Interleukin-1 receptor antagonist expression in human endothelial cells and atherosclerosis.
The proinflammatory cytokine interleukin (IL)-1 is expressed mainly within the endothelium of atherosclerotic plaques and may be linked with inflammatory mechanisms of atherogenesis. IL-1 action is complex and regulated in part by its naturally occurring inhibitor, the IL-1 receptor antagonist (IL-1ra). Therefore, we studied differential and specific isoform expression of IL-1ra in the endothelium of diseased coronary arteries and in endothelial cells (ECs) stimulated under defined conditions. In view of an association with IL-1ra gene (IL-1RN) polymorphism, the influence of endothelial cell genotype at IL-1RN on IL-1ra protein production was also examined. Secreted IL-1ra and intracellular IL-1ra mRNAs were detected by semiquantitative reverse transcription-polymerase chain reaction in human atherosclerotic and dilated cardiomyopathic coronary arteries; protein expression appeared increased in atherosclerotic compared with dilated cardiomyopathic arteries, where IL-1ra appeared to be confined to the endothelium. Only intracellular IL-1ra type I mRNA was detected in human umbilical vein ECs (HUVECs) and human coronary artery ECs (HCAECs) when they were stimulated with bacterial lipopolysaccharide/phorbol myristate acetate and transforming growth factor-beta. IL-1beta and IL-1alpha were without effect. IL-1ra protein was detected in HUVECs (intracellular IL-1ra), HCAECs (intracellular IL-1ra), and human coronary artery smooth muscle cells (intracellular IL-1ra) by immunoprecipitation and Western blot. IL-1ra was detected in HUVEC cell lysates by ELISA and appeared to be influenced by the genotype of the IL-1RN variable number tandem repeat, an 86-bp repeat polymorphism in intron 2 of the IL-1ra gene, with lower levels of IL-1ra produced by IL-1RN allele 2-containing cells (ratio of IL-1ra to total protein: for 1,1 homozygotes, 1.38+/-0.28x10(-9) [n=15]; for 1,2 heterozygotes, 0.81+/-0.17x10(-9) [n=8]; and for 2,2 homozygotes, 0.63+/-0.19x10(-9) [n=5]; P<0.05 compared with 1,1 homozygotes). This is the first demonstration of IL-1ra in human diseased arteries, stimulated HUVECs, and HCAECs and indicates the endothelial cell as an important source. Endothelial IL-1ra production may be controlled by the endothelial IL-1RN genotype. These data further support the role of the IL-1 system of cytokines in the pathogenesis of atherosclerosis.
Dewberry
R
R
Cardiovascular Medicine, Division of Clinical Sciences, Northern General Hospital, University of Sheffield, Sheffield, UK.
Holden
H
H
Crossman
D
D
Francis
S
S
D001161:Arteriosclerosis / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D002478:Cells, Cultured; D003324:Coronary Artery Disease / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D003331:Coronary Vessels / Q000737:chemistry / Q000378:metabolism; D004730:Endothelium, Vascular / Q000737:chemistry / Q000378:metabolism* / Q000473:pathology*; D005838:Genotype; D006801:Humans; D053590:Interleukin 1 Receptor Antagonist Protein; D009131:Muscle, Smooth, Vascular; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D017472:Receptors, Interleukin-1 / Q000032:analysis / Q000037:antagonists & inhibitors* / Q000096:biosynthesis* / Q000235:genetics; D012795:Sialoglycoproteins / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D014471:Umbilical Veins
C507763:IL1RN protein, human; D053590:Interleukin 1 Receptor Antagonist Protein; D012333:RNA, Messenger; D017472:Receptors, Interleukin-1; D012795:Sialoglycoproteins