Molecular Biology Reports

, Volume 40, Issue 8, pp 5123–5128

Evaluation of polymorphism, hypermethylation and expression pattern of CTLA4 gene in a sample of Iranian patients with schizophrenia

  • Dor Mohammad Kordi-Tamandani
  • Shahram Vaziri
  • Nahid Dahmardeh
  • Adam Torkamanzehi


The cytotoxic T lymphocyte-associated antigen 4 gene (CTLA4) has a crucial role in regulation of T cell proliferation and mediates T cell apoptosis by encoding the T cell receptor. Schizophrenia (SCZ) patients often have abnormalities in terms of the function and development of the immune system. The aim of the present study was to investigate promoter variation and expression profile of the CTLA4 gene in patients with SCZ. We isolated genomic DNA from peripheral blood of 94 individuals with SCZ and 99 healthy control subjects. Genotypic analysis of CTLA4 (−318) was done by Tetra-ARMS-PCR. Methylation-specific polymerase chain reaction (MS-PCR) was used to estimate promoter hypermethylation of the CTLA4 gene. In addition, we investigated CTLA4 mRNA levels in 34 blood samples from cases and healthy controls using real-time reverse transcription PCR. The CT genotype of CTLA4 has a significantly protective effect on the risk to SCZ (OR = 0.44; 95 % CI 0.18–1.06, P = 0.007) in comparison with the wild CC genotype. Promoter methylation of the CTLA4 gene increased the risk of disease statistically (OR = 3.82, 95 % CI 1.34–10.9, P = 0.015) in cases when compared to healthy controls in blood samples. The mRNA expression level results showed statistically significant differences (P < 0.0001) between cases (n = 17) and healthy controls (n = 17). To the best of our knowledge, this is the first evidence showing that promoter methylation of the CTLA4 gene along with transition of C to T was linked to a significantly higher expression of CTLA4 mRNA levels in patients with SCZ.


Methylation Expression CTLA4 Polymorphism 


  1. 1.
    Brown AS (2006) Prenatal infection as a risk factor for schizophrenia. Schizophr Bull 32(2):200–202PubMedCrossRefGoogle Scholar
  2. 2.
    Balbi G, Ferrera F, Rizzi M, Piccioli P, Morabito A, Cardamone L, Ghio M, Palmisano GL, Carrara P, Pedemonte S, Sessarego M, De Angioletti M, Notaro R, Indiveri F, Pistillo MP (2007) Association of −318 C/T and +49 A/G cytotoxic T lymphocyte antigen-4 (CTLA-4) gene polymorphisms with a clinical subset of Italian patients with systemic sclerosis. Clin Exp Immunol 149(1):40–47. doi:10.1111/j.1365-2249.2007.03394.x PubMedCrossRefGoogle Scholar
  3. 3.
    Bird AP (1986) CpG-rich islands and the function of DNA methylation. Nature 321(6067):209–213. doi:10.1038/321209a0 PubMedCrossRefGoogle Scholar
  4. 4.
    Brand O, Gough S, Heward J (2005) HLA, CTLA-4 and PTPN22: the shared genetic master-key to autoimmunity? Expert Rev Mol Med 7(23):1–15. doi:10.1017/S1462399405009981 PubMedCrossRefGoogle Scholar
  5. 5.
    Braun J, Donner H, Siegmund T, Walfish PG, Usadel KH, Badenhoop K (1998) CTLA-4 promoter variants in patients with Graves’ disease and Hashimoto’s thyroiditis. Tissue Antigens 51(5):563–566PubMedCrossRefGoogle Scholar
  6. 6.
    Cannon M, Jones PB, Murray RM (2002) Obstetric complications and schizophrenia: historical and meta-analytic review. Am J Psychiatry 159(7):1080–1092PubMedCrossRefGoogle Scholar
  7. 7.
    Carreno BM, Bennett F, Chau TA, Ling V, Luxenberg D, Jussif J, Baroja ML, Madrenas J (2000) CTLA-4 (CD152) can inhibit T cell activation by two different mechanisms depending on its level of cell surface expression. J Immunol 165(3):1352–1356PubMedGoogle Scholar
  8. 8.
    Connor CM, Akbarian S (2008) DNA methylation changes in schizophrenia and bipolar disorder. Epigenetics 3(2):55–58PubMedCrossRefGoogle Scholar
  9. 9.
    Coon H, Myles-Worsley M, Tiobech J, Hoff M, Rosenthal J, Bennett P, Reimherr F, Wender P, Dale P, Polloi A, Byerley W (1998) Evidence for a chromosome 2p13–14 schizophrenia susceptibility locus in families from Palau, Micronesia. Mol Psychiatry 3(6):521–527PubMedCrossRefGoogle Scholar
  10. 10.
    Dariavach P, Mattei MG, Golstein P, Lefranc MP (1988) Human Ig superfamily CTLA-4 gene: chromosomal localization and identity of protein sequence between murine and human CTLA-4 cytoplasmic domains. Eur J Immunol 18(12):1901–1905. doi:10.1002/eji.1830181206 PubMedCrossRefGoogle Scholar
  11. 11.
    Eaton WW, Byrne M, Ewald H, Mors O, Chen C-Y, Agerbo E, Mortensen PB (2006) Association of schizophrenia and autoimmune diseases: linkage of Danish national registers. Am J Psychiatry 163(3):521–528PubMedCrossRefGoogle Scholar
  12. 12.
    Egen JG, Allison JP (2002) Cytotoxic T lymphocyte antigen-4 accumulation in the immunological synapse is regulated by TCR signal strength. Immunity 16(1):23–35PubMedCrossRefGoogle Scholar
  13. 13.
    Faraone SV, Matise T, Svrakic D, Pepple J, Malaspina D, Suarez B, Hampe C, Zambuto CT, Schmitt K, Meyer J, Markel P, Lee H, Harkavy Friedman J, Kaufmann C, Cloninger CR, Tsuang MT (1998) Genome scan of European-American schizophrenia pedigrees: results of the NIMH genetics initiative and millennium consortium. Am J Med Genet 81(4):290–295. doi:10.1002/(SICI)1096-8628(19980710)81:4<290:AID-AJMG3>3.0.CO;2-Y PubMedCrossRefGoogle Scholar
  14. 14.
    Gogas H, Dafni U, Koon H, Spyropoulou-Vlachou M, Metaxas Y, Buchbinder E, Pectasides E, Tsoutsos D, Polyzos A, Stratigos A, Markopoulos C, Panagiotou P, Fountzilas G, Castana O, Skarlos P, Atkins MB, Kirkwood JM (2010) Evaluation of six CTLA-4 polymorphisms in high-risk melanoma patients receiving adjuvant interferon therapy in the He13A/98 multicenter trial. J Transl Med 8:108. doi:10.1186/1479-5876-8-108 PubMedCrossRefGoogle Scholar
  15. 15.
    Gogas H, Dafni U, Koon H, Spyropoulou-Vlachou M, Metaxas Y, Buchbinder E, Pectasides E, Tsoutsos D, Polyzos A, Stratigos A, Markopoulos C, Panagiotou P, Fountzilas G, Castana O, Skarlos P, Atkins MB, Kirkwood JM (2010) Evaluation of six CTLA-4 polymorphisms in high-risk melanoma patients receiving adjuvant interferon therapy in the He13A/98 multicenter trial. J Transl Med 8:108PubMedCrossRefGoogle Scholar
  16. 16.
    Gough SC, Walker LS, Sansom DM (2005) CTLA4 gene polymorphism and autoimmunity. Immunol Rev 204:102–115. doi:10.1111/j.0105-2896.2005.00249.x PubMedCrossRefGoogle Scholar
  17. 17.
    Jones AL, Holliday EG, Mowry BJ, McLean DE, McGrath JJ, Pender MP, Greer JM (2009) CTLA-4 single-nucleotide polymorphisms in a Caucasian population with schizophrenia. Brain Behav Immun 23(3):347–350. doi:10.1016/j.bbi.2008.09.008 PubMedCrossRefGoogle Scholar
  18. 18.
    Jun TY, Pae CU, Chae JH, Bahk WM, Kim KS, Han H (2002) Polymorphism of CTLA-4 gene at position 49 of exon 1 may be associated with schizophrenia in the Korean population. Psychiatry Res 110(1):19–25PubMedCrossRefGoogle Scholar
  19. 19.
    Karanikas EP (2011) Psycho-immunological mechanisms in schizophrenia. Psychiatrike 22(1):43–52PubMedGoogle Scholar
  20. 20.
    Kinney DK, Hintz K, Shearer EM, Barch DH, Riffin C, Whitley K, Butler R (2010) A unifying hypothesis of schizophrenia: abnormal immune system development may help explain roles of prenatal hazards, post-pubertal onset, stress, genes, climate, infections, and brain dysfunction. Med Hypotheses 74(3):555–563PubMedCrossRefGoogle Scholar
  21. 21.
    Kordi-Tamandani DM, Hashemi M, Birjandian E, Bahari A, Valizadeh J, Torkamanzehi A (2011) Lack of association of GSTT1 and GSTP1 genes methylation and their expression profiles with risk of NAFLD in a sample of Iranian patients. Clin Res Hepatol Gastroenterol 35(5):387–392. doi:10.1016/j.clinre.2011.01.015 PubMedCrossRefGoogle Scholar
  22. 22.
    Kordi-Tamandani DM, Hashemi M, Sharifi N, Kaykhaei MA, Torkamanzehi A (2011) Association between paraoxonase-1 gene polymorphisms and risk of metabolic syndrome. Mol Biol Rep. doi:10.1007/s11033-011-0819-x PubMedGoogle Scholar
  23. 23.
    Kordi-Tamandani DM, Moazeni-Roodi A-K, Rigi-Ladiz M-A, Hashemi M, Birjandian E, Torkamanzehi A (2010) Promoter hypermethylation and expression profile of MGMT and CDH1 genes in oral cavity cancer. Arch Oral Biol 55(10):809–814PubMedCrossRefGoogle Scholar
  24. 24.
    Kordi-Tamandani DM, Moazeni-Roodi A, Rigi Ladez MA, Hashemi M, Birjandian E, Torkamanzehi A (2010) Analysis of methylation patterns and expression profiles of p14ARF gene in patients with oral squamous cell carcinoma. Int J Biol Markers 25(2):99–103PubMedGoogle Scholar
  25. 25.
    Kordi-Tamandani DM, Moazeni-Roodi AK, Rigi-Ladiz MA, Hashemi M, Birjandian E, Torkamanzehi A (2010) Promoter hypermethylation and expression profile of MGMT and CDH1 genes in oral cavity cancer. Arch Oral Biol 55(10):809–814. doi:10.1016/j.archoralbio.2010.06.017 PubMedCrossRefGoogle Scholar
  26. 26.
    Kronfol Z, House JD (1989) Lymphocyte mitogenesis, immunoglobulin and complement levels in depressed patients and normal controls. Acta Psychiatr Scand 80(2):142–147PubMedCrossRefGoogle Scholar
  27. 27.
    Ligers A, Teleshova N, Masterman T, Huang WX, Hillert J (2001) CTLA-4 gene expression is influenced by promoter and exon 1 polymorphisms. Genes Immun 2(3):145–152. doi:10.1038/sj.gene.6363752 PubMedCrossRefGoogle Scholar
  28. 28.
    Lindsten T, Lee KP, Harris ES, Petryniak B, Craighead N, Reynolds PJ, Lombard DB, Freeman GJ, Nadler LM, Gray GS et al (1993) Characterization of CTLA-4 structure and expression on human T cells. J Immunol 151(7):3489–3499PubMedGoogle Scholar
  29. 29.
    Linsley PS, Nadler SG, Bajorath J, Peach R, Leung HT, Rogers J, Bradshaw J, Stebbins M, Leytze G, Brady W et al (1995) Binding stoichiometry of the cytotoxic T lymphocyte-associated molecule-4 (CTLA-4). A disulfide-linked homodimer binds two CD86 molecules. J Biol Chem 270(25):15417–15424PubMedCrossRefGoogle Scholar
  30. 30.
    Liu J, Li J, Li T, Wang T, Li Y, Zeng Z, Li Z, Chen P, Hu Z, Zheng L, Ji J, Lin H, Feng G, Shi Y (2011) CTLA-4 confers a risk of recurrent schizophrenia, major depressive disorder and bipolar disorder in the Chinese Han population. Brain Behav Immun 25(3):429–433. doi:10.1016/j.bbi.2010.10.024 PubMedCrossRefGoogle Scholar
  31. 31.
    Masterma T, Ligers A, Olerup O, Vrethem M, Hillert J (1999) CTLA-4 dimorphisms in gammopathy-associated peripheral neuropathy. Eur J Neurol 6(4):491–493PubMedCrossRefGoogle Scholar
  32. 32.
    Muller N, Schwarz M (2006) Schizophrenia as an inflammation-mediated dysbalance of glutamatergic neurotransmission. Neurotox Res 10(2):131–148PubMedCrossRefGoogle Scholar
  33. 33.
    Pentcheva-Hoang T, Egen JG, Wojnoonski K, Allison JP (2004) B7–1 and B7–2 selectively recruit CTLA-4 and CD28 to the immunological synapse. Immunity 21(3):401–413PubMedCrossRefGoogle Scholar
  34. 34.
    Quan N, Banks WA (2007) Brain-immune communication pathways. Brain Behav Immun 21(6):727–735. doi:10.1016/j.bbi.2007.05.005 PubMedCrossRefGoogle Scholar
  35. 35.
    Riedel M, Spellmann I, Schwarz MJ, Strassnig M, Sikorski C, Moller HJ, Muller N (2007) Decreased T cellular immune response in schizophrenic patients. J Psychiatr Res 41(1–2):3–7. doi:10.1016/j.jpsychires.2005.11.007 PubMedCrossRefGoogle Scholar
  36. 36.
    Rothermundt M, Arolt V, Bayer TA (2001) Review of immunological and immunopathological findings in schizophrenia. Brain Behav Immun 15(4):319–339. doi:10.1006/brbi.2001.0648 PubMedCrossRefGoogle Scholar
  37. 37.
    Sedlackova L, Nguyen TT, Zlacka D, Sosna A, Hromadnikova I (2009) Cell surface and relative mRNA expression of heat shock protein 70 in human synovial cells. Autoimmunity 42(1):17–24. doi:10.1080/08916930802227466 PubMedCrossRefGoogle Scholar
  38. 38.
    Shiratori T, Miyatake S, Ohno H, Nakaseko C, Isono K, Bonifacino JS, Saito T (1997) Tyrosine phosphorylation controls internalization of CTLA-4 by regulating its interaction with clathrin-associated adaptor complex AP-2. Immunity 6(5):583–589PubMedCrossRefGoogle Scholar
  39. 39.
    Silva JM, Dominguez G, Villanueva MJ, Gonzalez R, Garcia JM, Corbacho C, Provencio M, Espana P, Bonilla F (1999) Aberrant DNA methylation of the p16INK4a gene in plasma DNA of breast cancer patients. Br J Cancer 80(8):1262–1264. doi:10.1038/sj.bjc.6690495 PubMedCrossRefGoogle Scholar
  40. 40.
    Soderlund J, Schroder J, Nordin C, Samuelsson M, Walther-Jallow L, Karlsson H, Erhardt S, Engberg G (2009) Activation of brain interleukin-1beta in schizophrenia. Mol Psychiatry 14(12):1069–1071. doi:10.1038/mp.2009.52 PubMedCrossRefGoogle Scholar
  41. 41.
    St Clair D, Xu M, Wang P, Yu Y, Fang Y, Zhang F, Zheng X, Gu N, Feng G, Sham P, He L (2005) Rates of adult schizophrenia following prenatal exposure to the Chinese famine of 1959–1961. JAMA 294(5):557–562. doi:10.1001/jama.294.5.557 PubMedCrossRefGoogle Scholar
  42. 42.
    Strous RD, Shoenfeld Y (2006) Schizophrenia, autoimmunity and immune system dysregulation: a comprehensive model updated and revisited. J Autoimmun 27(2):71–80PubMedCrossRefGoogle Scholar
  43. 43.
    Sullivan PF, Kendler KS, Neale MC (2003) Schizophrenia as a complex trait: evidence from a meta-analysis of twin studies. Arch Gen Psychiatry 60(12):1187–1192. doi:10.1001/archpsyc.60.12.1187 PubMedCrossRefGoogle Scholar
  44. 44.
    Ueda H, Howson JMM, Esposito L, Heward J, Snook H, Chamberlain G, Rainbow DB, Hunter KMD, Smith AN, Di Genova G, Herr MH, Dahlman I, Payne F, Smyth D, Lowe C, Twells RCJ, Howlett S, Healy B, Nutland S, Rance HE, Everett V, Smink LJ, Lam AC, Cordell HJ, Walker NM, Bordin C, Hulme J, Motzo C, Cucca F, Hess JF, Metzker ML, Rogers J, Gregory S, Allahabadia A, Nithiyananthan R, Tuomilehto-Wolf E, Tuomilehto J, Bingley P, Gillespie KM, Undlien DE, Ronningen KS, Guja C, Ionescu-Tirgoviste C, Savage DA, Maxwell AP, Carson DJ, Patterson CC, Franklyn JA, Clayton DG, Peterson LB, Wicker LS, Todd JA, Gough SCL (2003) Association of the T-cell regulatory gene CTLA4 with susceptibility to autoimmune disease. Nature 423(6939):506–511PubMedCrossRefGoogle Scholar
  45. 45.
    Viviani B, Gardoni F, Marinovich M (2007) Cytokines and neuronal ion channels in health and disease. Int Rev Neurobiol 82:247–263. doi:10.1016/S0074-7742(07)82013-7 PubMedCrossRefGoogle Scholar
  46. 46.
    Wang XB, Zhao X, Giscombe R, Lefvert AK (2002) A CTLA-4 gene polymorphism at position −318 in the promoter region affects the expression of protein. Genes Immun 3(4):233–234. doi:10.1038/sj.gene.6363869 PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2013

Authors and Affiliations

  • Dor Mohammad Kordi-Tamandani
    • 1
  • Shahram Vaziri
    • 2
  • Nahid Dahmardeh
    • 1
  • Adam Torkamanzehi
    • 1
  1. 1.Department of BiologyUniversity of Sistan and BaluchestanZahedanIran
  2. 2.Department of PsychologyIslamic Azad University, Roodehen BranchTehranIran

Personalised recommendations