Interleukin (<em>IL</em>)-1 is an important neuroimmunomodulator and a key mediator of inflammation during brain disorders. It acts on neuronal and glial cells via binding to the <em>IL</em>-1 type 1 receptor and <em>IL</em>-1 receptor accessory protein (<em>IL</em>-<em>1RAcP</em>). More recently, a neuronal-specific isoform of <em>IL</em>-<em>1RAcP</em>, named <em>IL</em>-<em>1RAcP</em>b, has been identified. Our aim was to determine the role of <em>IL</em>-<em>1RAcP</em>b in <em>IL</em>-1 actions in neuronal and glial cells, and to further explore the signaling mechanisms of <em>IL</em>-1 in neurons. We found that <em>IL</em>-<em>1RAcP</em>b deletion had no effect on <em>IL</em>-1α- and <em>IL</em>-1β-induced activation of the extracellular signal-regulated kinase 1/2 or <em>IL</em>-6 release in glial cultures, although <em>IL</em>-6 release in response to high <em>IL</em>-1α concentration (30 IU/ml) was significantly reduced. We identified the p38 kinase as a key signaling element in <em>IL</em>-1α- and <em>IL</em>-1β-induced <em>IL</em>-6 synthesis and release in neuronal cultures. <em>IL</em>-<em>1RAcP</em>b deletion had no effect on <em>IL</em>-1α- and <em>IL</em>-1β-induced <em>IL</em>-6 release in neurons, but significantly reduced <em>IL</em>-1α- but not <em>IL</em>-1β-induced p38 phosphorylation. Our data demonstrate that the p38 signaling pathway plays an important role in <em>IL</em>-1 actions in neurons, and that <em>IL</em>-<em>1RAcP</em> may regulate some, but not all, neuronal activities in response to <em>IL</em>-1α.

Interleukin-33 (<em>IL</em>-33) is one of the members of the <em>IL</em>-1 family of cytokines and a ligand of ST2 and <em>IL</em>-1 receptor accessory protein (<em>IL</em>-<em>1RAcP</em>) that is known to affect Th2 inflammatory response with partial effects on Th1 responses. This cytokine is released by epithelial and smooth muscle cells of the airway system during their injury by several environmental stimuli, such as allergens, viruses, helminths, and pollutants. <em>IL</em>-33 is an alarmin that acts as an endogenous danger signal, and it has been known to affect various types of cells, such as mast cells, basophils, eosinophils, T cells, and specific subsets of innate lymphoid cells (<em>IL</em>Cs). In recent findings, this cytokine is believed to have a critical role in several types of cancers, such as lung cancer, liver cancer, and head and neck squamous cell cancer. The expression of <em>IL</em>-33/ST2 in cancer tissues shows a close association with tumor growth and tumor progression in several types of cancer, suggesting the <em>IL</em>-33/ST2 pathway as a potential target for therapy.

The <em>IL</em>-1R accessory protein (<em>IL</em>-<em>1RAcP</em>) plays a role in <em>IL</em>-1R signaling by forming a complex with <em>IL</em>-1RI bound to the <em>IL</em>-1 ligand. We identified four hydrophilic peptide regions of the extracellular <em>IL</em>-<em>1RAcP</em> that may be available for complex formation (peptide 1, 71-83 domain I; peptide 2, 204-211 domain II; peptide 3, 282-292 domain III; and peptide 4, 304-314 domain III). These peptides were synthesized, coupled to keyhole limpet hemocyanin, and used to produce rabbit antisera. Each affinity-purified antiserum showed specificity for the respective peptide without cross-reactivity. Anti-peptide 2, 3, and 4 recognized surface expression of <em>IL</em>-<em>1RAcP</em> on the Th2 D10S cells by FACS and inhibited <em>IL</em>-1-driven proliferation. Anti-peptide 4 recognized intact <em>IL</em>-<em>1RAcP</em> and soluble <em>IL</em>-<em>1RAcP</em>. Anti-<em>IL</em>-<em>1RAcP</em>-peptide 4, which targets the terminal segment of domain III, inhibited 80% of <em>IL</em>-1 beta-driven proliferation of D10S cells. However, these <em>IL</em>-<em>1RAcP</em> Abs had no effect on the activity of human or mouse <em>IL</em>-1 alpha. Whereas <em>IL</em>-1 beta down-regulated <em>IL</em>-1RI surface expression (p < 0.05), there was no change in the surface expression of <em>IL</em>-<em>1RAcP</em>. Moreover, murine <em>IL</em>-10 increased surface expression of <em>IL</em>-1RI, but did not affect expression of <em>IL</em>-<em>1RAcP</em> or the proliferation of D10S cells. Steady state levels of mRNA for <em>IL</em>-<em>1RAcP</em> and <em>IL</em>-1RI in D10S cells showed a similar pattern to that of surface expression of the respective receptors. We conclude that 1) blocking <em>IL</em>-<em>1RAcP</em> inhibits <em>IL</em>-1 signaling in D10S cells, 2) domains-II and III may be involved in complex formation with <em>IL</em>-1RI, 3) <em>IL</em>-<em>1RAcP</em> is not regulated as is <em>IL</em>-1RI in the same cells, and 4) <em>IL</em>-1 responsiveness is dependent on the expression of <em>IL</em>-1RI, not <em>IL</em>-<em>1RAcP</em>.

A panel of monoclonal antibodies to human <em>IL</em>-1 beta has been used to probe its conformational and functional characteristics. Real time antibody-protein interaction was assessed by surface plasmon resonance with a BIAcore apparatus, in order to determine the kinetic and thermodynamic parameters of the interaction and to map the recognition sites of the antibodies on the <em>IL</em>-1 beta surface. Topological analysis was thus compared to the inhibitory capacity of antibodies for <em>IL</em>-1 beta bioactivity and binding to the activating receptor <em>IL</em>-1RI. This functional mapping analysis allows the following hypothesis. At least two discrete areas of <em>IL</em>-1 beta, located within the sequences 133-147 and 177-186 (as defined by mAbs MhC1 and BRhD2, respectively), are apparently involved in <em>IL</em>-1RI-independent agonist activity, and thus possibly take part in the interaction with the receptor accessory protein <em>IL</em>-<em>1RAcP</em>. Another area in the 133-147 sequence (defined by mAb BRhC3) is involved in agonist binding to its receptor CDw121a (<em>IL</em>-1RI), whereas a site recognized by mAb BRhG5 within the sequence 218-243 is selectively responsible for non-agonist binding to the activating receptor. The loop between the 4th and the 5th beta-strand, at the open end of the <em>IL</em>-1 beta-barrel structure, may possibly take part in both non-agonist binding to <em>IL</em>-1RI and in the interaction with <em>IL</em>-<em>1RAcP</em>.

<em>IL</em>-33 (<em>IL</em>-1F11) is a member of <em>IL</em>-1 family ligand, which stimulates the production of inflammatory cytokines. <em>IL</em>-33 receptor complex is comprised of <em>IL</em>-1 receptor accessory protein (<em>IL</em>-<em>1RAcP</em>) and ST2 that are activated by <em>IL</em>-33 ligand binding. ST2 is a ligand-binding chain of the <em>IL</em>-33 receptor component, and the soluble ST2 form possesses antagonistic activity. Here, we expressed the extracellular domain of ST2-fused to the immunoglobulin of IgG1 constant region in order to generate a soluble recombinant Fc-ST2. Human and mouse recombinant Fc-ST2 protein were expressed in Chinese hamster ovary cells and purified using a mini-protein A affinity chromatography. The recombinant Fc-ST2 protein was used to examine inhibitory function in <em>IL</em>-33-induced cytokine production in different cell types. The human Fc-ST2 abolished <em>IL</em>-33-induced <em>IL</em>-8 production in human mast cells, but mouse Fc-ST2 failed to inhibit <em>IL</em>-33-induced TNFα production in mouse Raw 264.7 macrophage cells. We further investigated the expression of <em>IL</em>-33 receptor component with various cell lines. <em>IL</em>-33 receptors expression pattern and Fc-ST2 inhibitory activity in different cell types suggest that <em>IL</em>-<em>1RAcP</em> and ST2 are necessary but insufficient for <em>IL</em>-33 activity. Our results suggest that an additional receptor component may participate in the biological activity of <em>IL</em>-33.

Interleukin-1 beta is a key molecule in brain-immune interactions that, apart from its immune effects, stimulates the hypothalamo-pituitary-adrenal (HPA) axis and induces behavioral alterations. However, its physiological role during stress responses remain to be elucidated. The possible mechanisms involved in <em>IL</em>-1-mediated stimulation of the HPA axis during stress were assessed by using different approaches. They were first studied in mice deficient for the <em>IL</em>-1 beta-converting enzyme (ICE) gene. Mature <em>IL</em>-1 beta derives from a precursor, the pro-<em>IL</em>-1 beta, devoid of any conventional signal sequence that is mainly processed by ICE. After immune or stress stimulation, ICE-deficient mice were shown to have a hyperactive HPA axis and to able to produce immunoreactive <em>IL</em>-1 beta. This indicates that the greater reactivity of the HPA axis could result from a higher sensitivity to non-ICE-matured <em>IL</em>-1 beta, as suggested by a higher basal transcription of hypothalamic <em>IL</em>-1 receptor type I (<em>IL</em>-1 RI) in ICE-deficient mice. The biological effects of <em>IL</em>-1 beta are mediated by <em>IL</em>-1 RI associated with the <em>IL</em>-1 receptor accessory protein (<em>IL</em>-<em>1RAcP</em>). <em>IL</em>-<em>1RAcP</em> is an essential component for <em>IL</em>-1 action at the periphery, but its role in the brain is not well known. Therefore, the effects of i.c.v. <em>IL</em>-1 beta were studied in <em>IL</em>-<em>1RAcP</em>-deficient mice. In normal mice, i.c.v. <em>IL</em>-1 beta depresses peripheral immune responses, induces the production of plasma <em>IL</em>-6, and stimulates the HPA axis. None of these effects were observed in <em>IL</em>-<em>1RAcP</em>-deficient mice, indicating that <em>IL</em>-<em>1RAcP</em> is necessary for the induction of the main neuroendocrine and immune effects of central <em>IL</em>-1 beta. In normal mice, the role of <em>IL</em>-1 beta was assessed by pretreating the animal with the <em>IL</em>-1 receptor antagonist (<em>IL</em>-1Ra). <em>IL</em>-1Ra did modify the activation of the HPA axis observed during stress, except when the animals were previously sympathectomized. This suggests that the sympathetic nervous system can downregulate the <em>IL</em>-1 beta-induced stimulation of the HPA axis. Finally, the modulation of the production and physiological activities of <em>IL</em>-1 were studied in normal mice, taking advantage of interindividual differences in brain-immune interactions linked to cerebral lateralization. Behavioral/brain lateralization was shown to be related to behavioral response to peripheral administration of <em>IL</em>-1, and to the production of <em>IL</em>-1 and <em>IL</em>-6 in response to LPS. This suggests that cytokines, and especially <em>IL</em>-1 beta, may represent one of the factors responsible for interindividual differences in brain-immune interactions.

We investigated the role of SHPS-1/SIRPalpha1 in <em>IL</em>-1beta- and TNFalpha-dependent signaling that leads to the activation of Erk 1/2 and Akt. Treatment of Balb3T3 cells with <em>IL</em>-1beta or TNFalpha activated tyrosine phosphorylation of SHPS-1, its association with SHP-2 and the phosphorylation of Erk 1/2 and Akt. PP1, a specific inhibitor for the Src family protein tyrosine kinases, strongly inhibited tyrosine phosphorylation of SHPS-1 and complex formation of SHPS-1 with SHP-2 by <em>IL</em>-1beta. In addition, PP1 substantially inhibited the <em>IL</em>-2beta- and TNFalpha-dependent activation of Erk 1/2 and Akt. Exogenous expression of either SHPS-1 mutants that lack SHP-2 binding function or a dominant negative mutant of SHP-2 markedly inhibited the activation of Erk 1/2 and Akt by <em>IL</em>-1beta, whereas wild type SHPS-1 did not. Moreover, <em>IL</em>-1beta-stimulation induced association of SHPS-1 with <em>IL</em>-<em>1RAcP</em>, a second subunit of <em>IL</em>-1 receptor, whereas expression of SHPS-1 mutant that lack SHP-2 binding function clearly blocked the association and tyrosine phosphorylation of endogenous SHPS-1. Taken together, our results strongly suggest that activation of Erk 1/2 and Akt by proinflammatory cytokines requires tyrosine phosphorylation of SHPS-1 and subsequent association of SHPS-1 with SHP-2.

The <em>IL</em>-33/ST2 signaling pathway plays an important role in coronary artery disease (CHD); however, few studies have explored how variants in <em>IL</em>-33/ST2 genes influence CHD risk. Here, we examined the association between genetic variants in <em>IL</em>-33, ST2, and <em>IL</em>-<em>1RAcP</em> of the <em>IL</em>-33/ST2 axis and the risk of CHD. We conducted a case-controlled study with 1146 CHD cases and 1146 age- and sex-frequency-matched controls. Twenty-eight single nucleotide polymorphisms (SNPs) in <em>IL</em>-33, ST2, and <em>IL</em>-<em>1RAcP</em> were genotyped by Sequenom MassArray and TaqMan assay. Logistic regression was used to analyze these associations. The SNP rs4624606 in <em>IL</em>-<em>1RAcP</em> was nominally associated with CHD risk. The AA genotype was associated with a 1.85-fold increased risk of CHD (95% confidence interval (CI) = 1.01-3.36; p = 0.045) compared to the TT genotype. Further analysis showed that AA carriers also had a higher risk of CHD than TT + TA carriers (odds ratio (OR) = 1.85; 95% CI = 1.85-3.35; p = 0.043). However, no significant association was observed between variants in <em>IL</em>-33/ST2 genes and CHD risk. Further studies are needed to replicate our results in other ethnic groups with larger sample size.

Cytokine-amplified functional CD8⁺ T cells ensure effective eradication of tumors. Interleukin 36α (<em>IL</em>-36α), <em>IL</em>-36β, and <em>IL</em>-36γ share the same receptor complex, composed of the <em>IL</em>-36 receptor (<em>IL</em>-36R), and <em>IL</em>-<em>1RAcP</em>. Recently, we revealed that <em>IL</em>-36γ greatly promoted CD8⁺ T cell activation, contributing to antitumor immune responses. However, the underlying mechanism of <em>IL</em>-36-mediated CD8⁺ T cell activation remains understood. In the current study, we proved that <em>IL</em>-36β had the same effect on CD8⁺ T cell as <em>IL</em>-36γ, and uncovered that <em>IL</em>-36β significantly activated mammalian target of rapamycin complex 1 (mTORC1) of CD8⁺ T cells. When mTORC1 was inhibited by rapamycin, <em>IL</em>-36β-stimulated CD8⁺ T cell activation and expansion was drastically downregulated. Further, we elucidated that <em>IL</em>-36β-mediated mTORC1 activation was dependent on the pathway of phosphatidylinositol 3 kinase (PI3K)/Akt, IκB kinase (IKK) and myeloid differentiation factor 88 (MyD88). Inhibition of PI3K or IKK by inhibitor, or deficiency of MyD88, respectively, suppressed mTORC1 signal, causing arrest of CD8⁺ T cell activation. Additionally, it was validated that <em>IL</em>-36β significantly promoted mTORC1 activation and antitumor function of CD8⁺ tumor-infiltrating lymphocytes (T<em>IL</em>s) <i>in vivo</i>, resulting in inhibition of tumor growth and prolongation of survival of tumor-bearing mice. Taken together, we substantiated that <em>IL</em>-36β could promote CD8⁺ T cell activation through activating mTORC1 dependent on PI3K/Akt, IKK and MyD88 pathways, leading to enhancement of antitumor immune responses, which laid the foundations for applying <em>IL</em>-36β into tumor immunotherapy.

Aluminum (Al) is an inorganic pollutant that induces nerve cells apoptosis and necroptosis, thereby causing depression and neurodegenerative diseases. <em>IL</em>-1β/JNK signaling pathway can regulate apoptosis and necroptosis. However, it remains unclear whether <em>IL</em>-1β/JNK signaling pathway is involving in the regulation of Al-induced hippocampal neural cells apoptosis and necroptosis. To investigate the mechanism of Al on neural cells apoptosis and necroptosis, rats were orally exposed to different doses of AlCl₃ for 90 days. The open-field test results showed that AlCl₃ caused depressive behavior in rats. Histopathological evidence showed that AlCl₃ induced hippocampal neural cells apoptosis and necrosis. Moreover, Bax/Bcl-2 mRNA expression ratio, caspase-3 activity and mRNA expression and TUNEL positive rates were upregulated, meanwhile, TNF-α mRNA and protein expression levels, TNFR1, RIP1, RIP3 and MLKL proteins levels were increased, while caspase-8 protein level was decreased in the hippocampus of Al-exposed groups. These results proved that AlCl₃ induced hippocampal neural cells apoptosis and necroptosis. Combined with histopathology and correlation analysis, we deduced that hippocampal neural cells were more likely to undergo necroptosis at high doses (450 mg/kg) of AlCl₃, while <150 mg/kg AlCl₃ tended to induce apoptosis. Finally, AlCl₃ increased the proteins level of <em>IL</em>-1β, <em>IL</em>-1RI, <em>IL</em>-<em>1RAcP</em>, JNK and p-JNK, indicating that AlCl₃ activated <em>IL</em>-1β/JNK signaling pathway. However, the application of <em>IL</em>-1 receptor antagonist (<em>IL</em>-1Ra) inhibited the phosphorylation of JNK and the related genes expression of apoptosis and necroptosis caused by AlCl₃. Thus, we concluded that AlCl₃ induced hippocampal neural cells death and depression-like behavior in rats by activating <em>IL</em>-1β/JNK signaling pathway.

BACKGROUND

The inflammatory response following myocardial infarction (MI) is prerequisite for proper healing of infarcted tissue, but can also have detrimental effects on cardiac function. Interleukin (IL)-1α and IL-1β are potent inflammatory mediators and their bioactivity is tightly regulated by IL-1 receptor antagonist (IL-1ra) and soluble (s) IL-1 receptors (R). We aimed to examine whether levels of soluble regulators of IL-1 signalling are changed during ST-elevation MI (STEMI) and their associations with parameters of cardiac injury and ventricular remodelling.

METHODS

Plasma levels of IL-1Ra, sIL-1R1, sIL-1R2 and sIL-1R accessory protein (sIL-1RAcP) were measured by immunoassays in repeated samples from patients with STEMI (n = 255) and compared to healthy controls (n = 65).

RESULTS

IL-1Ra, sIL-1R1 and sIL-1R2 levels were all significantly elevated after STEMI, while levels of sIL-1RAcP were lower compared to controls. sIL-1R2 levels (at different time points) correlated positively with C-reactive protein, myocardial infarct size and change in indexed left ventricular end-diastolic and end-systolic volume (LVEDVi and LVESVi) measured by cardiac MR acutely and after 4 months, and negatively with LV ejection fraction. Patients with>>median levels of sIL-1R2 in the acute phase were more likely to have increased change in LVEDVi and LVESVi. Importantly, sIL-1R2 remained significantly associated with change in LVEDVi and LVESVi also after adjustment for clinical covariates.

CONCLUSIONS

Levels of sIL-1R2 are independently associated with parameters of LV adverse remodelling following STEMI.

Interleukin-33 (<em>IL</em>-33) is a novel identified chromatin-associated cytokine of <em>IL</em>-1 family cytokines. It signals through a heterodimer comprised of ST2L and <em>IL</em>-<em>1RAcp</em>, and plays a crucial role in many diseases. However, very little is known about the role and underlying intricate mechanisms of <em>IL</em>-33 in recurrent neonatal seizure (RNS). To determine whether <em>IL</em>-33 plays an important regulatory role, we established a neonatal seizure model in this study. Rats were subjected to recurrent seizures induced by inhaling volatile flurothyl. Recombinant <em>IL</em>-33 or PBS were also administered by intraperitoneally (IP) before surgery, respectively. Here, our current results indicated that RNS contributed to a significant reduction in <em>IL</em>-33 and its specific receptor (ST2L) expressions in cortex. While, in hippocampus, RNS induced an increase in <em>IL</em>-33 and ST2L evidently, compared with Sham group. After injection with <em>IL</em>-33, however, a remarkable increase in total <em>IL</em>-33 was detected both in brain cortex and hippocampus. In addition, <em>IL</em>-33 was mainly co-localized in the nuclear of GFAP+ astrocytes and the cytoplasm of the Iba-1+ microglia and <em>IL</em>-33+/NeuN+ merged cells. In parallel, ST2L was expressed mainly in the membrane of GFAP+ astrocytes, Iba-1+ microglia and NeuN+ neurons, respectively. Furthermore, administration of <em>IL</em>-33 improved RNS-induced behavioral deficits, promoted bodyweight gain, and ameliorated spatial learning and memory ability. Moreover, <em>IL</em>-33 pretreatment blocked the activation of NF-κB, resisted inflammatory cytokines <em>IL</em>-1β and TNF-α increase, as well as suppressed apoptosis and autophagy activation after RNS. Collectively, <em>IL</em>-33 provides potential neuroprotection through suppressing apoptosis, autophagy and at least in part by NF-κB-mediated inflammatory pathways after RNS.

Since recent work has identified an apoptotic pathway in sympathetic neurons that is mediated by autocrine interleukin-1 (<em>IL</em>-1), we investigated whether cultured sympathetic neurons possess functional <em>IL</em>-1 receptors. Cultured sympathetic neurons express levels of <em>IL</em>-1RI and <em>IL</em>-<em>1RAcP</em> mRNAs consistent with signal transduction. Neurons stimulated with <em>IL</em>-1 demonstrate enhanced p65 NF-kappaB nuclear translocation and enhanced NF-kappaB DNA binding activity, with at least p65 and p50 subunits participating in the DNA binding activity. RNA differential display identified several neuronal mRNAs regulated by <em>IL</em>-1, including a member of the reticulon family. We conclude that <em>IL</em>-1 stimulates a potential component of a neuronal secretory pathway.

<em>IL</em>-1 family members regulate innate immune responses, are produced by gestation-associated tissues, and have a role in healthy and adverse pregnancy outcomes. To better understand their role at the materno-fetal interface we used a human tissue explant model to map lipopolysaccharide (LPS)-stimulated production of <em>IL</em>-1α, <em>IL</em>-1β, <em>IL</em>-18, <em>IL</em>-33, <em>IL</em>-1Ra, <em>IL</em>-18BPa, ST2 and <em>IL</em>-<em>1RAcP</em> by placenta, choriodecidua and amnion. Caspase-dependent processing of <em>IL</em>-1α, <em>IL</em>-1β, <em>IL</em>-18, and <em>IL</em>-33 and the ability of <em>IL</em>-1α, <em>IL</em>-1β, <em>IL</em>-18, and <em>IL</em>-33 to regulate the production of <em>IL</em>-1RA, <em>IL</em>-18BPa, ST2 and <em>IL</em>-<em>1RAcP</em> was also determined. LPS acted as a potent inducer of <em>IL</em>-1 family member expression especially in the placenta and choriodecidua with the response by the amnion restricted to <em>IL</em>-1β. Caspases-1, 4 and 8 contributed to LPS-stimulated production of <em>IL</em>-1β and <em>IL</em>-18, whereas calpain was required for <em>IL</em>-1α production. Exogenous administration of <em>IL</em>-1α, <em>IL</em>-1β, <em>IL</em>-18, and <em>IL</em>-33 lead to differential expression of <em>IL</em>-1Ra, <em>IL</em>-18BPa, ST2 and <em>IL</em>-<em>1RAcP</em> across all tissues examined. Most notable were the counter-regulatory effect of LPS on <em>IL</em>-1β and <em>IL</em>-1Ra in the amnion and the broad responsiveness of the amnion to <em>IL</em>-1 family cytokines for increased production of immunomodulatory peptides and soluble receptors. The placenta and membranes vary not only in their output of various <em>IL</em>-1 family members but also in their counter-regulatory mechanisms through endogenous inhibitory peptides, processing enzymes and soluble decoy receptors. This interactive network of inflammatory mediators likely contributes to innate defence mechanisms at the materno-fetal interface to limit, in particular, the detrimental effects of microbial invasion.

Interleukin-1 (<em>IL</em>-1) is an important pro-inflammatory cytokine that exerts its diverse biological functions by binding to a receptor complex consisting of two transmembrane proteins, type I <em>IL</em>-1 receptor (<em>IL</em>-1RI) and <em>IL</em>-1 receptor accessory protein (<em>IL</em>-<em>1RAcP</em>). Both receptor chains are indispensable for <em>IL</em>-1 signaling, but only <em>IL</em>-1RI is able to bind the cytokine. The effects of <em>IL</em>-<em>1RAcP</em> on <em>IL</em>-1 binding are unclear. This study shows that although expression of <em>IL</em>-<em>1RAcP</em> does not alter the equilibrium dissociation constant of <em>IL</em>-1, it has an effect on the non-equilibrium binding modus, most likely due to changes in on/off rates. This defines an additional function of <em>IL</em>-<em>1RAcP</em>: it stabilizes the active <em>IL</em>-1 receptor complex.

The ligation of interleukin-1 receptor (<em>IL</em>-1R) or tumor necrosis factor receptor 1 (TNFR1) induces the recruitment of adaptor proteins and their concomitant ubiquitination to the proximal receptor signaling complex, respectively. Such are upstream signaling events of IKK that play essential roles in NF-κB activation. Thus, the discovery of a substance that would modulate the recruitment of key proximal signaling elements at the upstream level of IKK has been impending in this field of study. Here, we propose that brazilin, an active compound of Caesalpinia sappan L. (Leguminosae), is a potent NF-κB inhibitor that selectively disrupts the formation of the upstream <em>IL</em>-1R signaling complex. Analysis of upstream signaling events revealed that brazilin markedly abolished the <em>IL</em>-1β-induced polyubiquitination of IRAK1 and its interaction with IKK-γ counterpart. Notably, pretreatment of brazilin drastically interfered the recruitment of the receptor-proximal signaling components including IRAK1/4 and TRAF6 onto MyD88 in <em>IL</em>-1R-triggerd NF-κB activation. Interestingly, brazilin did not affect the TNF-induced RIP1 ubiquitination and the recruitment of RIP1 and TRAF2 to TNFR1, suggesting that brazilin is effective in selectively suppressing the proximal signaling complex formation of <em>IL</em>-1R, but not that of TNFR1. Moreover, our findings suggest that such a disruption of <em>IL</em>-1R-proximal complex formation by brazilin is not mediated by affecting the heterodimerization of <em>IL</em>-1R and <em>IL</em>-<em>1RAcP</em>. Taken together, the results suggest that the anti-IKK activity of brazilin is induced by targeting IKK upstream signaling components and subsequently disrupting proximal <em>IL</em>-1 receptor signaling complex formation.

Brain astrocytes play a pivotal role in the brain response to inflammation. They express <em>IL</em>-1 receptors including the type I <em>IL</em>-1 receptor (<em>IL</em>-1RI) that transduces <em>IL</em>-1 signals in cooperation with the <em>IL</em>-1 receptor accessory protein (<em>IL</em>-<em>1RAcP</em>) and the type II <em>IL</em>-1 receptor (<em>IL</em>-1RII) that functions as a decoy receptor. As glucocorticoid receptors are expressed on astrocytes, we hypothesized that glucocorticoids regulate <em>IL</em>-1 receptors expression. <em>IL</em>-1beta-activated mouse primary astrocytes were treated with 10(-6) M dexamethasone, and <em>IL</em>-1 receptors were studied at the mRNA and protein levels. Using RT-PCR, <em>IL</em>-1RI and <em>IL</em>-1RII but not <em>IL</em>-<em>1RAcP</em> mRNAs were found to be up-regulated by dexamethasone in a time-dependent manner. Dexamethasone (Dex), but not progesterone, had no effect on <em>IL</em>-1RI but strongly increased <em>IL</em>-1RII mRNA expression. Binding studies revealed an increase in the number of <em>IL</em>-1RII binding sites under the effect of Dex, but no change in affinity. These findings support the concept that glucocorticoids have important regulatory effect on the response of astrocytes to <em>IL</em>-1.

The role of interleukin-1 (<em>IL</em>-1), a pro-inflammatory cytokine, in parturition is typically noted by changes in its concentrations. Studying the expression of its receptor family, <em>IL</em>-1 receptor (<em>IL</em>-1R) 1, <em>IL</em>-1R2, <em>IL</em>-1R accessory protein (<em>IL</em>-<em>1RAcP</em>), and its predominantly brain isoform, <em>IL</em>-<em>1RAcP</em>b, during late gestation in the uterus in the Long-Evans rat is another. We assessed changes in their mRNA and protein relative abundance in the uterus and compared <em>IL</em>-<em>1RAcP</em> and <em>IL</em>-<em>1RAcP</em>b mRNA abundance in uterus, cervix, ovaries, placenta, and whole blood of Long-Evans rats during late gestation or in RU486 and progesterone-treated dams using quantitative real-time PCR and western immunoblotting. <em>IL</em>-1R1, <em>IL</em>-<em>1RAcP</em>, and <em>IL</em>-<em>1RAcP</em>b mRNA abundance significantly increased in the uterus at delivery whereas <em>IL</em>-1R2 mRNA abundance significantly decreased. <em>IL</em>-1R1 protein increased at term and <em>IL</em>-1R2 protein decreased at term compared to nonpregnant uteri. <em>IL</em>1-RAcPb mRNA abundance was less than <em>IL</em>-<em>1RAcP</em>, but in the lower uterine segment it was the highest of all tissues examined. RU486 stimulated preterm delivery and an increase in <em>IL</em>-1R1 mRNA abundance whereas progesterone administration extended pregnancy and suppressed the increase in <em>IL</em>-1R1. These data suggest that changes in uterine sensitivity to <em>IL</em>-1 occur during late gestation and suggest another level of regulation for the control of delivery. The roles for <em>IL</em>-<em>1RAcP</em> and <em>IL</em>-<em>1RAcP</em>b need to be determined, but may relate to different intracellular signaling pathways.

The interleukin (<em>IL</em>)-1 system plays a major role in immune responses and inflammation. The <em>IL</em>-1 system components include <em>IL</em>-1α, <em>IL</em>-1β, <em>IL</em>-1 receptor type 1 and <em>IL</em>-1 receptor type 2 (decoy receptor), <em>IL</em>-1 receptor accessory protein (<em>IL</em>-<em>1RAcP</em>), and <em>IL</em>-1 receptor antagonist (<em>IL</em>-1Ra). These components have been shown to play a role in pregnancy, specifically in embryo-maternal communication for implantation, placenta development, and protection against infections. As gestation advances, maternal tissues experience increasing fetal demand and physical stress and <em>IL</em>-1β is induced. Dependent on the levels of <em>IL</em>-1Ra, which regulates <em>IL</em>-1β activity, a pro-inflammatory response may or may not occur. If there is an inflammatory response, prostaglandins are synthesized which may lead to myometrial contractions and the initiation of labor. Many studies have examined the role of the <em>IL</em>-1 system in pregnancy by independently measuring plasma, cervical and amniotic fluid <em>IL</em>-1β or <em>IL</em>-1Ra levels. Other studies have tested for polymorphisms in <em>IL</em>-1β and <em>IL</em>-1Ra genes in women experiencing pregnancy complications such as early pregnancy loss, in vitro fertilization (IVF) failure, pre-eclampsia and preterm delivery. Data from those studies suggest a definite role for the <em>IL</em>-1 system in successful pregnancy outcomes. However, as anticipated, the results varied among different experimental models, ethnicities, and disease states. Here, we review the current literature and propose that measurement of <em>IL</em>-1Ra in relation to <em>IL</em>-1 may be useful in predicting the risk of poor pregnancy outcomes.

Keywords: IL-1; IVF; cytokine; high-risk pregnancy; immune system; implantation; in vitro fertilization; infertility; interleukin; parturition; placenta; preeclampsia; preterm delivery.

<i>Helicobacter pylori</i> colonizes human gastric epithelial cells and contributes to the development of several gastrointestinal disorders. Interleukin (<em>IL</em>)-33 is involved in various immune responses, with reported proinflammatory and anti-inflammatory effects, which may be associated with colitis and colitis-associated cancer. <em>IL</em>-33 induces the inflammatory cascade through its receptor, suppression of tumorigenicity-2 (ST-2). Binding of <em>IL</em>-33 to membrane-bound ST-2 (mST-2) recruits the <em>IL</em>-1 receptor accessory protein (<em>IL</em>-<em>1RAcP</em>) and activates intracellular signaling pathways. However, whether <em>IL</em>-33/ST-2 is triggered by <i>H. pylori</i> infection and whether this interaction occurs in lipid rafts remain unclear. Our study showed that both <em>IL</em>-33 and ST-2 expression levels were significantly elevated in <i>H. pylori</i>-infected cells. Confocal microscopy showed that ST-2 mobilized into the membrane lipid rafts during infection. Depletion of membrane cholesterol dampened <i>H. pylori</i>-induced <em>IL</em>-33 and <em>IL</em>-8 production. Furthermore, in vivo studies revealed <em>IL</em>-33/ST-2 upregulation, and severe leukocyte infiltration was observed in gastric tissues infected with <i>H. pylori</i>. Together, these results demonstrate that ST-2 recruitment into the lipid rafts serves as a platform for <em>IL</em>-33-dependent <i>H. pylori</i> infection, which aggravates inflammation in the stomach.

Interleukin-33 (<em>IL</em>-33) is recognized to transmit a signal through a heterodimeric receptor complex ST2/interleukin-1 receptor accessory protein (<em>IL</em>-<em>1RAcP</em>) bearing activation of myeloid differentiation factor 88 (MyD88). High-frequency stimulation to the Schaffer collateral induced long-term potentiation (LTP) in the CA1 region of hippocampal slices from wild-type control mice. Schaffer collateral/CA1 LTP in <em>IL</em>-33-deficient mice was significantly suppressed, which was neutralized by application with <em>IL</em>-33. Similar suppression of the LTP was found with MyD88-deficient mice but not with ST2-deficient mice. In the water maze test, the acquisition latency in <em>IL</em>-33-deficient and MyD88-deficient mice was significantly prolonged as compared with that in wild-type control mice. Moreover, the retention latency in MyD88-deficient mice was markedly prolonged. In contrast, the acquisition and retention latencies in ST2-deficient mice were not affected. Taken together, these results show that <em>IL</em>-33 acts to express Schaffer collateral/CA1 LTP relevant to spatial learning and memory in a MyD88-dependent manner and that the LTP might be expressed through an <em>IL</em>-1R1/<em>IL</em>-<em>1RAcP</em>-MyD88 pathway in the absence of ST2.

The cell-surface receptor ST2L triggers cytokine release by immune cells upon exposure to its ligand <em>IL</em>-33. To study the effect of ST2L-dependent signaling in different cell types, we generated antagonist antibodies that bind different receptor domains. We sought to characterize their activities in vitro using both transfected cells as well as basophil and mast cell lines that endogenously express the ST2L receptor. We found that antibodies binding Domain 1 versus Domain 3 of ST2L differentially impacted <em>IL</em>-33-induced cytokine release by mast cells but not the basophilic cell line. Analysis of gene expression in each cell type in the presence and absence of the Domain 1 and Domain 3 mAbs revealed distinct signaling pathways triggered in response to <em>IL</em>-33 as well as to each anti-ST2L antibody. We concluded that perturbing the ST2L/<em>IL</em>-33/<em>IL</em>-<em>1RAcP</em> complex using antibodies directed to different domains of ST2L have a cell-type-specific impact on cytokine release, and may indicate the association of additional receptors to the ST2L/<em>IL</em>-33/<em>IL</em>-<em>1RAcP</em> complex in mast cells.

The use of cytokines as adjuvants in poultry is promising because they may enhance immune responses to antigens. In this study, we created two mutants, chicken interleukin-1 beta (Ch<em>IL</em>-1β) Q19A and R140A, which exhibited significantly increased in vivo biological activity compared with wild-type Ch<em>IL</em>-1β. The potential mucosal adjuvant activity of the mutants Q19A and R140A was evaluated in chickens through the intranasal coadministration of a single dose of the Newcastle disease virus (NDV) vaccine with Q19A or R140A. Compared with chickens vaccinated with only the NDV vaccine or the NDV vaccine plus wild-type recombinant Ch<em>IL</em>-1β, chickens vaccinated with Q19A or R140A had significantly increased serum hemagglutination-inhibition antibody titers and anti-NDV-specific IgA antibody levels 1 week later, a high amount of interferon-γ secretion from splenocytes, and increased secretory IgA accumulated in nasal tissues. In addition, molecular dynamics simulations of the mutant R140A bound to its receptor (<em>IL</em>-1RI) and receptor accessory protein (<em>IL</em>-<em>1RAcP</em>) were more energetically favorable than the analogous wild-type ternary complex resulting in a decreased energy, which may stabilize the R140A/<em>IL</em>-1RI/<em>IL</em>-<em>1RAcP</em> complex. In conclusion, the mutants Q19A and R140A are effective adjuvants that accelerate and enhance chicken mucosal immunity when co-administered with one dose of the NDV vaccine.

Soluble or cell-bound <em>IL</em>-1 receptor accessory protein (<em>IL</em>-<em>1RAcP</em>) does not bind <em>IL</em>-1 but rather forms a complex with <em>IL</em>-1 and <em>IL</em>-1 receptor type I (<em>IL</em>-1RI) resulting in signal transduction. Synthetic peptides to various regions in the Ig-like domains of <em>IL</em>-<em>1RAcP</em> were used to produce antibodies and these antibodies were affinity-purified using the respective antigens. An anti-peptide-4 antibody which targets domain III inhibited 70% of <em>IL</em>-1beta-induced productions of <em>IL</em>-6 and PGE(2) from 3T3-L1 cells. Anti-peptide-2 or 3 also inhibited <em>IL</em>-1-induced <em>IL</em>-6 production by 30%. However, anti-peptide-1 which is directed against domain I had no effect. The antibody was more effective against <em>IL</em>-1beta compared to <em>IL</em>-1alpha. <em>IL</em>-1-induced <em>IL</em>-6 production was augmented by coincubation with PGE(2). The COX inhibitor ibuprofen blocked <em>IL</em>-1-induced <em>IL</em>-6 and PGE(2) production. These results confirm that <em>IL</em>-<em>1RAcP</em> is essential for <em>IL</em>-1 signaling and that increased production of <em>IL</em>-6 by <em>IL</em>-1 needs the co-induction of PGE(2). However, the effect of PGE(2) is independent of expressions of <em>IL</em>-1RI and <em>IL</em>-<em>1RAcP</em>. Our data suggest that domain III of <em>IL</em>-<em>1RAcP</em> may be involved in the formation or stabilization of the <em>IL</em>-1RI/<em>IL</em>-1 complex by binding to epitopes on domain III of the <em>IL</em>-1RI created following <em>IL</em>-1 binding to the <em>IL</em>-1RI.