title: MicroRNA-10a regulation of proinflammatory phenotype in athero-susceptible endothelium in vivo and in vitro.
abstract: A chronic proinflammatory state precedes pathological change in arterial endothelial cells located within regions of susceptibility to atherosclerosis. The potential contributions of regulatory microRNAs to this disequilibrium were investigated by artery site-specific profiling in normal adult swine. Expression of endothelial microRNA10a (miR-10a) was lower in the athero-susceptible regions of the inner aortic arch and aorto-renal branches than elsewhere. Expression of Homeobox A1 (HOXA1), a known miR-10a target, was up-regulated in the same locations. Endothelial transcriptome microarray analysis of miR-10a knockdown in cultured human aortic endothelial cells (HAEC) identified IkappaB/NF-kappaB-mediated inflammation as the top category of up-regulated biological processes. Phosphorylation of IkappaBalpha, a prerequisite for IkappaBalpha proteolysis and NF-kappaB activation, was significantly up-regulated in miR-10a knockdown HAEC and was accompanied by increased nuclear expression of NF-kappaB p65. The inflammatory biomarkers monocyte chemotactic protein 1 (MCP-1), IL-6, IL-8, vascular cell adhesion molecule 1 (VCAM-1), and E-selectin were elevated following miR-10a knockdown. Conversely, knockin of miR-10a (a conservative 25-fold increase) inhibited the basal expression of VCAM-1 and E-selectin in HAEC. Two key regulators of IkappaBalpha degradation--mitogen-activated kinase kinase kinase 7 (MAP3K7; TAK1) and beta-transducin repeat-containing gene (betaTRC)--contain a highly conserved miR-10a binding site in the 3' UTR. Both molecules were up-regulated by miR-10a knockdown and suppressed by miR-10a knockin, and evidence of direct miR-10a binding to the 3' UTR was demonstrated by luciferase assay. Comparative expression studies of endothelium located in athero-susceptible aortic arch and athero-protected descending thoracic aorta identified significantly up-regulated MAP3K7, betaTRC, phopho-IkappaBalpha, and nuclear p65 expression suggesting that the differential expression of miR-10a contributes to the regulation of proinflammatory endothelial phenotypes in athero-susceptible regions in vivo.
authors.affiliation: Institute for Medicine and Engineering, Center for Bioinformatics, and Department of Genetics, University of Pennsylvania, Philadelphia, PA 19104, USA.
authors.forename: Yun
authors.initials: Y
authors.lastname: Fang
authors.forename: Congzhu
authors.initials: C
authors.lastname: Shi
authors.forename: Elisabetta
authors.initials: E
authors.lastname: Manduchi
authors.forename: Mete
authors.initials: M
authors.lastname: Civelek
authors.forename: Peter F
authors.initials: PF
authors.lastname: Davies
mesh_terms: D000818:Animals; D001013:Aorta, Thoracic / Q000378:metabolism / Q000473:pathology; D050197:Atherosclerosis / Q000235:genetics; D015153:Blotting, Western; D002460:Cell Line; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000378:metabolism / Q000473:pathology; D004727:Endothelium / Q000378:metabolism* / Q000473:pathology; D020869:Gene Expression Profiling; D055785:Gene Knockdown Techniques; D020022:Genetic Predisposition to Disease; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D020672:I-kappa B Proteins / Q000378:metabolism; D017404:In Situ Hybridization, Fluorescence; D007249:Inflammation / Q000235:genetics; D020930:MAP Kinase Kinase Kinases / Q000235:genetics / Q000378:metabolism; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D010641:Phenotype; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013552:Swine; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D044844:beta-Transducin Repeat-Containing Proteins / Q000235:genetics / Q000378:metabolism; D000099261:Homeobox A1 Protein; D000099151:MAP Kinase Kinase Kinase 7
chemical_list: D018398:Homeodomain Proteins; D020672:I-kappa B Proteins; D020930:MAP Kinase Kinase Kinases; D035683:MicroRNAs; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B; D014157:Transcription Factors; D044844:beta-Transducin Repeat-Containing Proteins; D000099261:Homeobox A1 Protein; D000099151:MAP Kinase Kinase Kinase 7; C471644:BTRC protein, human; C000606204:NFKBIA protein, human