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Feasibility of up-regulating CD4(+)CD25(+) Tregs by IFN-γ in myasthenia gravis patients.

Huang S, Wang W, Chi L - BMC Neurol (2015)

Bottom Line: It shows the percentages of CD4(+)CD25(+) T cells among CD4(+) T cells have no significant difference in MG patients compared with those in HCs.This subject will further reveal the role of IFN-γ in the pathogenesis of MG from a new perspective.It will also provide the scientific basis for the clinical targeted therapy of MG.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology, The First Affiliated Hospital of Harbin Medical University, Harbin, 150080, P.R. China. cecile_huang@163.com.

ABSTRACT

Background: In myasthenia gravis (MG) patients, the dysfunction of CD4(+)CD25(+) regulatory T cells (CD4(+)CD25(+) Tregs) may be one of the important pathogenesis of MG. Currently, the role of IFN-γ in autoimmune diseases is still controversial and needs further exploration. In this study, whether IFN-γ can induce CD4(+)CD25(-) T cells into CD4(+)CD25(+) Tregs in MG in vitro was investigated systematically.

Methods: Flow cytometry was used to analyze the number of CD4(+)CD25(+) Tregs in MG patients and healthy controls (HCs). CD4(+)CD25(-) T cells were separated from the peripheral blood mononuclear cells of MG patients and HCs, and the CD4(+)CD25(+) Tregs were separated from HCs by Magnetic cell sorting (MACS). IFN-γ with different concentrations was used to stimulate CD4(+)CD25(-) T cells. The percentages of the induced CD4(+)CD25(+) T cells were detected by flow cytometry. The FoxP3 expression of the induced CD4(+)CD25(+) T cells in MG patients was detected by real-time PCR at mRNA level. The induced CD4(+)CD25(+) T cells were co-cultured with autologous CD4(+)CD25(-) T cells to estimate the suppressive ability of the induced CD4(+)CD25(+) T cells to CD4(+)CD25(-) T cells.

Results: It shows the percentages of CD4(+)CD25(+) T cells among CD4(+) T cells have no significant difference in MG patients compared with those in HCs. There is also merely no difference in the percentages of CD4(+)CD25(+) T cells between thymectomized and non-thymectomized MG patients. CD4(+)CD25(-) T cells can be induced to CD4(+)CD25(+) T cells after applying IFN-γ in MG patients and HCs. The proportion and FoxP3 expression of the induced CD4(+)CD25(+) T cells are the highest at the level of 40 ng/ml IFN-γ, and the suppressive function of the CD4(+)CD25(+) T cells induced by 40 ng/ml IFN-γ is the strongest in MG patients.

Conclusions: This subject will further reveal the role of IFN-γ in the pathogenesis of MG from a new perspective. It will also provide the scientific basis for the clinical targeted therapy of MG.

No MeSH data available.


Related in: MedlinePlus

Suppressive function of the induced CD4+CD25+ T cells on the proliferation of autologous CD4+CD25ˉ T cells in MG patients. a The induced CD4+CD25+ T cells were co-cultured with autologous CD4+CD25ˉT cells at a 1:1 ratio to assay their suppressive function on the proliferation of CD4+CD25ˉ T cells. Results are expressed as mean cpm ± SD, n = 16. *, P < 0.05 vs. CD4+CD25ˉ T cells cultured alone. &, P < 0.05 vs. co-cultures of CD4+CD25ˉT cells and nTregs. #, P < 0.05 vs. 40 ng/mL induced CD4+CD25+ T cells. b IFN-γ dose-dependent induction of FoxP3 mRNA expression and the suppressive percentage of the induced CD4+CD25+T cells in suppressing proliferation of co-cultured autologous CD4+CD25ˉT cells. There is linear regression between them (r2 = 0.917, p = 0.029)
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Fig5: Suppressive function of the induced CD4+CD25+ T cells on the proliferation of autologous CD4+CD25ˉ T cells in MG patients. a The induced CD4+CD25+ T cells were co-cultured with autologous CD4+CD25ˉT cells at a 1:1 ratio to assay their suppressive function on the proliferation of CD4+CD25ˉ T cells. Results are expressed as mean cpm ± SD, n = 16. *, P < 0.05 vs. CD4+CD25ˉ T cells cultured alone. &, P < 0.05 vs. co-cultures of CD4+CD25ˉT cells and nTregs. #, P < 0.05 vs. 40 ng/mL induced CD4+CD25+ T cells. b IFN-γ dose-dependent induction of FoxP3 mRNA expression and the suppressive percentage of the induced CD4+CD25+T cells in suppressing proliferation of co-cultured autologous CD4+CD25ˉT cells. There is linear regression between them (r2 = 0.917, p = 0.029)

Mentions: CD4+CD25+ Tregs have the ability to suppress autologous CD4+CD25− T cells. So the suppressive function of the induced CD4+CD25+ T cells was evaluated to determine whether they are similar to nTregs functionally. In the presence of anti-CD3 antibody and irradiated APCs, the induced CD4+CD25+ T cells were co-cultured with autologous CD4+CD25− T cells at a ratio of 1:1. In order to make a reasonable comparison, CD4+CD25− T cells were cultured independently or co-cultured with nTregs (as the references). The suppressive function of the induced CD4+CD25+ T cells is estimated through the proliferative ability of autologous CD4+CD25− T cells which is expressed as cpm (Fig. 5a). The value of cpm decreases when the induced CD4+CD25+ T cells are added to CD4+CD25− T cells. The value of cpm is the highest when CD4+CD25− T cells are cultured alone. The value is the lowest when the induced CD4+CD25+ T cells induced by 40 ng/ml IFN-γ are added and it is different from that of co-cultured CD4+CD25− T cells and nTregs (33 ± 4.2) (P < 0.05). The value of cpm is equal to the proliferation of CD4+CD25− T cells while this is opposite to the suppressive function of CD4+CD25+ Tregs. So this explains the CD4+CD25+ T cells induced by 40 ng/ml IFN-γ have the most powerful suppressive function but it is still lower than the function of nTregs.Fig. 5


Feasibility of up-regulating CD4(+)CD25(+) Tregs by IFN-γ in myasthenia gravis patients.

Huang S, Wang W, Chi L - BMC Neurol (2015)

Suppressive function of the induced CD4+CD25+ T cells on the proliferation of autologous CD4+CD25ˉ T cells in MG patients. a The induced CD4+CD25+ T cells were co-cultured with autologous CD4+CD25ˉT cells at a 1:1 ratio to assay their suppressive function on the proliferation of CD4+CD25ˉ T cells. Results are expressed as mean cpm ± SD, n = 16. *, P < 0.05 vs. CD4+CD25ˉ T cells cultured alone. &, P < 0.05 vs. co-cultures of CD4+CD25ˉT cells and nTregs. #, P < 0.05 vs. 40 ng/mL induced CD4+CD25+ T cells. b IFN-γ dose-dependent induction of FoxP3 mRNA expression and the suppressive percentage of the induced CD4+CD25+T cells in suppressing proliferation of co-cultured autologous CD4+CD25ˉT cells. There is linear regression between them (r2 = 0.917, p = 0.029)
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Fig5: Suppressive function of the induced CD4+CD25+ T cells on the proliferation of autologous CD4+CD25ˉ T cells in MG patients. a The induced CD4+CD25+ T cells were co-cultured with autologous CD4+CD25ˉT cells at a 1:1 ratio to assay their suppressive function on the proliferation of CD4+CD25ˉ T cells. Results are expressed as mean cpm ± SD, n = 16. *, P < 0.05 vs. CD4+CD25ˉ T cells cultured alone. &, P < 0.05 vs. co-cultures of CD4+CD25ˉT cells and nTregs. #, P < 0.05 vs. 40 ng/mL induced CD4+CD25+ T cells. b IFN-γ dose-dependent induction of FoxP3 mRNA expression and the suppressive percentage of the induced CD4+CD25+T cells in suppressing proliferation of co-cultured autologous CD4+CD25ˉT cells. There is linear regression between them (r2 = 0.917, p = 0.029)
Mentions: CD4+CD25+ Tregs have the ability to suppress autologous CD4+CD25− T cells. So the suppressive function of the induced CD4+CD25+ T cells was evaluated to determine whether they are similar to nTregs functionally. In the presence of anti-CD3 antibody and irradiated APCs, the induced CD4+CD25+ T cells were co-cultured with autologous CD4+CD25− T cells at a ratio of 1:1. In order to make a reasonable comparison, CD4+CD25− T cells were cultured independently or co-cultured with nTregs (as the references). The suppressive function of the induced CD4+CD25+ T cells is estimated through the proliferative ability of autologous CD4+CD25− T cells which is expressed as cpm (Fig. 5a). The value of cpm decreases when the induced CD4+CD25+ T cells are added to CD4+CD25− T cells. The value of cpm is the highest when CD4+CD25− T cells are cultured alone. The value is the lowest when the induced CD4+CD25+ T cells induced by 40 ng/ml IFN-γ are added and it is different from that of co-cultured CD4+CD25− T cells and nTregs (33 ± 4.2) (P < 0.05). The value of cpm is equal to the proliferation of CD4+CD25− T cells while this is opposite to the suppressive function of CD4+CD25+ Tregs. So this explains the CD4+CD25+ T cells induced by 40 ng/ml IFN-γ have the most powerful suppressive function but it is still lower than the function of nTregs.Fig. 5

Bottom Line: It shows the percentages of CD4(+)CD25(+) T cells among CD4(+) T cells have no significant difference in MG patients compared with those in HCs.This subject will further reveal the role of IFN-γ in the pathogenesis of MG from a new perspective.It will also provide the scientific basis for the clinical targeted therapy of MG.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology, The First Affiliated Hospital of Harbin Medical University, Harbin, 150080, P.R. China. cecile_huang@163.com.

ABSTRACT

Background: In myasthenia gravis (MG) patients, the dysfunction of CD4(+)CD25(+) regulatory T cells (CD4(+)CD25(+) Tregs) may be one of the important pathogenesis of MG. Currently, the role of IFN-γ in autoimmune diseases is still controversial and needs further exploration. In this study, whether IFN-γ can induce CD4(+)CD25(-) T cells into CD4(+)CD25(+) Tregs in MG in vitro was investigated systematically.

Methods: Flow cytometry was used to analyze the number of CD4(+)CD25(+) Tregs in MG patients and healthy controls (HCs). CD4(+)CD25(-) T cells were separated from the peripheral blood mononuclear cells of MG patients and HCs, and the CD4(+)CD25(+) Tregs were separated from HCs by Magnetic cell sorting (MACS). IFN-γ with different concentrations was used to stimulate CD4(+)CD25(-) T cells. The percentages of the induced CD4(+)CD25(+) T cells were detected by flow cytometry. The FoxP3 expression of the induced CD4(+)CD25(+) T cells in MG patients was detected by real-time PCR at mRNA level. The induced CD4(+)CD25(+) T cells were co-cultured with autologous CD4(+)CD25(-) T cells to estimate the suppressive ability of the induced CD4(+)CD25(+) T cells to CD4(+)CD25(-) T cells.

Results: It shows the percentages of CD4(+)CD25(+) T cells among CD4(+) T cells have no significant difference in MG patients compared with those in HCs. There is also merely no difference in the percentages of CD4(+)CD25(+) T cells between thymectomized and non-thymectomized MG patients. CD4(+)CD25(-) T cells can be induced to CD4(+)CD25(+) T cells after applying IFN-γ in MG patients and HCs. The proportion and FoxP3 expression of the induced CD4(+)CD25(+) T cells are the highest at the level of 40 ng/ml IFN-γ, and the suppressive function of the CD4(+)CD25(+) T cells induced by 40 ng/ml IFN-γ is the strongest in MG patients.

Conclusions: This subject will further reveal the role of IFN-γ in the pathogenesis of MG from a new perspective. It will also provide the scientific basis for the clinical targeted therapy of MG.

No MeSH data available.


Related in: MedlinePlus