iii18 Ann Rheum Dis 2001;60:iii18–iii24 Ann Rheum Dis: first published as 10.1136/ard.60.90003.iii18 on 1 November 2001. Downloaded from Novel targets for interleukin 18 binding protein C A Dinarello Abstract clear that IL18 is a proinflammatory cytokine Background—Interleukin 18 (IL18) is re- and that its mechanism of action can be lated to the IL1 family by structure, independent of its ability to induce IFNã. receptors, signalling molecules, and func- IL18 is related to IL1â more than any other tion. IL18 induces gene expression and cytokine. The similarities between IL1â and synthesis of tumour necrosis factor IL18 exist at several levels. Firstly, the IL18 (TNF), IL1, Fas ligand, several chemo- precursor form (proIL18), like proIL1â, does kines, and vascular adhesion molecules. not contain a signal peptide, and proIL18 Similar to IL1â, IL18 is synthesised as a requires cleavage to an active cytokine by the biologically inactive precursor molecule IL1â converting enzyme (ICE, caspase-1). lacking a signal peptide. The IL18 precur- IL1â and IL18 are structurally related because sor requires cleavage into an active, both cytokines are primarily all â-pleated sheet 2 mature molecule by the intracellular folded molecules. This structural relationship cysteine protease, IL1â converting enzyme is significant because there are very few all-â (ICE, or caspase-1). Inhibitors of ICE sheet molecules among the diVerent cytokines. activity limit the biological activity of IL18 At the receptor level, the activity of IL18 is in animals and may be useful in reducing through a heterodimeric complex, the IL18 the activity of IL18 in human disease. receptor (IL18R) complex. The IL18R binding However, a constitutively secreted IL18 chain is termed IL18Rá. IL18Rá is a member of the IL1 receptor family,3 previously identi- binding protein (IL18BP) exists which 4 functions as a natural inhibitor of IL18 fied as the IL1R related protein (IL1Rrp). A activity. IL18BP binds IL18 with a high signalling chain (IL18Râ), also termed acces- sory protein-like (AcPL),5 aYnity (Kd of 400 pM) and, at equimolar is related to the IL1R accessory protein.6 Although similar to ratios, inhibits 50–70% of IL18; at twofold the IL1 receptor accessory protein in that the molar excess, IL18BP neutralises nearly IL18Râ does not itself bind its ligand in all IL18 activity. solution, the IL18Râ chain is part of the IL18 Method—IL18 was investigated for its role receptor complex. After binding of IL18 to the in human myocardial function. An IL18Rá, the IL18R AcPL binds to form a high ischaemia/reperfusion(I/R)modelofsupra- aYnity heterodimeric complex with the ligand. fused human atrial myocardium was used The high aYnity IL18R complex recruits the to assess myocardial contractile force. IL1 receptor activating kinase (IRAK), result- Results—The addition of IL18BP to the ing in phosphorylation of nuclear factor êB perfusate during and after I/R resulted in (NF B)-inducing kinase (NIK) with subse- ê http://ard.bmj.com/ improved post-I/R contractile function quent translocation of NFêB to the nucleus.7 from 35% of control to 76% with IL18BP. Initially identified as part of the IL1R signalling Also, IL18BP treatment preserved intra- events,8 IRAK is recruited to the IL1R complex cellular tissue creatine kinase levels (by after exposure to IL1. In cells possessing both 420%). Because active IL18 requires cleav- the IL18Rá and â chains, nuclear translocation age of its precursor form by ICE, inhibi- of NFêB is seen after incubation with IL18, tion of ICE attenuated the depression in and this property helps to explain the pleo- contractile force after I/R (from 35% of tropic nature of IL18. In IL18 deficient mice, on October 1, 2021 by guest. Protected copyright. control compared with 75.8% in treated production of IFNã and cytotoxic T cells is atrial muscle, p<0.01). markedly diminished despite ample amounts Conclusion—Myocardial ischaemia is a of IL12.9 A similar finding exists in mice target for IL18BP and use of IL18BP may deficient in ICE.10 11 The role of IL12 in IFNã thereby reduce ischaemia-induced myo- production therefore seems to require IL18. cardial dysfunction. (Ann Rheum Dis 2001;60:iii18–iii24) Constitutive production from Interleukin (IL) 18 was first described as an monocyte/macrophage interferon (IFN) ã-inducing factor1 which The production of IL18 in freshly obtained Department of circulated during endotoxaemia in mice pre- human peripheral blood mononuclear cells Medicine, Division conditioned with an infection of Propionibacte- (PBMC) and in splenic macrophages from Infectious Diseases, 12 B168, University rium acnes. Because of its ability to induce non-preconditioned mice has been studied. Colorado Health IFNã, IL18 is by default a member of the T cell Reverse transcriptase polymerase chain reac- Sciences Centre, 4200 helper type I (Th1)-inducing family of cyto- tion and western blot analysis were used to East Ninth Avenue, kines (IFNã, IL2, IL12, IL15). However, compare the production of IL18 with that of Denver, CO 80262, because antibodies to IL18 also reduced the IL1â in the same preparations. USA C A Dinarello hepatotoxicity of endotoxaemia, IL18 was con- There is no constitutive gene expression for sidered to possess other biological properties IL1â in freshly obtained human PBMC from Accepted 27 June 2001 beyond that of inducing IFNã. It has become healthy donors using over 40 cycles of PCR.13 14 www.annrheumdis.com Interleukin 18 binding protein iii19 Ann Rheum Dis: first published as 10.1136/ard.60.90003.iii18 on 1 November 2001. Downloaded from Surprisingly, the same PBMC expressed con- IL18 induced IL8 gene expression and synthe- stitutive mRNA for IL18. This was also seen sis.20 When IL1 receptor antagonist was used to using western blot analysis for proIL18 in block IL1 activity in these cells, IL18-induced lysates from the same PBMC which contained IL8 production was reduced by 40%. When no proIL1â in the same cells. Constitutive tumour necrosis factor (TNF) activity was IL18 gene expression and the presence of inhibited, IL18-induced IL8 production was proIL18 protein were also observed in freshly reduced by 80%, suggesting that the primary obtained murine splenocytes.12 The promoter action of IL18 was through a TNF dependent regions for IL1â and IL18 gene expression pathway. The source of TNF was the CD3+ have been studied and may provide an insight and the natural killer cells. The induction of into these observations. The promoter for IL18 TNF can be seen at the level of gene expression is TATA-less and IL18 promoter activity within two hours after exposure to IL18. IL18 upstream of exon 2 acts constitutively.15 There- also stimulates the synthesis of macrophage fore, it is not unexpected that IL18 mRNA is chemoattractant protein 1 in these cultures.20 constitutively expressed even in whole blood Using neutralising anti-IL18 antibodies, freshly obtained from healthy donors where no Netea et al compared the myeloperoxidase artefact is introduced. The additional finding activity in lungs and livers of mice after that the 3' untranslated region of human IL18 endotoxaemia.23 ICE deficient mice were com- lacks the AUUUA destabilisation sequence15 is pletely resistant to lethal endotoxaemia in- also consistent with these observations. This duced by lipopolysaccharide (LPS) derived would allow for more sustained levels of the from either Escherichia coli or Salmonella polyadenylated species and translation into typhimurium. IFNã deficient mice were not protein. Daily injections of IL12 into mice resistant to S typhimurium LPS, suggesting an result in high circulating levels of IFNã, which IFNã-independent role for IL18. Anti-IL18 is completely prevented by prior treatment protected mice against a lethal injection of with anti-IL18 antibodies, or absent in mice either LPS and also reduced neutrophil deficient in ICE.11 In this model, IL12 may accumulation in liver and lungs. The increased activate ICE. Osteoclasts also produce IL18,16 survival was accompanied by decreased levels and regulation of bone density may be a prop- of chemokine, macrophage inflammatory pro- erty of IL18, as it is for IL1â. tein 2.23 In addition to monocyte/macrophages, the epidermal cells from mouse skin produce IL18 IL18 up regulates expression of Fas ligand constitutively.17 The keratinocyte is the major IL18 enhances Fas ligand expression and source of the IL18. Moreover, contact sensitis- induces apoptosis in Fas-expressing human ers increased IL18 synthesis, whereas skin irri- myelomonocytic KG-1 cells.22 KG-1 cells are tants did not. A murine keratinocyte cell line monocyte-like and also produce the intercellu- (PAM 212) also produced IL18 constitu- lar adhesion molecule-1 (ICAM-1). IL18 tively.17 IL18 has also been found constitutively increases ICAM-1 expression in these macro- expressed in colonic specimens and isolated phagic cells.24 These cells also express constitu- mucosal cell populations from patients with tively Fas antigen (CD95) and, after exposure Crohn’s disease.18 Similar to IL1, IL18 is found to IL18, KG-1 cells became apoptotic. Fas lig- in the rat adrenal cortex and pituitary gland.19 and is up regulated in T cells and NK cells after http://ard.bmj.com/ The adrenal IL18 was increased by cold stress, incubation with IL18.25 In the liver, cytotoxic T whereas the pituitary IL18 was not. Two tran- cells are thought to contribute to the destruc- scripts of adrenal IL18 were sequenced: an tion of hepatic cells, particularly when infected abundant form was clearly the rat homologue by viruses. Cytotoxic T cells and NK cells iso- of mouse IL18 and shared 91% amino acid lated from the liver and then stimulated with homology. The other transcript was less abun- IL18 exhibit enhanced killing of Jurkat T 26 dant. It had a 59 base pair deletion and the cells.