Abstract 139: Promoter Methylation Changes in ALOX12, AIRE1, and NETO1 : Novel Epigenetic Markers for Atherosclerosis

Abstract

Background and Objective: We aimed to identify and validate specific genes developing changes in promoter methylation in atherosclerotic plaques. Methods: We initially profiled DNA methylation signatures via a high-throughput sequencing platform from 5 carotid endarterectomy (CEA) plaques to identify genes with altered promoter methylation. The significances of the identified gene-specific promoter methylations were validated by the comparison between 1) plaques and non-plaque intima of the common carotid artery harvested from 20 cadavers, 2) cellular components composing atherosclerosis including plaques and buffy coats of blood of 27 CEA patients, 5 human umbilical vein endothelial cells (HUVEC), 2 human aortic endothelial (HAEC) and 2 smooth muscle (HASMC) cell lines, and, 3) monocytes, T- and B-cell circulating in blood of 19 ischemic stroke patients. Finally, the localization pattern the genes was compared by the inflammatory cell types infiltrated into atherosclerotic plaque and non-plaque intima using immunofluorescence stain. Results: Upon the profiling, five genes ( AIRE1 , ALOX12 , FANK1 , NETO1 , and SERHL2 ) displayed changes in promoter methylation. Of these, AIRE1 , ALOX12, and NETO1 displayed higher methylation in plaques than in non-plaque intima, but lower than buffy coat of blood. In vascular cells, AIRE1 and NETO1 methylation was lower in HUVEC, HAEC and HASMC, but, ALOX12 methylation was higher in HUVEC and HAEC, but, lower in HASMC than CEA plaques. Of inflammatory cells, methylation of the three genes was significantly lower in monocytes than T- and B-cells. On immunofluorescence statin for ALOX12, CD14(+)-monocytes was 3 times more frequent than CD4(+)-T-cell of total ALOX12-stained cells in plaques than non-plaque intima. Conclusions: The present study profiled and validated the promoter methylation changes of AIRE1 , ALOX12, and NETO1 as novel epigenetic markers related with atherosclerosis. In particular, epigenetic augmentation of the 3 genes occurred in monocytes of the cellular components composing atherosclerotic plaques.