Propolis Immunomodulatory Action In Vivo on Toll-Like Receptors 2 and 4 Expression and on Pro-Inflammatory Cytokines Production in Mice

Propolis is a bee product and its immunomodulatory action has been the subject of intense investigation lately. The recent discovery and characterization of the family of Toll-like receptors (TLR) have triggered a great deal of interest in the field of innate immunity due to their crucial role in microbial recognition and development of the adaptive immune response. This work aimed to evaluate propolis's effect on TLR-2 and TLR-4 expression and on the production of pro-inflammatory cytokines (IL-1 beta and IL-6). Male BALB/c mice were treated with propolis (200 mg/kg) for three consecutive days, and TLR-2 and TLR-4 expression as well as IL-1 beta and IL-6 production were assessed in peritoneal macrophages and spleen cells. Basal IL-1 beta production and TLR-2 and TLR-4 expression were increased in peritoneal macrophages of propolis-treated mice. TLR-2 and TLR-4 expression and IL-1 beta and IL-6 production were also upregulated in the spleen cells of propolis-treated mice. One may conclude that propolis activated the initial steps of the immune response by upregulating TLRs expression and the production of pro-inflammatory cytokines in mice, modulating the mechanisms of the innate immunity. Copyright (C) 2009 John Wiley & Sons, Ltd.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP

Paracoccidioides brasiliensis interferes on dendritic cells maturation by inhibiting PGE(2) production

Paracoccidioidomycosis (PCM) is a systemic mycosis, endemic in most Latin American countries, especially in Brazil, whose etiologic agent is the thermodimorphic fungus of the genus Paracoccidioides, comprising cryptic species of Paracoccidioides brasiliensis, S1, PS2, PS3 and Paracoccidioides lutzii. The mechanisms involved in the initial interaction of the fungus with cells of the innate immune response, as dendritic cells (DCs), deserve to be studied. Prostaglandins (PGs) are eicosanoids that play an important role in modulating functions of immune cells including DCs. Here we found that human immature DCs derived from the differentiation of monocytes cultured with GM-CSF and IL-4 release substantial concentrations of PGE2, which, however, were significantly inhibited after challenge with P. brasiliensis. In vitro blocking of pattern recognition receptors (PRRs) by monoclonal antibodies showed the involvement of mannose receptor (MR) in PGE2 inhibition by the fungus. In addition, phenotyping assays showed that after challenge with the fungus, DCs do not change their phenotype of immature cells to mature ones, as well as do not produce IL-12 p70 or adequate concentrations of TNF-alpha. Assays using exogenous PGE2 confirmed an association between PGE2 inhibition and failure of cells to phenotypically mature in response to P. brasiliensis. We conclude that a P. brasiliensis evasion mechanism exists associated to a dysregulation on DC maturation. These findings may provide novel information for the understanding of the complex interplay between the host and this fungus.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP

Human neutrophils produce IL-12, IL-10, PGE2 and LTB4 in response to Paracoccidioides brasiliensis. Involvement of TLR2, mannose receptor and dectin-1

The functions of phagocytic cells against pathogens are initiated by the interaction between membrane receptors and molecular structures which compose the cell wall of these microorganisms. Thus our study aimed to identify the neutrophil receptors involved in the recognition of different strains of Paracoccidioides brasiliensis and the consequent modulation of immune response through the production of cytokines and inflammatory mediators. Neutrophils did not produce TNF-alfa in response to both strains. However, these cells produce IL-12, mainly in tesponse to Pb 265, with participation of TLR2 and dectin-1. These cells also produce L-10, whose levels were higher for Pb 18 with involvement of TLR2 and MR and only TLR2 for Pb 265. The production of PGE2 and LTB4 was detected similarly for the two strains. For PGE2, MR and dectin-1 were involved, while in relation to LTB4, none of them. In summary, we demonstrated that neutrophils have a dynamic role during host immune response to P. brasiliensis, since in addition to their role as effector cells of innate immunity; they have the capacity to modulate innate and adaptative immune response against this fungus by producing cytokines and lipidic mediators. This modulation may be toward a pro- or anti-inflammatory pattern in a dependence of P. brasiliensis strains and PRR involved in fungus recognition by these cells. (C) 2014 Elsevier Ltd. All rights reserved.Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES

TNF-α production by DCs challenged with Pb18 or Pb265 or activated by LPS for 48 h, and measured by ELISA.

<p>The results are expressed as mean ± SD of independent experiments performed with cells obtained from 4 subjects. Statistically significant differences between groups are indicated: *p< 0.05.</p

Effect of exogenous PGE₂ on percentage of cells, mean of fluorescence intensity (MFI) relative to the expression of CD40, CD80 (A) CD83, CD86 (B), HLA-DR, CXCR4 (C) CCR5, and CCR7 (D) by DCs after challenge or not with Pb18 and Pb265.

<p>The results are expressed in mean ± SD of experiments performed with cells obtained from 4 subjects. Statistically significant differences between groups are indicated: *p< 0.05 versus without PGE₂.</p

Effect of exogenous PGE₂ on TNF-α production by DCs challenged with Pb18 or Pb265 by 48 h and evaluated by ELISA.

<p>The results are expressed in mean ± SD of experiments performed with cells obtained from 4 subjects. Statistically significant differences between groups are indicated: *p< 0.05 versus without PGE₂.</p

Involvement of MR, TLR2, Dectin-1 and DC-SIGN on PGE₂ production inhibition induced by <i>P</i>. <i>brasiliensis</i> in human DCs.

<p>Cells were incubated with anti-MR, anti-TLR2, anti-Dectin-1 and/or anti-DC-SIGN monoclonal antibodies for 1 h, challenged with Pb18 or Pb265 (DCs/yeast ratio 5:1) for 4 and 24 h, and evaluated for PGE₂ production by ELISA. The results are expressed in mean ± SD of independent experiments performed with cells obtained from 4 subjects. Statistically significant differences between groups are indicated: *p< 0.05 versus control DCs and other available receptors.</p

PGE₂ production by DCs activated with LPS or challenged with Pb18 or Pb265 (DCs/Pb ratio 5:1) for different periods.

<p>The results are expressed in mean ± SD of experiments performed with cells from 5 subjects. Statistically significant differences between groups in the same period are indicated: *p< 0.05; ** p< 0.01; ***p<0.001 versus control DC; o p< 0.05 versus Pb18.</p