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Differential effects of the Toll-like receptor 2 agonists, PGN and Pam3CSK4 on anti-IgE induced human mast cell activation.

Yu Y, Yip KH, Tam IY, Sam SW, Ng CW, Zhang W, Lau HY - PLoS ONE (2014)

Bottom Line: Pretreatment of PGN and Pam3CSK4 inhibited anti-IgE induced calcium mobilization and degranulation without down-regulation of FcεRI expression.Pam3CSK4 but not PGN acted in synergy with anti-IgE for IL-8 release when the TLR2 agonist was added simultaneously with anti-IgE.Studies with inhibitors of key enzymes implicated in mast cell signaling revealed that the synergistic release of IL-8 induced by Pam3CSK4 and anti-IgE involved ERK and calcineurin signaling cascades.

View Article: PubMed Central - PubMed

Affiliation: Shenzhen Key Laboratory for Translational Medicine of Dermatology, Biomedical Research Institute, Shenzhen Peking University-The Hong Kong University of Science and Technology Medical Center, Shenzhen, Guangdong Province, China; School of Biomedical Sciences, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong, SAR, China.

ABSTRACT
Mast cells are pivotal in the pathogenesis of allergy and inflammation. In addition to the classical IgE-dependent mechanism involving crosslinking of the high-affinity receptor for IgE (FcεRI), mast cells are also activated by Toll-like receptors (TLRs) which are at the center of innate immunity. In this study, we demonstrated that the response of LAD2 cells (a human mast cell line) to anti-IgE was altered in the presence of the TLR2 agonists peptidoglycan (PGN) and tripalmitoyl-S-glycero-Cys-(Lys)4 (Pam3CSK4). Pretreatment of PGN and Pam3CSK4 inhibited anti-IgE induced calcium mobilization and degranulation without down-regulation of FcεRI expression. Pam3CSK4 but not PGN acted in synergy with anti-IgE for IL-8 release when the TLR2 agonist was added simultaneously with anti-IgE. Studies with inhibitors of key enzymes implicated in mast cell signaling revealed that the synergistic release of IL-8 induced by Pam3CSK4 and anti-IgE involved ERK and calcineurin signaling cascades. The differential modulations of anti-IgE induced mast cell activation by PGN and Pam3CSK4 suggest that dimerization of TLR2 with TLR1 or TLR6 produced different modulating actions on FcεRI mediated human mast cell activation.

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Effect of TLR2 ligands on anti-IgE induced degranulation and IL-8 release from LAD2 cells.(A, B) LAD2 cells were incubated with only PGN or Pam3CSK4 for 30 min (○). LAD2 cells were incubated with anti-IgE (2 µg/ml) at the same time (▴) or after 24 h pre-incubation (•) with PGN or Pam3CSK4. The levels of β-hex release induced by anti-IgE alone and in the present of TLR2 ligands were compared with one-way ANOVA and Dunnett’s multiple comparison tests. *p<0.05, **p<0.01, ***p<0.001 (n = 3–5). (C, D) LAD2 cells were incubated alone with anti-IgE (2 µg/ml, •), PGN/Pam3CSK4 (▪) or combination of anti-IgE with PGN/Pam3CSK4 (▴) for 24 h. Two-way ANOVA and Bonferroni posttests were applied to compare the actual amount of IL-8 released with the predicted value obtained by adding the individual amounts released by anti-IgE and PGN or Pam3CSK4 (○). **p<0.01 (n = 5).
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pone-0112989-g001: Effect of TLR2 ligands on anti-IgE induced degranulation and IL-8 release from LAD2 cells.(A, B) LAD2 cells were incubated with only PGN or Pam3CSK4 for 30 min (○). LAD2 cells were incubated with anti-IgE (2 µg/ml) at the same time (▴) or after 24 h pre-incubation (•) with PGN or Pam3CSK4. The levels of β-hex release induced by anti-IgE alone and in the present of TLR2 ligands were compared with one-way ANOVA and Dunnett’s multiple comparison tests. *p<0.05, **p<0.01, ***p<0.001 (n = 3–5). (C, D) LAD2 cells were incubated alone with anti-IgE (2 µg/ml, •), PGN/Pam3CSK4 (▪) or combination of anti-IgE with PGN/Pam3CSK4 (▴) for 24 h. Two-way ANOVA and Bonferroni posttests were applied to compare the actual amount of IL-8 released with the predicted value obtained by adding the individual amounts released by anti-IgE and PGN or Pam3CSK4 (○). **p<0.01 (n = 5).

Mentions: PGN and Pam3CSK4 did not induce significant release of β-hex on their own (Fig. 1A, B) while anti-IgE induced a release of around 30% of total β-hex after a 30 min incubation with LAD2 cells. When added to LAD2 cells simultaneously with anti-IgE, PGN had no significant effect on anti-IgE induced β-hex release while Pam3CSK4 inhibited the release only at the highest concentration of 20 µg/ml tested. In contrast, dose-dependent inhibition of anti-IgE induced β-hex release from LAD2 cells was observed with both TLR2 ligands when LAD2 cells were activated after pre-incubation with the TLR2 ligands for 24 h. (Fig. 1A, B).


Differential effects of the Toll-like receptor 2 agonists, PGN and Pam3CSK4 on anti-IgE induced human mast cell activation.

Yu Y, Yip KH, Tam IY, Sam SW, Ng CW, Zhang W, Lau HY - PLoS ONE (2014)

Effect of TLR2 ligands on anti-IgE induced degranulation and IL-8 release from LAD2 cells.(A, B) LAD2 cells were incubated with only PGN or Pam3CSK4 for 30 min (○). LAD2 cells were incubated with anti-IgE (2 µg/ml) at the same time (▴) or after 24 h pre-incubation (•) with PGN or Pam3CSK4. The levels of β-hex release induced by anti-IgE alone and in the present of TLR2 ligands were compared with one-way ANOVA and Dunnett’s multiple comparison tests. *p<0.05, **p<0.01, ***p<0.001 (n = 3–5). (C, D) LAD2 cells were incubated alone with anti-IgE (2 µg/ml, •), PGN/Pam3CSK4 (▪) or combination of anti-IgE with PGN/Pam3CSK4 (▴) for 24 h. Two-way ANOVA and Bonferroni posttests were applied to compare the actual amount of IL-8 released with the predicted value obtained by adding the individual amounts released by anti-IgE and PGN or Pam3CSK4 (○). **p<0.01 (n = 5).
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4232580&req=5

pone-0112989-g001: Effect of TLR2 ligands on anti-IgE induced degranulation and IL-8 release from LAD2 cells.(A, B) LAD2 cells were incubated with only PGN or Pam3CSK4 for 30 min (○). LAD2 cells were incubated with anti-IgE (2 µg/ml) at the same time (▴) or after 24 h pre-incubation (•) with PGN or Pam3CSK4. The levels of β-hex release induced by anti-IgE alone and in the present of TLR2 ligands were compared with one-way ANOVA and Dunnett’s multiple comparison tests. *p<0.05, **p<0.01, ***p<0.001 (n = 3–5). (C, D) LAD2 cells were incubated alone with anti-IgE (2 µg/ml, •), PGN/Pam3CSK4 (▪) or combination of anti-IgE with PGN/Pam3CSK4 (▴) for 24 h. Two-way ANOVA and Bonferroni posttests were applied to compare the actual amount of IL-8 released with the predicted value obtained by adding the individual amounts released by anti-IgE and PGN or Pam3CSK4 (○). **p<0.01 (n = 5).
Mentions: PGN and Pam3CSK4 did not induce significant release of β-hex on their own (Fig. 1A, B) while anti-IgE induced a release of around 30% of total β-hex after a 30 min incubation with LAD2 cells. When added to LAD2 cells simultaneously with anti-IgE, PGN had no significant effect on anti-IgE induced β-hex release while Pam3CSK4 inhibited the release only at the highest concentration of 20 µg/ml tested. In contrast, dose-dependent inhibition of anti-IgE induced β-hex release from LAD2 cells was observed with both TLR2 ligands when LAD2 cells were activated after pre-incubation with the TLR2 ligands for 24 h. (Fig. 1A, B).

Bottom Line: Pretreatment of PGN and Pam3CSK4 inhibited anti-IgE induced calcium mobilization and degranulation without down-regulation of FcεRI expression.Pam3CSK4 but not PGN acted in synergy with anti-IgE for IL-8 release when the TLR2 agonist was added simultaneously with anti-IgE.Studies with inhibitors of key enzymes implicated in mast cell signaling revealed that the synergistic release of IL-8 induced by Pam3CSK4 and anti-IgE involved ERK and calcineurin signaling cascades.

View Article: PubMed Central - PubMed

Affiliation: Shenzhen Key Laboratory for Translational Medicine of Dermatology, Biomedical Research Institute, Shenzhen Peking University-The Hong Kong University of Science and Technology Medical Center, Shenzhen, Guangdong Province, China; School of Biomedical Sciences, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong, SAR, China.

ABSTRACT
Mast cells are pivotal in the pathogenesis of allergy and inflammation. In addition to the classical IgE-dependent mechanism involving crosslinking of the high-affinity receptor for IgE (FcεRI), mast cells are also activated by Toll-like receptors (TLRs) which are at the center of innate immunity. In this study, we demonstrated that the response of LAD2 cells (a human mast cell line) to anti-IgE was altered in the presence of the TLR2 agonists peptidoglycan (PGN) and tripalmitoyl-S-glycero-Cys-(Lys)4 (Pam3CSK4). Pretreatment of PGN and Pam3CSK4 inhibited anti-IgE induced calcium mobilization and degranulation without down-regulation of FcεRI expression. Pam3CSK4 but not PGN acted in synergy with anti-IgE for IL-8 release when the TLR2 agonist was added simultaneously with anti-IgE. Studies with inhibitors of key enzymes implicated in mast cell signaling revealed that the synergistic release of IL-8 induced by Pam3CSK4 and anti-IgE involved ERK and calcineurin signaling cascades. The differential modulations of anti-IgE induced mast cell activation by PGN and Pam3CSK4 suggest that dimerization of TLR2 with TLR1 or TLR6 produced different modulating actions on FcεRI mediated human mast cell activation.

Show MeSH
Related in: MedlinePlus