Abstract 8783: Methylation of FOXP3 In Regulatory T Cells is Related to the Severity of Coronary Atherosclerosis

Abstract

Objective: Regulatory T (Treg) cells play a protective role in experimental atherosclerosis. However, the role of Treg in the development of human coronary artery disease (CAD) is inconsistent. Demethylation in the DNA that encodes transcription factor forkhead box P3 ( FOXP3 ) is a prerequisite for the stable maintenance of suppressive properties in Tregs, which is also identified as the most specific marker for human Tregs. We aim to evaluate Treg levels in CAD patients by a novel method based on Treg-specific DNA demethylation within FOXP3 gene. Methods and Results: We included patients with acute myocardial infarction (AMI) and unstable angina (UA), and control subjects with normal coronary artery on angiography (NCA) in our study. Peripheral blood mononuclear cells were collected to determine Treg levels by both flow cytometry and PCR based DNA methylation analysis. We found that Treg levels that were quantified by the demethylation rate of Treg-specific demethylated region in FOXP3 gene, but not by flow cytometry, were univocally decreased in UA and AMI patients (NCA 4.22% ± 1.92%; UA 3.29% ± 1.45%; AMI 2.86% ± 1.26%). This decline was inversely related to the severity of CAD ( r =−0.308, P <0.05). After adjustment for traditional risk factors, the association was still significant ( β =−0.336, P <0.05). Furthermore, we detected the expression of DNA methyltransferases in CD4 + CD25 + Tregs sorted from included subjects. A significant increase of DNA methyltransferase3b was observed in CAD patients compared with normal controls. Treatment of CD4 + CD25 + Tregs with ox-LDL induced a 41% increase in the methylation of FOXP3 , accompanied with a more than 3-fold reduction of FOXP3 mRNA expression and increases in the expressions of DNA methyltransferase 3a and 3b. Moreover, the effect of ox-LDL on FOXP3 can be reversed by methyltransferase inhibitor (-)-epigallocatechin-3-gallate. Conclusion: Our data demonstrate that Treg cells are decreased in CAD patients and its reduction are associated with the severity of atherosclerosis. Ox-LDL decreases the expression of FOXP3 by DNA methylation. Thus, we confirm the important role of Treg cells in human CAD and suggest potential epigenetic regulation in the progression of atherosclerosis.