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Heat Shock Protein 60 in Eggs Specifically Induces Tregs and Reduces Liver Immunopathology in Mice with Schistosomiasis Japonica.

Zhou S, Jin X, Chen X, Zhu J, Xu Z, Wang X, Liu F, Hu W, Zhou L, Su C - PLoS ONE (2015)

Bottom Line: In addition, we identified that SjHSP60 derived from SEA significantly induces Tregs in vivo and in vitro.SjHSP60 specially induced Tregs in vivo and in vitro without inducing other CD4+ T sub-populations including Th1, Th2 and Th17 cells.Furthermore, we showed that the SjHSP60-depleted SEA almost lost the ability in vitro and displayed a significant impaired ability to induce Tregs in vivo.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathogen Biology and Immunology, Jiangsu Key Laboratory of Pathogen Biology, Nanjing Medical University, Nanjing, Jiangsu, China.

ABSTRACT

Background: Parasitic helminths need to suppress the host immune system to establish chronic infections. Paradoxically, immunosuppression induced by the worm also benefits the host by limiting excessive inflammation and tissue damage, which remains the major cause leading to serious morbidity and mortality. Regulatory T cells (Tregs) are key immune regulators of this mutualism. The successive rise in Tregs during schistosome infection plays a critical role in immunoregulation. We and others previously showed that Schistosoma japonicum (S. japonicum) egg antigens (SEA) induce Tregs both in vitro and in vivo. In addition, we identified that SjHSP60 derived from SEA significantly induces Tregs in vivo and in vitro. However, the contribution of SjHSP60 in SEA to Treg induction and the related mechanisms of the Treg induction have not yet been identified.

Methodology/principal findings: In this study, we showed that S. japonicum stress protein HSP60 (SjHSP60) was constitutively and extensively expressed in eggs of S. japonicum. SjHSP60 specially induced Tregs in vivo and in vitro without inducing other CD4+ T sub-populations including Th1, Th2 and Th17 cells. Furthermore, we showed that the SjHSP60-depleted SEA almost lost the ability in vitro and displayed a significant impaired ability to induce Tregs in vivo. Finally, our study illustrated that the mechanisms of SjHSP60-mediated induction of Tregs are through both conversion of CD4+CD25- T cells into CD4+CD25+Foxp3+ Tregs and expansion of preexisting CD4+CD25+Foxp3+ Tregs in a TLR4-dependent manner.

Conclusions/significance: Collectively, our findings identify SjHSP60 as a major parasitic contributor of Treg induction in S. japonicum egg antigens, which not only contributes to the better understanding of the mechanism of immunoregulation during helminth infection, but also suggests its potential as a therapeutic target for control of immunopathology, allergic and autoimmune diseases.

No MeSH data available.


Related in: MedlinePlus

TLR4 signaling is necessary for SjHSP60-mediated Treg increase.(A and B) TLR2-/- or TLR4-/- mice or their WT control littermates were immunized with SjHSP60 or PBS. Ten days after the final injection, CD4+CD25+Foxp3+ Tregs in spleen of individual mice were analyzed by FCM. (C) Purified CD4+CD25- T cells were co-cultured with purified APCs (DC, Mφ, TLR2-/- Mφ, TLR4-/- Mφ, or Mφ pre-treated with anti-TLR2, anti-TLR4 or their isotype control antibodies) at a 2:1 ratio of T cell:APC (DC or Mφ) with or without stimulation of SjHSP60 (0.2 μg/ml). After 3 days, Tregs in co-cultures were analyzed by FCM. (D-F) Graphs of CD4+CD25+Foxp3+ Treg frequencies. (G and H) CD4+CD25+ Tregs purified from mice deficient for TLR2, TLR4 or their control littermates were cultured in medium alone or stimulated with 1 μg/ml anti-CD3 and/or 0.5 μg/ml SjHSP60. On day 3, proliferation of CD4+CD25+ Tregs was determined by [3H]thymidine incorporation. FCM plots are representative of one of three independent experiments. Cells were gated on CD4+ T cells. The bar graph of average percentages ± SD of CD4+CD25+Foxp3+ Tregs are from 5 mice or triplicate cultures representative of one of three independent experiments. **P < 0.01, ***P < 0.001.
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pone.0139133.g005: TLR4 signaling is necessary for SjHSP60-mediated Treg increase.(A and B) TLR2-/- or TLR4-/- mice or their WT control littermates were immunized with SjHSP60 or PBS. Ten days after the final injection, CD4+CD25+Foxp3+ Tregs in spleen of individual mice were analyzed by FCM. (C) Purified CD4+CD25- T cells were co-cultured with purified APCs (DC, Mφ, TLR2-/- Mφ, TLR4-/- Mφ, or Mφ pre-treated with anti-TLR2, anti-TLR4 or their isotype control antibodies) at a 2:1 ratio of T cell:APC (DC or Mφ) with or without stimulation of SjHSP60 (0.2 μg/ml). After 3 days, Tregs in co-cultures were analyzed by FCM. (D-F) Graphs of CD4+CD25+Foxp3+ Treg frequencies. (G and H) CD4+CD25+ Tregs purified from mice deficient for TLR2, TLR4 or their control littermates were cultured in medium alone or stimulated with 1 μg/ml anti-CD3 and/or 0.5 μg/ml SjHSP60. On day 3, proliferation of CD4+CD25+ Tregs was determined by [3H]thymidine incorporation. FCM plots are representative of one of three independent experiments. Cells were gated on CD4+ T cells. The bar graph of average percentages ± SD of CD4+CD25+Foxp3+ Tregs are from 5 mice or triplicate cultures representative of one of three independent experiments. **P < 0.01, ***P < 0.001.

Mentions: Studies have demonstrated that HSP60 is a natural ligand for TLR2 and TLR4 [35]. To further investigate whether TLR2 and/or TLR4 are involved in SjHSP60-mediated Treg increase, we injected TLR2-/-, TLR4-/- or WT mice with SjHSP60 or PBS and then examined Treg frequency. FCM analysis showed that Tregs were significantly increased in TLR2-/- and WT mice, but not in TLR4-/- mice (Fig 5A and 5B).


Heat Shock Protein 60 in Eggs Specifically Induces Tregs and Reduces Liver Immunopathology in Mice with Schistosomiasis Japonica.

Zhou S, Jin X, Chen X, Zhu J, Xu Z, Wang X, Liu F, Hu W, Zhou L, Su C - PLoS ONE (2015)

TLR4 signaling is necessary for SjHSP60-mediated Treg increase.(A and B) TLR2-/- or TLR4-/- mice or their WT control littermates were immunized with SjHSP60 or PBS. Ten days after the final injection, CD4+CD25+Foxp3+ Tregs in spleen of individual mice were analyzed by FCM. (C) Purified CD4+CD25- T cells were co-cultured with purified APCs (DC, Mφ, TLR2-/- Mφ, TLR4-/- Mφ, or Mφ pre-treated with anti-TLR2, anti-TLR4 or their isotype control antibodies) at a 2:1 ratio of T cell:APC (DC or Mφ) with or without stimulation of SjHSP60 (0.2 μg/ml). After 3 days, Tregs in co-cultures were analyzed by FCM. (D-F) Graphs of CD4+CD25+Foxp3+ Treg frequencies. (G and H) CD4+CD25+ Tregs purified from mice deficient for TLR2, TLR4 or their control littermates were cultured in medium alone or stimulated with 1 μg/ml anti-CD3 and/or 0.5 μg/ml SjHSP60. On day 3, proliferation of CD4+CD25+ Tregs was determined by [3H]thymidine incorporation. FCM plots are representative of one of three independent experiments. Cells were gated on CD4+ T cells. The bar graph of average percentages ± SD of CD4+CD25+Foxp3+ Tregs are from 5 mice or triplicate cultures representative of one of three independent experiments. **P < 0.01, ***P < 0.001.
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pone.0139133.g005: TLR4 signaling is necessary for SjHSP60-mediated Treg increase.(A and B) TLR2-/- or TLR4-/- mice or their WT control littermates were immunized with SjHSP60 or PBS. Ten days after the final injection, CD4+CD25+Foxp3+ Tregs in spleen of individual mice were analyzed by FCM. (C) Purified CD4+CD25- T cells were co-cultured with purified APCs (DC, Mφ, TLR2-/- Mφ, TLR4-/- Mφ, or Mφ pre-treated with anti-TLR2, anti-TLR4 or their isotype control antibodies) at a 2:1 ratio of T cell:APC (DC or Mφ) with or without stimulation of SjHSP60 (0.2 μg/ml). After 3 days, Tregs in co-cultures were analyzed by FCM. (D-F) Graphs of CD4+CD25+Foxp3+ Treg frequencies. (G and H) CD4+CD25+ Tregs purified from mice deficient for TLR2, TLR4 or their control littermates were cultured in medium alone or stimulated with 1 μg/ml anti-CD3 and/or 0.5 μg/ml SjHSP60. On day 3, proliferation of CD4+CD25+ Tregs was determined by [3H]thymidine incorporation. FCM plots are representative of one of three independent experiments. Cells were gated on CD4+ T cells. The bar graph of average percentages ± SD of CD4+CD25+Foxp3+ Tregs are from 5 mice or triplicate cultures representative of one of three independent experiments. **P < 0.01, ***P < 0.001.
Mentions: Studies have demonstrated that HSP60 is a natural ligand for TLR2 and TLR4 [35]. To further investigate whether TLR2 and/or TLR4 are involved in SjHSP60-mediated Treg increase, we injected TLR2-/-, TLR4-/- or WT mice with SjHSP60 or PBS and then examined Treg frequency. FCM analysis showed that Tregs were significantly increased in TLR2-/- and WT mice, but not in TLR4-/- mice (Fig 5A and 5B).

Bottom Line: In addition, we identified that SjHSP60 derived from SEA significantly induces Tregs in vivo and in vitro.SjHSP60 specially induced Tregs in vivo and in vitro without inducing other CD4+ T sub-populations including Th1, Th2 and Th17 cells.Furthermore, we showed that the SjHSP60-depleted SEA almost lost the ability in vitro and displayed a significant impaired ability to induce Tregs in vivo.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathogen Biology and Immunology, Jiangsu Key Laboratory of Pathogen Biology, Nanjing Medical University, Nanjing, Jiangsu, China.

ABSTRACT

Background: Parasitic helminths need to suppress the host immune system to establish chronic infections. Paradoxically, immunosuppression induced by the worm also benefits the host by limiting excessive inflammation and tissue damage, which remains the major cause leading to serious morbidity and mortality. Regulatory T cells (Tregs) are key immune regulators of this mutualism. The successive rise in Tregs during schistosome infection plays a critical role in immunoregulation. We and others previously showed that Schistosoma japonicum (S. japonicum) egg antigens (SEA) induce Tregs both in vitro and in vivo. In addition, we identified that SjHSP60 derived from SEA significantly induces Tregs in vivo and in vitro. However, the contribution of SjHSP60 in SEA to Treg induction and the related mechanisms of the Treg induction have not yet been identified.

Methodology/principal findings: In this study, we showed that S. japonicum stress protein HSP60 (SjHSP60) was constitutively and extensively expressed in eggs of S. japonicum. SjHSP60 specially induced Tregs in vivo and in vitro without inducing other CD4+ T sub-populations including Th1, Th2 and Th17 cells. Furthermore, we showed that the SjHSP60-depleted SEA almost lost the ability in vitro and displayed a significant impaired ability to induce Tregs in vivo. Finally, our study illustrated that the mechanisms of SjHSP60-mediated induction of Tregs are through both conversion of CD4+CD25- T cells into CD4+CD25+Foxp3+ Tregs and expansion of preexisting CD4+CD25+Foxp3+ Tregs in a TLR4-dependent manner.

Conclusions/significance: Collectively, our findings identify SjHSP60 as a major parasitic contributor of Treg induction in S. japonicum egg antigens, which not only contributes to the better understanding of the mechanism of immunoregulation during helminth infection, but also suggests its potential as a therapeutic target for control of immunopathology, allergic and autoimmune diseases.

No MeSH data available.


Related in: MedlinePlus