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Fingolimod increases CD39-expressing regulatory T cells in multiple sclerosis patients.

Muls N, Dang HA, Sindic CJ, van Pesch V - PLoS ONE (2014)

Bottom Line: As ATP has numerous proinflammatory effects, its degradation by CD39 has anti-inflammatory influence.B cells, CD4+ cells and Treg proportions were significantly reduced by this treatment, but remaining CD4+ T cells were enriched in FOXP3+ cells and in CD39-expressing Tregs.In addition to the decrease in circulating CD4+ T cells and CD19+ B cells, our findings highlight additional immunoregulatory mechanisms induced by fingolimod.

View Article: PubMed Central - PubMed

Affiliation: Neurochemistry Unit, Institute of Neuroscience, Université catholique de Louvain, Brussels, Belgium.

ABSTRACT

Background: Multiple sclerosis (MS) likely results from an imbalance between regulatory and inflammatory immune processes. CD39 is an ectoenzyme that cleaves ATP to AMP and has been suggested as a novel regulatory T cells (Treg) marker. As ATP has numerous proinflammatory effects, its degradation by CD39 has anti-inflammatory influence. The purpose of this study was to explore regulatory and inflammatory mechanisms activated in fingolimod treated MS patients.

Methods and findings: Peripheral blood mononuclear cells (PBMCs) were isolated from relapsing-remitting MS patients before starting fingolimod and three months after therapy start. mRNA expression was assessed in ex vivo PBMCs. The proportions of CD8, B cells, CD4 and CD39-expressing cells were analysed by flow cytometry. Treg proportion was quantified by flow cytometry and methylation-specific qPCR. Fingolimod treatment increased mRNA levels of CD39, AHR and CYP1B1 but decreased mRNA expression of IL-17, IL-22 and FOXP3 mRNA in PBMCs. B cells, CD4+ cells and Treg proportions were significantly reduced by this treatment, but remaining CD4+ T cells were enriched in FOXP3+ cells and in CD39-expressing Tregs.

Conclusions: In addition to the decrease in circulating CD4+ T cells and CD19+ B cells, our findings highlight additional immunoregulatory mechanisms induced by fingolimod.

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Related in: MedlinePlus

Regulatory T cells in PBMCs of fingolimod-treated patients.Regulatory T cells of MS patients before (t0, n = 16) or after three months of treatment by fingolimod (t3, n = 16) and healthy controls (HC, n = 10) were analysed ex vivo by MethylS-qPCR (A) to quantify nTregs with demethylated FOXP3i1. The proportion of circulating CD4+CD25hiFOXP3+ Tregs was also measured by flow cytometry and expressed as a percentage of lymphoid cells (B). The proportion of CD25hiFOXP3+ cells was also expressed as a percentage of CD4+ T cells (C). nTreg proportion among total PBMCs was decreased by the treatment, as well as the proportion of Tregs within lymphoid cells. CD25hiFOXP3+ Tregs were however enriched within the CD4+ cell population in 12 out of 16 patients. The horizontal lines represent the median value in all subgroups. ** indicates a p-value of ≤0.01.
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pone-0113025-g004: Regulatory T cells in PBMCs of fingolimod-treated patients.Regulatory T cells of MS patients before (t0, n = 16) or after three months of treatment by fingolimod (t3, n = 16) and healthy controls (HC, n = 10) were analysed ex vivo by MethylS-qPCR (A) to quantify nTregs with demethylated FOXP3i1. The proportion of circulating CD4+CD25hiFOXP3+ Tregs was also measured by flow cytometry and expressed as a percentage of lymphoid cells (B). The proportion of CD25hiFOXP3+ cells was also expressed as a percentage of CD4+ T cells (C). nTreg proportion among total PBMCs was decreased by the treatment, as well as the proportion of Tregs within lymphoid cells. CD25hiFOXP3+ Tregs were however enriched within the CD4+ cell population in 12 out of 16 patients. The horizontal lines represent the median value in all subgroups. ** indicates a p-value of ≤0.01.

Mentions: As regulatory T cells play a central role in maintaining self-tolerance, we then analysed the Treg proportion within total PBMCs or CD4+ T cells. Natural Tregs were quantified by analysing the epigenetic status of the first intron of FOXP3 using methylation specific PCR (Figure 4A). CD4+CD25hiFOXP3+ Treg phenotype was also examined by flow cytometry (Figure 4B). Using both MethylS-qPCR (p = 0.041) and flow cytometry (p = 0.0029), we showed that the proportion of Tregs among total PBMCs was decreased by the treatment. However, the proportion of CD25hiFoxp3+ Tregs within the CD4+ cell population increased in 12 patients out of 16 (Figure 4C, Table 1). Only 3 patients displayed a reduction in their Treg proportion while on treatment by fingolimod and one patient did not show change in this parameter.


Fingolimod increases CD39-expressing regulatory T cells in multiple sclerosis patients.

Muls N, Dang HA, Sindic CJ, van Pesch V - PLoS ONE (2014)

Regulatory T cells in PBMCs of fingolimod-treated patients.Regulatory T cells of MS patients before (t0, n = 16) or after three months of treatment by fingolimod (t3, n = 16) and healthy controls (HC, n = 10) were analysed ex vivo by MethylS-qPCR (A) to quantify nTregs with demethylated FOXP3i1. The proportion of circulating CD4+CD25hiFOXP3+ Tregs was also measured by flow cytometry and expressed as a percentage of lymphoid cells (B). The proportion of CD25hiFOXP3+ cells was also expressed as a percentage of CD4+ T cells (C). nTreg proportion among total PBMCs was decreased by the treatment, as well as the proportion of Tregs within lymphoid cells. CD25hiFOXP3+ Tregs were however enriched within the CD4+ cell population in 12 out of 16 patients. The horizontal lines represent the median value in all subgroups. ** indicates a p-value of ≤0.01.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4239031&req=5

pone-0113025-g004: Regulatory T cells in PBMCs of fingolimod-treated patients.Regulatory T cells of MS patients before (t0, n = 16) or after three months of treatment by fingolimod (t3, n = 16) and healthy controls (HC, n = 10) were analysed ex vivo by MethylS-qPCR (A) to quantify nTregs with demethylated FOXP3i1. The proportion of circulating CD4+CD25hiFOXP3+ Tregs was also measured by flow cytometry and expressed as a percentage of lymphoid cells (B). The proportion of CD25hiFOXP3+ cells was also expressed as a percentage of CD4+ T cells (C). nTreg proportion among total PBMCs was decreased by the treatment, as well as the proportion of Tregs within lymphoid cells. CD25hiFOXP3+ Tregs were however enriched within the CD4+ cell population in 12 out of 16 patients. The horizontal lines represent the median value in all subgroups. ** indicates a p-value of ≤0.01.
Mentions: As regulatory T cells play a central role in maintaining self-tolerance, we then analysed the Treg proportion within total PBMCs or CD4+ T cells. Natural Tregs were quantified by analysing the epigenetic status of the first intron of FOXP3 using methylation specific PCR (Figure 4A). CD4+CD25hiFOXP3+ Treg phenotype was also examined by flow cytometry (Figure 4B). Using both MethylS-qPCR (p = 0.041) and flow cytometry (p = 0.0029), we showed that the proportion of Tregs among total PBMCs was decreased by the treatment. However, the proportion of CD25hiFoxp3+ Tregs within the CD4+ cell population increased in 12 patients out of 16 (Figure 4C, Table 1). Only 3 patients displayed a reduction in their Treg proportion while on treatment by fingolimod and one patient did not show change in this parameter.

Bottom Line: As ATP has numerous proinflammatory effects, its degradation by CD39 has anti-inflammatory influence.B cells, CD4+ cells and Treg proportions were significantly reduced by this treatment, but remaining CD4+ T cells were enriched in FOXP3+ cells and in CD39-expressing Tregs.In addition to the decrease in circulating CD4+ T cells and CD19+ B cells, our findings highlight additional immunoregulatory mechanisms induced by fingolimod.

View Article: PubMed Central - PubMed

Affiliation: Neurochemistry Unit, Institute of Neuroscience, Université catholique de Louvain, Brussels, Belgium.

ABSTRACT

Background: Multiple sclerosis (MS) likely results from an imbalance between regulatory and inflammatory immune processes. CD39 is an ectoenzyme that cleaves ATP to AMP and has been suggested as a novel regulatory T cells (Treg) marker. As ATP has numerous proinflammatory effects, its degradation by CD39 has anti-inflammatory influence. The purpose of this study was to explore regulatory and inflammatory mechanisms activated in fingolimod treated MS patients.

Methods and findings: Peripheral blood mononuclear cells (PBMCs) were isolated from relapsing-remitting MS patients before starting fingolimod and three months after therapy start. mRNA expression was assessed in ex vivo PBMCs. The proportions of CD8, B cells, CD4 and CD39-expressing cells were analysed by flow cytometry. Treg proportion was quantified by flow cytometry and methylation-specific qPCR. Fingolimod treatment increased mRNA levels of CD39, AHR and CYP1B1 but decreased mRNA expression of IL-17, IL-22 and FOXP3 mRNA in PBMCs. B cells, CD4+ cells and Treg proportions were significantly reduced by this treatment, but remaining CD4+ T cells were enriched in FOXP3+ cells and in CD39-expressing Tregs.

Conclusions: In addition to the decrease in circulating CD4+ T cells and CD19+ B cells, our findings highlight additional immunoregulatory mechanisms induced by fingolimod.

Show MeSH
Related in: MedlinePlus