Differential Role of an NF-κB Transcriptional Response Element in Endothelial Versus Intimal Cell VCAM-1 Expression. Issue 2 (3rd July 2015)

• Record Type: Journal Article
• Authors: Milstone, David S.; Ilyama, Motoi; Chen, Mian; O'Donnell, Peter; Davis, Vannessa M.; Plutzky, Jorge; Brown, Jonathan D.; Haldar, Saptarsi M.; Siu, Allan; Lau, Andrew C.; Zhu, Su-Ning; Basheer, Mayada F.; Collins, Tucker; Jongstra-Bilen, Jenny; Cybulsky, Myron I.
• Is Part Of: Circulation research. Volume 117:Issue 2(2015)
• Publication Date: 2015-07-03
• Subjects: atherosclerosis -- chromatin immunoprecipitation -- endothelial cells -- gene expression -- NF-κB -- tunica intima -- vascular cell adhesion molecule-1; Cardiovascular system -- Periodicals; Blood -- Circulation -- Periodicals; Blood Circulation; Cardiovascular System; Vascular Diseases; Sang -- Circulation -- Périodiques; Appareil cardiovasculaire -- Périodiques; 612.1
• DOI: 10.1161/CIRCRESAHA.117.306666
• Languages: English
• ISSNs: 0009-7330

Abstract

Abstract : Rationale: : Human and murine Vcam1 promoters contain 2 adjacent nuclear factor-κB (NF-κB)–binding elements. Both are essential for cytokine-induced transcription of transiently transfected promoter–reporter constructs. However, the relevance of these insights to regulation of the endogenous Vcam1 gene and to pathophysiological processes in vivo remained unknown. Objective: : Determine the role of the 5′ NF-κB–binding element in expression of the endogenous Vcam1 gene. Methods and Results: : Homologous recombination in embryonic stem cells was used to inactivate the 5′ NF-κB element in the Vcam1 promoter and alter 3 nucleotides in the 5′ untranslated region to allow direct comparison of wild-type versus mutant allele RNA expression and chromatin configuration in heterozygous mice. Systemic treatment with inflammatory cytokines or endotoxin (lipopolysaccharide) induced lower expression of the mutant allele relative to wild-type by endothelial cells in the aorta, heart, and lungs. The mutant allele also showed lower endothelial expression in 2-week atherosclerotic lesions in Vcam1 heterozygous/low-density lipoprotein receptor-deficient mice fed a cholesterol-rich diet. In vivo chromatin immunoprecipitation assays of heart showed diminished lipopolysaccharide-induced association of RNA polymerase 2 and NF-κB p65 with the mutant promoter. In contrast, expression of mutant and wild-type alleles was comparable in intimal cells of wire-injured carotid artery and 4- to 12-week atherosclerotic lesions. Conclusions: : This study highlights differences between in vivo and in vitro promoter analyses, and reveals a differential role for a NF-κB transcriptional response element in endothelial vascular cell adhesion molecule-1 expression induced by inflammatory cytokines or a cholesterol-rich diet versus intimal cell expression in atherosclerotic lesions and injured arteries. Abstract : Supplemental Digital Content is available in the text.