Dura����o do efeito analg��sico e express��o de diferentes metaloproteinases, do col��geno tipo IV e da interleucina-10, em c��rneas de coelhos tratadas com morfina, ap��s ceratectomia lamelar

Funda����o de Amparo �� Pesquisa do Estado de S��o Paulo (FAPESP)

N��veis de merc��rio, prolactina e interleucina 10 em mulheres em idade reprodutiva e pu��rperas dos munic��pios de Itaituba e Ananindeua, Par��

Ao merc��rio tem sido atribu��da a capacidade de interferir nos sistemas org��nicos imunol��gico e hormonal, al��m dos sistemas nervoso e renal frequentemente atingidos por esse agente t��xico. Mulheres em idade f��rtil ou gr��vidas constituem um grupo vulner��vel a esses efeitos, em rela����o a si mesmas e seus conceptos. Foi avaliada a exposi����o ao merc��rio (Hg) e os n��veis de prolactina (PRL) e interleucina-10 (IL-10) em 144 mulheres (no p��s-parto e cerca de um ano depois) de Itaituba, ��rea sob impacto ambiental do merc��rio e em mulheres de munic��pios da ��rea metropolitana de Bel��m, sobretudo Ananindeua, ��rea sem impacto conhecido do merc��rio (156 pu��rperas e 156 n��o pu��rperas). As an��lises de merc��rio total (Hg-t) em sangue foram feitas por Espectrometria de Absor����o At��mica por Vapor Frio. As an��lises s��ricas de PRL foram feitas por Ensaio Imunoenzim��tico com detec����o final em fluoresc��ncia e as determina����es de IL-10 foram realizadas por Ensaio Imunoenzim��tico de Fase S��lida. Dados demogr��ficos e epidemiol��gicos foram obtidos atrav��s de question��rio semi-estruturado. As pu��rperas de Itaituba apresentaram m��dia de Hg-t, PRL e IL-10 de 13,93 ��g/l, 276,20 ng/ml e 39,54 pg/ml, respectivamente. Nas pu��rperas de Ananindeua as respectivas m��dias foram 3,67 ��g/l, 337,70 ng/ml e 4,90 pg/ml. As mulheres n��o pu��rperas de Itaituba apresentaram m��dia de Hg-t de 12,68 ��g/l, m��dia de PRL de 30,75 ng/ml e m��dia de IL-10 de 14,20 pg/ml. As m��dias de Hg-t, PRL e IL-10 das mulheres de Ananindeua foram 2,73 ��g/l, 17,07 ng/ml e 1,49 pg/ml, respectivamente. Os n��veis de Hg-t, PRL e IL-10 foram maiores em Itaituba (p<0,0001), exceto em rela����o �� PRL das pu��rperas, maior em Ananindeua. Os n��veis semelhantes de Hg-t nas duas avalia����es das mulheres de Itaituba (p=0,7056) e a correla����o moderada sugerem continuidade da exposi����o (r=0,4736, p<0,0001). A principal vari��vel preditora dos n��veis de merc��rio foi o consumo de peixe nos modelos de regress��o m��ltipla linear e log��stica. A paridade e os n��veis de IL-10 apresentaram associa����o positiva com a PRL nas pu��rperas de Itaituba e o peso do rec��m-nascido e a IL-10, associa����o positiva com a PRL em pu��rperas de Ananindeua. A IL-10 apresentou associa����o negativa com a PRL nas mulheres n��o pu��rperas de Itaituba (p=0,0270) e positiva nas mulheres de Ananindeua (p=0,0266). Os n��veis de Hg-t estavam associados negativamente com a PRL nas pu��rperas (p=0,0460) e positivamente com o trabalho em garimpo (p=0,0173) (este tamb��m importante para as n��o pu��rperas) em Itaituba, segundo os modelos log��sticos. A IL-10 esteve associada positivamente �� morbidade recente nas pu��rperas de Itaituba (p=0,0210), negativamente ao consumo de bebida alco��lica (p=0,0178) e positivamente ao trabalho em garimpo nas mulheres n��o pu��rperas (p=0,0199). A exposi����o cr��nica ao Hg das mulheres de Itaituba, a diferen��a nos n��veis dos fatores imunoend��crinos avaliados em rela����o ��s mulheres n��o expostas e a associa����o com vari��veis epidemiol��gicas relevantes, sugerem a possibilidade de impactos da exposi����o no perfil imunoend��crino das mulheres de Itaituba, chamando aten����o para a import��ncia da vigil��ncia da sa��de dessa popula����o e o poss��vel uso de bioindicadores como a PRL em sua avalia����o.

Participa����o das citocinas Interleucina-12, Interferon-��, Interleucina-4 e Interleucina-10 e investiga����o de polimorfismos nos genes do INF-�� (IFNG+874) e da IL-10 (IL10-1082) na mal��ria causada por Plasmodium vivax

A resposta imune na mal��ria �� complexa, e os mecanismos de ativa����o e regula����o de linf��citos T efetores e de mem��ria ainda s��o pouco compreendidos. No presente estudo, determinamos a concentra����o das citocinas Interferon-�� (IFN-��), Interleucina-10 (IL-10), Interleucina-4 (IL-4) e Interleucina-12 (IL-12) no soro de indiv��duos infectados por Plasmodium vivax, investigamos os polimorfismos no gene do IFN-�� (IFNG+874) e da IL-10 (IL10-1082) e analisamos a associa����o destes polimorfismos com a concentra����o das citocinas e com a densidade parasit��ria. A concentra����o das citocinas foi determinada por ELISA, e a genotipagem dos polimorfismos IFNG+874 e IL10-1082 foi realizada pelas t��cnicas de ASO-PCR e PCR-RFLP, respectivamente. Os indiv��duos infectados apresentaram n��veis s��ricos de IFN-�� e IL-10 aumentados. A produ����o de IFN-�� foi maior nos indiv��duos primoinfectados, por��m n��o foi associada com a redu����o da parasitemia. A produ����o de IL-10 foi alta e associada com altas parasitemias. As citocinas IL-4 e IL-12 n��o foram detectadas. As freq����ncias dos gen��tipos homozigoto mutante AA, heterozigoto AT e selvagem TT do gene do IFN-�� foram 0,51, 0,39 e 0,10, respectivamente. As freq����ncias dos gen��tipos homozigoto mutante AA, heterozigoto AG e selvagem GG para IL10 foram 0,49, 0,43 e 0,08, respectivamente. Apenas o polimorfismo do IFN-�� foi associado com n��veis reduzidos desta citocina. Na mal��ria causada por P. vivax, houve produ����o de citocina que caracteriza o perfil Th1 (IFN-��), com poss��vel participa����o da IL-10 na imunorregula����o.

Associa����o entre express��o de receptores TLR2 e TLR4 e produ����o de TNF-alpha e IL-10 por mon��citos de gestantes portadoras de pr��-ecl��mpsia

Preeclampsia (PE) is a pregnancy specific syndrome characterized by a systemic inflammatory response, with higher intensity than that observed in normal pregnancy. Cells of the immune system, such as monocytes and granulocytes are endogenously activated and secrete high levels of free radicals and inflammatory cytokines. The objective of this study was to assess the activation state of monocytes from pregnant women with preeclampsia by endogenous expression of TLR2 e TLR4 receptors and to correlate the expression of TLR2 and TLR4 on monocytes surface of pregnant women with PE with the production of tumor necrosis factor-alpha (TNF-������ and interleukin-10 (IL-10) by these cells stimulated or not with peptidoglycan (PG) and lipopolysaccharide (LPS), as agonists agents of TLR2 and TLR4, respectively. We evaluated 15 pregnant women with PE, 15 normotensive pregnant women (NT) and 15 non-pregnant (NP). Peripheral blood monocytes were incubates in the presence or absence of LPS or PG. The supernatant obtained after 18h of culture was aspirated and used for TNF-������ and IL-10 determination by enzyme immunoassay (ELISA). The endogenous expression of TLR2 and TLR4 receptors was evaluated by flow cytometry. Our results showed significant highly concentrations of TNF-������ and TLR4 expression in monocytes of preeclamptic women when compared with NT and NP. Normal pregnant women presented higher levels of IL-10 in comparison with PE and NP groups. TLR2 expression was similar in the three groups studied. Therefore, our study highlights the important role of TLR4 in PE and the consequent high production of TNF-������ by monocytes of these patients, as well as the potential mechanism involving low levels of IL-10 in the pathophysiology of the disease. These observations demonstrate the strong link between the pathology of PE and the immune system of these patients

Early IL-10 production is essential for syngeneic graft acceptance

We performed a comparative study and evaluated cellular infiltrates and anti-inflammatory cytokine production at different time-points after syngeneic or allogeneic skin transplantation. We observed an early IL-10 production in syngeneic grafts compared with allografts. This observation prompted us to investigate the role of IL-10 in isograft acceptance. For this, we used IL-10 KO and WT mice to perform syngeneic transplantation, where IL-10 was absent in the graft or in the recipient. The majority of syngeneic grafts derived from IL-10 KO donors did not engraft or was only partially accepted, whereas IL-10 KO mice transplanted with skin from WT donors accepted the graft. We evaluated IL-10 producers in the transplanted skin and observed that epithelial cells were the major source. Taken together, our data show that production of IL-10 by donor cells, but not by the recipient, is determinant for graft acceptance and strongly suggest that production of this cytokine by keratinocytes immediately upon transplantation is necessary for isograft survival. J. Leukoc. Biol. 92: 259-264; 2012.

Exploring the function of IL-10, BTLA and DCs as regulators of immune responses

This thesis focuses on different aspects of immune regulation, both at the cellular and molecular levels. More specifically, this work concentrates on the importance of Interleukin-10, B and T Lymphocyte Attenuator (BTLA), and dendritic cells in respect to immune regulation, with special emphasis on autoimmunity. In this thesis, we show that the cellular source of IL10 production can dramatically influence the outcome of an autoimmune response. We show that T cell-derived IL10 plays an important role in controlling the viability of recently activated T cells, allowing them to become fully functional T effector cells. T cell-specific IL10-deficient mice failed to induce EAE when immunized with MOG peptide. Furthermore, when re-challenged with MOG or other stimuli, these T cells exhibited increased apoptosis rates. Here we report for the first time the generation of a novel mouse model that allows the conditional over-expression of BTLA. We show that BTLA can negatively regulate CD4+ T cells responses, when expressed by the T cells themselves. BTLA over-expression by CD8+ T cells or dendritic cells, however, resulted in enhanced viral clearance. In this study, we show that depletion of DCs, either early on from birth or later in adulthood, does not prevent EAE induction, but instead leads to a lower state of tolerance and stronger immune response. We also show that DCs are responsible for the upregulation of PD-1 on antigen-specific T cells and subsequently induce the formation of Tregs during immune responses.

Donor treatment with pegylated G-CSF augments the generation of IL-10-producing regulatory T cells and promotes transplantation tolerance

We investigated whether the protection from graft-versus-host disease (GVHD) afforded by donor treatment with granulocyte colony-stimulating factor (G-CSF) could be enhanced by dose escalation. Donor treatment with human G-CSIF prevented GVHD in the B6 --&gt; B6D2F1 murine model in a dose-dependent fashion, and murine G-CSF provided equivalent protection from GVHD at 10-fold lower doses. Donor pretreatment with a single dose of pegylated G-CSF (peg-G-CSF) prevented GVHD to a significantly greater extent than standard G-CSIF (survival, 75% versus 11%, P &lt; .001). Donor T cells from peg-G-CSF-treated donors failed to proliferate to alloantigen and inhibited the responses of control T cells in an interleukin 10 (IL-10)-dependent-fashion in vitro. T cells from peg-GCSF-treated IL-10(-/-) donors induced lethal GVHD; T cells from peg-G-CSF-treated wild-type (wt) donors promoted long-term survival. Whereas T cells from peg-G-CSF wt donors were able to regulate GVHD induced by T cells from control-treated donors, T cells from G-CSF-treated wt donors and peg-G-CSF-treated IL-10(-/-) donors did not prevent mortality. Thus, peg-G-CSF is markedly superior to standard G-CSF for the prevention of GVHD following allogeneic stem cell transplantation (SCT), due to the generation of IL-10-producing regulatory T cells. These data support prospective clinical trials of peg-G-CSF-mobilized allogeneic blood SCT. (C) 2004 by The American Society of Hematology.

IL-10-producing Regulatory B Cell Development in Human Autoimmune Disease

<p>B cell abnormalities contribute to the development and progress of autoimmune disease. Traditionally, the role of B cells in autoimmune disease was thought to be predominantly limited to the production of autoantibodies. Nevertheless, in addition to autoantibody production, B cells have other functions potentially relevant to autoimmunity. Such functions include antigen presentation to and activation of T cells, expression of costimulatory molecules and cytokine production. Recently, the ability of B cells to negatively regulate cellular immune responses and inflammation has been described and the concept of ���regulatory B cells��� has emerged. A variety of cytokines produced by regulatory B cell subsets have been reported with interleukin-10 (IL-10) being the most studied. IL-10-producing regulatory B cells predominantly localize within a rare CD1dhiCD5+ B cell subset in mice and the CD24hiCD27+ B cell subset in adult humans. This specific IL-10-producing subset of regulatory B cells have been named ���B10 cells��� to highlight that the regulatory function of these rare B cells is primarily mediated by IL-10, and to distinguish them from other regulatory B cell subsets that regulate immune responses through different mechanisms. B10 cells have been studies in a variety of animal models with autoimmune disease and clinical settings of human autoimmunity. There are many unsolved questions related to B10 cells including their surface phenotype, their origin and development in vivo, and their role in autoimmunity.</p><p>In Chapter 3 of this dissertation, the role of the B cell receptor (BCR) in B10 cell development is highlighted. First, the BCR repertoire of mouse peritoneal cavity B10 cells is examined by single cell sequencing; peritoneal cavity B10 cells have clonally diverse germline BCRs that are predominantly unmutated. Second, mouse B10 cells are shown to have higher frequencies of ��+ BCRs compared to non-B10 cells which may indicate the involvement of BCR light chain editing early in the process of B10 cell development in vivo. Third, human peripheral blood B10 cells are examined and are also found to express higher frequencies of �� chains compared to non-b10 cells. Therefore, B10 cell BCRs are clonally diverse and enriched for unmutated germline sequences and �� light chains.</p><p>In Chapter 4 of this dissertation, B10 cells are examined in the healthy developing human across the entire age range of infancy, childhood and adolescence, and in a large cohort of children with autoimmunity. The study of B10 cells in the developing human documents a massive transient expansion during middle childhood when up to 30% of blood B cells were competent to produce IL-10. The surface phenotype of pediatric B10 cells was variable and reflective of overall B cell development. B10 cells down-regulated CD4+ T cell interferon-gamma (IFN-��) production through IL-10-dependent pathways and IFN-�� inhibited whereas interleukin-21 (IL-21) promoted B cell IL-10 competency in vitro. Children with autoimmunity had a contracted B10 cell compartment, along with increased IFN-�� and decreased IL-21 serum levels compared to age-matched healthy controls. The decreased B10 cell frequencies and numbers in children with autoimmunity may be partially explained by the differential regulation of B10 cell development by IFN-�� and IL-21 and alterations in serum cytokine levels. The age-related changes of the B10 cell compartment during normal human development provide new insights into immune tolerance mechanisms involved in inflammation and autoimmunity.</p><p>These studies collectively demonstrate that BCR signals are the most important early determinant of B10 cell development in vivo, that human B10 cells are not a surface phenotype defined developmental B cell subset but a functionally defined regulatory B cell subset that regulates CD4+ T IFN-�� production through IL-10-dependent pathways and that human B10 cell development can be regulated by soluble factors in vivo such as the cytokine milieu. The findings of these studies provide new insights into immune tolerance mechanisms involved in human autoimmunity and the potent effects of IL-21 on human B cell IL-10 competence in vitro open new horizons in the development of autologous B10 cell-based therapies as an approach to treat human autoimmune disease in the future.</p>

Relationship between IL-10 gene -819C/T polymorphism and the risk of inflammatory bowel disease: a meta-analysis.

Background: The -819C/T polymorphism in interleukin 10 (IL-10) gene has been reported to be associated with inflammatory bowel disease (IBD) ,but the previous results are conflicting. Materials and Methods: The present study aimed at investigating the association between this polymorphism and risk of IBD using a meta-analysis.PubMed,Web of Science,EMBASE,google scholar and China National Knowledge Infrastructure (CNKI) databases were systematically searched to identify relevant publications from their inception to April 2016.Pooled odds ratio (OR) with 95% confidence interval (CI) was calculated using fixed- or random-effects models. Results: A total of 7 case-control studies containing 1890 patients and 2929 controls were enrolled into this meta-analysis, and our results showed no association between IL-10 gene -819C/T polymorphism and IBD risk(TT vs. CC:OR=0.81,95%CI 0.64- 1.04;CT vs. CC:OR=0.92,95%CI 0.81-1.05; Dominant model: OR=0.90,95%CI 0.80-1.02; Recessive model: OR=0.84,95%CI 0.66-1.06). In a subgroup analysis by nationality, the -819C/T polymorphism was not associated with IBD in both Asians and Caucasians. In the subgroup analysis stratified by IBD type, significant association was found in Crohn���s disease(CD)(CT vs. CC:OR=0.68,95%CI 0.48-0.97). Conclusion: In summary, the present meta-analysis suggests that the IL-10 gene -819C/T polymorphism may be associated with CD risk.

An Efficient Protocol For The Generation Of Monocyte Derived Dendritic Cells Using Serum-free Media For Clinical Applications In Post Remission Aml Patients.

Protocols for the generation of dendritic cells (DCs) using serum as a supplementation of culture media leads to reactions due to animal proteins and disease transmissions. Several types of serum-free media (SFM), based on good manufacture practices (GMP), have recently been used and seem to be a viable option. The aim of this study was to evaluate the results of the differentiation, maturation, and function of DCs from Acute Myeloid Leukemia patients (AML), generated in SFM and medium supplemented with autologous serum (AS). DCs were analyzed by phenotype characteristics, viability, and functionality. The results showed the possibility of generating viable DCs in all the conditions tested. In patients, the X-VIVO 15 medium was more efficient than the other media tested in the generation of DCs producing IL-12p70 (p=0.05). Moreover, the presence of AS led to a significant increase of IL-10 by DCs as compared with CellGro (p=0.05) and X-Vivo15 (p=0.05) media, both in patients and donors. We concluded that SFM was efficient in the production of DCs for immunotherapy in AML patients. However, the use of AS appears to interfere with the functional capacity of the generated DCs.

Monitoramento de par��metros imunol��gicos em atletas adolescentes de basquetebol durante e ap��s a temporada esportiva= : Monitoring of immunological parameters in adolescent basketball athletes during and after a sports season

Universidade Estadual de Campinas . Faculdade de Educa����o F��sica

Paracoccidioides brasilinsis-Induced Migration of Dendritic Cells and Subsequent T-Cell Activation in the Lung-Draining Lymph Nodes

Paracoccidioidomycosis is a mycotic disease caused by a dimorphic fungus, Paracoccidioides brasiliensis (Pb), that starts with inhalation of the fungus; thus, lung cells such as DC are part of the first line of defense against this microorganism. Migration of DC to the lymph nodes is the first step in initiating T cell responses. The mechanisms involved in resistance to Pb infection are poorly understood, but it is likely that DC play a pivotal role in the induction of effector T cells that control Pb infection. In this study, we showed that after Pb Infection, an important modification of lung DC receptor expression occurred. We observed an increased expression of CCR7 and CD103 on lung DC after infection, as well as MHC-II. After Pb infection, bone marrow-derived DC as well lung DC, migrate to lymph nodes. Migration of lung DC could represent an important mechanism of pathogenesis during PCM infection. In resume our data showed that Pb induced DC migration. Furthermore, we demonstrated that bone marrow-derived DC stimulated by Pb migrate to the lymph nodes and activate a T helper (Th) response. To the best of our knowledge, this is the first reported data showing that Pb induces migration of DC and activate a T helper (Th) response.

Prolonged Survival of Allografts Induced by Mycobacterial Hsp70 Is Dependent on CD4+CD25+Regulatory T Cells

Background: Heat shock proteins (Hsps) are stress induced proteins with immunomodulatory properties. The Hsp70 of Mycobacterium tuberculosis (TBHsp70) has been shown to have an anti-inflammatory role on rodent autoimmune arthritis models, and the protective effects were demonstrated to be dependent on interleukin-10 (IL-10). We have previously observed that TBHsp70 inhibited maturation of dendritic cells (DCs) and induced IL-10 production by these cells, as well as in synovial fluid cells. Methodology/Principal Findings: We investigated if TBHsp70 could inhibit allograft rejection in two murine allograft systems, a transplanted allogeneic melanoma and a regular skin allograft. In both systems, treatment with TBHsp70 significantly inhibited rejection of the graft, and correlated with regulatory T cells (Tregs) recruitment. This effect was not tumor mediated because injection of TBHsp70 in tumor-free mice induced an increase of Tregs in the draining lymph nodes as well as inhibition of proliferation of lymph node T cells and an increase in IL-10 production. Finally, TBHsp70 inhibited skin allograft acute rejection, and depletion of Tregs using a monoclonal antibody completely abolished this effect. Conclusions/Significance: We present the first evidence for an immunosuppressive role for this protein in a graft rejection system, using an innovative approach - immersion of the graft tissue in TBHsp70 solution instead of protein injection. Also, this is the first study that demonstrates dependence on Treg cells for the immunosuppressive role of TBHsp70. This finding is relevant for the elucidation of the immunomodulatory mechanism of TBHsp70. We propose that this protein can be used not only for chronic inflammatory diseases, but is also useful for organ transplantation management.

Severe Plasmodium vivax malaria exhibits marked inflammatory imbalance

Background: Despite clinical descriptions of severe vivax malaria cases having been reported, data regarding immunological and inflammatory patterns are scarce. In this report, the inflammatory and immunological status of both mild and severe vivax malaria cases are compared in order to explore immunopathological events in this disease. Methods and Results: Active and passive malaria case detections were performed during 2007 in Buritis, Rondonia, in the Brazilian Amazon. A total of 219 participants enrolled the study. Study individuals were classified according to the presence of Plasmodium vivax infection within four groups: non-infected (n = 90), asymptomatic (n = 60), mild (n = 50) and severe vivax infection (n = 19). A diagnosis of malaria was made by microscopy and molecular assays. Since at present no clear criteria define severe vivax malaria, this study adapted the consensual criteria from falciparum malaria. Patients with severe P. vivax infection were younger, had lived for shorter time in the endemic area, and recalled having experienced less previous malaria episodes than individuals with no malaria infection and with mild or asymptomatic infection. Strong linear trends were identified regarding increasing plasma levels of C reactive protein (CRP), serum creatinine, bilirubins and the graduation of disease severity. Plasma levels of tumour necrosis factor (TNF), interferon-gamma(IFN-gamma) and also IFN-gamma/interleukin-10 ratios were increased and exhibited a linear trend with gradual augmentation of disease severity. Both laboratory parameters of organ dysfunction and inflammatory cytokines were reduced during anti-parasite therapy in those patients with severe disease. Conclusion: Different clinical presentations of vivax malaria infection present strong association with activation of pro-inflammatory responses and cytokine imbalance. These findings are of utmost importance to improve current knowledge about physiopathological concepts of this serious widespread disease.

Hepatitis B Infection Is Associated with Asymptomatic Malaria in the Brazilian Amazon

Background: Areas that are endemic for malaria are also highly endemic for hepatitis B virus (HBV) infection. Nevertheless, it is unknown whether HBV infection modifies the clinical presentation of malaria. This study aimed to address this question. Methodology and Findings: An observational study of 636 individuals was performed in Rondonia, western Amazon, Brazil between 2006 and 2007. Active and passive case detections identified Plasmodium infection by field microscopy and nested Polymerase Chain Reaction (PCR). HBV infections were identified by serology and confirmed by real-time PCR. Epidemiological information and plasma cytokine profiles were studied. The data were analyzed using adjusted multinomial logistic regression. Plasmodium-infected individuals with active HBV infection were more likely to be asymptomatic (OR: 120.13, P < 0.0001), present with lower levels of parasitemia and demonstrate a decreased inflammatory cytokine profile. Nevertheless, co-infected individuals presented higher HBV viremia. Plasmodium parasitemia inversely correlated with plasma HBV DNA levels (r=-0.6; P=0.0003). Conclusion: HBV infection diminishes the intensity of malaria infection in individuals from this endemic area. This effect seems related to cytokine balance and control of inflammatory responses. These findings add important insights to the understanding of the factors affecting the clinical outcomes of malaria in endemic regions.