Methotrexate Restores Regulatory T Cell Function Through Demethylation of the FoxP3 Upstream Enhancer in Patients With Rheumatoid Arthritis

被引:125
作者
Cribbs, Adam P. [1 ]
Kennedy, Alan [1 ]
Penn, Henry [2 ]
Amjadi, Parisa [1 ]
Green, Patricia [1 ]
Read, Jordan E. [1 ]
Brennan, Fionula
Gregory, Bernard [1 ]
Williams, Richard O. [1 ]
机构
[1] Univ Oxford, Kennedy Inst Rheumatol, Oxford OX3 7FY, England
[2] Northwick Pk Hosp & Clin Res Ctr, Harrow HA1 3UJ, Middx, England
关键词
DNA METHYLTRANSFERASES DNMT3A; LOW-DOSE METHOTREXATE; TNF-ALPHA THERAPY; PERIPHERAL-BLOOD; S-ADENOSYLMETHIONINE; MULTIPLE-SCLEROSIS; GENE-EXPRESSION; SYNOVIAL-FLUID; METHYLATION; MODULATION;
D O I
10.1002/art.39031
中图分类号
R5 [内科学];
学科分类号
1002 ; 100201 ;
摘要
Objective. We have previously shown, in a cohort of untreated rheumatoid arthritis (RA) patients, that the suppressive function of Treg cells is defective. However, other studies in cohorts of patients with established RA have shown that Treg cell function is normal. We hypothesized that treatment may restore Treg cell function and lead to reduced disease activity. The aim of this study was to investigate whether treatment with methotrexate (MTX) can result in epigenetic changes that lead to restoration of the Treg cell suppressive function in RA. Methods. Peripheral blood samples from RA patients were assessed using 3 H-thymidine incorporation to measure Treg cell suppression of T cell proliferation, and by enzyme-linked immunosorbent assay to determine Treg cell suppression of interferon-gamma production. CTLA-4 and FoxP3 expression was measured by flow cytometry and quantitative polymerase chain reaction (qPCR) in Treg cells from healthy individuals and RA patients. CD4+ T cells isolated from healthy individuals were cultured with interleukin-2 (IL-2), IL-6, and tumor necrosis factor a in the presence or absence of MTX, and FoxP3 expression was determined using qPCR and flow cytometry. Methylation of the FOXP3 upstream enhancer was analyzed by bisulfite sequencing PCR. Results. Defective Treg cell function was observed only in RA patients who had not been treated with MTX, whereas Treg cells from MTX-exposed RA patients had restored suppressive function. This restored suppression was associated with increased expression of FoxP3 and CTLA-4 in Treg cells. Bisulfite sequencing PCR of Treg cells cultured in MTX revealed a significant reduction in methylation of the FOXP3 upstream enhancer. Conclusion. This study identifies a novel mechanism of action of MTX, in which treatment of RA patients with MTX restores defective Treg cell function through demethylation of the FOXP3 locus, leading to a subsequent increase in FoxP3 and CTLA-4 expression.
引用
收藏
页码:1182 / 1192
页数:11
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