Skip to main content

Advertisement

SpringerLink
Log in

DNA hypermethylation of PTPN22 gene promoter in children and adolescents with Hashimoto thyroiditis

  • Original Article
  • Published:
Journal of Endocrinological Investigation Aims and scope Submit manuscript

Abstract

Purpose

Protein tyrosine phosphatase non-receptor type 22 (PTPN22) is an inhibitor of T-cell activation, regulating intracellular signal transduction and thereby being implicated in the pathogenesis of autoimmune thyroid disease (AITD). The exact molecular mechanisms have not been fully elucidated. The aim of the present study was to quantitate DNA methylation within the PTPN22 gene promoter in children and adolescents with AITD and healthy controls.

Methods

60 Patients with Hashimoto thyroiditis (HT), 25 patients with HT and type 1 diabetes (HT + T1D), 9 patients with Graves’ disease (GD) and 55 healthy controls without any individual or family history of autoimmune disease were enrolled. Whole blood DNA extraction, DNA modification using sodium bisulfate and quantification of DNA methylation in the PTPN22 gene promoter, based on melting curve analysis of the selected DNA fragment using a Real-Time PCR assay, were implemented.

Results

DNA methylation in the PTPN22 gene promoter was found to be significantly higher in HT patients (39.9 ± 3.1%) in comparison with other study groups (20.3 ± 2.4% for HT + T1D, 32.6 ± 7.8% for GD, 27.1 ± 2.4% for controls, p < 0.001). PTPN22 gene promoter DNA methylation was also associated marginally with thyroid autoimmunity in general (p = 0.059), as well as considerably with thyroid volume (p = 0.004) and the presence of goiter (p = 0.001) but not thyroid function tests.

Conclusions

This study demonstrates for the first time that a relationship between autoimmune thyroiditis and PTPN22 gene promoter DNA methylation state is present, thus proposing another possible etiological association between thyroiditis and abnormalities of PTPN22 function. Further expression studies are required to confirm these findings.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3

Similar content being viewed by others

Data availability

The datasets generated and analysed during the current study are available from the corresponding author on reasonable request.

References

  1. Coppedè F (2017) Epigenetics and autoimmune thyroid diseases. Front Endocrinol (Lausanne) 8:149. https://doi.org/10.3389/fendo.2017.00149

    Article  Google Scholar 

  2. Effraimidis G, Wiersinga WM (2014) Mechanisms in endocrinology: autoimmune thyroid disease: old and new players. Eur J Endocrinol 170(6):R241–R252. https://doi.org/10.1530/EJE-14-0047

    Article  CAS  PubMed  Google Scholar 

  3. Tomer Y (2014) Mechanisms of autoimmune thyroid diseases: from genetics to epigenetics. Annu Rev Pathol 9:147–156. https://doi.org/10.1146/annurev-pathol-012513-104713

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Jones PA (2012) Functions of DNA methylation: islands, start sites, gene bodies and beyond. Nat Rev Genet 13(7):484–492. https://doi.org/10.1038/nrg3230

    Article  CAS  PubMed  Google Scholar 

  5. Fournier A, Sasai N, Nakao M, Defossez PA (2012) The role of methyl-binding proteins in chromatin organization and epigenome maintenance. Brief Funct Genomics 11(3):251–264. https://doi.org/10.1093/bfgp/elr040

    Article  CAS  PubMed  Google Scholar 

  6. Cohen S, Dadi H, Shaoul E, Sharfe N, Roifman CM (1999) Cloning and characterization of a lymphoid-specific, inducible human protein tyrosine phosphatase, Lyp. Blood 93(6):2013–2024

    Article  CAS  Google Scholar 

  7. Mustelin T, Bottini N, Stanford SM (2019) The contribution of PTPN22 to rheumatic disease. Arthritis Rheumatol 71(4):486–495. https://doi.org/10.1002/art.40790

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Burn GL, Svensson L, Sanchez-Blanco C, Saini M, Cope AP (2011) Why is PTPN22 a good candidate susceptibility gene for autoimmune disease? FEBS Lett 585(23):3689–3698. https://doi.org/10.1016/j.febslet.2011.04.032

    Article  CAS  PubMed  Google Scholar 

  9. Simera I, Moher D, Hoey J, Schulz KF, Altman DG (2010) A catalogue of reporting guidelines for health research. Eur J Clin Investig 40(1):35–53. https://doi.org/10.1111/j.1365-2362.2009.02234.x

    Article  CAS  Google Scholar 

  10. Caturegli P, De Remigis A, Rose NR (2014) Hashimoto thyroiditis: clinical and diagnostic criteria. Autoimmun Rev 13(4–5):391–397. https://doi.org/10.1016/j.autrev.2014.01.007

    Article  CAS  PubMed  Google Scholar 

  11. Mayer-Davis EJ, Kahkoska AR, Jefferies C, Dabelea D, Balde N, Gong CX, Aschner P, Craig ME (2018) ISPAD Clinical Practice Consensus Guidelines 2018: definition, epidemiology, and classification of diabetes in children and adolescents. Pediatr Diabetes 19(Suppl 27):7–19. https://doi.org/10.1111/pedi.12773

    Article  PubMed  PubMed Central  Google Scholar 

  12. Alisch RS, Barwick BG, Chopra P, Myrick LK, Satten GA, Conneely KN, Warren ST (2012) Age-associated DNA methylation in pediatric populations. Genome Res 22(4):623–632. https://doi.org/10.1101/gr.125187.111

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. The Children’s Hospital of Philadelphia: Research Institute Pediatric Z-Score Calculator. https://zscore.research.chop.edu/. Accessed 8 March 2020

  14. Ma J, Dempsey AA, Stamatiou D, Marshall KW, Liew CC (2007) Identifying leukocyte gene expression patterns associated with plasma lipid levels in human subjects. Atherosclerosis 191(1):63–72. https://doi.org/10.1016/j.atherosclerosis.2006.05.032

    Article  CAS  PubMed  Google Scholar 

  15. Wettinger SB, Doggen CJ, Spek CA, Rosendaal FR, Reitsma PH (2005) High throughput mRNA profiling highlights associations between myocardial infarction and aberrant expression of inflammatory molecules in blood cells. Blood 105(5):2000–2006. https://doi.org/10.1182/blood-2004-08-3283

    Article  CAS  PubMed  Google Scholar 

  16. Smith E, Jones ME, Drew PA (2009) Quantitation of DNA methylation by melt curve analysis. BMC Cancer 9:123. https://doi.org/10.1186/1471-2407-9-123

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Arroyo-Jousse V, Garcia-Diaz DF, Codner E, Pérez-Bravo F (2016) Epigenetics in type 1 diabetes: TNFa gene promoter methylation status in Chilean patients with type 1 diabetes mellitus. Br J Nutr 116(11):1861–1868. https://doi.org/10.1017/S0007114516003846

    Article  CAS  PubMed  Google Scholar 

  18. Kyrgios I, Fragou A, Kotanidou EP, Mouzaki K, Efraimidou S, Tzimagiorgis G, Galli-Tsinopoulou A (2020) DNA methylation analysis within the IL2RA gene promoter in youth with autoimmune thyroid disease. Eur J Clin Investig 50(3):e13199. https://doi.org/10.1111/eci.13199

    Article  CAS  Google Scholar 

  19. The Li Lab MethPrimer. https://www.urogene.org/methprimer/. Accessed 8 March 2020

  20. Li LC, Dahiya R (2002) MethPrimer: designing primers for methylation PCRs. Bioinformatics 18(11):1427–1431. https://doi.org/10.1093/bioinformatics/18.11.1427

    Article  CAS  PubMed  Google Scholar 

  21. The UCSC Genomics Institute Genome Browser. https://genome.ucsc.edu/index.html. Accessed 8 March 2020

  22. Kurdyukov S, Bullock M (2016) DNA methylation analysis: choosing the right method. Biology (Basel). https://doi.org/10.3390/biology5010003

    Article  Google Scholar 

  23. Lorente A, Mueller W, Urdangarín E, Lázcoz P, von Deimling A, Castresana JS (2008) Detection of methylation in promoter sequences by melting curve analysis-based semiquantitative real time PCR. BMC Cancer 8:61. https://doi.org/10.1186/1471-2407-8-61

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Wojdacz TK, Dobrovic A (2007) Methylation-sensitive high resolution melting (MS-HRM): a new approach for sensitive and high-throughput assessment of methylation. Nucleic Acids Res 35(6):e41. https://doi.org/10.1093/nar/gkm013

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Amornpisutt R, Sriraksa R, Limpaiboon T (2012) Validation of methylation-sensitive high resolution melting for the detection of DNA methylation in cholangiocarcinoma. Clin Biochem 45(13–14):1092–1094. https://doi.org/10.1016/j.clinbiochem.2012.04.027

    Article  CAS  PubMed  Google Scholar 

  26. The NCBI BLAST. https://blast.ncbi.nlm.nih.gov. Accessed 8 March 2020

  27. Dell RB, Holleran S, Ramakrishnan R (2002) Sample size determination. ILAR J 43(4):207–213. https://doi.org/10.1093/ilar.43.4.207

    Article  CAS  PubMed  Google Scholar 

  28. Center for Biomathematics Biomath. https://www.biomath.info. Accessed 8 March 2020

  29. Brčić L, Barić A, Gračan S, Brekalo M, Kaličanin D, Gunjača I, Torlak Lovrić V, Tokić S, Radman M, Škrabić V, Miljković A, Kolčić I, Štefanić M, Glavaš-Obrovac L, Lessel D, Polašek O, Zemunik T, Barbalić M, Punda A, Boraska Perica V (2019) Genome-wide association analysis suggests novel loci for Hashimoto’s thyroiditis. J Endocrinol Investig 42(5):567–576. https://doi.org/10.1007/s40618-018-0955-4

    Article  CAS  Google Scholar 

  30. Bottini N, Musumeci L, Alonso A, Rahmouni S, Nika K, Rostamkhani M, MacMurray J, Meloni GF, Lucarelli P, Pellecchia M, Eisenbarth GS, Comings D, Mustelin T (2004) A functional variant of lymphoid tyrosine phosphatase is associated with type I diabetes. Nat Genet 36(4):337–338. https://doi.org/10.1038/ng1323

    Article  CAS  PubMed  Google Scholar 

  31. Dultz G, Matheis N, Dittmar M, Röhrig B, Bender K, Kahaly GJ (2009) The protein tyrosine phosphatase non-receptor type 22 C1858T polymorphism is a joint susceptibility locus for immunthyroiditis and autoimmune diabetes. Thyroid 19(2):143–148. https://doi.org/10.1089/thy.2008.0301

    Article  CAS  PubMed  Google Scholar 

  32. Giza S, Goulas A, Gbandi E, Effraimidou S, Papadopoulou-Alataki E, Eboriadou M, Galli-Tsinopoulou A (2013) The role of PTPN22 C1858T gene polymorphism in diabetes mellitus type 1: first evaluation in Greek children and adolescents. Biomed Res Int 2013:721604. https://doi.org/10.1155/2013/721604

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Velaga MR, Wilson V, Jennings CE, Owen CJ, Herington S, Donaldson PT, Ball SG, James RA, Quinton R, Perros P, Pearce SH (2004) The codon 620 tryptophan allele of the lymphoid tyrosine phosphatase (LYP) gene is a major determinant of Graves’ disease. J Clin Endocrinol Metab 89(11):5862–5865. https://doi.org/10.1210/jc.2004-1108

    Article  CAS  PubMed  Google Scholar 

  34. Heward JM, Brand OJ, Barrett JC, Carr-Smith JD, Franklyn JA, Gough SC (2007) Association of PTPN22 haplotypes with Graves’ disease. J Clin Endocrinol Metab 92(2):685–690. https://doi.org/10.1210/jc.2006-2064

    Article  CAS  PubMed  Google Scholar 

  35. Lee YH, Rho YH, Choi SJ, Ji JD, Song GG, Nath SK, Harley JB (2007) The PTPN22 C1858T functional polymorphism and autoimmune diseases—a meta-analysis. Rheumatology (Oxf) 46(1):49–56. https://doi.org/10.1093/rheumatology/kel170

    Article  CAS  Google Scholar 

  36. Lefvert AK, Zhao Y, Ramanujam R, Yu S, Pirskanen R, Hammarstrom L (2008) PTPN22 R620W promotes production of anti-AChR autoantibodies and IL-2 in myasthenia gravis. J Neuroimmunol 197(2):110–113. https://doi.org/10.1016/j.jneuroim.2008.04.004

    Article  CAS  PubMed  Google Scholar 

  37. Vang T, Congia M, Macis MD, Musumeci L, Orrú V, Zavattari P, Nika K, Tautz L, Taskén K, Cucca F, Mustelin T, Bottini N (2005) Autoimmune-associated lymphoid tyrosine phosphatase is a gain-of-function variant. Nat Genet 37(12):1317–1319. https://doi.org/10.1038/ng1673

    Article  CAS  PubMed  Google Scholar 

  38. Cai TT, Muhali FS, Song RH, Qin Q, Wang X, Shi LF, Jiang WJ, Xiao L, Li DF, Zhang JA (2015) Genome-wide DNA methylation analysis in Graves’ disease. Genomics 105(4):204–210. https://doi.org/10.1016/j.ygeno.2015.01.001

    Article  CAS  PubMed  Google Scholar 

  39. Limbach M, Saare M, Tserel L, Kisand K, Eglit T, Sauer S, Axelsson T, Syvänen AC, Metspalu A, Milani L, Peterson P (2016) Epigenetic profiling in CD4+ and CD8+ T cells from Graves’ disease patients reveals changes in genes associated with T cell receptor signaling. J Autoimmun 67:46–56. https://doi.org/10.1016/j.jaut.2015.09.006

    Article  CAS  PubMed  Google Scholar 

  40. Chandra A, Senapati S, Roy S, Chatterjee G, Chatterjee R (2018) Epigenome-wide DNA methylation regulates cardinal pathological features of psoriasis. Clin Epigenet 10(1):108. https://doi.org/10.1186/s13148-018-0541-9

    Article  CAS  Google Scholar 

  41. Shalaby SM, Mackawy AMH, Atef DM, Atef RM, Saeed J (2019) Promoter methylation and expression of intercellular adhesion molecule 1 gene in blood of autoimmune thyroiditis patients. Mol Biol Rep 46(5):5345–5353. https://doi.org/10.1007/s11033-019-04990-6

    Article  CAS  PubMed  Google Scholar 

  42. Liu T, Sun J, Wang Z, Yang W, Zhang H, Fan C, Shan Z, Teng W (2017) Changes in the DNA methylation and hydroxymethylation status of the intercellular adhesion molecule 1 gene promoter in thyrocytes from autoimmune thyroiditis patients. Thyroid 27(6):838–845. https://doi.org/10.1089/thy.2016.0576

    Article  CAS  PubMed  Google Scholar 

  43. Morita E, Watanabe M, Inoue N, Hashimoto H, Haga E, Hidaka Y, Iwatani Y (2018) Methylation levels of the TNFA gene are different between Graves’ and Hashimoto’s diseases and influenced by the TNFA polymorphism. Autoimmunity 51(3):118–125. https://doi.org/10.1080/08916934.2018.1448078

    Article  CAS  PubMed  Google Scholar 

  44. Hashimoto H, Watanabe M, Inoue N, Hirai N, Haga E, Kinoshita R, Hidaka Y, Iwatani Y (2019) Association of IFNG gene methylation in peripheral blood cells with the development and prognosis of autoimmune thyroid diseases. Cytokine 123:154770. https://doi.org/10.1016/j.cyto.2019.154770

    Article  CAS  PubMed  Google Scholar 

  45. Xin Z, Hua L, Shi TT, Tuo X, Yang FY, Li Y, Cao X, Yang JK (2018) A genome-wide DNA methylation analysis in peripheral blood from patients identifies risk loci associated with Graves’ orbitopathy. J Endocrinol Investig 41(6):719–727. https://doi.org/10.1007/s40618-017-0796-6

    Article  CAS  Google Scholar 

  46. Wojciechowska-Durczynska K, Krawczyk-Rusiecka K, Zygmunt A, Stawerska R, Lewinski A (2016) In children with autoimmune thyroiditis CTLA4 and FCRL3 genes—but not PTPN22—are overexpressed when compared to adults. Neuroendocrinol Lett 37(1):65–69

    CAS  PubMed  Google Scholar 

  47. Tokić S, Štefanić M, Glavaš-Obrovac L, Kishore A, Navratilova Z, Petrek M (2018) miR-29a-3p/T-bet regulatory circuit is altered in T cells of patients with Hashimoto’s thyroiditis. Front Endocrinol (Lausanne) 9:264. https://doi.org/10.3389/fendo.2018.00264

    Article  Google Scholar 

Download references

Funding

This study was supported by the Research Committee of the Aristotle University of Thessaloniki (Grant Number 89650).

Author information

Authors and Affiliations

  1. 4th Department of Pediatrics, Papageorgiou General Hospital, School of Medicine, Faculty of Health Sciences, Aristotle University of Thessaloniki, Thessaloniki, Greece

    I. Kyrgios & S. Giza

  2. Laboratory of Biological Chemistry, School of Medicine, Faculty of Health Sciences, Aristotle University of Thessaloniki, Thessaloniki, Greece

    A. Fragou & G. Tzimagiorgis

  3. 2nd Department of Pediatrics, AHEPA General University Hospital, School of Medicine, Faculty of Health Sciences, Aristotle University of Thessaloniki, St. Kiriakidi 1, Thessaloniki, 54636, Greece

    A. Galli-Tsinopoulou

Authors
  1. I. Kyrgios
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. S. Giza
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. A. Fragou
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. G. Tzimagiorgis
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. A. Galli-Tsinopoulou
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

IK, GT and AGT conceived and designed the study. IK and SG performed collection of samples and clinical data. IK and AF performed laboratory analyses. IK and AF analyzed and interpreted the data. IK and SG drafted the manuscript. AF, GT and AGT revised it critically for important intellectual content. All authors read and approved the final manuscript.

Corresponding author

Correspondence to A. Galli-Tsinopoulou.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

All procedures performed in studies involving human participants were in accordance with the Ethical Standards of the Institutional and/or National Research Committee and with the 1964 Helsinki Declaration and its later amendments or comparable ethical standards. The study was approved by the Bioethics Committee of the Medical School of the Aristotle University of Thessaloniki (No. 62/17-2-2014).

Informed consent

Written informed consent was obtained from the parents.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Kyrgios, I., Giza, S., Fragou, A. et al. DNA hypermethylation of PTPN22 gene promoter in children and adolescents with Hashimoto thyroiditis. J Endocrinol Invest 44, 2131–2138 (2021). https://doi.org/10.1007/s40618-020-01463-7

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s40618-020-01463-7

Keywords

Search

Navigation