Limits...
The involvement of TLR2 and TLR4 in cytokine and nitric oxide production in visceral leishmaniasis patients before and after treatment with anti-leishmanial drugs.

Gatto M, de Abreu MM, Tasca KI, de Assis Golim M, da Silva LD, Simão JC, Fortaleza CM, de Campos Soares ÂM, Calvi SA - PLoS ONE (2015)

Bottom Line: Before treatment, the production of TNF-α and NO was associated with TLR2 and TLR4 expression, while IL-10 production was only associated with TLR2 expression.After treatment, both receptors were associated with the production of TNF-α, IFN-γ, IL-10 and NO, while the production of IL-17 was associated only with TLR4 expression.The results presented in this study suggest that both TLR2 and TLR4 participate in the modulation of cytokine and NO production in VL patients, contributing to the pathogenesis of VL prior to treatment and the protective immune response after treatment.

View Article: PubMed Central - PubMed

Affiliation: Tropical Diseases Department, Botucatu School of Medicine-UNESP, Botucatu, São Paulo, Brazil.

ABSTRACT
Toll-like receptors (TLRs) have significant involvement in Leishmania infection, although little is known about the relationship between these receptors, cytokines and nitric oxide (NO) in patients with visceral leishmaniasis (VL) before or after treatment with anti-leishmanial drugs. The goal of this study was to evaluate the expression of TLR2 and TLR4 in CD3+ and CD14+ cells and the production of TNF-α, IFN-γ, IL-17, IL-10, TGF-β and NO in peripheral blood mononuclear cells (PBMCs) from VL patients pre- and post-treatment with anti-leishmanial drugs. In addition, we investigated whether these receptors were involved in the production of these cytokines and NO. In the active VL patients, increased TLR2 and TLR4 expression in lymphocytes and monocytes, increased production of TNF-α, IL-10 and TGF-β and decreased production of IFN-γ, IL-17 and NO were observed. After treatment, TLR2 and TLR4 were still expressed in lymphocytes and monocytes, the TNF-α and IL-10 levels were lower, the production of IFN-γ, IL-17 and NO was higher, and the TGF-β level remained high. Before treatment, the production of TNF-α and NO was associated with TLR2 and TLR4 expression, while IL-10 production was only associated with TLR2 expression. After treatment, both receptors were associated with the production of TNF-α, IFN-γ, IL-10 and NO, while the production of IL-17 was associated only with TLR4 expression. The results presented in this study suggest that both TLR2 and TLR4 participate in the modulation of cytokine and NO production in VL patients, contributing to the pathogenesis of VL prior to treatment and the protective immune response after treatment.

Show MeSH

Related in: MedlinePlus

NO production after stimulation with LPS and PGN agonists.Levels of NO in supernatants were analyzed after 24 hours of PBMCs cultured (1x106 cells/ml), obtained from patients with VL pre-treatment, post-treatment and control subjects, and stimulated or not with LPS ultra-purified (1 μg/ml), PGN (5 μg/ml) and LPS+PGN. Lowercase letters represent significant differences among agonists in the same group: p<0.05 a—PBMC vs. LPS, PGN, LPS+PGN. Capital letters represent significant differences in the same agonists among groups: p<0.05 A—pre-treatment vs. post-treatment; B—pre-treatment vs. control individuals. Each dot represent a different patient and each bar represents the median. NO levels were measured by the levels of nitrite/nitrate and the data are representative of triplicates.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4338033&req=5

pone.0117977.g003: NO production after stimulation with LPS and PGN agonists.Levels of NO in supernatants were analyzed after 24 hours of PBMCs cultured (1x106 cells/ml), obtained from patients with VL pre-treatment, post-treatment and control subjects, and stimulated or not with LPS ultra-purified (1 μg/ml), PGN (5 μg/ml) and LPS+PGN. Lowercase letters represent significant differences among agonists in the same group: p<0.05 a—PBMC vs. LPS, PGN, LPS+PGN. Capital letters represent significant differences in the same agonists among groups: p<0.05 A—pre-treatment vs. post-treatment; B—pre-treatment vs. control individuals. Each dot represent a different patient and each bar represents the median. NO levels were measured by the levels of nitrite/nitrate and the data are representative of triplicates.

Mentions: NO production was measured in the supernatants of cultures of PBMC from patients with VL pre- and post-treatment (Fig. 3). We demonstrated that unstimulated cells obtained from patients prior to treatment produced significantly lower levels of NO (p = 0.04) compared with cells obtained post-treatment. When LPS and PGN were used as stimuli, PBMCs from pre-treatment patients produced lower NO levels (p = 0.002, p = 0.02) compared with control individuals. Furthermore, PBMCs from pre-treatment patients stimulated with PGN produced lower NO levels (p = 0.03) than PBMCs from post-treatment patients. The involvement of TLR2 and TLR4 in NO production following stimulation with the agonists PGN and LPS, respectively, was also verified. The results showed that stimulation with LPS, PGN and LPS+PGN significantly increased (p<0.05) the levels of NO compared with unstimulated cells in pre- and post-treatment patients and control individuals, suggesting that both TLR2 and TLR4 are involved in the production this metabolite.


The involvement of TLR2 and TLR4 in cytokine and nitric oxide production in visceral leishmaniasis patients before and after treatment with anti-leishmanial drugs.

Gatto M, de Abreu MM, Tasca KI, de Assis Golim M, da Silva LD, Simão JC, Fortaleza CM, de Campos Soares ÂM, Calvi SA - PLoS ONE (2015)

NO production after stimulation with LPS and PGN agonists.Levels of NO in supernatants were analyzed after 24 hours of PBMCs cultured (1x106 cells/ml), obtained from patients with VL pre-treatment, post-treatment and control subjects, and stimulated or not with LPS ultra-purified (1 μg/ml), PGN (5 μg/ml) and LPS+PGN. Lowercase letters represent significant differences among agonists in the same group: p<0.05 a—PBMC vs. LPS, PGN, LPS+PGN. Capital letters represent significant differences in the same agonists among groups: p<0.05 A—pre-treatment vs. post-treatment; B—pre-treatment vs. control individuals. Each dot represent a different patient and each bar represents the median. NO levels were measured by the levels of nitrite/nitrate and the data are representative of triplicates.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4338033&req=5

pone.0117977.g003: NO production after stimulation with LPS and PGN agonists.Levels of NO in supernatants were analyzed after 24 hours of PBMCs cultured (1x106 cells/ml), obtained from patients with VL pre-treatment, post-treatment and control subjects, and stimulated or not with LPS ultra-purified (1 μg/ml), PGN (5 μg/ml) and LPS+PGN. Lowercase letters represent significant differences among agonists in the same group: p<0.05 a—PBMC vs. LPS, PGN, LPS+PGN. Capital letters represent significant differences in the same agonists among groups: p<0.05 A—pre-treatment vs. post-treatment; B—pre-treatment vs. control individuals. Each dot represent a different patient and each bar represents the median. NO levels were measured by the levels of nitrite/nitrate and the data are representative of triplicates.
Mentions: NO production was measured in the supernatants of cultures of PBMC from patients with VL pre- and post-treatment (Fig. 3). We demonstrated that unstimulated cells obtained from patients prior to treatment produced significantly lower levels of NO (p = 0.04) compared with cells obtained post-treatment. When LPS and PGN were used as stimuli, PBMCs from pre-treatment patients produced lower NO levels (p = 0.002, p = 0.02) compared with control individuals. Furthermore, PBMCs from pre-treatment patients stimulated with PGN produced lower NO levels (p = 0.03) than PBMCs from post-treatment patients. The involvement of TLR2 and TLR4 in NO production following stimulation with the agonists PGN and LPS, respectively, was also verified. The results showed that stimulation with LPS, PGN and LPS+PGN significantly increased (p<0.05) the levels of NO compared with unstimulated cells in pre- and post-treatment patients and control individuals, suggesting that both TLR2 and TLR4 are involved in the production this metabolite.

Bottom Line: Before treatment, the production of TNF-α and NO was associated with TLR2 and TLR4 expression, while IL-10 production was only associated with TLR2 expression.After treatment, both receptors were associated with the production of TNF-α, IFN-γ, IL-10 and NO, while the production of IL-17 was associated only with TLR4 expression.The results presented in this study suggest that both TLR2 and TLR4 participate in the modulation of cytokine and NO production in VL patients, contributing to the pathogenesis of VL prior to treatment and the protective immune response after treatment.

View Article: PubMed Central - PubMed

Affiliation: Tropical Diseases Department, Botucatu School of Medicine-UNESP, Botucatu, São Paulo, Brazil.

ABSTRACT
Toll-like receptors (TLRs) have significant involvement in Leishmania infection, although little is known about the relationship between these receptors, cytokines and nitric oxide (NO) in patients with visceral leishmaniasis (VL) before or after treatment with anti-leishmanial drugs. The goal of this study was to evaluate the expression of TLR2 and TLR4 in CD3+ and CD14+ cells and the production of TNF-α, IFN-γ, IL-17, IL-10, TGF-β and NO in peripheral blood mononuclear cells (PBMCs) from VL patients pre- and post-treatment with anti-leishmanial drugs. In addition, we investigated whether these receptors were involved in the production of these cytokines and NO. In the active VL patients, increased TLR2 and TLR4 expression in lymphocytes and monocytes, increased production of TNF-α, IL-10 and TGF-β and decreased production of IFN-γ, IL-17 and NO were observed. After treatment, TLR2 and TLR4 were still expressed in lymphocytes and monocytes, the TNF-α and IL-10 levels were lower, the production of IFN-γ, IL-17 and NO was higher, and the TGF-β level remained high. Before treatment, the production of TNF-α and NO was associated with TLR2 and TLR4 expression, while IL-10 production was only associated with TLR2 expression. After treatment, both receptors were associated with the production of TNF-α, IFN-γ, IL-10 and NO, while the production of IL-17 was associated only with TLR4 expression. The results presented in this study suggest that both TLR2 and TLR4 participate in the modulation of cytokine and NO production in VL patients, contributing to the pathogenesis of VL prior to treatment and the protective immune response after treatment.

Show MeSH
Related in: MedlinePlus