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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

FoxP3 expression at mRNA levels in MG patients. FoxP3 mRNA expression of CD4+CD25ˉ T cells (before stimulating), the induced CD4+CD25+ T cells from MG patients and nTregs from HCs were assessed by real-time PCR (all the cells are purified). Results are expressed as mean ± SD, n = 15. *, P < 0.05 vs. nTregs. &, P < 0.05 vs. CD4+CD25ˉ T cells (before stimulating) from MG patients. #, P < 0.05 vs. 40 ng/mL induced CD4+CD25+ T cells
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Fig4: FoxP3 expression at mRNA levels in MG patients. FoxP3 mRNA expression of CD4+CD25ˉ T cells (before stimulating), the induced CD4+CD25+ T cells from MG patients and nTregs from HCs were assessed by real-time PCR (all the cells are purified). Results are expressed as mean ± SD, n = 15. *, P < 0.05 vs. nTregs. &, P < 0.05 vs. CD4+CD25ˉ T cells (before stimulating) from MG patients. #, P < 0.05 vs. 40 ng/mL induced CD4+CD25+ T cells

Mentions: As described before, the CD4+ T cells which express the higher level of CD25 as the induced CD4+CD25+ T cells were selected [19]. But it should be noted that CD25 is not a specific marker of CD4+CD25+ Tregs. For example, Hong J and Leonard C found in that effector CD4+ T cells, CD25 can also be detected [20]. FoxP3 is the most specific marker in comparison with the other markers. Thus, besides CD25, FoxP3 is also included in our experiment detection to further study whether the induced CD4+CD25+ T cells are similar to nTregs phenotypically. Figure 4 shows the relationship between the FoxP3 expression of the induced CD4+CD25+ T cells at mRNA level and the concentration of IFN-γ in MG patients. It can be found that IFN-γ is able to induce FoxP3 expression. Even though the concentration of IFN-γ is lower (20 ng/ml), obvious induction of FoxP3 is observed. The FoxP3 expression reaches the highest when IFN-γ is 40 ng/ml (22 ± 3.1). But it is still lower than that in nTregs of HCs (25 ± 4.2) (P < 0.05).Fig. 4


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

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

FoxP3 expression at mRNA levels in MG patients. FoxP3 mRNA expression of CD4+CD25ˉ T cells (before stimulating), the induced CD4+CD25+ T cells from MG patients and nTregs from HCs were assessed by real-time PCR (all the cells are purified). Results are expressed as mean ± SD, n = 15. *, P < 0.05 vs. nTregs. &, P < 0.05 vs. CD4+CD25ˉ T cells (before stimulating) from MG patients. #, P < 0.05 vs. 40 ng/mL induced CD4+CD25+ T cells
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4562356&req=5

Fig4: FoxP3 expression at mRNA levels in MG patients. FoxP3 mRNA expression of CD4+CD25ˉ T cells (before stimulating), the induced CD4+CD25+ T cells from MG patients and nTregs from HCs were assessed by real-time PCR (all the cells are purified). Results are expressed as mean ± SD, n = 15. *, P < 0.05 vs. nTregs. &, P < 0.05 vs. CD4+CD25ˉ T cells (before stimulating) from MG patients. #, P < 0.05 vs. 40 ng/mL induced CD4+CD25+ T cells
Mentions: As described before, the CD4+ T cells which express the higher level of CD25 as the induced CD4+CD25+ T cells were selected [19]. But it should be noted that CD25 is not a specific marker of CD4+CD25+ Tregs. For example, Hong J and Leonard C found in that effector CD4+ T cells, CD25 can also be detected [20]. FoxP3 is the most specific marker in comparison with the other markers. Thus, besides CD25, FoxP3 is also included in our experiment detection to further study whether the induced CD4+CD25+ T cells are similar to nTregs phenotypically. Figure 4 shows the relationship between the FoxP3 expression of the induced CD4+CD25+ T cells at mRNA level and the concentration of IFN-γ in MG patients. It can be found that IFN-γ is able to induce FoxP3 expression. Even though the concentration of IFN-γ is lower (20 ng/ml), obvious induction of FoxP3 is observed. The FoxP3 expression reaches the highest when IFN-γ is 40 ng/ml (22 ± 3.1). But it is still lower than that in nTregs of HCs (25 ± 4.2) (P < 0.05).Fig. 4

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