Genome-Wide Methylation Analysis Identifies Novel Cpg Loci For Perimembranous Ventricular Septal Defects In Human

Abstract

Aim: Congenital heart diseases are the most common birth defects worldwide and leading cause of infant mortality. The perimembranous ventricular septal defect is most prevalent. Epigenetics may provide an underlying mechanism of the gene–environment interactions involved. Materials & methods: We examined epigenome-wide DNA methylation using the Illumina HumanMethylation450 BeadChip in 84 case children and 196 control children. Results: We identified differential methylation of a CpG locus (cg17001566) within the PRDM16 gene after Bonferroni correction (p = 9.17 × 10^-8). This was validated by bisulfite pyrosequencing. PRDM16 functions as a repressor of TGF-β signaling controlling tissue morphogenesis crucial during cardiogenesis. At 15% false-discovery rate, we identified seven additional CpG loci. Conclusion: These findings provide novel insights in the pathogenesis of perimembranous ventricular septal defect, which is of interest for future prediction and prevention.

Keywords::

• bisulfite pyrosequencing
• case–control studies
• DNA methylation
• epigenetics
• epigenome-wide
• Illumina 450k
• perimembranous ventricular septal defects

All authors critically revised and approved the manuscript. KPJ Wijnands participated in the design of the study, laboratory preparations, statistical analysis of the data and wrote the first draft and revisions of the manuscript. J Chen performed the statistical analysis, drafted the statistical methods part and was involved in interpretation of the data. L Liang and X Lin participated in the statistical analysis. MMPJ Verbiest was involved in the laboratory work of the 450k arrays and bisulfite pyrosequencing. WA Helbing was clinically involved in enrollment of cases. AC Gittenberger dan Groot contributed to the interpretation of the data. PJ van der Spek performed the pathway analysis and was involved in interpretation of the data. AG Uitterlinden supervised the laboratory logistics. RPM Steegers-Theunissen initiated and designed the study, supervised all aspects of the study and contributed to all versions of the manuscript.

Acknowledgements

The authors are grateful to all participating families. J Ottenkamp of the departments of Paediatric Cardiology of Leiden University Medical Center, Academic Medical Center Amsterdam and VU University Medical Center, and FMH Siebel of the child health centers of Careyn, are acknowledged for their involvement in recruitment of cases and controls, respectively. The authors thank the PhD students of the HAVEN study for data acquisition and recruitment.

Financial & competing interests disclosure

The authors have no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties.

No writing assistance was utilized in the production of this manuscript.

Ethical conduct of research

The authors state that they have obtained appropriate institutional review board approval or have followed the principles outlined in the Declaration of Helsinki for all human or animal experimental investigations. In addition, for investigations involving human subjects, informed consent has been obtained from the participants involved.