma and interleukin-10 were measured. RESULTS: For non-stimulated cultures, the interferon-gamma levels produced by the preoperative period and the late postoperative period cultures were lower than the levels produced by the control group cultures. The interferon-gamma levels after stimulation with concanavalin A were higher in the late postoperative period cultures than in the preoperative evaluation cultures. Stimulation with Bacille Calmette-Guerin led to the production of similar levels of interferon-gamma and interleukin-10 by all cultures; thus, stimulation increased the levels of interferon-gamma produced by both the preoperative and postoperative cultures relative to the levels produced by the corresponding unstimulated cultures. CONCLUSION: Patients with advanced supraglottic laryngeal cancer exhibit an in vitro deficiency in interferongamma secretion by mononuclear cells. Stimulated cells seem to recover this function during the postoperative period.

Expresión de IL-10, IL-4 e IFN- γ en lesiones activas de piel en niños con urticaria papular por picadura de pulga / Expression of IL-10, IL-4 and IFN-γ in active skin lesions of children with papular urticaria

Introducción. La urticaria papular por picadura de pulga se conoce como una enfermedad alérgica. Sin embargo, las investigaciones no muestran una clara relación con las enfermedades alérgicas. Objetivo. Estudiar la expresión de IL-10, IL-4 e IFN-γ, como marcadores de la respuesta efectora de células T en lesiones de piel de pacientes con urticaria papular por picadura de pulga. Materiales y métodos. Se incluyeron 14 biopsias de lesiones de piel de niños con diagnóstico de urticaria papular por picadura de pulga y 5 biopsias de piel sana obtenidas de niños sometidos a cirugía por enfermedades no inflamatorias. Todas las muestras se obtuvieron de niños menores de 12 años. Se extrajo ARN con trizol y se cuantificaron los niveles de expresión de las citocinas con la técnica de reacción en cadena de la polimerasa en tiempo real. Resultados. En los pacientes con urticaria papular por picadura de pulga, se encontró amplia diversidad en los niveles de expresión de IFN-γ e IL-10, y valores bajos constantes para IL-4. Se observaron tres perfiles que no corresponden a un patrón común en los pacientes. Las muestras obtenidas de tejidos sanos no presentaron expresión de las citocinas. Conclusiones. Los datos corresponden a la primera descripción de citocinas que median la respuesta inmunitaria en el sitio de la lesión cutánea en niños con con urticaria papular por picadura de pulga, lo cual indica que la respuesta local es mixta ya que no se encuentra predominio de un fenotipo específico en ninguno de los pacientes.

Introduction: Papular urticaria caused by the bites of fleas traditionally has been defined as a chronic allergic disease. However, currently no clear relationship has been described between this pathology and common allergic diseases. Objective: The expression of IL-10, IL-4 and IFN-γ as markers of effector T cell responses was examined in skin lesions of patients with papular urticaria by flea bite. Materials and methods: Fourteen skin lesion biopsies were sampled from children with a clinical diagnosis of papular urticaria by flea bite and were compared with 5 healthy skin biopsies of children with no history of the disease. All children were under 12 years old. RNA was extracted with trizol and the expression levels of cytokines were analyzed by real time PCR technique. Results: A wide range in the expression levels of IFN-γ and IL-10 was noted as well as constant low values of IL-4. Three distinct profiles were observed, but which did not correspond to a recognizable pattern among the patients. The samples obtained from healthy tissues showed no expression of any of the cytokines. Conclusions: This is the first characterization of cytokines that mediate the immune response at the site of the skin lesion in children with papular urticaria by flea bite. The data indicated that the local response was mixed and that a single phenotype is not predominant among the patients.

Schistosoma mansoni antigens alter the cytokine response in vitro during cutaneous leishmaniasis

Schistosoma mansoni infection or associated products are able to down-modulate the type 1 CD4+ T cell inflammatory response characteristic of autoimmune diseases. In this study, we evaluated how S. mansoni antigens altered the immune response that was induced by the soluble Leishmania antigen (SLA) from cutaneous leishmaniasis (CL) patients. Cytokines were measured from the supernatants of peripheral blood mononuclear cell cultures stimulated with SLA. This was performed using the sandwich enzyme linked immunosorbent assay technique in the presence or absence of S. mansoni recombinant antigens Sm29, SmTSP-2 and PIII. The addition of S. mansoni antigens to the cultures resulted in the reduction of interferon gamma (IFN-γ) levels in 37-50 percent of patients. Although to a lesser extent, the antigens were also able to decrease the production of tumour necrosis factor-alpha (TNF-α). We compared patients that either had or did not have reduction in IFN-γ and TNF-α production in cultures stimulated with SLA in the presence of S. mansoni antigens. We found that there was no significant difference in the levels of interleukin (IL)-10 and IL-5 in response to S. mansoni antigens between the groups. The antigens used in this study down-modulated the in vitro proinflammatory response induced by SLA in a group of CL patients through a currently undefined mechanism.

Effect of TNF-α production inhibitors on the production of pro-inflammatory cytokines by peripheral blood mononuclear cells from HTLV-1-infected individuals

Human T lymphotropic virus type 1 (HTLV-1) is the causal agent of myelopathy/tropical spastic paraparesis (HAM/TSP), a disease mediated by the immune response. HTLV-1 induces a spontaneous proliferation and production of pro-inflammatory cytokines by T cells, and increasing interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α) levels are potentially involved in tissue damage in diseases related to HTLV-1. This exaggerated immune response is also due to an inability of the natural regulatory mechanisms to down-modulate the immune response in this group of patients. TNF-α inhibitors reduce inflammation and have been shown to improve chronic inflammatory diseases in clinical trials. The aim of this study was to evaluate the ability of pentoxifylline, forskolin, rolipram, and thalidomide to decrease in vitro production of TNF-α and IFN-γ in cells of HTLV-1-infected subjects. Participants of the study included 19 patients with HAM/TSP (mean age, 53 ± 11; male:female ratio, 1:1) and 18 HTLV-1 carriers (mean age, 47 ± 11; male:female ratio, 1:2.6). Cytokines were determined by ELISA in supernatants of mononuclear cell cultures. Pentoxifylline inhibited TNF-α and IFN-γ synthesis with the minimum dose used (50 µM). The results with forskolin were similar to those observed with pentoxifylline. The doses of rolipram used were 0.01-1 µM and the best inhibition of TNF-α production was achieved with 1 µM and for IFN-γ production it was 0.01 µM. The minimum dose of thalidomide used (1 µM) inhibited TNF-α production but thalidomide did not inhibit IFN-γ production even when the maximum dose (50 µM) was used. All drugs had an in vitro inhibitory effect on TNF-α production and, with the exception of thalidomide, all of them also decreased IFN-γ production.

Influence of GB virus C on IFN-γ and IL-2 production and CD38 expression in T lymphocytes from chronically HIV-infected and HIV-HCV-co-infected patients

This study was designed to assess the effect of GB virus (GBV)-C on the immune response to human immunodeficiency virus (HIV) in chronically HIV-infected and HIV- hepatitis C virus (HCV)-co-infected patients undergoing antiretroviral therapy. A cohort of 159 HIV-seropositive patients, of whom 52 were HCV-co-infected, was included. Epidemiological data were collected and virological and immunological markers, including the production of interferon gamma (IFN-γ) and interleukin (IL)-2 by CD4, CD8 and Tγδ cells and the expression of the activation marker, CD38, were assessed. A total of 65 patients (40.8 percent) presented markers of GBV-C infection. The presence of GBV-C did not influence HIV and HCV replication or TCD4 and TCD8 cell counts. Immune responses, defined by IFN-γ and IL-2 production and CD38 expression did not differ among the groups. Our results suggest that neither GBV-C viremia nor the presence of E2 antibodies influence HIV and HCV viral replication or CD4 T cell counts in chronically infected patients. Furthermore, GBV-C did not influence cytokine production or CD38-driven immune activation among these patients. Although our results do not exclude a protective effect of GBV-C in early HIV disease, they demonstrate that this effect may not be present in chronically infected patients, who represent the majority of patients in outpatient clinics.

Immune and inflammatory responses to Leishmania amazonensis isolated from different clinical forms of human leishmaniasis in CBA mice

Leishmania amazonensis causes different diseases depending on the host and parasitic virulence factors. In this study, CBA mice were infected with L. amazonensis isolates from patients with localized (Ba125), diffuse cutaneous (Ba276) or visceral leishmaniasis (Ba109). Mice infected with Ba125 and Ba276 progressed rapidly and lesions displayed an infiltrate rich in parasitized macrophages and were necrotic and ulcerated. Ba109 induced smaller lesions and a mixed inflammatory infiltrate without necrosis or ulceration. Ba109 induced an insidious disease with lower parasite load in CBA mice, similar to human disease. Levels of IFN-γ, IL-4 and IL-10 did not differ among the groups. Because all groups were unable to control the infection, expression of IL-4 associated with low production of IFN-γ in the early phase of infection may account for susceptibility, but others factors may contribute to the differences observed in inflammatory responses and infection progression. Evaluation of some parasitic virulence factors revealed that Ba276 exhibits higher ecto-ADPase and 5'-nucleotidase activities compared to the Ba109 and Ba125 strains. Both Ba276 and Ba125 had higher arginase activity in comparison to Ba109. Finally, these data suggest that the differences in enzyme activities among parasites can account for differences in host inflammatory responses and infection progression.

Protective effects of basic fibroblast growth factor in the development of emphysema induced by interferon-gamma

Recent clinical evidence indicates that the non-eosinophilic subtype of severe asthma is characterized by fixed airway obstruction, which may be related to emphysema. Transgenic studies have demonstrated that high levels of IFN-gamma in the airways induce emphysema. Fibroblast growth factor 2 (FGF2), which is the downstream mediator of TGF-beta, is important in wound healing. We investigated the role of FGF2 in IFN-gamma-induced emphysema and the therapeutic effects of recombinant FGF2 in the prevention of emphysema in a severe non-eosinophilic asthma model. To evaluate the role of FGF2 in IFN-gamma-induced emphysema, lung targeted IFN-gamma transgenic mice were cross-bred with FGF2-deficient mice. A severe non-eosinophilic asthma model was generated by airway application of LPS-containing allergens twice a week for 4 weeks. To evaluate protective effects of FGF2, recombinant FGF2 (10 microg) was injected subcutaneously during allergen challenge in the severe asthma model. We found that non-eosinophilic inflammation and emphysema induced by transgenic overexpression of IFN-gamma in the airways were aggravated by the absence of FGF2. Airway challenge with LPS-containing allergens induced more inflammation in mice sensitized with LPS-containing allergens compared to challenge with allergens alone. In addition, LPS-induced lung inflammation and emphysema depended on IFN-gamma but not on IL-13. Interestingly, emphysema in the severe asthma model was significantly inhibited by treatment with recombinant FGF2 during allergen challenge, whereas lung inflammation was unaffected. Therefore, our present data suggest that FGF2 may help protect against IFN-gamma-induced emphysema, and that recombinant FGF2 may help lessen the severity of emphysema.

Heterogeneidade de resposta por IFN-γ a cepas clínicas de Mycobacterium tuberculosis em humanos / Heterogeneity in human IFN-γ responses to clinical Mycobacterium tuberculosis strains

Mycobacterium tuberculosis é um dos mais bem sucedidos patógenos do homem. As cepas virulentas são mais facilmente transmitidas, induzindo respostas imunes variáveis. Avaliamos a resposta celular tipo Th1, através da produção de IFN-γ, como resposta a cepas com padrões diversos em voluntários sadios. Nossos resultados mostraram que indivíduos com teste tuberculínico (TT) negativo já tiveram contato com algumas das cepas testadas, ao passo que indivíduos com TT positivo não responderam a todas as cepas testadas. Cepas resistentes induziram uma média menor de produção de IFN-γ que aquelas sensíveis. Uma possível aplicação prática disto seria que a produção de IFN-γ, em relação a uma cepa isolada específica, poderia auxiliar na previsão da resposta ao tratamento dos pacientes.

Mycobacterium tuberculosis is one of the most successful human pathogens. Highly virulent strains, which are more easily transmitted than are less virulent strains, elicit variable immune responses. We evaluated the Th1 responses (IFN-γ production) in healthy volunteers after stimulation with various strains. Our results show that the individuals with negative tuberculin skin test (TST) results were not necessarily naive to all of the strains tested, whereas individuals with positive TST results did not respond to all of the strains tested. Drug-resistant strains induced a lower mean level of IFN-γ production than did drug-sensitive strains. One possible practical application of this finding would be for the prediction of responses to treatment, in which it might be advantageous to have knowledge of the estimated IFN-γ production elicited by a specific isolated strain.

Decreased production of TNF-alpha by lymph node cells indicates experimental autoimmune encephalomyelitis remission in Lewis rats

Experimental autoimmune encephalomyelitis (EAE) is mediated by CD4+ Th1 cells that mainly secrete IFN-γ and TNF-α, important cytokines in the pathophysiology of the disease. Spontaneous remission is, in part, attributed to the down regulation of IFN-γ and TNF-α by TGF-β. In the current paper, we compared weight, histopathology and immunological parameters during the acute and recovery phases of EAE to establish the best biomarker for clinical remission. Female Lewis rats were immunised with myelin basic protein (MBP) emulsified with complete Freund's adjuvant. Animals were evaluated daily for clinical score and weight prior to euthanisation. All immunised animals developed the expected characteristics of EAE during the acute phase, including significant weight loss and high clinical scores. Disease remission was associated with a significant reduction in clinical scores, although immunised rats did not regain their initial weight values. Brain inflammatory infiltrates were higher during the acute phase. During the remission phase, anti-myelin antibody levels increased, whereas TNF-α and IFN-γ production by lymph node cells cultured with MBP or concanavalin A, respectively, decreased. The most significant difference observed between the acute and recovery phases was in the induction of TNF-α levels in MBP-stimulated cultures. Therefore, the in vitro production of this cytokine could be used as a biomarker for EAE remission.

Tuberculose em pacientes com artrite reumatoide: a dificuldade no diagnóstico da forma latente / Tuberculosis in rheumatoid arthritis patients: the difficulty in making the diagnosis of latent infection

Desde o início do uso de drogas anti-TNF para o tratamento da artrite reumatoide e outras doenças inflamatórias, casos de tuberculose pulmonar e extrapulmonar vêm sendo notificados em pacientes submetidos a tal tratamento. Na maioria das vezes, a doença se desenvolve durante as seis primeiras infusões. Todo paciente deve ser avaliado para tuberculose latente antes do início do uso de um bloqueador de TNF; no entanto, o diagnóstico de tuberculose latente é um desafio. A prova tuberculínica, o único teste disponível para a detecção de tuberculose latente por quase um século, apresenta uma série de limitações. Testes baseados na detecção da produção de IFN-γ in vitro por células mononucleares ativadas por antígenos específicos parecem ser mais acurados e vêm sendo pesquisados em pacientes com artrite reumatoide.

Since the beginning of the use of anti-TNF in the treatment of rheumatoid arthritis and other inflammatory diseases, cases of pulmonary tuberculosis and extrapulmonary tuberculosis have been reported in patients receiving such treatment. In most cases, the disease develops by the time the patient has received the sixth infusion. Every patient should be evaluated for latent tuberculosis infection prior to the use of a TNF inhibitor. However, the diagnosis of latent tuberculosis infection is a challenge. The tuberculin test, which was the only test available to detect latent tuberculosis infection for nearly a century, presents a number of limitations. Tests based on the detection of the in-vitro production of IFN-γ by mononuclear cells activated by specific antigens appear to be more accurate and have been studied in patients with rheumatoid arthritis.

O desafio da tuberculose na faixa etária pediátrica frente a novas técnicas diagnósticas: [revisão] / The challenge of pediatric tuberculosis in face of new diagnostic techniques: [review]

OBJETIVOS: Apresentar uma revisão atualizada sobre os novos métodos para o diagnóstico da tuberculose baseados na produção in vitro de interferon-gama (IFN-γ) por células T dos pacientes sob investigação, comparando-os com a tradicional prova tuberculínica. FONTES DOS DADOS: Revisão de literatura utilizando os bancos de dados MEDLINE e LILACS (2000-2008) utilizando as palavras-chave tuberculose, interferon-gama, quantiFERON, ELISPOT e T-SPOT.TB. SÍNTESE DOS DADOS: Esses novos testes mostraram-se, de um modo geral, tão ou mais sensíveis e específicos que a prova tuberculínica, tanto em adultos como em crianças e imunossuprimidos, para o diagnóstico da infecção latente e da doença ativa, apresentando vantagens como a menor interferência da vacinação prévia pelo BCG, menor influência de estados anérgicos e melhor acurácia em crianças menores. Nos Estados Unidos, já estão sendo utilizados em substituição à prova tuberculínica, e apesar dos custos ainda elevados, a Organização Mundial de Saúde vai priorizar a sua viabilidade econômica. CONCLUSÕES: Sempre levando em conta a importância da história clínica e epidemiológica, os novos testes baseados na produção de IFN-γ apresentam resultados promissores e deverão ser considerados na investigação de tuberculose em qualquer paciente, mas especialmente nos grupos de risco, como as crianças e os imunossuprimidos.

OBJECTIVES: To present an updated review concerning new assays for diagnosing tuberculosis based on in vitro interferon-gamma production by host T cells, and compare them with tuberculin skin test. SOURCES: A literature review was carried out based on Medline and LILACS databases (2000-2008) searching for the following keywords: tuberculosis, interferon-gamma, quantiFERON, ELISPOT and T-SPOT.TB. SUMMARY OF THE FINDINGS: These new assays proved to have, in general, equal or superior sensitivity and specificity than the tuberculin skin test not only in adults but also in children and immunosuppressed patients for the diagnosis of both latent tuberculosis infection or active disease, with some advantages such as less cross-reactivity as a result of previous BCG vaccination, less influence of anergy and better accuracy in small children. In the United States, these assays have been used instead of the tuberculin skin test and, although still very expensive, the World Health Organization will be making its economic viability a priority. CONCLUSIONS: Always having in mind the importance of clinical and epidemiological histories, these new assays based on interferon-gamma release present promising results and should be considered in tuberculosis investigation procedures for all patients, however with a special concern in the risk groups (i.e., children and immunosuppressed patients).

Lactobacillus delbrueckii UFV-H2b20 induces type 1 cytokine production by mouse cells in vitro and in vivo

Lactobacillus delbrueckii UFV-H2b20 has been shown to increase clearance of bacteria injected into the blood of germ-free mice. Moreover, it induces the production of type 1 cytokines by human peripheral mononuclear cells. The objective of the present study was to investigate the production of inflammatory cytokines [interleukin-12 (IL-12 p40), tumor necrosis factor-α (TNF-α), and interferon-γ (IFN-γ)] triggered in vitro by live, heat-killed or lysozyme-treated L. delbrueckii UFV-H2b20 and in vivo by a live preparation. Germ-free, L. delbrueckii-monoassociated and lipopolysaccharide (LPS)-resistant C3H/HeJ mice were used as experimental models. UFV-H2b20 induced the production of IL-12 p40 and TNF-α by peritoneal cells and IFN-γ by spleen cells from germ-free or monoassociated Swiss/NIH mice and LPS-hyporesponsive mice (around 40 ng/mL for IL-12 p40, 200 pg/mL for TNF-α and 10 ng/mL for IFN-γ). Heat treatment of L. delbrueckii did not affect the production of these cytokines. Lysozyme treatment decreased IL-12 p40 production by peritoneal cells from C3H/HeJ mice, but did not affect TNF-α production by these cells or IFN-γ production by spleen cells from the same mouse strain. TNF-α production by peritoneal cells from Swiss/NIH L. delbrueckii-monoassociated mice was inhibited by lysozyme treatment. When testing IL-12 p40 and IFN-γ levels in sera from germ-free or monoassociated Swiss/NIH mice systemically challenged with Escherichia coli we observed that IL-12 p40 was produced at marginally higher levels by monoassociated mice than by germ-free mice (40 vs 60 ng/mL), but IFN-γ was produced earlier and at higher levels by monoassociated mice (monoassociated 4 and 14 ng/mL 4 and 8 h after infection, germfree 0 and 7.5 ng/mL at the same times). These results show that L. delbrueckii UFV-H2b20 stimulates the production of type 1 cytokines in vitro and in vivo, therefore suggesting...

Immunological imbalance between IFN-³ and IL-10 levels in the sera of patients with the cardiac form of Chagas disease

The immune response is crucial for protection against disease; however, immunological imbalances can lead to heart and digestive tract lesions in chagasic patients. Several studies have evaluated the cellular and humoral immune responses in chagasic patients in an attempt to correlate immunological findings with clinical forms of Chagas disease. Moreover, immunoglobulins and cytokines are important for parasitic control and are involved in lesion genesis. Here, cytokine and IgG isotype production were studied, using total epimastigote antigen on sera of chagasic patients with indeterminate (IND, n = 27) and cardiac (CARD, n = 16) forms of the disease. Samples from normal, uninfected individuals (NI, n = 30) were use as controls. The results showed that sera from both IND and CARD patients contained higher levels of Trypanosoma cruzi-specific IgG1 (IgG1) antibodies than sera from NI. No difference in IgG2 production levels was observed between NI, IND and CARD patients, nor was a difference in IL-10 and IFN-³ production detected in the sera of IND, CARD and NI patients. However, IND patients displayed a positive correlation between IL-10 and IFN-³ levels in serum, while CARD patients showed no such correlation, indicating an uncontrolled inflammatory response in CARD patients. These findings support the hypothesis that a lack of efficient regulation between IFN-³ and IL-10 productions in CARD patients may lead to cardiac immunopathology.

Recombinant in vitro assembled hepatitis C virus core particles induce strong specific immunity enhanced by formulation with an oil-based adjuvant

In the present work, immunogenicity of recombinant in vitro assembled hepatitis C virus core particles, HCcAg.120-VLPs, either alone or in combination with different adjuvants was evaluated in BALB/c mice. HCcAg.120-VLPs induced high titers of anti-HCcAg.120 antibodies and virus-specific cellular immune responses. Particularly, HCcAg.120-VLPs induced specific delayed type hypersensitivity, and generated a predominant T helper 1 cytokine pro file in immunized mice. In addition, HCcAg.120-VLPs prime splenocytes proliferate in vitro against different HCcAg.120-specific peptides, depending on either the immunization route or the adjuvant used. Remarkably, immunization with HCcAg.120-VLPs/Montanide ISA888 formulation resulted in a significant control of vaccinia virus titer in mice after challenge with a recombinant vaccinia virus expressing HCV core protein, vvCore. Animals immunized with this formulation had a marked increase in the number of IFN-γ producing spleen cells, after stimulation with P815 cells infected with vvCore. These results suggest the use of recombinant HCV core particles as components of therapeutic or preventive vaccine candidates against HCV.

Aureobasidium-Derived Soluble Branched (1,3-1,6) beta-Glucan (Sophy beta-glucan) Enhances Natural Killer Activity in Leishmania amazonensis-Infected Mice

The beta-glucans derived from yeast cell walls have been reported for having many immunomodulatory activities in vivo and in vitro. In this study, Aureobasidium-derived soluble branched (1,3-1,6) beta-glucan (Sophy beta-glucan) was checked for natural killer (NK) activity and for the production of IFN-gamma and IL-4 in Leishmania amazonensis infection. The main experiment was performed with a group of female C57BL/6 and BALB/c mice, orally supplemented with 5% of Sophy beta-glucan and infected with promastogotes of L. amazonensis (1 x 10(7)) into the footpad. Increase in the footpad thickness with time was observed in BALB/c mice in spite of the oral Sophy beta-glucan supplement, but it was less in C57BL/6 mice. The difference in overall mean footpad thickness between 'infection only' versus 'infection + glucan' groups was statistically significant (P < 0.001). High NK activity in C57BL/6 than BALB/c mice was observed in 'glucan only' group compared to the control group and also in 'infection + glucan' group compared to 'infection only' group. The difference in the NK activity among these groups was significant (P < 0.05). The IFN-gamma level increased at weeks 7 and 8 post-infection in C57BL/6 mice and was significantly high in 'infection + glucan' group compared to the 'infection only' group (P < 0.05). IL-4 levels did not increase up to detectable levels throughout the study. The results led a conclusion that Sophy beta-glucan enhances NK activity and cellular immunity in L. amazonensis-infected mice.

Selective addition of CXCR3+CCR4-CD4+ Th1 cells enhances generation of cytotoxic T cells by dendritic cells in vitro

Increasing importance is being given to the stimulation of Th1 response in cancer immunotherapy because its presence can shift the direction of adaptive immune responses toward protective immunity. Based on chemokine receptor expression, CXCR3+CCR4-CD4+ T cells as Th1-type cells were investigated its capacity in monocyte-derived dendritic cell (DC) maturation and polarization, and induction of antigen specific cytotoxic T lymphocytes (CTL) in vitro. The levels of IL-4, IL-5 and IL-10 were decreased to the basal level compared with high production of IFN-gamma, TNF-alpha, and IL-2 in CXCR3+CCR4-CD4+ T cells stimulated with anti-CD3 and anti-CD28 antibodies. Co-incubation of activated CD4+ or CXCR3+CCR4-CD4+ T cells with DC (CD4+/DC or CXCR3+CD4+/DC, respectively) particularly up-regulated IL-12 and CD80 expression compared with DC matured with TNF-alpha and LPS (mDC). Although there was no significant difference between the effects of the CXCR3+CCR4-CD4+ and CD4+ T cells on DC phenotype expression, CXCR3+CD4+/DC in CTL culture were able to expand number of CD8+ T cells and increased frequencies of IFN-gamma secreting cells and overall cytolytic activity against tumor antigen WT-1. These results demonstrated that the selective addition of CXCR3+CCR4-CD4+ T cells to CTL cultures could enhance the induction of CTLs by DC in vitro, and implicated on a novel strategy for adoptive T cell therapy.

Differential response of cytokines induced by Leptospira interrogans, serogroup Pomona, serovar Pomona, in mouse and human cell lines.

Leptospira interrogans, the causative agent of leptospirosis, is an important zoonotic bacterium. The mechanisms and roles of cytokine induction in both humans and animals remain unclear. Therefore, the IFN-gamma, IL-6, IL-10 and IL-12 levels were measured by enzyme-linked immunosorbent assay (ELISA) in human THP-1 and mouse RAW 264.7 monocyte cell lines following stimulation with heat-killed Leptospira interrogans serogroup Pomona serovar Pomona, L. biflexa, E. coli or Salmonella group B. The production of IL-6 and IL-12 were higher and rose more rapidly in the RAW 264.7 cells with all bacteria. The IL-10 was not detected in the RAW 264.7 cells when induced by leptospires. The IFN-gamma level in human peripheral blood mononuclear cells (PBMCs) induced by leptospires was also significantly lower than with other bacteria. When IL-10 and IL-12 mRNA expressions were detected in hamster's spleen, their patterns were similar to what was observed in THP-1 in that IL-12 was only slightly increased while IL-10 expression was high. Moreover, the IFN-gamma expression could not be detected in hamsters. The more potent cytokine responses in the RAW 264.7 cells may indirectly reflect the disease outcome in mice which render them to be a good reservoir of leptospirosis. Whether these cytokines have contributed to immunoprotection during the L. interrogans infection remains to be further investigated.