The Effects Of Hcy And Ox-LDL On The Expression And The Methylation Modification Of ER And LDLR Gene

Atherosclerosis is a common disease, it is the major cause of death in developed countries and also the most rapidly increased risk factor for death in most developing countries. Being a most common type of cardiovascular diseases, the incidence rate of AS increases quite fast in our country. Because many details of the pathologenical mechanisms of AS are still obscure, the advanced preventive and curative measures still is badly needed.DNA methylation is an epigenetic process leading to the chemical modification of genome. Accumulating evidence has shown aberrant DNA methylation patterns in various diseases, including cancer, certain X-linked genetic diseases ,autoimmune diseases , aging, etc. Until now, the estrogen receptorα(ERα) gene is the only gene known to have aberrant hypermethylation in its promoter region in atherosclerosis.High levels of homocysteine and oxidized low density lipoprotein (ox-LDL) have been proved as the independent risk factors for atherosclerosis and many other cardiovascular diseases. So what are the effects of these two risk factors on the methylation modification in atherosclerosis-related genes and the As development? The low density lipoprotein receptor(LDLR) and estrogen receptor are both directly related to atherogenesis, so to investigate the aberrant DNA methylation patterns in LDLR and ERαgenes in high levels of homocysteine and ox-LDL and their correlation with atherosclerosis development may reveal some missing knowledge links between risk factors and atherosclerosis development.If the two important risk factors could exert a profound influence for atherosclerosis on the methylation pattern of the ERαgene and LDLR gene, it would be most helpful in our understanding of the mechanisms of atherosclerosis development. We therefore designed the current study to investigate the potential effects of Hcy and ox-LDL with various concentrations and treating times on the methylation pattern and their expression of the ERαgene and LDLR gene.Cultured smooth muscle cells of humans umbilical vessel (HUSMCs) and THP-1 cells were treated by Hcy and ox-LDL with different concentrations for different periods of time. The DNA methylation status was assayed by nested methylation-specific polymERαse chain reaction (nMS-PCR) ; the expression of ERαof the cells were detected by immunohistochemical assay; the function of LDLR of THP-1 cells were dected by enzyme linked immunosorbent assay; the gene expression of ERαand LDLR were detected by Semiquantitative reverse transcription polymERαse chain reaction (RT-PCR) ; MTT assay was used to observe the prolifERαtion of HUSMCs.The results showed that ox-LDL in modERαte concentrations (10-40 mg/L) induced de novo methylation in the promoter region of the ERαgene of the cells. However, high concentrations (50 mg/L) of ox-LDL, resulted in demethylation of ERα. The Hcy treatment resulted in de novo methylation in the promoter region of the ERαgene with a concentration- and treating time-dependent manner. And the hypermethylation of the ERαgene lead to the decreased expression of ERαand the prolifERαtion of SMCs. The Hcy and ox-LDL treatment have no obvious effect on the methylation modification in the promoter region of the LDLR gene.These data indicated that the two risk factors for atherosclerosis had the function of inducing de novo methylation in the promoter region of the ERαgene of SMCs. However, high concentrations (50mg/L) of ox-LDL induced demethylation, indicating that different risk factors of atherosclerosis with different potency might cause different aberrant methylation patterns in the promoter region of the ERαgene. The atherogenic mechanism of Hcy might involve the hypermethylation of the ERαgene, leading to the prolifERαtion of SMCs in atherosclerotic lesions. But the methylation pattern of LDLR gene seems not to be modifyed by Hcy and ox-LDL treatment.