Fig. 5.

Pulsatile shear stress (PS) and oscillatory shear stress (OS) regulation of DNA methylation of genes involved in the endothelial-to-mesenchymal transition. A and B: human umbilical vein endothelial cells were pretreated with or without EUK-134 (1 μmol/l) for 5 h before stimulation with PS or OS for 24 h. Genomic DNA was converted to nonmethylated uracil, and DNA methylation levels of promoter regions of von Willebrand factor (vWF), CD31, cadherin 5 (CDH5), α-smooth muscle actin (α-SMA), cadherin 2 (CDH2), fibroblast-specific protein 1 (FSP1), and vimentin genes were measured by methylation-specific quantitative PCR. C and D: human umbilical vein endothelial cells were treated with or without 5-aza-2′-deoxycytidine (5-Aza; 5 μmol/l) for 12 h and then stimulated with OS for an additional 12 h. C: mRNA levels of vWF, CD31, and CDH5 were quantified by real-time quantitative PCR. D: modifications of DNA methylation on the promoter region of the vWF, CD31, and CDH5 genes were measured by methylation-specific quantitative PCR. E and F: the intima was collected from thoracic aorta (TA) or aortic arch (AA) areas of C57BL/6 mice (n = 18). Genomic DNA was converted to nonmethylated uracil, and the DNA methylation of the indicated genes was measured accordingly. Ctrl, control. Data are means ± SE from three independent experiments. *P < 0.05 between the indicated groups.