Original Article
  

Analysis of Methylation and Expression Profile of Foxp3 Gene in Patients with Behçet’s Syndrome

Abstract

Forkhead box P3 (Foxp3) gene is an important means in the Treg cells function, in both maintenances of immune tolerance and regulation of response. Epigenetic modifications of the foxp3 gene at its regulatory regions control the chromatin accessibility for the transcription factors and other transcriptional regulators in order to control Foxp3 expression. In addition, the methylation status of CpG islands within the Foxp3 promoter and regulatory elements regulate the expression of Foxp3. This study was performed to assess the role of the foxp3 gene in patients with Behçet’s syndrome (BS). Venous blood samples were collected from all participants and peripheral blood mononuclear cells (PBMC) were extracted through Ficoll-Hypaque method. Genomic DNA was randomly sheared by sonication and immunoprecipitated with a monoclonal antibody. The status methylation of the foxp3 gene was estimated in 108 blood samples of active BS patients and healthy individuals (controls); using methylation DNA immunoprecipitation (MeDIP) technique. Expression analysis was carried out; using Real-time PCR. The expression of foxp3 gene in the patients' group (mean±SD: 1.79±1.12) was significantly lower than the healthy group (mean±SD: 2.73±1.33) (p<001). Also, the methylation levels of Foxp3 promoter showed that its level in patients (mean±SD: 2.3±1.16) was higher than the healthy group (mean±SD: 1.85±0.59). However, this increase was not statistically significant (p>0.05). Also, these results indicated that increasing the amount of methylation of the foxp3 gene by reducing its expression leads to an increase and intensifying of the disease. The decrease in Foxp3 expression is possibly associated with hypermethylation of the gene, and it can be considered as a risk factor for BS. Future studies may be needed to identify the capability of specific DNA methylation alterations in this syndrome.

1. Hirohata S, Kikuchi H. Behçet's disease. Arthritis Res Ther 2003; 5(3):139.
2. Saadoun D, Wechsler B. Behçet's disease. Orphanet J Rare Dis 2012; 7(1):20.
3. Demirelli S, Degirmenci H, Inci S, Arisoy A. Cardiac manifestations in Behcet's disease. ntractable Rare Dis Res 2015; 4(2):70-5.
4. Wang C, Ye Z, Kijlstra A, Zhou Y, Yang P. Decreased expression of the aryl hydrocarbon receptor
in ocular Behcet’s disease. Mediators Inflamm 2014; 2014:195994.
5. Lourenço EV, La Cava A. Natural regulatory T cells in autoimmunity. Autoimmunity 2011;
4(1):33-42.
6. Hori S, Nomura T, Sakaguchi S. Control of regulatory T cell development by the transcription factor Foxp3. Science 2003; 299(5609):1057-61.
7. Bennett CL, Christie J, Ramsdell F, Brunkow ME, Ferguson PJ, Whitesell L, et al. The immune dysregulation, polyendocrinopathy, enteropathy, X-linked syndrome (IPEX) is caused by mutations of FOXP3. Nat Genet 2001; 27(1):20-1.
8. Wildin RS, Ramsdell F, Peake J, Faravelli F, Casanova J-L, Buist N, et al. X-linked neonatal diabetes mellitus, enteropathy and endocrinopathy syndrome is the human equivalent of mouse scurfy. Nat Genet 2001; 27(1):18.
9. Oda JMM, Hirata BKB, Guembarovski RL, Watanabe MAE. Genetic polymorphism in FOXP3 gene: imbalance in regulatory T-cell role and development of human diseases. J Genet 2013; 92(1):163-71.
10. Morikawa H, Ohkura N, Vandenbon A, Itoh M, Nagao-Sato S, Kawaji H, et al. Differential roles of epigenetic changes and Foxp3 expression in regulatory T cell-specific transcriptional regulation. Proc Natl Acad Sci U S A 2014; 111(14):5289-94.
11. Gavin M, Rudensky A. Control of immune homeostasis by naturally arising regulatory CD4+ T cells. Curr Opin Immunol 2003; 15(6):690-6.
12. Wan YY, Flavell RA. Regulatory T-cell functions are subverted and converted owing to attenuated Foxp3 expression. Nature 2007; 445(7129):766-70.
13. Samstein RM, Arvey A, Josefowicz SZ, Peng X, Reynolds A, Sandstrom R, et al. Foxp3 exploits a pre-existent enhancer landscape for regulatory T cell lineage specification. Cell 2012; 151(1):153-66.
14. Lal G, Bromberg JS. Epigenetic mechanisms of regulation of Foxp3 expression. Blood 2009; 114(18):3727-35.
15. Bettini ML, Pan F, Bettini M, Finkelstein D, Rehg JE, Floess S, et al. Loss of epigenetic modification driven by the Foxp3 transcription factor leads to regulatory T cell insufficiency. Immunity 2012; 36(5):717-30.
16. Nie J, Li YY, Zheng SG, Tsun A, Li B. FOXP3+ Treg cells and gender bias in autoimmune diseases. Front Immunol 2015; 6:493.
17. Alipour, S., et al., Hypermethylation of IL‐10 gene is responsible for its low mRNA expression in Behçet's disease. Journal of cellular biochemistry 2018.
18. Shahram F, Khabbazi A, Nadji A, Ziaie N, Banihashemi AT, Davatchi F. Comparison of existing disease activity indices in the follow-up of patients with Behcet’s disease. Mod Rheumatol 2009;
19(5):536-41.
19. Flores-Borja F, Jury EC, Mauri C, Ehrenstein MR. Defects in CTLA-4 are associated with abnormal regulatory T cell function in rheumatoid arthritis. Proc Natl Acad Sci U S A 2008; 105(49):19396-401.
20. Komatsu N, Mariotti-Ferrandiz ME, Wang Y, Malissen B, Waldmann H, Hori S. Heterogeneity of natural Foxp3+ T cells: a committed regulatory T-cell lineage and an uncommitted minor population retaining plasticity. Proc Natl Acad Sci U S A 2009; 106(6):1903-8.
21. Miyara M, Yoshioka Y, Kitoh A, Shima T, Wing K, Niwa A, et al. Functional delineation and differentiation dynamics of human CD4+ T cells expressing the FoxP3 transcription factor. Immunity 2009; 30(6):899-911.
22. Maruyama T, Kono K, Mizukami Y, Kawaguchi Y, Mimura K, Watanabe M, et al. Distribution of Th17 cells and FoxP3 (+) regulatory T cells in tumor‐infiltrating lymphocytes, tumor‐draining lymph nodes and peripheral blood lymphocytes in patients with gastric cancer. Cancer Sci 2010; 101(9):1947-54.
23. Polansky JK, Kretschmer K, Freyer J, Floess S, Garbe A, Baron U, et al. DNA methylation controls Foxp3 gene expression. Eur J Immunol 2008; 38(6):1654-63.
24. Lal G, Zhang N, Van Der Touw W, Ding Y, Ju W, Bottinger EP, et al. Epigenetic regulation of Foxp3 expression in regulatory T cells by DNA methylation. J Immunol 2009; 182(1):259-73.
25. Kim H-P, Leonard WJ. CREB/ATF-dependent T cell receptor–induced FoxP3 gene expression: a role for DNA methylation. J Exp Med 2007; 204(7):1543-51.
26. Janson PC, Winerdal ME, Marits P, Thörn M, Ohlsson R, Winqvist O. FOXP3 promoter demethylation reveals the committed Treg population in humans. PloS one 2008; 3(2):e1612.
27. Barzaghi F, Passerini L, Gambineri E, Mannurita SC, Cornu T, Kang E, et al. Demethylation analysis of the FOXP3 locus shows quantitative defects of regulatory T cells in IPEX-like syndrome. J Autoimmun. 2012; 38(1):49-58.
28. Zhang P, Zhao M, Liang G, Yin G, Huang D, Su F, et al. Whole-genome DNA methylation in skin lesions from patients with psoriasis vulgaris. J Autoimmun 2013; 41:17-24.
29. Valencia X, Stephens G, Goldbach-Mansky R, Wilson M, Shevach EM, Lipsky PE. TNF downmodulates the function of human CD4+ CD25 hi T-regulatory cells. Blood 2006; 108(1):253-61.
30. Long SA, Cerosaletti K, Bollyky PL, Tatum M, Shilling H, Zhang S, et al. Defects in IL-2R signaling contribute to diminished maintenance of FOXP3 expression in CD4+ CD25+ regulatory T-cells of type 1 diabetic subjects. diabetes 2010; 59(2):407-15.
31. Long SA, Buckner JH. CD4+ FOXP3+ T regulatory cells in human autoimmunity: more than a numbers game. J Immunol 2011; 187(5):2061-6.
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IssueVol 19, No 1 (2020) QRcode
SectionOriginal Article(s)
DOI https://doi.org/10.18502/ijaai.v19i1.2409
PMID32245315
Keywords
Behçet’s syndrome DNA methylation Foxp3 DIP
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How to Cite
1.
Farhadi J, Nouri M, Khabbazi A, Samadi N, Babaloo Z, Azad M, Abolhasani S, Alipour S, Jadideslam G, Seydi Shirvani S, Sakhinia E. Analysis of Methylation and Expression Profile of Foxp3 Gene in Patients with Behçet’s Syndrome. Iran J Allergy Asthma Immunol. 2019;19(1):1-8.