title: IL-32-dependent effects of IL-1beta on endothelial cell functions.
abstract: Increasing evidence demonstrates that interleukin (IL)-32 is a pro-inflammatory cytokine, inducing IL-1alpha, IL-1beta, IL-6, tumor necrosis factor (TNF)-alpha, and chemokines via nuclear factor (NF)-kappaB, p38 mitogen-activated protein kinase (MAPK), and activating protein (AP)-1 activation. Here we report that IL-32 is expressed and is also functional in human vascular endothelial cells (EC) of various origins. Compared with primary blood monocytes, high levels of IL-32 are constitutively produced in human umbilical vein EC (HUVEC), aortic macrovascular EC, and cardiac as well as pulmonary microvascular EC. At concentrations as low as 0.1 ng/ml, IL-1beta stimulated IL-32 up to 15-fold over constitutive levels, whereas 10 ng/ml of TNFalpha or 100 ng/ml of lipopolysaccharide (LPS) were required to induce similar quantities of IL-32. IL-1beta-induced IL-32 was reduced by inhibition of the IkappaB kinase-beta/NF-kappaB and ERK pathways. In addition to IL-1beta, pro-coagulant concentrations of thrombin or fresh platelets increased IL-32 protein up to 6-fold. IL-1beta and thrombin induced an isoform-switch in steady-state mRNA levels from IL-32alpha/gamma to beta/epsilon. Adult EC responded in a similar fashion. To prove functionality, we silenced endogenous IL-32 with siRNA, decreasing intracellular IL-32 protein levels by 86%. The knockdown of IL-32 resulted in reduction of constitutive as well as IL-1beta-induced intercellular adhesion molecule-1 (ICAM-1) (of 55% and 54%, respectively), IL-1alpha (of 62% and 43%), IL-6 (of 53% and 43%), and IL-8 (of 46% and 42%). In contrast, the anti-inflammatory/anti-coagulant CD141/thrombomodulin increased markedly when IL-32 was silenced. This study introduces IL-32 as a critical regulator of endothelial function, expanding the properties of this cytokine relevant to coagulation, endothelial inflammation, and atherosclerosis.
authors.affiliation: Department of Medicine, University of Colorado Denver, Aurora, CO 80045, USA.
authors.forename: Claudia A
authors.initials: CA
authors.lastname: Nold-Petry
authors.forename: Marcel F
authors.initials: MF
authors.lastname: Nold
authors.forename: Jarod A
authors.initials: JA
authors.lastname: Zepp
authors.forename: Soo-Hyun
authors.initials: SH
authors.lastname: Kim
authors.forename: Norbert F
authors.initials: NF
authors.lastname: Voelkel
authors.forename: Charles A
authors.initials: CA
authors.lastname: Dinarello
mesh_terms: D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism; D042783:Endothelial Cells / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation / Q000187:drug effects; D020868:Gene Silencing / Q000187:drug effects; D006801:Humans; D051550:I-kappa B Kinase / Q000378:metabolism; D018799:Intercellular Adhesion Molecule-1 / Q000378:metabolism; D053583:Interleukin-1beta / Q000378:metabolism* / Q000494:pharmacology; D007378:Interleukins / Q000235:genetics / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000494:pharmacology; D016328:NF-kappa B / Q000378:metabolism; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D013917:Thrombin / Q000494:pharmacology; D018180:Thrombomodulin / Q000378:metabolism; D014471:Umbilical Veins / Q000166:cytology
chemical_list: C496971:IL32 protein, human; D053583:Interleukin-1beta; D007378:Interleukins; D008070:Lipopolysaccharides; D016328:NF-kappa B; D020033:Protein Isoforms; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D018180:Thrombomodulin; D018799:Intercellular Adhesion Molecule-1; D051550:I-kappa B Kinase; D013917:Thrombin