Leishmania infantum induces high phagocytic capacity and intracellular nitric oxide production by human proinflammatory monocyte

BACKGROUND The mechanism of resistance to SbIII in Leishmania is complex, multifactorial and involves not only biochemical mechanisms, but also other elements, such as the immune system of the host. OBJECTIVES In this study, putative changes in the immunological profile of human monocytes infected with wild-type (WT) and antimony (SbIII)-resistant Leishmania (Viannia) braziliensis and Leishmania (Leishmania) infantum lines were evaluated. METHODS Susceptibility assays WT and SbIII-resistant L. braziliensis and L. infantum were performed using lines THP-1 human monocytic lineage. Phagocytic capacity, cytokine profile, intracellular nitric oxide (NO) production and surface carbohydrate residues profile were performed in peripheral blood monocytes by flow cytometry. FINDINGS The phagocytic capacity and intracellular NO production by classical (CD14++CD16-) and proinflammatory (CD14++CD16+) monocytes were higher in the presence of L. infantum lines compared to L. braziliensis lines. The results also highlight proinflammatory monocytes as the cellular subpopulation of major relevance in a phagocytosis event and NO expression. It is important to note that L. infantum induced a proinflammatory cytokine profile characterised by higher levels of TNF-α in culture supernatant than L. braziliensis. Conversely, both Leishmania lines induce high levels of IL-6 in culture supernatant. Analysis of the expression profile of surface carbohydrates showed that L. braziliensis presents 4.3-fold higher expression of galactose(β1,4)N-acetylglucosamine than L. infantum line. Interestingly, the expression level of α-N-acetylgalactosamine residues was 2-fold lower in the SbIII-resistant L. braziliensis line than its counterpart WT line, indicating differences in surface glycoconjugates between these lines. MAIN CONCLUSIONS Our results showed that L. braziliensis and L. infantum induce different innate immune responses and a highly inflammatory profile, which is characteristic of infection by L. infantum, the species associated with visceral disease.

Effect of fresh frozen plasma on the in vitro activation of U937 monocytes: a potential role for the age of blood donors and their underlying cytokine profile

BACKGROUND: Fresh frozen plasma (FFP) administration may increase the risk of nosocomial infections in parallel with the development of immune modulation. This could be driven by soluble mediators, possibly influencing the in vitro activation of human U937 monocyte cells, in a manner dependent on the age of the donors. METHODS: FFP donors were stratified into groups of 19-30 years, 31-40 years or 41-50 years, and U937 cells were cultured with FFP (alone or plus lipopolysaccharide-LPS) for 24 h. Both in FFP and supernatants, TNF, IL-1ß, IL-6, and IL-10 levels were measured by ELISA. Additionally, CD11B, TLR2, and CASP3 gene expression were measured by qtPCR in U937 cells. Total phagocytic activity was also assayed. RESULTS: Elevated IL-10, but low TNF and IL-1ß levels were measured in FFP from individuals aged 19-40 years, whereas in individuals aged 41-50 years FFP were characterized by equalized TNF and IL-10 levels. Elevated IL-6 levels were found in all FFP samples, especially in those from the oldest individuals. FFP stimulation was associated with striking modifications in cytokine production in an age-dependent way. Exposure to FFP attenuates the response to LPS. TLR2 and CD11B expression were enhanced regardless of the age of plasma donors, although CASP3 expression was increased only when FFP from individuals aged 19-40 years were tested. Phagocytosis decreased after exposure to FFP regardless of donor age. CONCLUSION: Our results suggest that soluble mediators in FFP may modulate the functioning of monocytes. Interestingly, this effect appears to be partially influenced by the age of donors.

Infecção por leishmania modula a ativação de Beta-1 integrinas e altera a cinética do espalhamento celular de monócitos sobre fibronectina/Leishmania infection modulates the activation of Beta-1 integrins and alters the kinetics of monocyte cell spread on fibronectin / Leishmania infection modulates the activation of beta-1 integrins and alters the kinetics of monocyte cell spread on fibronectin / Leishmania infection modulates the activation of beta-1 integrins and alters the kinetics of monocyte cell spread on fibronectin

A leishmaniose é considerada como um grupo de doenças, causadas por parasitos do gênero Leishmania. O gênero inclui mais de 20 espécies de parasitos que são transmitidas ao homem pela picada da fêmea, infectada, dos insetos da família dos flebotomíneos e as diferentes espécies estão associadas a diferentes manifestações clínicas da doença. Uma característica marcante na infecção é a presença de macrófagos infectados no sítio de inoculação do parasito e em órgãos internos do hospedeiro. Estudos prévios, demonstraram que a infecção de fagócitos mononucleares com Leishmania amazonensis, L. infantum ou L. braziliensis promove uma diminuição na adesão celular ao tecido conjuntivo inflamado da pele. Esta diminuição da adesão é decorrente, principalmente, de mecanismos envolvidos na regulação da molécula VLA-4, uma integrina da família beta 1 que se liga à VCAM-1...

Leishmaniasis is considered a group of diseases, caused by parasites of the genus Leishmania. The genus includes more than 20 species of parasites that are transmitted to humans by the bite of vector female insect, of the phlebotomine family, and the different species are associated with different clinical manifestations of the disease. A striking feature of infection is the presence of infected macrophages at the site of inoculation of the parasite and internal organs of the host. Previous studies have demonstrated that infection of mononuclear phagocytes with Leishmania amazonensis, L. infantum or L. braziliensis promotes a decrease in cellular adhesion to the inflamed connective tissue of the skin. This decrease in adhesion results mainly from mechanisms involved in the regulation of the VLA-4 molecule, a beta 1 integrin that binds to VCAM-1...

Activation and cytokine profile of monocyte derived dendritic cells in leprosy: in vitro stimulation by sonicated Mycobacterium leprae induces decreased level of IL-12p70 in lepromatous leprosy

Dendritic cells (DCs) play a pivotal role in the connection of innate and adaptive immunity of hosts to mycobacterial infection. Studies on the interaction of monocyte-derived DCs (MO-DCs) using Mycobacterium leprae in leprosy patients are rare. The present study demonstrated that the differentiation of MOs to DCs was similar in all forms of leprosy compared to normal healthy individuals. In vitro stimulation of immature MO-DCs with sonicated M. leprae induced variable degrees of DC maturation as determined by the increased expression of HLA-DR, CD40, CD80 and CD86, but not CD83, in all studied groups. The production of different cytokines by the MO-DCs appeared similar in all of the studied groups under similar conditions. However, the production of interleukin (IL)-12p70 by MO-DCs from lepromatous (LL) leprosy patients after in vitro stimulation with M. lepraewas lower than tuberculoid leprosy patients and healthy individuals, even after CD40 ligation with CD40 ligand-transfected cells. The present cumulative findings suggest that the MO-DCs of LL patients are generally a weak producer of IL-12p70 despite the moderate activating properties ofM. leprae. These results may explain the poor M. leprae-specific cell-mediated immunity in the LL type of leprosy.

Percepção dos graduandos de enfermagem sobre o seu envelhecimento / Percepción de los estudiantes de enfermería sobre lo proprio envejecimiento / Perception of nursing undergraduate students on self-aging

RESUMO Objetivo: analisar a percepção dos graduandos de enfermagem sobre o próprio envelhecimento. Método: pesquisa de abordagem qualitativa, realizada em agosto e setembro de 2011, com 18 graduandos de enfermagem de uma Universidade pública de Salvador (Bahia). Os depoimentos foram analisados por meio da Análise de Conteúdo. Resultados: apreendeu-se o núcleo temático: Percepção do graduando de enfermagem sobre o próprio envelhecimento e, a partir deste, emergiram duas subcategorias: A) O Não Pensar; B) O contexto influenciando no processo. Conclusão: os graduandos revelam que o envelhecimento está intrínseco ao desenvolvimento humano, e possui o vínculo familiar, a espiritualidade e atividade física como ferramentas fundamentais para um envelhecimento ativo. Entretanto, os mesmos relatam que, o modo de vida acelerado e estressante vivido na sociedade possibilita inserir hábitos considerados inadequados, como o consumo de “fast food” e álcool, que trazem influências negativas para o próprio processo de envelhecimento. .

RESUMEN Objetivo: analizar la percepción de los estudiantes de enfermería sobre su proprio envejecimiento. Método: estudio cualitativo, realizado en agosto y septiembre de 2011, con 18 estudiantes de enfermería de una universidad pública en Salvador/Bahia. Los datos fueron analizados através de análisis de contenido. Resultado: incautados el tema central: Percepción de alumnos de enfermería sobre su propio envejecimiento y de esto surgieron dos subcategorías: A) No creo; B) El contexto influye en el proceso. Conclusión: los estudiantes revelan que el envejecimiento es intrínseco al desarrollo humano, y tiene los vínculos familiares, la espiritualidad y la actividad física como herramienta clave para el envejecimiento activo. Sin embargo, el mismo informe que, debido a la forma de vida que se vive en la sociedad de ritmo rápido y estresante permite insertar hábitos considerados inadecuados, como el consumo de “comida rápida” y el alcohol y convertirse en influencias negativas para su propio proceso tuvo como objetivo analizar de los estudiantes de enfermería su propio envejecimiento. .

ABSTRACT Objective: to analyze the perceptions of nursing undergraduate students on their self-aging process. Method: qualitative study carried out between August and September, 2011 with 18 nursing undergraduate students of a public university in Salvador, Bahia. The interviews were analyzed by means of the Content Analysis method. Results: the following thematic concept was apprehended: Perceptions of nursing undergraduates on their self-aging, which generated two subcategories: A) The “don’t think about it” process; B) The context infl uencing the process. Conclusion: undergraduates reveal that the aging process is an intrinsic factor to human development. Family ties, spirituality and physical activity would be key mechanisms toward active aging. However, students also reported that their accelerated and stressed social lifestyles led to inadequate habits, such as the consumption of fast food and alcohol, which become negative infl uences in their aging process. .

Avaliação das subpopulações de monócitos em pacientes com tuberculose pulmonar / Evaluation of subpopulations of monocytes in patients with pulmonary tuberculosis

Os macrófagos são componentes importantes da resposta imune inata contra o Mycobaterium tuberculosis (Mtb) e podem desempenhar um papel importante na patogênese da tuberculose (TB). Macrófagos são derivados dos monócitos, os quais são classificados em subpopulações a partir da expressão da molécula de superfície CD14 e CD16. São denominados de clássicos, intermediários e não clássicos, e possuem diferenças funcionais e fenotípicas. Os fatores que levam ao desenvolvimento de TB ativa ainda não são claros. Um desequilíbrio entre subpopulações de monócitos circulantes pode estar envolvido na imunopatogênese da TB, uma vez que macrófagos são células importantes da resposta imune inicial da doença. Assim, neste estudo avaliou-se os subgrupos de monócitos em pacientes com TB ativa e latente (TBL). Voluntários com TB ativa, TBL e indivíduos saudáveis foram recrutados para avaliação de frequência, níveis de ativação e produção de citocinas dos subgrupos de monócitos circulantes e após a estimulação antigênica por citometria de fluxo. Nossos resultados não demonstraram diferenças significativas nas frequências, níveis de ativação e produções de citocinas das subpopulações de monócitos entre os grupos estudados. No entanto, pacientes com TB ativa tiveram um aumento na frequência dos monócitos clássicos ativados após estimulação antigênica comparados com os controles saudáveis. Não observou-se uma expansão das subpopulações CD16+ em pacientes TB. Por outro lado, se observou uma expansão dos monócitos CD16...

Macrophages are important components of the innate immune response against Mycobacterium tuberculosis (Mtb) and may play an important role in the pathogenesis of tuberculosis (TB). Macrophages are derived from monocytes, which are classified into subpopulations from the expression of CD14 and CD16 surface molecule. They are denominated classics, intermediate and non-classical, and have functional and phenotypic differences. The factors that lead to the development of active tuberculosis are not clear yet. However, an imbalance between subpopulations of monocytes may be involved in the immunopathogenesis of TB, since macrophages are important cells in the initial immune responses of the disease. In this study we evaluated the monocyte subsets in patients with active and latent TB (ILTB). Volunteers with active TB, ILTB and healthy subjects were recruited to evaluate the frequency, levels of activation and cytokine production of blood monocytes subsets circulating and after the antigenic stimulation by flow cytometry. Our results did not show significant differences in the frequency, activation levels and cytokine production of monocytes subsets between studies groups. However, patients with active TB have an increased of frequency and activated levels of classical monocytes after antigenic stimulation compared to healthy controls. An expansion of CD16+ in monocytes subsets of TB patient was not observed. Moreover, it was observed an expansion and increased activation of CD16...

Instituição de longa permanência para idosos: um lugar de cuidado para quem não tem opção? / Long-stay institutions for the elderly: a place of care for those who have no choice? / Institución de larga estadía para las personas mayores: ¿un lugar de atención para aquellos que no tienen otra opción?

Pesquisa descritiva, qualitativa, com abordagem compreensiva, que teve por objetivo compreender o significado da instituição de longa permanência para idosos institucionalizados. Os dados foram coletados com 13 idosos institucionalizados, no período de 5 de abril a 25 de maio de 2013 por meio da entrevista narrativa, e submetidos a análise de conteúdo, na modalidade de análise temática. Os resultados indicam que ser idoso institucionalizado significa ter suas necessidades de cuidado atendidas, no que concerne a suas necessidades básicas; ao acesso a serviços e recursos de saúde, e a ter um lugar onde possam envelhecer e morrer. O estudo permitiu concluir que a instituição aparece como um lugar ambíguo para os idosos, pois ao mesmo tempo em que os acolhe, abriga e atende suas necessidades, é um ambiente que inviabiliza a vida independente e autônoma.

This is a descriptive, qualitative research, with comprehensive approach, which aimed to understand the meaning that the longterm institution has to institutionalized elderly. Data were collected with 13 institutionalized elderly in the period from April 5 to May 25, 2013, through narrative interview, and subjected to content analysis, in the form of thematic analysis. The results indicated that being elderly institutionalized means having their care needs met, with respect to their basic needs; access to health services and resources, and to have a place where they can grow old and die. The study concluded that the institution appears as an ambiguous place for the elderly because, even embracing and housing them and meeting their needs, is an environment that prevents the independent and autonomous life.

Investigación descriptiva, cualitativa con enfoque comprehensivo, que tuve como objetivo comprender el significado de la institución a largo plazo tiene para ancianos institucionalizados. Los datos fueron recolectados con 13 ancianos institucionalizados en el periodo comprendido entre el 5 abril-25 mayo de 2013, a través de entrevista narrativa, y tratados mediante análisis de contenido, en la modalidad de análisis temático. Los resultados indican que estar en edad avanzada y estar institucionalizado significa tener sus atendidas sus necesidades básicas; el acceso a los servicios de salud y los recursos, y tener un lugar donde pueden envejecer y morir. El estudio llegó a la conclusión de que la institución se presenta como un lugar ambiguo para las personas mayores, ya que si bien les acoja, albergue y atiendan sus necesidades, es un ambiente que impide la vida independiente y autónoma.

Social and immunological differences among uninfected Brazilians exposed or unexposed to human immunodeficiency virus-infected partners

Understanding the social conditions and immunological characteristics that allow some human immunodeficiency virus (HIV)-exposed patients to remain uninfected represents an on-going challenge. In this study, the socio-demographic and sexual behaviour characteristics and immune activation profiles of uninfected individuals exposed to HIV-infected partners were investigated. A confidential and detailed questionnaire was administered and venous blood was tested using HIV-1/enzyme immunoassays, plasma HIV-1 RNA levels/bDNA and immunophenotyping/flow cytometry to determine the frequencies of CD4 and CD8 T cells expressing activation markers. The data analysis showed significant differences (p < 0.05) for immune parameters in individuals who were uninfected, albeit exposed to HIV-infected partners, compared with unexposed individuals. In particular, the exposed, uninfected individuals had a higher frequency (median, minimum-maximum) of CD4+HLA-DR+ (4.2, 1.8-6.1), CD8+HLA-DR+ (4.6, 0.9-13.7), CD4+CD45RO+ (27.5, 14.2-46.6), CD4+CD45RO+CD62L+ (46.7, 33.9-67.1), CD8+CD45RA+HLA-DR+ (12.1, 3.4-35.8) and CD8+CD45RO+HLA-DR+ (9.0, 3.2-14.8) cells, a decreased percentage of CD8+CD28+ cells (11.7, 4.5-24.0) and a lower cell-surface expression of Fcγ-R/CD16 on monocytes (56.5, 22.0-130.0). The plasma HIV-1 RNA levels demonstrated detectable RNA virus loads in 57% of the HIV-1+ female partners. These findings demonstrate an activation profile in both CD4 and CD8 peripheral T cells from HIV-1 exposed seronegative individuals of serodiscordant couples from a referral centre in Belo Horizonte, state of Minas Gerais.

TLR-2 and TLR-4 expression in monocytes of newborns with late-onset sepsis, / Expressão do TLR-2 e do TLR-4 em monócitos de recém-nascidos a termo com sepse tardia

Objective: To analyze toll-like receptor (TLR)-2 and TLR-4 expression in monocytes of newborns with late-onset sepsis. Methods: This prospective study included 27 full-term newborns aged 8 to 29 days, with clinical and laboratory diagnosis of late-onset sepsis. Ten newborns (37%) had positive cultures. Cytokines were measured by cytometric bead array in peripheral blood, while TLR-2, TLR-4 expression, and median fluorescence intensity (MFI) were determined by immunophenotyping peripheral whole blood monocytes, and were analyzed with a BD FACSDiva flow cytometer (Becton, Dickinson and Company, USA). A comparison was performed with healthy adults. Results: Microorganisms were identified in 37% of these septic newborns, and all of them had high levels of pro-inflammatory cytokines (IL-8, IL-6, IL-1β) and anti-inflammatory cytokine (IL-10) corroborating the inflammatory/septic process. In monocytes, the frequency of TLR-4 expression was higher in infected newborns (p = 0.01). Conclusion: This study investigated the innate immune response in septic newborns. Septic newborns that relied almost exclusively on the innate immune system showed little in vivo response at monocyte activation, suggesting impaired immune response and increased susceptibility to infection. .

Objetivos: Analisar a expressão dos TLR-2 e TLR-4 em monócitos de recém-nascidos com sepse tardia. Métodos: Trata-se de um estudo prospectivo com 27 recém-nascidos a termo entre 8 e 29 dias de vida com diagnóstico clínico e laboratorial de sepse tardia dos quais dez (37%) apresentaram cultura positiva. As citocinas foram determinadas por teste de CBA em sangue periférico enquanto que a expressão e MFI (mediana de intensidade de fluorescência) dos TLR-2 e TLR-4 foi determinado por imunofenotipagem em monócitos de sangue periférico total através de análise pelo citômetro de fluxo BD FACSDiva. O grupo usado para comparação foi de adultos saudáveis. Resultados: Microrganismos foram identificados em 37% dos pacientes e estes juntamente com os pacientes com sepse clínica tiveram níveis elevados de citocinas pró-inflamatórias (IL-8, IL-6, IL-1β) e de citocina anti-inflamatória (IL-10) corroborando o processo inflamatório/infeccioso. No monócito, a frequência de expressão do TLR-4 foi mais elevada (p = 0,01). Conclusões: Este estudo analisou a resposta imune inata no recém-nascido com sepse. Recémnascidos sépticos que dependem quase exclusivamente do sistema imune inato apresentaram pouca resposta in vivo na ativação de monócitos o que sugere uma resposta imune deficiente e maior susceptibilidade à infecção. .

Leukocytosis, muscle damage and increased lymphocyte proliferative response after an adventure sprint race

The effect of an adventure sprint race (ASR) on T-cell proliferation, leukocyte count and muscle damage was evaluated. Seven young male runners completed an ASR in the region of Serra do Espinhaço, Brazil. The race induced a strong leukocytosis (6.22±2.04×103 cells/mm3 before vs 14.81±3.53×103 cells/mm3 after the race), marked by a significant increase of neutrophils and monocytes (P<0.05), but not total lymphocytes, CD3+CD4+ or CD3+CD8+ cells. However, the T-cell proliferative response to mitogenic stimulation was increased (P=0.025) after the race, which contradicted our hypothesis that ASR, as a high-demand competition, would inhibit T-cell proliferation. A positive correlation (P=0.03, r=0.79) was observed between the proliferative response of lymphocytes after the race and the time to complete the race, suggesting that the proliferative response was dependent on exercise intensity. Muscle damage was evident after the race by increased serum levels of aspartate amino transferase (24.99±8.30 vs 50.61±15.76 U/L, P=0.003). The results suggest that humoral factors and substances released by damaged muscle may be responsible for lymphocyte activation, which may be involved in muscle recovery and repair.

Kinetics of IL-23 and IL-12 Secretion in Response to Toxoplasma gondii Antigens from THP-1 Monocytic Cells

IL-23 and IL-12 are structurally similar and critical for the generation of efficient cellular immune responses. Toxoplasma gondii induces a strong cell-mediated immune response. However, little is known about IL-23 secretion profiles in T. gondii-infected immune cells in connection with IL-12. We compared the patterns of IL-23 and IL-12 production by THP-1 human monocytic cells in response to stimulation with live or heat-killed T. gondii tachyzoites, or with equivalent quantities of either T. gondii excretory/secretory proteins (ESP) or soluble tachyzoite antigen (STAg). IL-23 and IL-12 were significantly increased from 6 hr after stimulation with T. gondii antigens, and their secretions were increased with parasite dose-dependent manner. IL-23 concentrations were significantly higher than those of IL-12 at the same multiplicity of infection. IL-23 secretion induced by live parasites was significantly higher than that by heat-killed parasites, ESP, or STAg, whereas IL-12 secretion by live parasite was similar to those of ESP or STAg. However, the lowest levels of both cytokines were at stimulation with heat-killed parasites. These data indicate that IL-23 secretion patterns by stimulation with various kinds of T. gondii antigens at THP-1 monocytic cells are similar to those of IL-12, even though the levels of IL-23 induction were significantly higher than those of IL-12. The detailed kinetics induced by each T. gondii antigen were different from each other.

Impaired neutrophil and monocyte chemotaxis in chronic and aggressive periodontitis and effects of periodontal therapy.

Background and Objectives: Depressed chemotactic activity of polymorphoneutrophil (PMN) and monocyte (MN) appears to be one of the significant risk factors in the development of periodontal disease. Although bacteria are the primary etiologic factor in periodontal disease, the patient's host response is a determinant of disease susceptibility. Depressed chemotaxis of PMN and MN could lead to periodontal destruction by altering the host response i.e. impairment of the normal host response in neutralizing infection and alterations that result in destruction of the surrounding periodontal tissues. Materials and Methods: Thirty patients (10 healthy subjects, 10 chronic periodontitis, and 10 with aggressive periodontitis) participated in this study. Clinical parameters like plaque index, gingival index, probing pocket depth, and radiographic assessment were done. The peripheral blood PMNs and MNs were isolated from the patient and the chemotactic response was studied. Statistical analysis was performed using post-hoc Newman-Keul range test. Results: PMN and MN chemotaxis was found to be statistically significant (P<0.05) at baseline and three months after periodontal therapy in chronic and aggressive periodontitis group compared to healthy subjects. However on comparison between chronic and aggressive periodontitis group statistical significance was not found (P>0.05).Comparision between chronic periodontitis, aggressive periodontitis with healthy subjects, PMN and MN chemotaxis showed statistical significance (P<0.05) at baseline and three months after periodontal therapy, Whereas statistically there was no difference when chronic periodontitis was compared with aggressive periodontitis Interpretation and Conclusion: Depressed chemotaxis of PMN and MN results in increased periodontal destruction. In this study, depressed PMN and MN chemotaxis is seen in both aggressive periodontitis group and chronic periodontitis group and the response was altered although to a lesser degree after periodontal therapy in both groups indicating that effect of treatment does exist.

Avaliação dos monócitos entre as formas clínicas crônicas da doença de Chagas antes e após estímulo in vitro com os antígenos recombinantes CRA e FRA de Trypanosoma cruzi / Assessing of monocytes between the clinical forms of chronic Chagas' disease before and after in vitro stimulation with CRA and FRA recombinant antigens of Trypanosoma cruzi

As interações moleculares entre os monócitos e os linfócitos são de extrema importância para a produção de uma resposta imune eficiente. Nesse contexto nos propomos avaliar a expressão da molécula de superfície HLA-DR+ em monócitos CD14+, e das moléculas co-estimulatórias CD80+ e CD86+ e a produção das citocinas IL-10, IFN-gama, TNF-alfa e IL-12 por monócitos CD14+ HLA-DR+ de portadores de formas clínicas crônicas da doença de Chagas. O grupo de indivíduos acometidos cronicamente pela infecção chagásica foi composto por 10 portadores da forma cardíaca (FC) sem dilatação cardíaca (FC1), 14 portadores da forma cardíaca com dilatação cardíaca (FC2) e 7 portadores da forma indeterminada (FI). O sangue total destes indivíduos foi submetido à cultura na presença dos antígenos CRA ou FRA, específicos ao Trypanosoma cruzi. Observamos que o aumento na expressão da molécula co-estimulatória CD80+ associado à diminuição da expressão de CD86+ no grupo de indivíduos FC, quando comparamos os contextos ex vivo e após estímulo com CRA ou FRA, aponta um possível envolvimento desses antígenos nos mecanismos de fuga do sistema imune do hospedeiro pelo T. cruzi. Correlações positivas entre citocinas inflamatórias e anti-inflamatórias foram observadas entre os portadores da FI e FC1, indicando um controle do parasitismo, concomitantemente a uma modulação da resposta inflamatória. No grupo de pacientes FC2 observamos uma predominância de uma correlação positiva entre as citocinas inflamatórias, bem como de correlação positiva entre citocinas Th1 e Th2, indicando que há predomínio de um perfil do tipo Th1, mas com indícios de mecanismo de regulação da resposta inflamatória exacerbada. Nossos resultados sugerem que os antígenos CRA e FRA, por estarem presentes no parasito, poderiam atuar no processo de apresentação e ativação celular durante a cronicidade da doença, sobretudo nos indivíduos portadores da FC.

Granulocyte macrophage colony-stimulating factor enhances the modulatory effect of cytokines on monocyte-derived multinucleated giant cell formation and fungicidal activity against Paracoccidioides brasiliensis

Multinucleated giant cells (MGC) are cells present in characteristic granulomatous inflammation induced by intracellular infectious agents or foreign materials. The present study evaluated the modulatory effect of granulocyte macrophage colony-stimulating factor (GM-CSF) in association with other cytokines such as interferon-gamma (IFN-γ), tumour necrosis factor-alpha, interleukin (IL)-10 or transforming growth factor beta (TGF-β1) on the formation of MGC from human peripheral blood monocytes stimulated with Paracoccidioides brasiliensis antigen (PbAg). The generation of MGC was determined by fusion index (FI) and the fungicidal activity of these cells was evaluated after 4 h of MGC co-cultured with viable yeast cells of P. brasiliensis strain 18 (Pb18). The results showed that monocytes incubated with PbAg and GM-CSF plus IFN-γ had a significantly higher FI than in all the other cultures, while the addition of IL-10 or TGF-β1 had a suppressive effect on MGC generation. Monocytes incubated with both pro and anti-inflammatory cytokines had a higher induction of foreign body-type MGC rather than Langhans-type MGC. MGC stimulated with PbAg and GM-CSF in association with the other cytokines had increased fungicidal activity and the presence of GM-CSF also partially inhibited the suppressive effects of IL-10 and TGF-β1. Together, these results suggest that GM-CSF is a positive modulator of PbAg-stimulated MGC generation and on the fungicidal activity against Pb18.

Interleukin-15 augments oxidative metabolism and fungicidal activity of human monocytes against Paracoccidioides brasiliensis

Interleukin (IL)-15 is a pleiotropic cytokine that regulates the proliferation and survival of many cell types. IL-15 is produced by monocytes and macrophages against infectious agents and plays a pivotal role in innate and adaptive immune responses. This study analyzed the effect of IL-15 on fungicidal activity, oxidative metabolism and cytokine production by human monocytes challenged in vitro with Paracoccidioides brasiliensis (Pb18), the agent of paracoccidioidomycosis. Peripheral blood monocytes were pre-incubated with IL-15 and then challenged with Pb18. Fungicidal activity was assessed by viable fungi recovery from cultures after plating on brain-heart infusion-agar. Superoxide anion (O2-), hydrogen peroxide (H2O2), tumour necrosis factor-alpha (TNF-α), IL-6, IL-15 and IL-10 production by monocytes were also determined. IL-15 enhanced fungicidal activity against Pb18 in a dose-dependent pattern. This effect was abrogated by addition of anti-IL-15 monoclonal antibody. A significant stimulatory effect of IL-15 on O2- and H2O2 release suggests that fungicidal activity was dependent on the activation of oxidative metabolism. Pre-treatment of monocytes with IL-15 induced significantly higher levels of TNF-α, IL-10 and IL-15 production by cells challenged with the fungus. These results suggest a modulatory effect of IL-15 on pro and anti-inflammatory cytokine production, oxidative metabolism and fungicidal activity of monocytes during Pb18 infection.

Lipopolysaccharide-induced expression of cell surface receptors and cell activation of neutrophils and monocytes in whole human blood

Lipopolysaccharide (LPS) activates neutrophils and monocytes, inducing a wide array of biological activities. LPS rough (R) and smooth (S) forms signal through Toll-like receptor 4 (TLR4), but differ in their requirement for CD14. Since the R-form LPS can interact with TLR4 independent of CD14 and the differential expression of CD14 on neutrophils and monocytes, we used the S-form LPS from Salmonella abortus equi and the R-form LPS from Salmonella minnesota mutants to evaluate LPS-induced activation of human neutrophils and monocytes in whole blood from healthy volunteers. Expression of cell surface receptors and reactive oxygen species (ROS) and nitric oxide (NO) generation were measured by flow cytometry in whole blood monocytes and neutrophils. The oxidative burst was quantified by measuring the oxidation of 2',7'-dichlorofluorescein diacetate and the NO production was quantified by measuring the oxidation of 4-amino-5-methylamino-2',7'-difluorofluorescein diacetate. A small increase of TLR4 expression by monocytes was observed after 6 h of LPS stimulation. Monocyte CD14 modulation by LPS was biphasic, with an initial 30 percent increase followed by a 40 percent decrease in expression after 6 h of incubation. Expression of CD11b was rapidly up-regulated, doubling after 5 min on monocytes, while down-regulation of CXCR2 was observed on neutrophils, reaching a 50 percent reduction after 6 h. LPS induced low production of ROS and NO. This study shows a complex LPS-induced cell surface receptor modulation on human monocytes and neutrophils, with up- and down-regulation depending on the receptor. R- and S-form LPS activate human neutrophils similarly, despite the low CD14 expression, if the stimulation occurs in whole blood.

Human monocytes tolerant to LPS retain the ability to phagocytose bacteria and generate reactive oxygen species

Tolerance to lipopolysaccharide (LPS) occurs when animals or cells exposed to LPS become hyporesponsive to a subsequent challenge with LPS. This mechanism is believed to be involved in the down-regulation of cellular responses observed in septic patients. The aim of this investigation was to evaluate LPS-induced monocyte tolerance of healthy volunteers using whole blood. The detection of intracellular IL-6, bacterial phagocytosis and reactive oxygen species (ROS) was determined by flow cytometry, using anti-IL-6-PE, heat-killed Staphylococcus aureus stained with propidium iodide and 2',7'-dichlorofluorescein diacetate, respectively. Monocytes were gated in whole blood by combining FSC and SSC parameters and CD14-positive staining. The exposure to increasing LPS concentrations resulted in lower intracellular concentration of IL-6 in monocytes after challenge. A similar effect was observed with challenge with MALP-2 (a Toll-like receptor (TLR)2/6 agonist) and killed Pseudomonas aeruginosa and S. aureus, but not with flagellin (a TLR5 agonist). LPS conditioning with 15 ng/mL resulted in a 40 percent reduction of IL-6 in monocytes. In contrast, phagocytosis of P. aeruginosa and S. aureus and induced ROS generation were preserved or increased in tolerant cells. The phenomenon of tolerance involves a complex regulation in which the production of IL-6 was diminished, whereas the bacterial phagocytosis and production of ROS was preserved. Decreased production of proinflammatory cytokines and preserved or increased production of ROS may be an adaptation to control the deleterious effects of inflammation while preserving antimicrobial activity.

Dengue-2 infection and the induction of apoptosis in human primary monocytes

Monocytes/macrophages are important targets for dengue virus (DENV) replication; they induce inflammatory mediators and are sources of viral dissemination in the initial phase of the disease. Apoptosis is an active process of cellular destruction genetically regulated, in which a complex enzymatic pathway is activated and may be trigged by many viral infections. Since the mechanisms of apoptotic induction in DENV-infected target cells are not yet defined, we investigated the virus-cell interaction using a model of primary human monocyte infection with DENV-2 with the aim of identifying apoptotic markers. Cultures analyzed by flow cytometry and confocal microscopy yielded DENV antigen positive cells with rates that peaked at the second day post infection (p.i.), decayed afterwards and produced the apoptosis-related cytokines TNF-á and IL-10. Phosphatidylserine, an early marker for apoptosis, was increased at the cell surface and the Fas death receptor was upregulated at the second day p.i. at significantly higher rates in DENV infected cell cultures than controls. However, no detectable changes were observed in the expression of the anti-apoptotic protein Bcl-2 in infected cultures. Our data support virus modulation of extrinsic apoptotic factors in the in vitro model of human monocyte DENV-2 infection. DENV may be interfering in activation and death mechanisms by inducing apoptosis in target cells.

Hormonas sexuales y respuesta inmunológica / Sex hormones and immune system

Existe una evidente interrelación entre el sistema endócrino y el sistema inmunológico. Un ejemplo de esto es el efecto que las hormonas sexuales ejercen sobre las distintas poblaciones de leucocitos (linfocitos T y B, Células NK, granulocitos y macrófagos), así como sobre la producción y liberación de citoquinas y proteínas inmunoreguladoras. Tanto en las mujeres como en las hembras de otras especies, los estrógenos y la progesterona harían que primase una respuesta inmune humoral, lo cual resultaría beneficioso para la gestación, pero al mismo tiempo favorecería la aparición de ciertas enfermedades autoinmunes. Contrariamente, la testoterona haría que en los machos predominase la respuesta inmune celular. El siguiente trabajo es una revisión de distintos estudios referentes a la acción que las hormonas sexuales esteroideas ejercen sobre distintos componentes del sistema inmunológico.