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Ubiquitous Over-Expression of Chromatin Remodeling Factor SRG3 Ameliorates the T Cell-Mediated Exacerbation of EAE by Modulating the Phenotypes of both Dendritic Cells and Macrophages.

Lee SW, Park HJ, Jeon SH, Lee C, Seong RH, Park SH, Hong S - PLoS ONE (2015)

Bottom Line: We found that the outcome of EAE development was completely different depending on the expression pattern of SRG3.SRG3 over-expression not only reduced pro-inflammatory cytokine production by DCs but also shifted macrophages from the inducible nitric oxide synthase (iNOS)-expressing M1 phenotype to the arginase-1-expressing M2 phenotype during EAE.Taken together, our results provide the first evidence supporting the development of a new therapeutic strategy for EAE involving the modulation of SRG3 expression to induce M2 and Th2 polarization, thereby inhibiting inflammatory immune responses.

View Article: PubMed Central - PubMed

Affiliation: Dept. of Bioscience and Biotechnology, Institute of Bioscience, Sejong University, Seoul 143-747, Korea; School of Life Sciences and Biotechnology, Korea University, Seoul 136-701, Korea.

ABSTRACT
Although SWI3-related gene (SRG3), a chromatin remodeling factor, is critical for various biological processes including early embryogenesis and thymocyte development, it is unclear whether SRG3 is involved in the differentiation of CD4+ T cells, the key mediator of adaptive immune responses. Because it is known that experimental autoimmune encephalomyelitis (EAE) development is determined by the activation of CD4+ T helper cells, here, we investigated the role of SRG3 in EAE development using SRG3 transgenic mouse models exhibiting two distinct SRG3 expression patterns: SRG3 expression driven by either the CD2 or β-actin promoter. We found that the outcome of EAE development was completely different depending on the expression pattern of SRG3. The specific over-expression of SRG3 using the CD2 promoter facilitated EAE via the induction of Th1 and Th17 cells, whereas the ubiquitous over-expression of SRG3 using the β-actin promoter inhibited EAE by promoting Th2 differentiation and suppressing Th1 and Th17 differentiation. In addition, the ubiquitous over-expression of SRG3 polarized CD4+ T cell differentiation towards the Th2 phenotype by converting dendritic cells (DCs) or macrophages to Th2 types. SRG3 over-expression not only reduced pro-inflammatory cytokine production by DCs but also shifted macrophages from the inducible nitric oxide synthase (iNOS)-expressing M1 phenotype to the arginase-1-expressing M2 phenotype during EAE. In addition, Th2 differentiation in β-actin-SRG3 Tg mice during EAE was associated with an increase in the basophil and mast cell populations and in IL4 production. Furthermore, the increased frequency of Treg cells in the spinal cord of β-actin-SRG3 Tg mice might induce the suppression of and accelerate the recovery from EAE symptoms. Taken together, our results provide the first evidence supporting the development of a new therapeutic strategy for EAE involving the modulation of SRG3 expression to induce M2 and Th2 polarization, thereby inhibiting inflammatory immune responses.

No MeSH data available.


Related in: MedlinePlus

During EAE development, SRG3 over-expression driven by the CD2 promoter enhances Th1 and Th17 differentiation, whereas SRG3 over-expression driven by the β-actin promoter increases Th2 differentiation but decreases Th1 and Th17 differentiation.(A-C) Both MBP TCR Tg B10.PL mice and CD2-SRG3/MBP TCR double Tg B10.PL mice (upper panel in Fig 5A, 5B, and 5C) or both MBP TCR Tg B10.PL mice and β-actin-SRG3/MBP TCR double Tg B10.PL mice (lower panel in Fig 5A, 5B, and 5C) were either non-immunized or s.c. immunized with the MBP-Ac1-11 peptide in CFA. (A) Purified CD4+ splenocytes from the four groups were activated using plate-bound anti-CD3 (10 μg/ml) and anti-CD28 (1 μg/ml) mAbs for 16 hrs and subsequently stimulated with PMA/ionomycin for 2 hrs in the presence of brefeldin A (10 μg/ml). The intracellular expression of IFNγ, IL17, IL4, and IL10 was analyzed via flow cytometry. The mean values ± SD are shown (n = 5; *P<0.05, **P<0.01). (B) The expression of T-bet, RORγt, and GATA-3 in splenic CD4+ T cells purified from the four groups was analyzed via flow cytometry. The mean values ± SD are shown (n = 5; *P<0.05). (C) Mononuclear cells (MNCs) were isolated from the spinal cord of the four groups using a Percoll gradient on day 21 or 24 after EAE induction. The isolated MNCs were incubated for 2 hrs in brefeldin A (10 μg/ml), and subsequently, the intracellular expression of the cytokines IFNγ and IL17 was analyzed via flow cytometry. The mean values ± SD are shown (n = 5; *P<0.05, **P<0.01).
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pone.0132329.g005: During EAE development, SRG3 over-expression driven by the CD2 promoter enhances Th1 and Th17 differentiation, whereas SRG3 over-expression driven by the β-actin promoter increases Th2 differentiation but decreases Th1 and Th17 differentiation.(A-C) Both MBP TCR Tg B10.PL mice and CD2-SRG3/MBP TCR double Tg B10.PL mice (upper panel in Fig 5A, 5B, and 5C) or both MBP TCR Tg B10.PL mice and β-actin-SRG3/MBP TCR double Tg B10.PL mice (lower panel in Fig 5A, 5B, and 5C) were either non-immunized or s.c. immunized with the MBP-Ac1-11 peptide in CFA. (A) Purified CD4+ splenocytes from the four groups were activated using plate-bound anti-CD3 (10 μg/ml) and anti-CD28 (1 μg/ml) mAbs for 16 hrs and subsequently stimulated with PMA/ionomycin for 2 hrs in the presence of brefeldin A (10 μg/ml). The intracellular expression of IFNγ, IL17, IL4, and IL10 was analyzed via flow cytometry. The mean values ± SD are shown (n = 5; *P<0.05, **P<0.01). (B) The expression of T-bet, RORγt, and GATA-3 in splenic CD4+ T cells purified from the four groups was analyzed via flow cytometry. The mean values ± SD are shown (n = 5; *P<0.05). (C) Mononuclear cells (MNCs) were isolated from the spinal cord of the four groups using a Percoll gradient on day 21 or 24 after EAE induction. The isolated MNCs were incubated for 2 hrs in brefeldin A (10 μg/ml), and subsequently, the intracellular expression of the cytokines IFNγ and IL17 was analyzed via flow cytometry. The mean values ± SD are shown (n = 5; *P<0.05, **P<0.01).

Mentions: Because CD2-SRG3 over-expression facilitates the development of EAE, we examined whether CD2-SRG3 over-expression polarizes CD4+ T cells toward the Th1 and Th17 phenotypes since it has been accepted that EAE pathogenesis is primarily mediated by both Th1 and Th17 immune responses [29]. Thus, we measured the cytokine expression pattern of CD4+ T cells in both the spleen and the spinal cord under either naive (no MBP immunization) or EAE conditions (MBP immunization) in CD2-SRG3/MBP TCR double Tg mice. We found that the expression of both IFNγ and IL17 was significantly increased but that the expression of both IL4 and IL10 was comparable in splenic CD4+ T cells from CD2-SRG3-over-expressing mice compared to those from MBP TCR single Tg B10.PL mice (Fig 5A, upper panel; Fig A in S5 Fig). Next, we hypothesized that β-actin promoter-driven SRG3 over-expression influences CD4+ T cell differentiation during EAE development due to differential effects of the SRG3 expression pattern on EAE pathogenesis. To test this hypothesis, we examined the cytokine expression pattern of T helper cells after immunization. We found that the frequency of Th1 and Th17 cells was dramatically decreased in β-actin-SRG3/MBP TCR double Tg mice compared to the control mice and that IL4-producing Th2 cells were increased in both the naive (non-immunization) and EAE (immunization) groups in these double Tg mice compared to the control mice (Fig 5A, lower panel; Fig B in S5 Fig). Based on these cytokine profiles, we evaluated the Th1/Th2 ratio among splenic CD4+ T cells and found that it significantly increased in CD2-SRG3/MBP TCR double Tg mice but dramatically decreased in β-actin-SRG3/MBP TCR double Tg mice compared to the control mice (Fig A in S6 Fig). In addition, we compared the intracellular expression of specific transcription factors that orchestrate the differentiation of T helper subsets in CD4+ T cells between the CD2-SRG3 or β-actin-SRG3 Tg mice and the WT mice. We examined T-bet for Th1 cells, GATA-3 for Th2 cells, and RORγt for Th17 cells [30]. The CD2-SRG3 Tg mice showed increased numbers of CD4+ T cells expressing T-bet and RORγt but not GATA-3 (Fig 5B, upper panel). As expected, the ubiquitous expression of SRG3 down-regulated T-bet and RORγt expression but up-regulated GATA-3 expression; these results were consistent with the cytokine profiles of T helper cells (Fig 5B, lower panel). Finally, we observed a dramatic increase in the frequencies of Th1 and Th17 cells that infiltrated into the spinal cord in the CD2-SRG3/MBP TCR double Tg mice subjected to EAE compared to the control mice (Fig 5C, upper panel). Next, we examined whether β-actin promoter-mediated SRG3 over-expression restricts the infiltration of pathogenic effector T cells in the spinal cord during EAE development. We found that the infiltration of Th1 and Th17 cells into the spinal cord was significantly reduced in the β-actin-SRG3/MBP TCR double Tg mice subjected to EAE compared to the control mice (Fig 5C, lower panel). Taken together, our results demonstrated that CD2-SRG3-overexpressing mice are much more vulnerable to the pathogenesis of autoimmune diseases such as EAE but that the over-expression of SRG3 in additional cell types inhibits EAE development.


Ubiquitous Over-Expression of Chromatin Remodeling Factor SRG3 Ameliorates the T Cell-Mediated Exacerbation of EAE by Modulating the Phenotypes of both Dendritic Cells and Macrophages.

Lee SW, Park HJ, Jeon SH, Lee C, Seong RH, Park SH, Hong S - PLoS ONE (2015)

During EAE development, SRG3 over-expression driven by the CD2 promoter enhances Th1 and Th17 differentiation, whereas SRG3 over-expression driven by the β-actin promoter increases Th2 differentiation but decreases Th1 and Th17 differentiation.(A-C) Both MBP TCR Tg B10.PL mice and CD2-SRG3/MBP TCR double Tg B10.PL mice (upper panel in Fig 5A, 5B, and 5C) or both MBP TCR Tg B10.PL mice and β-actin-SRG3/MBP TCR double Tg B10.PL mice (lower panel in Fig 5A, 5B, and 5C) were either non-immunized or s.c. immunized with the MBP-Ac1-11 peptide in CFA. (A) Purified CD4+ splenocytes from the four groups were activated using plate-bound anti-CD3 (10 μg/ml) and anti-CD28 (1 μg/ml) mAbs for 16 hrs and subsequently stimulated with PMA/ionomycin for 2 hrs in the presence of brefeldin A (10 μg/ml). The intracellular expression of IFNγ, IL17, IL4, and IL10 was analyzed via flow cytometry. The mean values ± SD are shown (n = 5; *P<0.05, **P<0.01). (B) The expression of T-bet, RORγt, and GATA-3 in splenic CD4+ T cells purified from the four groups was analyzed via flow cytometry. The mean values ± SD are shown (n = 5; *P<0.05). (C) Mononuclear cells (MNCs) were isolated from the spinal cord of the four groups using a Percoll gradient on day 21 or 24 after EAE induction. The isolated MNCs were incubated for 2 hrs in brefeldin A (10 μg/ml), and subsequently, the intracellular expression of the cytokines IFNγ and IL17 was analyzed via flow cytometry. The mean values ± SD are shown (n = 5; *P<0.05, **P<0.01).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4492541&req=5

pone.0132329.g005: During EAE development, SRG3 over-expression driven by the CD2 promoter enhances Th1 and Th17 differentiation, whereas SRG3 over-expression driven by the β-actin promoter increases Th2 differentiation but decreases Th1 and Th17 differentiation.(A-C) Both MBP TCR Tg B10.PL mice and CD2-SRG3/MBP TCR double Tg B10.PL mice (upper panel in Fig 5A, 5B, and 5C) or both MBP TCR Tg B10.PL mice and β-actin-SRG3/MBP TCR double Tg B10.PL mice (lower panel in Fig 5A, 5B, and 5C) were either non-immunized or s.c. immunized with the MBP-Ac1-11 peptide in CFA. (A) Purified CD4+ splenocytes from the four groups were activated using plate-bound anti-CD3 (10 μg/ml) and anti-CD28 (1 μg/ml) mAbs for 16 hrs and subsequently stimulated with PMA/ionomycin for 2 hrs in the presence of brefeldin A (10 μg/ml). The intracellular expression of IFNγ, IL17, IL4, and IL10 was analyzed via flow cytometry. The mean values ± SD are shown (n = 5; *P<0.05, **P<0.01). (B) The expression of T-bet, RORγt, and GATA-3 in splenic CD4+ T cells purified from the four groups was analyzed via flow cytometry. The mean values ± SD are shown (n = 5; *P<0.05). (C) Mononuclear cells (MNCs) were isolated from the spinal cord of the four groups using a Percoll gradient on day 21 or 24 after EAE induction. The isolated MNCs were incubated for 2 hrs in brefeldin A (10 μg/ml), and subsequently, the intracellular expression of the cytokines IFNγ and IL17 was analyzed via flow cytometry. The mean values ± SD are shown (n = 5; *P<0.05, **P<0.01).
Mentions: Because CD2-SRG3 over-expression facilitates the development of EAE, we examined whether CD2-SRG3 over-expression polarizes CD4+ T cells toward the Th1 and Th17 phenotypes since it has been accepted that EAE pathogenesis is primarily mediated by both Th1 and Th17 immune responses [29]. Thus, we measured the cytokine expression pattern of CD4+ T cells in both the spleen and the spinal cord under either naive (no MBP immunization) or EAE conditions (MBP immunization) in CD2-SRG3/MBP TCR double Tg mice. We found that the expression of both IFNγ and IL17 was significantly increased but that the expression of both IL4 and IL10 was comparable in splenic CD4+ T cells from CD2-SRG3-over-expressing mice compared to those from MBP TCR single Tg B10.PL mice (Fig 5A, upper panel; Fig A in S5 Fig). Next, we hypothesized that β-actin promoter-driven SRG3 over-expression influences CD4+ T cell differentiation during EAE development due to differential effects of the SRG3 expression pattern on EAE pathogenesis. To test this hypothesis, we examined the cytokine expression pattern of T helper cells after immunization. We found that the frequency of Th1 and Th17 cells was dramatically decreased in β-actin-SRG3/MBP TCR double Tg mice compared to the control mice and that IL4-producing Th2 cells were increased in both the naive (non-immunization) and EAE (immunization) groups in these double Tg mice compared to the control mice (Fig 5A, lower panel; Fig B in S5 Fig). Based on these cytokine profiles, we evaluated the Th1/Th2 ratio among splenic CD4+ T cells and found that it significantly increased in CD2-SRG3/MBP TCR double Tg mice but dramatically decreased in β-actin-SRG3/MBP TCR double Tg mice compared to the control mice (Fig A in S6 Fig). In addition, we compared the intracellular expression of specific transcription factors that orchestrate the differentiation of T helper subsets in CD4+ T cells between the CD2-SRG3 or β-actin-SRG3 Tg mice and the WT mice. We examined T-bet for Th1 cells, GATA-3 for Th2 cells, and RORγt for Th17 cells [30]. The CD2-SRG3 Tg mice showed increased numbers of CD4+ T cells expressing T-bet and RORγt but not GATA-3 (Fig 5B, upper panel). As expected, the ubiquitous expression of SRG3 down-regulated T-bet and RORγt expression but up-regulated GATA-3 expression; these results were consistent with the cytokine profiles of T helper cells (Fig 5B, lower panel). Finally, we observed a dramatic increase in the frequencies of Th1 and Th17 cells that infiltrated into the spinal cord in the CD2-SRG3/MBP TCR double Tg mice subjected to EAE compared to the control mice (Fig 5C, upper panel). Next, we examined whether β-actin promoter-mediated SRG3 over-expression restricts the infiltration of pathogenic effector T cells in the spinal cord during EAE development. We found that the infiltration of Th1 and Th17 cells into the spinal cord was significantly reduced in the β-actin-SRG3/MBP TCR double Tg mice subjected to EAE compared to the control mice (Fig 5C, lower panel). Taken together, our results demonstrated that CD2-SRG3-overexpressing mice are much more vulnerable to the pathogenesis of autoimmune diseases such as EAE but that the over-expression of SRG3 in additional cell types inhibits EAE development.

Bottom Line: We found that the outcome of EAE development was completely different depending on the expression pattern of SRG3.SRG3 over-expression not only reduced pro-inflammatory cytokine production by DCs but also shifted macrophages from the inducible nitric oxide synthase (iNOS)-expressing M1 phenotype to the arginase-1-expressing M2 phenotype during EAE.Taken together, our results provide the first evidence supporting the development of a new therapeutic strategy for EAE involving the modulation of SRG3 expression to induce M2 and Th2 polarization, thereby inhibiting inflammatory immune responses.

View Article: PubMed Central - PubMed

Affiliation: Dept. of Bioscience and Biotechnology, Institute of Bioscience, Sejong University, Seoul 143-747, Korea; School of Life Sciences and Biotechnology, Korea University, Seoul 136-701, Korea.

ABSTRACT
Although SWI3-related gene (SRG3), a chromatin remodeling factor, is critical for various biological processes including early embryogenesis and thymocyte development, it is unclear whether SRG3 is involved in the differentiation of CD4+ T cells, the key mediator of adaptive immune responses. Because it is known that experimental autoimmune encephalomyelitis (EAE) development is determined by the activation of CD4+ T helper cells, here, we investigated the role of SRG3 in EAE development using SRG3 transgenic mouse models exhibiting two distinct SRG3 expression patterns: SRG3 expression driven by either the CD2 or β-actin promoter. We found that the outcome of EAE development was completely different depending on the expression pattern of SRG3. The specific over-expression of SRG3 using the CD2 promoter facilitated EAE via the induction of Th1 and Th17 cells, whereas the ubiquitous over-expression of SRG3 using the β-actin promoter inhibited EAE by promoting Th2 differentiation and suppressing Th1 and Th17 differentiation. In addition, the ubiquitous over-expression of SRG3 polarized CD4+ T cell differentiation towards the Th2 phenotype by converting dendritic cells (DCs) or macrophages to Th2 types. SRG3 over-expression not only reduced pro-inflammatory cytokine production by DCs but also shifted macrophages from the inducible nitric oxide synthase (iNOS)-expressing M1 phenotype to the arginase-1-expressing M2 phenotype during EAE. In addition, Th2 differentiation in β-actin-SRG3 Tg mice during EAE was associated with an increase in the basophil and mast cell populations and in IL4 production. Furthermore, the increased frequency of Treg cells in the spinal cord of β-actin-SRG3 Tg mice might induce the suppression of and accelerate the recovery from EAE symptoms. Taken together, our results provide the first evidence supporting the development of a new therapeutic strategy for EAE involving the modulation of SRG3 expression to induce M2 and Th2 polarization, thereby inhibiting inflammatory immune responses.

No MeSH data available.


Related in: MedlinePlus