Identification of novel hyper- or hypomethylated CpG sites and genes associated with atherosclerotic plaque using 
an epigenome-wide association study

Yamada,Yoshiji; Horibe,Hideki; Oguri,Mitsutoshi; Sakuma,Jun; Takeuchi,Ichiro; Yasukochi,Yoshiki; Kato,Kimihiko; Sawabe,Motoji

doi:10.3892/ijmm.2018.3453

Identification of novel hyper- or hypomethylated CpG sites and genes associated with atherosclerotic plaque using an epigenome-wide association study

Authors:
- Yoshiji Yamada
- Hideki Horibe
- Mitsutoshi Oguri
- Jun Sakuma
- Ichiro Takeuchi
- Yoshiki Yasukochi
- Kimihiko Kato
- Motoji Sawabe
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Affiliations: Department of Human Functional Genomics, Advanced Science Research Promotion Center, Mie University, Tsu 514‑8507, Japan, Department of Cardiovascular Medicine, Gifu Prefectural Tajimi Hospital, Tajimi 507‑8522, Japan, CREST, Japan Science and Technology Agency, Kawaguchi 332‑0012, Japan, Section of Molecular Pathology, Graduate School of Health Care Sciences, Tokyo Medical and Dental University, Tokyo 113‑8510, Japan
Published online on: February 2, 2018 https://doi.org/10.3892/ijmm.2018.3453
Pages: 2724-2732
Copyright: © Yamada et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

DNA methylation is an important epigenetic modification that has been implicated in the pathogenesis of atherosclerosis. Although previous studies have identified various CpG sites and genes whose methylation is associated with atherosclerosis in populations with European or Mexican ancestry, the genome‑wide pattern of DNA methylation in the atherosclerotic human aorta is yet to be elucidated in Japanese individuals. In the present study, a genome‑wide analysis of DNA methylation at ~853,000 CpG sites was performed using 128 postmortem aortic intima specimens obtained from 64 Japanese patients. To avoid the effects of interindividual variation, intraindividual paired comparisons were performed between atheromatous plaque lesions and corresponding plaque‑free tissue for each patient. Bisulfite‑modified genomic DNA was analyzed using a specific microarray for DNA methylation. DNA methylation at each CpG site was calculated as the β value, where β = (intensity of the methylated allele)/(intensity of the methylated allele + intensity of the unmethylated allele + 100). Bonferroni's correction for statistical significance of association was applied to compensate for multiple comparisons. The methylation of 2,679 CpG sites differed significantly (P<5.86x10‑8) between atheromatous plaque lesions and the corresponding plaque‑free intima, with 2,272 and 407 CpG sites in atheromatous plaques being hyper‑ or hypomethylated, respectively. A total of 5 hypermethylated CpG sites in atheromatous plaques were demonstrated to have a difference in β value of >0.15 (plaque lesion‑plaque‑free intima) and 11 had a β ratio of >1.50 (plaque/plaque‑free intima). A further 15 and 17 hypomethylated CpG sites in atheromatous plaques were observed to have a difference in β value of <‑0.15 or a β ratio of <0.67, respectively. According to these limits, a total of 16 novel genes that were significantly hyper‑ or hypomethylated in atheromatous plaque lesions compared with the plaque‑free intima were identified in the present study. The results of the present study suggest that the methylation of these genes may contribute to the pathogenesis of atherosclerosis in the Japanese population.

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1	Lusis AJ: Atherosclerosis. Nature. 407:233–241. 2000. View Article : Google Scholar : PubMed/NCBI
2	Baccarelli A, Rienstra M and Benjamin EJ: Cardiovascular epigenetics: Basic concepts and results from animal and human studies. Circ Cardiovasc Genet. 3:567–573. 2010. View Article : Google Scholar : PubMed/NCBI
3	Neele AE, Van den Bossche J, Hoeksema MA and de Winther MP: Epigenetic pathways in macrophages emerge as novel targets in atherosclerosis. Eur J Pharmacol. 763:79–89. 2015. View Article : Google Scholar : PubMed/NCBI
4	Rakyan VK, Down TA, Balding DJ and Beck S: Epigenome-wide association studies for common human diseases. Nat Rev Genet. 12:529–541. 2011. View Article : Google Scholar : PubMed/NCBI
5	Handy DE, Castro R and Loscalzo J: Epigenetic modifications: Basic mechanisms and role in cardiovascular disease. Circulation. 123:2145–2156. 2011. View Article : Google Scholar : PubMed/NCBI
6	Tsai PC, Spector TD and Bell JT: Using epigenome-wide association scans of DNA methylation in age-related complex human traits. Epigenomics. 4:511–526. 2012. View Article : Google Scholar : PubMed/NCBI
7	Pfeiffer L, Wahl S, Pilling LC, Reischl E, Sandling JK, Kunze S, Holdt LM, Kretschmer A, Schramm K, Adamski J, et al: DNA methylation of lipid-related genes affects blood lipid levels. Circ Cardiovasc Genet. 8:334–342. 2015. View Article : Google Scholar : PubMed/NCBI
8	Rask-Andersen M, Martinsson D, Ahsan M, Enroth S, Ek WE, Gyllensten U and Johansson Å: Epigenome-wide association study reveals differential DNA methylation in individuals with a history of myocardial infarction. Hum Mol Genet. 25:4739–4748. 2016.
9	Wahl S, Drong A, Lehne B, Loh M, Scott WR, Kunze S, Tsai PC, Ried JS, Zhang W, Yang Y, et al: Epigenome-wide association study of body mass index, and the adverse outcomes of adiposity. Nature. 541:81–86. 2017. View Article : Google Scholar :
10	Li J, Zhu X, Yu K, Jiang H, Zhang Y, Deng S, Cheng L, Liu X, Zhong J, Zhang X, et al: Genome-wide analysis of DNA methylation and acute coronary syndrome. Circ Res. 120:1754–1767. 2017. View Article : Google Scholar : PubMed/NCBI
11	Fernández-Sanlés A, Sayols-Baixeras S, Subirana I, Degano IR and Elosua R: Association between DNA methylation and coronary heart disease or other atherosclerotic events: A systematic review. Atherosclerosis. 263:325–333. 2017. View Article : Google Scholar : PubMed/NCBI
12	Khyzha N, Alizada A, Wilson MD and Fish JE: Epigenetics of atherosclerosis: Emerging mechanisms and methods. Trends Mol Med. 23:332–347. 2017. View Article : Google Scholar : PubMed/NCBI
13	Deaton AM and Bird A: CpG islands and the regulation of transcription. Genes Dev. 25:1010–1022. 2011. View Article : Google Scholar : PubMed/NCBI
14	Nazarenko MS, Puzyreva VP, Lebedev IN, Frolov AV, Barbarash OL and Barbarash LS: Methylation profiling of DNA in the area of atherosclerotic plaque in humans. Mol Biol. 45:5612011. View Article : Google Scholar
15	Zaina S, Heyn H, Carmona FJ, Varol N, Sayols S, Condom E, Ramírez-Ruz J, Gomez A, Gonçalves I, Moran S and Esteller M: DNA methylation map of human atherosclerosis. Circ Cardiovasc Genet. 7:692–700. 2014. View Article : Google Scholar : PubMed/NCBI
16	Aavik E, Lumivuori H, Leppänen O, Wirth T, Häkkinen SK, Bräsen JH, Beschorner U, Zeller T, Braspenning M, van Criekinge W, et al: Global DNA methylation analysis of human atherosclerotic plaques reveals extensive genomic hypomethylation and reactivation at imprinted locus 14q32 involving induction of a miRNA cluster. Eur Heart J. 36:993–1000. 2015. View Article : Google Scholar
17	Kucher AN, Nazarenko MS, Markov AV, Koroleva IA and Barbarash OL: Variability of methylation profiles of CpG sites in microRNA genes in leukocytes and vascular tissues of patients with atherosclerosis. Biochemistry. 82:698–706. 2017.PubMed/NCBI
18	Castillo-Díaz SA, Garay-Sevilla ME, Hernández-González MA, Solís-Martínez MO and Zaina S: Extensive demethylation of normally hypermethylated CpG islands occurs in human atherosclerotic arteries. Int J Mol Med. 26:691–700. 2010.PubMed/NCBI
19	Yamada Y, Nishida T, Horibe H, Oguri M, Kato K and Sawabe M: Identification of hypo- and hypermethylated genes related to atherosclerosis by a genome-wide analysis of DNA methylation. Int J Mol Med. 33:1355–1363. 2014. View Article : Google Scholar : PubMed/NCBI
20	Moran S, Arribas C and Esteller M: Validation of a DNA meth-ylation microarray for 850,000 CpG sites of the human genome enriched in enhancer sequences. Epigenomics. 8:389–399. 2016. View Article : Google Scholar
21	Christensen BC, Houseman EA, Marsit CJ, Zheng S, Wrensch MR, Wiemels JL, Nelson HH, Karagas MR, Padbury JF, Bueno R, et al: Aging and environmental exposures alter tissue-specific DNA methylation dependent upon CpG island context. PLoS Genet. 5:e10006022009. View Article : Google Scholar : PubMed/NCBI
22	Bell JT, Pai AA, Pickrell JK, Gaffney DJ, Pique-Regi R, Degner JF, Gilad Y and Pritchard JK: DNA methylation patterns associate with genetic and gene expression variation in HapMap cell lines. Genome Biol. 12:R102011. View Article : Google Scholar : PubMed/NCBI
23	ENCODE Project Consortium: An integrated encyclopedia of DNA elements in the human genome. Nature. 489:57–74. 2012. View Article : Google Scholar : PubMed/NCBI
24	Lizio M, Harshbarger J, Shimoji H, Severin J, Kasukawa T, Sahin S, Abugessaisa I, Fukuda S, Hori F, Ishikawa-Kato S, et al: Gateways to the FANTOM5 promoter level mammalian expression atlas. Genome Biol. 16:222015. View Article : Google Scholar : PubMed/NCBI
25	Bibikova M, Barnes B, Tsan C, Ho V, Klotzle B, Le JM, Delano D, Zhang L, Schroth GP, Gunderson KL, et al: High density DNA methylation array with single CpG site resolution. Genomics. 98:288–295. 2011. View Article : Google Scholar : PubMed/NCBI
26	Libby P: Inflammation in atherosclerosis. Nature. 420:868–874. 2002. View Article : Google Scholar : PubMed/NCBI
27	Libby P: Mechanisms of acute coronary syndromes and their implications for therapy. N Engl J Med. 368:2004–2013. 2013. View Article : Google Scholar : PubMed/NCBI
28	Turunen MP, Aavik E and Ylä-Herttuala S: Epigenetics and atherosclerosis. Biochim Biophys Acta. 1790:886–891. 2009. View Article : Google Scholar : PubMed/NCBI
29	Hai Z and Zuo W: Aberrant DNA methylation in the pathogenesis of atherosclerosis. Clin Chim Acta. 456:69–74. 2016. View Article : Google Scholar : PubMed/NCBI
30	Fishbein GA and Fishbein MC: Arteriosclerosis: Rethinking the current classification. Arch Pathol Lab Med. 133:1309–1316. 2009.PubMed/NCBI
31	Peng Y, Meng K, Jiang L, Zhong Y, Yang Y, Lan Y, Zeng Q and Cheng L: Thymic stromal lymphopoietin-induced HOTAIR activation promotes endothelial cell proliferation and migration in atherosclerosis. Biosci Rep. 37:pii: BSR201703512017. View Article : Google Scholar
32	Shen T, Zhu Y, Patel J, Ruan Y, Chen B, Zhao G, Cao Y, Pang J, Guo H, Li H, et al: T-box20 suppresses oxidized low-density lipoprotein-induced human vascular endothelial cell injury by upregulation of PPAR-γ. Cell Physiol Biochem. 32:1137–1150. 2013. View Article : Google Scholar
33	Warren HR, Evangelou E, Cabrera CP, Gao H, Ren M, Mifsud B, Ntalla I, Surendran P, Liu C, Cook JP, et al: Genome-wide association analysis identifies novel blood pressure loci and offers biological insights into cardiovascular risk. Nat Genet. 49:403–415. 2017. View Article : Google Scholar : PubMed/NCBI
34	Graff M, Scott RA, Justice AE, Young KL, Feitosa MF, Barata L, Winkler TW, Chu AY, Mahajan A, Hadley D, et al: Genome-wide physical activity interactions in adiposity-a meta-analysis of 200,452 adults. PLoS Genet. 13:e10065282017. View Article : Google Scholar
35	Zhang J, Burridge KA and Friedman MH: In vivo differences between endothelial transcriptional profiles of coronary and iliac arteries revealed by microarray analysis. Am J Physiol Heart Circ Physiol. 295:H1556–H1561. 2008. View Article : Google Scholar : PubMed/NCBI
36	Ellinghaus E, Ellinghaus D, Krusche P, Greiner A, Schreiber C, Nikolaus S, Gieger C, Strauch K, Lieb W, Rosenstiel P, et al: Genome-wide association analysis for chronic venous disease identifies EFEMP1 and KCNH8 as susceptibility loci. Sci Rep. 7:456522017. View Article : Google Scholar : PubMed/NCBI
37	Astle WJ, Elding H, Jiang T, Allen D, Ruklisa D, Mann AL, Mead D, Bouman H, Riveros-Mckay F, Kostadima MA, et al: The allelic landscape of human blood cell trait variation and links to common complex disease. Cell. 167:1415–1429.e19. 2016. View Article : Google Scholar : PubMed/NCBI