IL-1 Bioactivity Research Articles

BIOACTIVE INTERLEUKIN-1 DETECTED IN IBD PATIENT INTESTINAL BIOPSIES IS A HALLMARK OF ULCERS AND CORRELATES WITH TRANSCRIPTOMIC ASSESSMENTS, INCLUDING AN ULCER-ASSOCIATED GENE MODULE

Abstract BACKGROUND Interleukin (IL)-1 and its post-translationally modified and released forms, IL-1α and IL-1β, have been identified as key molecules to maintain mucosal homeostasis and to drive inflammatory bowel disease (IBD). Deep cellular phenotypic, transcriptional, and in vitro data highlight the importance of IL-1-associated cytokine networks and IL-1 producing cells in severe and anti-TNF non-responsive IBD. Nevertheless, little is known about the presence of bioactive IL-1 proteins within the cell-free gastrointestinal (GI) mucosal environment and associated function in severe IBD and ulceration. METHODS We established a highly sensitive functional assay (~500-fold greater than ELISA) to assess bioactive extracellular IL-1 protein from previously cryopreserved GI biopsies. We performed an ex vivo study in a cohort of Crohn’s disease (CD, n=23), ulcerative colitis (UC, n=18) and non-IBD (n=17) patients, with biopsies from paired inflamed and uninflamed intestinal regions per patient (primarily colon but also ileal). We assessed differential IL-1 bioactivity, including IL-1α vs IL-1β contributions, and performed RNA-seq of the matched cellular compartments of the samples. An ulcer-associated gene signature derived from RNA-seq analysis was evaluated in a single-cell RNA-seq cohort (n=42) of very early onset IBD including monogenic disorders. RESULTS The extent of bioactive intestinal mucosal IL-1α and IL-1β corresponded with disease and ulcer severity in both CD and UC. The most extreme signals were seen in select CD patients with deep ulceration. Bioactive IL-1α was the predominant contributor to total IL-1 signal in most patients although several with ulcers displayed an IL-1β-predominant signal. Matched RNA-seq analysis demonstrated enhanced IL-1 transcripts, correlating with IL-1 bioactivity, and a weighted gene co-expression network analysis (WGCNA) revealed a compelling ulcer-specific module enriched in IL-1 signaling genes and wound repair. We validated our findings in several published (e.g., RISK cohort of ileal CD) as well as unpublished independent adult and pediatric datasets from our group. Deconvolution of the ulcer-specific gene module and projection onto an unpublished single-cell RNA-seq cohort of very early onset IBD patients (including IBD-causative mutations such as IL-10R deficiency characterized by deep ulceration) implicated specific stromal and myeloid populations in ulcer biology. CONCLUSION Mucosal ulceration in IBD is associated with bioactive IL-1α and IL-1β proteins, and transcriptomic evaluation of the same correlates with bioactive signal. An ulcer-associated gene module, validated in other datasets, sheds light on IL-1 biology in intestinal epithelial repair. Results strongly suggest the IL-1 signaling pathway being an attractive and precision-based therapeutic target in subsets of IBD patients.

OP17 IBD ulcers are characterized by bioactive interleukin-1 and transcriptomic hallmarks of stromal cell state reprogramming

Abstract Background The programs that perpetuate the inflammation and prevent epithelial repair in Inflammatory Bowel Disease (IBD) remain unclear. Interleukin (IL)-1 plays a role in the maintenance of mucosal homeostasis but also in IBD. Expansion of IL-1 expressing myeloid cells is a hallmark of IBD tissue, including severe and anti-TNF unresponsive disease, and IL-1 expression is highly localized to ulcer beds. However, little is known about the presence of bioactive IL-1 proteins in the cell-free mucosal environment of IBD, nor whether IL-1-driven programs affect epithelial regeneration in IBD ulcers. Methods We established a highly sensitive assay (~500-fold greater than ELISA) to assess bioactive extracellular IL-1 from previously cryopreserved IBD biopsies. We assessed a cohort of Crohn’s Disease (CD, n=23), Ulcerative Colitis (UC, n=18) and non-IBD (n=17) patients, with biopsies from paired inflamed/uninflamed regions (primarily colon but also ileal). We assessed IL-1 bioactivity, including IL-1α vs IL-1β contributions, and performed RNA-seq of samples’ matched cellular compartments. An ulcer-associated gene signature was interrogated in a single-cell (sc)RNA-seq cohort (n=42) of very early onset (VEO)IBD including monogenic disorders, as well as publicly available IBD bulk RNA-seq datasets and scRNA-seq of mouse models of colitis. Results IL-1α and IL-1β bioactivity corresponded with disease and ulcer severity in CD and UC. The most extreme signals were seen in select CD patients with deep ulceration. IL-1α was the predominant contributor to total IL-1 bioactivity in most patients although several with ulcers displayed IL-1β predominance. IL-1 bioactivity correlated with IL-1 transcripts in matched RNA-seq, and weighted gene co-expression network analysis revealed a compelling ulcer-specific module. This module contained transcription factors and genes related to cell state, validated in publicly available datasets (e.g. RISK cohort of ileal CD and scRNA-seq of murine DSS colitis), that when interrogated in a scRNA-seq dataset of VEOIBD (including patients with IL-10RA mutations characterized by deep ulcers), suggests an orchestrated IL-1-driven myeloid/stromal response to epithelial ulceration involving cell state reprogramming and loss of key myofibroblast populations. Conclusion Mucosal ulceration in IBD is associated with bioactive IL-1α and IL-1β proteins, and transcriptomic evaluation of the same correlates with bioactive signal. An ulcer-associated gene module, validated in other datasets, sheds light on IL-1 biology in intestinal epithelial repair. Results strongly suggest the IL-1 signaling pathway being an attractive and precision-based therapeutic target in subsets of IBD patients.

The course of action for effective anti-cytokine treatment in COVID-19

The course of action for effective anti-cytokine treatment in COVID-19

More than Inflammation: Interleukin-1β Polymorphisms and the Lipid Metabolism

More than Inflammation: Interleukin-1β Polymorphisms and the Lipid Metabolism

Early Production of IL-22 but Not IL-17 by Peripheral Blood Mononuclear Cells Exposed to live Borrelia burgdorferi: The Role of Monocytes and Interleukin-1

If insufficiently treated, Lyme borreliosis can evolve into an inflammatory disorder affecting skin, joints, and the CNS. Early innate immunity may determine host responses targeting infection. Thus, we sought to characterize the immediate cytokine storm associated with exposure of PBMC to moderate levels of live Borrelia burgdorferi. Since Th17 cytokines are connected to host defense against extracellular bacteria, we focused on interleukin (IL)-17 and IL-22. Here, we report that, despite induction of inflammatory cytokines including IL-23, IL-17 remained barely detectable in response to B. burgdorferi. In contrast, T cell-dependent expression of IL-22 became evident within 10 h of exposure to the spirochetes. This dichotomy was unrelated to interferon-γ but to a large part dependent on caspase-1 and IL-1 bioactivity derived from monocytes. In fact, IL-1β as a single stimulus induced IL-22 but not IL-17. Neutrophils display antibacterial activity against B. burgdorferi, particularly when opsonized by antibodies. Since neutrophilic inflammation, indicative of IL-17 bioactivity, is scarcely observed in Erythema migrans, a manifestation of skin inflammation after infection, protective and antibacterial properties of IL-22 may close this gap and serve essential functions in the initial phase of spirochete infection.

Post-natal Changes in Testicular Concentrations of Interleukin-1 Alpha and Beta and Interleukin-6 during Sexual Maturation in Bulls

Based on observations in laboratory animals interleukins could be regulators of testicular development. The objects of this study were to see if interleukins (IL-1 and IL-6) are present in the developing bull testis and to establish the temporal patterns of concentrations of IL-1 and IL-6 in the bovine testis during development. Separate groups of six bull calves were castrated every 4 weeks from 5 to 33 weeks of age, and at 56 weeks of age. Mean testicular IL-1 alpha concentrations decreased (p < 0.01) from 5 to 9 weeks of age and 13 to 21 weeks of age. Mean testicular IL-1 beta concentrations decreased (p < 0.01) from 13 to 17 weeks of age and from 29 to 33 weeks of age. Mean IL-1 bioactivity increased from 13 to 17 weeks of age, decreased to 21 weeks, increased to 25 weeks, decreased to 29 weeks and decreased from 33 to 56 weeks of age (p < 0.05). Mean testicular IL-6 concentrations decreased (p < 0.05) from 9 to 13 weeks of age, increased (p < 0.05) to 21 weeks, decreased (p < 0.05) to 25 weeks, increased (p < 0.05) to 29 weeks and decreased (p < 0.01) to 56 weeks of age. In conclusion, testicular IL-1 alpha, IL-1 beta and IL-6 were found in the bovine testis and concentrations were age dependent. Testicular IL-1 alpha and IL-1 beta concentrations were highest in the early post-natal period; however, IL-1 bioactivity and IL-6 concentrations were greatest in the immediate pre-pubertal period. These findings suggest a functional role for interleukins in testicular development in the bull.

Allele 2 of the interleukin-1 receptor antagonist gene (IL1RN*2) is associated with a decreased risk of primary lung cancer

IL-1 receptor antagonist (IL-1ra), a structural variant of IL-1, binds to the same IL-1 receptor and acts as a competitive inhibitor of IL-1 bioactivity. IL-1ra protein has been widely investigated and found to be associated with different human malignancies. In the second intron of the IL- 1RN gene, there is a functional polymorphism of a variable number of tandem repeats (VNTR), which is characterized as having an importance role in regulating the serum IL-1ra levels, human immune response and cancer risk. We genotyped this VNTR of IL- 1RN in a case-control study of 885 histologically confirmed lung cancer patients and 1024 cancer-free controls frequency-matched to the cases on age and sex in a Chinese population to evaluate the association of this variant and lung cancer risk. We found that the presence of the allele 2 of IL- 1RN ( IL-RN* 2) was associated with a 32% significantly decreased risk of lung cancer (adjusted OR, 0.68; 95% CI, 0.52–0.89). Stratified analyses revealed that the reduced risks associated with the genotypes with IL-RN*2allele (I/II and II/II) were more evident in non-smokers (adjusted OR, 0.53; 95% CI, 0.35–0.79) and in subjects with squamous cell carcinoma (adjusted OR, 0.49; 95% CI, 0.31–0.76). These findings support our priori hypothesis that the IL1RN* 2 allele may contribute to lung cancer risk in the Chinese population. More functional data for this IL1RN polymorphism are warranted to explore its role in lung carcinogenesis.

Genetic control of IL-1beta bioactivity through differential regulation of the IL-1 receptor antagonist.

Regulation of IL-1 bioactivity by the IL-1 receptor antagonist (IL-1Ra) is critical for the preservation of normal vascular structure and function in mice. In humans, IL-1 bioactivity may be further fine-tuned by genetic polymorphisms. We recently proposed that an intronic polymorphism of the human gene encoding the IL-1R antagonist (IL1RN) is a genetic risk factor for the development of chronic cardiac allograft rejection. Here we aimed to establish a physiological basis for this susceptibility. Plasma and peripheral blood mononuclear cells (PBMC) were obtained from 55 healthy human volunteers, whose genotypes for four polymorphisms of IL1 family genes (IL1B, IL1R1 and IL1RN) were determined by PCR. IL-1beta and IL-1Ra production and release in PBMC cultures were studied by flow cytometry and ELISA. Functional and genotypic data were pooled and analyzed first by multivariate and, subsequently, by univariate statistical tests. We were able to confirm our observed association of IL1RN genotype with chronic cardiac allograft rejection using multivariate statistics. IL1RN genotype also emerged as the principal regulator of both constitutive and stimulated IL-1Ra and IL-1beta release. Allele IL1RN*2 was consistently associated with higher IL-1Ra and lower IL-1beta release, in a dosage-dependent manner. Conversely, IL1RN*2 carriage was associated with reduced production of the intracellular isoform of IL-1Ra. These genotypic effects were only observed with prolonged culture prior to stimulation and were not appreciably influenced by the presence of exogenous modulators of IL-1beta production. We conclude that IL1RN genotype is the principal determinant of IL-1beta bioactivity within theIL1 gene cluster in humans and discuss its putative role in disease.

Production and secretion of interleukin-1α proteins by rat testis

The present study characterizes constitutively expressed rat testicular interleukin-1α (IL-1α) proteins. IL-1 bioactivity of crude testis protein was completely neutralized by IL-1α antiserum, IL-1 receptor antagonist, and soluble type I IL-1 receptor. Upon non-denaturating gel permeation chromatography, bioactive IL-1 eluted at molecular sizes of 45, 31, and 17 kDa and at charges of pH 5.7 and 6.0 after chromatofocusing. SDS–PAGE/Western blot analysis of proteins extracted from whole testis, seminiferous tubules, interstitial, and seminiferous tubule fluids all demonstrated IL-1α immunoreactivity at 45, 24, and 19 kDa. Activated macrophages and tissue proteins from endotoxin treated rats showed immunoreactive 31 and 19 kDa IL-1α. The results indicate that the testis produces three isoforms of IL-1α proteins that are secreted into the interstitial compartment and tubular lumen where they may exert paracrine functions. The testicular IL-1α isoforms may represent posttranslationally modified precursor, mature IL-1α, and a 24-kDa alternate splice form.

IL-1ra and IL-1 production in human oral mucosal epithelial cells in culture: differential modulation by TGF-β1 and IL-4

Regulatory cytokines mediate the participation of oral mucosal epithelial cells (OMEC) in local immune responses. The aim of this study was to characterize the isoforms of IL-1 receptor antagonist (IL-1ra) in cultured human primary OMECs and to compare its production with that of IL-1 alpha (IL-1alpha) and IL-1 beta (IL-1beta). Western blot analysis showed that IL-1ra was 22 kDa in size hence slightly smaller than monocyte IL-1ra (25 kDa). A minor form of 20 kDa was also found in unstimulated cell culture lysates. In culture supernatants, IL-1 bioactivity increased after IL-1ra neutralization, indicating that the baseline production of IL-1ra is biologically relevant. Immunohistochemistry showed a relation between IL-1ra and involucrin expressions, suggesting that intracytoplasmic IL-1ra may be involved in cell terminal differentiation. In unstimulated culture lysates, there was far more IL-1ra than IL-1alpha and IL-1beta. TGF-beta1 markedly increased the IL-1ra/IL-1beta ratio from 93.6 : 1 to 300 : 1. IL-4, which is generally described as an anti-inflammatory cytokine, increased IL-1 but not IL-1ra production. TNF-alpha increased intracellular production of the three IL-1 members. IL-1ra levels were lower in supernatants than in lysates of cultured cells. Our results show that human OMECs constitutively produce significant amounts of a biologically active form of IL-1ra. TGF-beta1 mu(p)-regulation points to a positive amplification loop and IL-4 to a down-regulation loop, both including Th2 cells and OMECs. They may be important in oral tolerance and IgA production, respectively.

Interleukin 1 and Chronic Rejection: Possible Genetic Links in Human Heart Allografts

Chronic rejection is a leading cause of graft loss in thoracic transplant recipients. Studies on the pathogenesis of chronic rejection have suggested a contributory role for certain cytokines and growth factors. The activity of these mediators is subject to genetic variation if a polymorphism alters expression, or function, of the ligand or its receptor. Here we have asked if certain cytokine and growth factor gene polymorphisms correlate with chronic rejection in recipients of thoracic allografts. In a retrospective analysis of 179 recipients of thoracic organ transplants (128 heart; 36 heart-lung; and 15 lung), polymorphisms in 8 genes that influence the inflammatory process, namely IL1B, IL1R1, IL1RN, IL6, IL10, TNFA, TGFB1 and FCGRIIA, were examined. Genotypic data from recipients who had either died or been re-transplanted as a result of chronic rejection (n = 96) were then compared to those of recipients who had a functioning graft for more than 11 years (n=83). In the heart graft recipients, only those polymorphisms that influenced expression of the IL1 receptor antagonist gene had a significant correlation with graft survival, with homozygosity for the IL1RN*1 allele being associated with rejection. The alternative, less frequent IL1RN alleles emerged as genomic predictors of long-term allograft survival. This association was especially strong when IL1 region haplotypes were considered, particularly when analysis was confined to heart transplant recipients who had had multiple acute rejection episodes (OR>20). This case-control study indicates that gene polymorphisms which influence IL1 bioactivity also influence the progression of chronic rejection in heart grafts.

ENDOTOXIN-INDUCED INTERLEUKIN 1 EXPRESSION IN TESTICULAR MACROPHAGES IS ACCOMPANIED BY DOWNREGULATION OF THE CONSTITUTIVE EXPRESSION IN SERTOLI CELLS

Interleukin-1α (IL-1α) is constitutively produced by Sertoli cells in adult rat testes. We demonstrate here that adult rats initiate expression of IL-1α and IL-1β in testicular macrophages and decrease plasma testosterone by 60%, 2h after administration of endotoxin. The macrophage activation was accompanied by downregulation of IL-1α mRNA expression in Sertoli cells. Despite increased tissue concentrations of IL-1α and IL-1β immunoreactive protein, the level of bioactive IL-1 in the testis remained unchanged. Testes from prepubertal rats responded similarly to endotoxin, but lacked constitutive expression of IL-1α. We conclude that endotoxin-induced inflammation involves the testis by local macrophage activation and cytokine secretion. The paracrine mechanisms regulating IL-1 bioactivity in the testis are unknown but may represent a means to protect germ cells from noxious effects of inflammation.

Porcine alveolar macrophages: poor accessory or effective suppressor cells for T-lymphocytes

Porcine Alv-Mø from bronchoalveolar lavages were tested for their function in an in vitro foot-and-mouth disease virus (FMDV)-specific lymphoproliferative recall response. The Alv-Mø were seen to be poor accessory cells when compared with peripheral blood monocytes. This poor capacity was evident despite an efficient expression of SLA-DR region antigens, and other co-stimulatory adhesion molecules. It was noted that Alv-Mø secrete relatively little interleukin 1 (IL-1β), with or without LPS induction, even though mRNA for the cytokine could be detected. In contrast, blood monocytes with their effective accessory activity were potent secretors of IL-1. Although this IL-1β would be important with respect to the accessory capacity of monocytic cells, it was noted that the absence of bioactive IL-1 from the Alv-Mø cultures was not solely responsible for their poor accessory function. In fact, the Alv-Mø produced factors which not only inhibited IL-1 bioactivity, but were also responsible for a clear suppression of lymphoproliferation. This suppressor activity was dependent on the type of monocytic cell present in the culture, being more prominent when “scavenger” phagocytes were present. Thus, the major role of Alv-Mø is not as an accessory cell akin to monocytes, but as both a scavenger cell, related to Mø derived from monocytes in the absence of inflammatory signals, and an immunoregulatory cell.

Acute systemic inflammation up-regulates secretory sphingomyelinase in vivo: a possible link between inflammatory cytokines and atherogenesis.

Inflammation plays a critical role in atherogenesis, yet the mediators linking inflammation to specific atherogenic processes remain to be elucidated. One such mediator may be secretory sphingomyelinase (S-SMase), a product of the acid sphingomyelinase gene. The secretion of S-SMase by cultured endothelial cells is induced by inflammatory cytokines, and in vivo data have implicated S-SMase in subendothelial lipoprotein aggregation, macrophage foam cell formation, and possibly other atherogenic processes. Thus, the goal of this study was to seek evidence for S-SMase regulation in vivo during a physiologically relevant inflammatory response. First, wild-type mice were injected with saline or lipopolysaccharide (LPS) as a model of acute systemic inflammation. Serum S-SMase activity 3 h postinjection was increased 2- to 2.5-fold by LPS (P < 0.01). To determine the role of IL-1 in the LPS response, we used IL-1 converting enzyme knockout mice, which exhibit deficient IL-1 bioactivity. The level of serum S-SMase activity in LPS-injected IL-1 converting enzyme knockout mice was approximately 35% less than that in identically treated wild-type mice (P < 0.01). In LPS-injected IL-1-receptor antagonist knockout mice, which have an enhanced response to IL-1, serum S-SMase activity was increased 1. 8-fold compared with LPS-injected wild-type mice (P < 0.01). Finally, when wild-type mice were injected directly with IL-1beta, tumor necrosis factor alpha, or both, serum S-SMase activity increased 1. 6-, 2.3-, and 2.9-fold, respectively (P < 0.01). These data show regulation of S-SMase activity in vivo and they raise the possibility that local stimulation of S-SMase may contribute to the effects of inflammatory cytokines in atherosclerosis.

Canine rheumatoid arthritis and inflammatory cytokines

Bioassays were developed to detect canine pro-inflammatory cytokines. These enabled characterisation of these cytokines and their isoforms and provided means for their assay in the joints of dogs with different naturally occurring arthropathies. Canine IL-1 was detected by its induction of proliferation of D10(N4)M cell line, whilst IL-6 had a proliferative effect on B9 cell line. TNFα had a cytotoxic effect on WEHI 164 (13) cells. Partial purification of the cytokines was achieved by FPLC ion-exchange chromatography and two isoforms of IL-1 were shown, possibly corresponding to IL-1α and IL-1β. TNFα only appeared as one isoform whereas IL-6 showed at least five isoforms, possibly corresponding to the other molecules in the IL-6 family, such as IL-11 and oncostatin M. Analysis of synovial fluids from dogs with osteoarthritis (OA) and rheumatoid arthritis (RA) showed that IL-1 and TNFα bioactivity was not readily detectable at increased levels in diseased joints but that IL-6 was significantly increased in both diseases. It is now important to determine the role of IL-6 in OA and RA in the dog, particularly in the induction of proteolytic enzymes which lead to cartilage loss.

Interleukin-1 in fish

It has been known for over a decade that IL-1 bioactivity exists in fish. Recently IL-1 has been cloned in several teleost species using two different approaches; homology cloning or a suppression subtractive PCR technique. One of the most important differences of the fish sequences compared with mammalian IL-1 is the lack of a clear ICE cut site, and it remains to be determined whether the fish molecule is processed to a mature peptide. The IL-1 gene organisation has also been determined for rainbow trout, and Southern blot analysis suggests at least one related gene is present in this species. Expression of the IL-1 transcript can be induced in vitro by stimulation of head kidney leucocytes with LPS or PHA, or in vivo by challenge with Gram-negative bacteria. A number of incompletely spliced transcripts have also been detected in tissues from challenged fish.

Rapid Expression of IL-1β by Intestinal Epithelial Cellsin Vitro

Intestinal epithelial cells have been shown to produce IL-1βin vivo.This gene expression is rapid and precedes most determinants of inflammation, suggesting a pivotal role for IL-1β in the early events leading to inflammation. To better understand the mechanisms leading to this IL-1β production, we have developed anin vitromodel system employing a nontransformed intestinal epithelial cell line that does not constitutively express IL-1β. Following detachment, these cells rapidly expressed IL-1β mRNA. This expression was enhanced, but not induced, by LPS. IL-1β protein was detected by immunoprecipitation in the culture medium from passaged IEC-18 but not intracellularly, suggesting an efficient secretion of the molecule following induction. Interestingly, culture supernatants from passaged cells were without IL-1 bioactivity, suggesting the presence of an inhibitor as well. RT-PCR and Western blot analysis showed expression of IL-1RII by IEC-18 following detachment, possibly explaining the observed lack of bioactivity. These results indicate a novel pathway for IL-1β production and suggest that proinflammatory effects of IEC-derived IL-1 may be modulated by the simultaneous production of IL-1 antagonists.

Extracellular release of the type I intracellular IL-1 receptor antagonist from human airway epithelial cells: differential effects of IL-4, IL-13, IFN-gamma, and corticosteroids.

Three IL-1R antagonists (IL-1Ra) exist: secreted IL-1Ra and intracellular IL-1Ra (icIL-1Ra) types I and II. We have previously reported that human airway epithelial cells (HAEC) express icIL-1Ra type I, which can be up-regulated by corticosteroids. This study assessed whether cytokines and corticosteroids differentially effect icIL-1Ra type I protein release from HAEC to the extracellular compartment. We report that icIL-1Ra type I mRNA and intracellular protein are up-regulated in NCI-H292 cells, a human pulmonary mucoepidermoid carcinoma cell line, in response to IL-4, IL-13, IFN-gamma, and dexamethasone. The icIL-1Ra type I protein was detected in concentrated cell culture supernatants from NCI-H292 cells and normal human bronchial epithelial cells. The release of biologically relevant concentrations of active IL-1Ra from normal human bronchial epithelial cells was demonstrated by the ability of a neutralizing anti-IL-1Ra Ab to augment IL-1beta-mediated IL-8 secretion. IL-4, IL-13, and IFN-gamma induced immunoreactive IL-1Ra release into supernatants from NCI-H292 cells. Dexamethasone inhibited constitutive and cytokine-induced release of immunoreactive IL-1Ra. The release of icIL-1Ra type I protein was not related to cytotoxicity, as measured by lactate dehydrogenase. We propose that icIL-1Ra type I release from HAEC represents a novel mechanism by which IL-1 bioactivity in the airway microenvironment may be modulated. Cytokine-mediated icIL-1Ra type I synthesis may increase both intracellular protein and release to the extracellular space, where cell surface IL-1R can be antagonized. In contrast, corticosteroid-induced increases in icIL-1Ra type I synthesis and inhibition of extracellular protein release promote accumulation of icIL-1Ra type I protein within the intracellular compartment.

Anthralin (dithranol)in vitroinhibits human monocytes to secrete IL-6, IL-8 and TNF-α, but not IL-1

Anthralin is a most widely used compound for topical treatment of psoriasis. Whereas numerous studies have ascertained anthralin as a safe and effective drug its mode of action still remains unclear. Previous studies demonstrated dose-dependent inhibition of a number of pro-inflammatory functions in human neutrophils and monocytes (MO). The aim of the present study was to investigate in stimulated MO the effect of anthralin on the secretion of cytokines which are of known importance for the psoriatic tissue reaction. Highly purified MO were incubated with anthralin (0.01-1.0 microgram/ml), its clinically inactive derivative danthrone (0.1 and 10 micrograms/ml), the solvent acetone, or medium alone. Culture supernatants were analysed for immunoreactivity for interleukin-1 beta, and -8 (IL-1 beta, IL-8), and tumour necrosis factor alpha (TNF-alpha) by specific ELISA. IL-6 bioactivity was determined using the B9-bioassay. Additionally, IL-1 bioactivity was measured by the D10[N4]M-bioassay. The results show a dose-dependent inhibition of MO IL-6, IL-8, and TNF-alpha release with a half-maximal inhibitory concentration of 0.25-0.6 microgram/ml of anthralin. There was no effect of danthrone or acetone on the secretion of these cytokines from MO. Secretion of IL-1 beta immunoreactivity measured by ELISA as well as determination of biological activity of IL-1 using the D10[N4]M-bioassay revealed a slight increase in IL-1 secretion with a maximum at an anthralin concentration of 0.1 microgram/ml. Danthrone at a concentration of 10 micrograms/ml and acetone (0.1%) similarly enhanced IL-1 secretion from human MO measured by both methods. Our results demonstrate a differential, dose-dependent inhibition of cytokine secretion from human MO by anthralin. The present data provide evidence that the anti-inflammatory and anti-proliferative activity of anthralin may at least in part be due to its inhibitory effect on pro-inflammatory cytokine secretion by MO.

Induced upregulation of IL-1, IL-1RA and IL-1R type I gene expression by Schwann cells

We analyzed two distinct phenotypes of Schwann cells (SC), non-differentiated and differentiated, for their ability to produce IL-1 α and IL-1 β, and to express message for IL-1 receptor antagonist (IL-1RA) and IL-1R type I, in vitro. SC were stimulated with: lipopolysaccharide (LPS), products of activated splenocytes (ASP), products from LPS stimulated SC (SCP), rat recombinant IL-1 β (rrIL-1 β) or dexamethasone. IL-1 α, IL-1 β and IL-1RA mRNA levels were highly upregulated after stimulation with LPS, ASP, SCP or rrIL-1 β. SC constitutively expressed low levels of message for IL-1 α and IL-1 β but not IL-1RA. Specific mRNAs for both IL-1 isotypes were highly upregulated 2 to 4 h after LPS stimulation and then decreased and were undetectable by 24 h. IL-1RA mRNA was detectable after 6 h of LPS stimulation and was maximally upregulated at 24 h. IL-1 gene expression was inducible in both SC phenotypes. IL-1 β could be detected by immunofluorescence in SC, one to three days after LPS stimulation. At the same time IL-1 bioactivity was maximal in SC supernatants. Treatment with either SCP, rrIL-1 β or dexamethasone induced upregulation of IL-1R type I mRNA with maximal expression between 2 to 4 h, in SC. Inducible expression of genes for IL-1 α, IL-1 β, IL-1RA and IL-1R type I in both differentiated and non-differentiated SC suggest autocrine and paracrine regulation of IL-1 by SC.