| Background and Aim:Coronary artery disease(CAD)based in the atherosclerosis and its complications such as thromboembolism,dysfunction of papillary muscle and cardiac rupture have posed a serious threat to human health.However,the mechanism of CAD is still unclear.A variety of cells involve in the development of CAD,including vascular endothelial cell,vascular smooth muscle cell and monocytes/macrophages.Monocytes can differentiate into macrophage which can result in forming atherosclerotic plaque by secretion of pro-inflammation factors,uptake of lipid and interaction with the other cells under ox-LDL stimulating.Many literature have reported that miRNA revolves in many cardiovascular diaseases,including CAD.Especially,miRNA can regulate the function of mononuclear macrophages.MiR-29a plays important roles in many biological process,such as inflammatory immunological reactions,glucose metabolism and extracellular matrix formation.Recent data showed miR-29a could regulate gene expression related to apoptosis and protein transport in macrophage and contribute to the development of tumor.Previously we have found that not only the expression of miR-29a,but also that of scavenger receptor SRA and CD 36 are enhanced in ox-LDL-stimulated dendritic cells.However,the function and mechanism of miR-29a in the macrophage under ox-LDL treatment have not been revealed yet.Thus we focus on the unresolved problems as mentioned above.Methods:Our study is based on the macrophage differentiated from THP-1 by treatment with PMD.Firstly,we observed the expression of miR-29a under ox-LDL stimulating.Moreover,we inhibited the scavenger receptor SRA,LOX-1 and CD36 involved in the uptake of ox-LDL and detected the expression of miR-29a.Then we found the transcription factors(TFs)by dual-luciferase reporter assay and the TF filter plate assay.Besides,we studied the mechanism of TFs regulating miR-29a transcription with bioinformatics analysis,DNA agarose gel electrophoresis,CHIP and co-IP.Finally,we observed the secretion of pro-inflammation factors and uptake of lipid after overexpression of miR-29a.Results:We use ox-LDL to stimulated macrophage differentiated from THP-1 and find the expression of miR-29 is elevated.The expression of miR-29a is decreased though inhibiting SRA and LOX-1,but that is not by CD36.We also find that miR-29a can promote the expression of SRA and that inhibiting miR-29a can lead to the decrease of SRA.Under ox-LDL stimulating,the 2,000bp upstream of the transcription initiation site of miR-29a in macrophage shows high activity.YY1 and STAT1 are the main TFs involved the miR-29 transcription.Besides,we inhibit YY1 and STAT1 respectively and find that they both can reduce the expression of miR-29a.We also find that the promoter region of miR-29a only has YY1 binding site through bioinformatics analysis,but not STAT1 binding site.We verify that wild type miR-29a can bind to YY1,while mutant one cannot.Not only the combination of YY1 and STAT1 with promoter region,but also the expression of YY1 and STAT1 increases under ox-LDL treatment.We also show that YY1 and ST ATI can combine with each other and this complex increases in the nucleus.After we inhibit the JAK1/2 kinase,the expression of YY1 and STAT1 is reduced.Besides,overexpression or knockdown miR-29a respectively shows that the expression of pro-inflammation factors IL-1?,IL-6 and TNF-? is elevated and reduced and that the uptake of lipid is increased or decreased.Conclusion:Our data indicate that ox-LDL can make the expression of YY1 and STAT1 elevated.YY1 and STAT1 can combine with each other,moreover,the complex can bind to the promoter region of miR-29a resulting in increasing miR-29a transcription.Besides,YY1 and STAT1 cooperatively induce miR-29a expression through JAK1/2 kinase activation.We also find miR-29a can positively regulate the secretion of pro-inflammation factors and lipid uptake. |
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