Is Interleukin Research Articles

Gene deletion of Interleukin-1α reduces ER stress-induced CHOP expression in macrophages and attenuates the progression of atherosclerosis in apoE-deficient mice

The pathophysiology of atherosclerosis initiation and progression involves many inflammatory cytokines, one of them is interleukin (IL)-1α that has been shown to be secreted by activated macrophages. We have previously shown that IL-1α from bone marrow-derived cells is critical for early atherosclerosis development in mice. It is known that endoplasmic reticulum (ER) stress in macrophages is involved in progression to more advanced atherosclerosis, but it is still unknown whether this effect is mediated through cytokine activation or secretion. We previously demonstrated that IL-1α is required in ER stress-induced activation of inflammatory cytokines in hepatocytes and in the associated induction of steatohepatitis. In the current study, we aimed to examine the potential role of IL-1α in ER stress-induced activation of macrophages, which is relevant to progression of atherosclerosis. First, we demonstrated that IL-1α is required for atherosclerosis development and progression in the apoE knockout (KO) mouse model of atherosclerosis. Next, we showed that ER stress in mouse macrophages results in the protein production and secretion of IL-1α in a dose-dependent manner, and that IL-1α is required in ER stress-induced production of the C/EBP homologous protein (CHOP), a critical step in ER stress-mediated apoptosis. We further demonstrated that IL-1α-dependent CHOP production in macrophages is specifically mediated through the PERK-ATF4 signaling pathway. Altogether, these findings highlight IL-1α as a potential target for prevention and treatment of atherosclerotic cardiovascular disease.

344 Foxo4 and AHR control inflammation-induced tissue damage via secretion of IL-22 in T cells

Epithelial barrier integrity is of outermost importance to protect the human body from external harm. One of the key cytokines involved in tissue regeneration is interleukin (IL)-22. During inflammation, IL-22 producing cells are recruited into barrier organs to restore inflammation mediated barrier damage. The aryl hydrocarbon receptor (AhR) has been shown to regulate IL-22 expression in T cells. Previous results from our group indicated that another transcription factor Forkhead box O (FOXO) 4 is expressed in human T cells.

Colchicine for COVID-19: targeting NLRP3 inflammasome to blunt hyperinflammation.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is capable of inducing the activation of NACHT, leucine-rich repeat, and pyrin domain-containing protein 3 (NLRP3) inflammasome, a macromolecular structure sensing the danger and amplifying the inflammatory response. The main product processed by NLRP3 inflammasome is interleukin (IL)-1β, responsible for the downstream production of IL-6, which has been recognized as an important mediator in coronavirus disease 2019 (COVID-19). Since colchicine is an anti-inflammatory drug with the ability to block NLRP3 inflammasome oligomerization, this may prevent the release of active IL-1β and block the detrimental effects of downstream cytokines, i.e. IL-6. To date, few randomized clinical trials and many observational studies with colchicine have been conducted, showing interesting signals. As colchicine is a nonspecific inhibitor of the NLRP3 inflammasome, compounds specifically blocking this molecule might provide increased advantages in reducing the inflammatory burden and its related clinical manifestations. This may occur through a selective blockade of different steps preceding NLRP3 inflammasome oligomerization as well as through a reduced release of the main cytokines (IL-1β and IL-18). Since most evidence is based on observational studies, definitive conclusion cannot be drawn and additional studies are needed to confirm preliminary results and further dissect how colchicine and other NLRP3 inhibitors reduce the inflammatory burden and evaluate the timing and duration of treatment.Supplementary InformationThe online version contains supplementary material available at 10.1007/s00011-022-01540-y.

Inhalant Mammal-Derived Lipocalin Allergens and the Innate Immunity.

A major part of important mammalian respiratory allergens belongs to the lipocalin family of proteins. By this time, 19 respiratory mammalian lipocalin allergens have been registered in the WHO/IUIS Allergen Nomenclature Database. Originally, lipocalins, small extracellular proteins (molecular mass ca. 20 kDa), were characterized as transport proteins but they are currently known to exert a variety of biological functions. The three-dimensional structure of lipocalins is well-preserved, and lipocalin allergens can exhibit high amino acid identities, in several cases more than 50%. Lipocalins contain an internal ligand-binding site where they can harbor small principally hydrophobic molecules. Another characteristic feature is their capacity to bind to specific cell-surface receptors. In all, the physicochemical properties of lipocalin allergens do not offer any straightforward explanations for their allergenicity. Allergic sensitization begins at epithelial barriers where diverse insults through pattern recognition receptors awaken innate immunity. This front-line response is manifested by epithelial barrier-associated cytokines which together with other components of immunity can initiate the sensitization process. In the following, the crucial factor in allergic sensitization is interleukin (IL)-4 which is needed for stabilizing and promoting the type 2 immune response. The source for IL-4 has been searched widely. Candidates for it may be non-professional antigen-presenting cells, such as basophils or mast cells, as well as CD4+ T cells. The synthesis of IL-4 by CD4+ T cells requires T cell receptor engagement, i.e., the recognition of allergen peptides, which also provides the specificity for sensitization. Lipocalin and innate immunity-associated cell-surface receptors are implicated in facilitating the access of lipocalin allergens into the immune system. However, the significance of this for allergic sensitization is unclear, as the recognition by these receptors has been found to produce conflicting results. As to potential adjuvants associated with mammalian lipocalin allergens, the hydrophobic ligands transported by lipocalins have not been reported to enhance sensitization while it is justified to suppose that lipopolysaccharide plays a role in it. Taken together, type 2 immunity to lipocalin allergens appears to be a harmful immune response resulting from a combination of signals involving both the innate and adaptive immunities.

Structural Understanding of Interleukin 6 Family Cytokine Signaling and Targeted Therapies: Focus on Interleukin 11.

Cytokines are small signaling proteins that have central roles in inflammation and cell survival. In the half-century since the discovery of the first cytokines, the interferons, over fifty cytokines have been identified. Amongst these is interleukin (IL)-6, the first and prototypical member of the IL-6 family of cytokines, nearly all of which utilize the common signaling receptor, gp130. In the last decade, there have been numerous advances in our understanding of the structural mechanisms of IL-6 family signaling, particularly for IL-6 itself. However, our understanding of the detailed structural mechanisms underlying signaling by most IL-6 family members remains limited. With the emergence of new roles for IL-6 family cytokines in disease and, in particular, roles of IL-11 in cardiovascular disease, lung disease, and cancer, there is an emerging need to develop therapeutics that can progress to clinical use. Here we outline our current knowledge of the structural mechanism of signaling by the IL-6 family of cytokines. We discuss how this knowledge allows us to understand the mechanism of action of currently available inhibitors targeting IL-6 family cytokine signaling, and most importantly how it allows for improved opportunities to pharmacologically disrupt cytokine signaling. We focus specifically on the need to develop and understand inhibitors that disrupt IL-11 signaling.

Fucoxanthin modulates the development of 7, 12-dimethyl benz (a) anthracene-induced skin carcinogenesis in swiss albino mice in vivo

Background: Fucoxanthin (Fx), an distinctive carotenoid occurs on brown seaweed, contains several benefits including anti-cancer effects. To investigate the chemopreventive effectual of Fx on 7,12-dimethylbenz[a] anthracene (DMBA)-alone skin tumor development in Swiss albino mice. Materials and Methods: The skin sarcoma being provoked on the hairless flipside of the mice's skin, twice weekly for 8 weeks through challenging through DMBA (25 μg on 0.1 ml acetone/mice). Thereafter, the mice were oral supplementation with 50 mg/kg body weight (BW), Fx for 25th week a frequency of three times/week. Tumor size, change in BW, the cumulative quality of papillomas and some oxidative stress-related parameters were measured. Results: Orally administered Fx were notably increased BW, delayed tumor incidence with no abnormal pathology in DMBA-induced skin tumor mice. Fx effectively modulates the level of xenobiotic enzymes, brought back to the statuses of lipid peroxidation (LPO) enzymes and increased antioxidant enzymes status in squamous cell carcinomas (SCC). A considerable decrease in the protein expression of proinflammatory markers that is interleukin (IL) 1-6, tumor necrosis factor-alpha (TNF-α), and IL-β and restored the expression status of inflammatory regulators such as nuclear factor-kappa B (NF-κB), cyclooxygenase-2 (COX-2), and inducible nitric oxide synthase (iNOS) on serum. These findings were supported through histopathological examinations. Hence, it was clear that Fx possessed a good anticancer activity against skin cancer. Conclusion: The chemopreventive prospective of Fx was eventually owing to its altering capacity on the levels of proinflammatory (IL 1-6, TNF-α, and IL-β) cytokines, inflammatory (NF-κB, COX-2, and iNOS) markers, LPO, antioxidants, and toxin-eliminating mediators in DMBA-provoked skin SCC. Thus, it can be concluded that the FX indicates the antitumor potential on DMBA challenged skin tumor proliferation model on mice.

Antiproliferative and Anti-invasion Effects of Carvacrol on PC3 Human Prostate Cancer Cells through Reducing pSTAT3, pAKT, and pERK1/2 Signaling Proteins.

Background:One of the most effective parameters in the progression of the prostate cancer is interleukin (IL)-6 through affecting pSTAT3, pERK1/2, and pAKT cell signaling proteins. Carvacrol is an herbal antioxidant with antitumor effects. The purpose of this study was to investigate the effects of carvacrol on IL-6 gene expression, pSTAT3, pAKT, pERK1/2 cellular signaling proteins, and invasion in human prostate cancer PC3 cells.Methods:PC3 cell viability was evaluated by MTT assay with different concentrations of carvacrol (0–800 μM). IL-6 gene expression and cellular concentration of pSTAT3, pERK1/2, and pAKT were investigated using the real-time reverse transcription quantitative polymerase chain reaction (RT-qPCR) and western blotting technic, respectively. PC3 cell invasion was determined by invasion assay test.Results:Carvacrol IC50 for PC3 prostate cancer cells was 360 μM. Carvacrol led to a significant reduction (P < 0.05) for IL-6 gene expression in a dose-dependent manner compared to control. IL-6 protein reduced 41.5% and 52.7% when compared with control cells at 360 and 420 μM of carvacrol, respectively. Carvacrol led to a decline in pSTAT3, pAKT, and pERK1/2 above 360 μM compared to control. PC3 potential invasion was significantly reduced after treatment with carvacrol in a dose-dependent manner.Conclusions:Decreased IL-6 protein level by carvacrol resulted in diminishing of pSTAT3, pERK1/2, and pAKT signaling proteins, which leads to the reduction of the cell survival, proliferation, and invasion in PC3 cells.

Benralizumab in the treatment of severe asthma: design, development and potential place in therapy.

Asthma is a widespread and heterogeneous inflammatory disease of the airways, which is characterized by several different phenotypes and endotypes. In particular, eosinophilic airway inflammation is a common pathologic trait of both allergic and nonallergic asthma. The key cytokine responsible for maturation, activation, recruitment, and survival of eosinophils is interleukin (IL)-5, which is mainly produced by T helper 2 (Th2) lymphocytes and group 2 innate lymphoid cells. Therefore, for uncontrolled patients with severe eosinophilic asthma, who are not fully responsive to corticosteroids, IL-5 represents a very important molecular target for add-on biological therapies. Among these new treatments, anti-IL-5 monoclonal antibodies such as mepolizumab and reslizumab have been developed and clinically evaluated. Furthermore, benralizumab is currently the only available biologic drug that specifically binds to the IL-5 receptor, thus preventing the interaction with its ligand and the consequent pro-inflammatory effects. The effectiveness of benralizumab in improving severe eosinophilic asthma has been well-documented by many randomized controlled trials.

Airway Inflammation and Inflammatory Biomarkers.

Severe asthma is a complex disease consisting of different endotypes with different inflammatory and clinicopathological characteristics due to the heterogeneity of immune responses and smooth muscle dysfunction. There is an unmet clinical need to develop and to validate biomarkers that can differentiate between the asthma endotypes and guide clinical management, particularly since the availability of biologicals directed against T2 cytokines. The presence of a "Th2 endotype" is currently assessed in clinical practice using markers, such as eosinophil count in sputum or blood, fraction of exhaled nitric oxide, and immunoglobulin E. Individually or in combination, they may help to identify, for example, if the dominant effector protein is interleukin (IL) 5, IL13, or IgE. There is no reliable marker of a "non-Th2 endotype" although sputum neutrophil may provide some indication. The unbiased systems biology approach to severe asthma endotyping which integrates omics signatures and clinical data using large cohort studies may provide more comprehensive information than simple cellular measurements. Novel imaging techniques, such as hyperpolarized noble gas magnetic resonance imaging and computed tomography parametric response maps and metabolomics profiling in breath and other body fluids are also being evaluated as potential biomarkers to guide therapy and to assess prognosis.

Association of the IL-15 and IL-15Rα genes with celiac disease

Celiac disease is a chronic autoimmune condition triggered by dietary gluten in genetically predisposed individuals and the treatment is a strict gluten-free diet. The major predisposing genes are HLA-DQA1 and HLA-DQB1, but these are not sufficient for disease development. One of the candidate genes worth studying is interleukin (IL)-15 gene, together with its specific receptor, IL-15Rα, as they participate in promoting lymphocyte signaling and survival, and the establishment of appropriate conditions for villous atrophy, then acting as key players in the immunopathogenesis of CD. Here we analyze IL-15 and IL-15Rα genes in samples from the Spanish Consortium for Genetics of Celiac Disease (CEGEC) collection, identifying two regulatory single-nucleotide polymorphisms (SNP) that might be associated with celiac disease: rs4956400 (p-value 0.0112, OR 1.21, 95% CI 1.04–1.40) and rs11100722 (p-value 0.0087, OR 1.24, 95% CI 1.06–1.45), both located upstream the IL15 gene. When the expression of both genes was assessed, these two SNPs were found to be correlated with IL-15 higher protein expression. Besides, rs8177655 from IL15RA was also associated to mRNA IL-15 expression in CD patients. Finally, three SNPs from IL15RA intronic regions, rs2296141, rs3136614 and rs3181148, and another from its 3’UTR region, rs2229135, could be related to the age of diagnosis of celiac disease patients.

A Model to Explain How the Bacille Calmette Guérin (BCG) Vaccine Drives Interleukin-12 Production in Neonates

The Bacille Calmette Guérin (BCG) vaccine is the only routine vaccination at birth that effectively induces neonatal T-helper 1 (Th1)-polarized immune responses. The primary cytokine that drives CD4+ T-cell Th1 differentiation is interleukin (IL)-12 p70, a heterodimeric cytokine composed of the IL-12 p35 and IL-12 p40 subunits. We therefore examined the mechanisms involved in BCG vaccine stimulation of IL-12 p35 and p40 production from human umbilical cord (neonatal) cells. We found that BCG bacilli did not upregulate IL-12 p35 mRNA production, but upregulated IL-12 p40 mRNA production in a Toll-like receptor (TLR)2-dependent manner, in human neonatal monocyte-derived dendritic cells (mdDCs). The combination of TLR2 signaling, Type I interferon (IFN), and Type II IFN induced maximal levels of IL-12 p35 and p40 mRNA production in human neonatal mdDCs. The cell-free supernatants of reconstituted BCG vaccine vials contained extracellular mycobacterial (BCG) DNA which could induce IFN-α (Type I IFN) production in human neonatal plasmacytoid dendritic cells (pDCs). BCG bacilli also stimulated human neonatal CD16lo natural killer (NK) cells to produce IFN-γ (Type II IFN) in a TLR2-dependent manner. We have therefore proposed a model where BCG vaccine could stimulate the combination of neonatal conventional DCs (cDCs), pDCs, and CD16lo NK cells to produce optimal neonatal IL-12 p35 and p40 (IL-12 p70) production and subsequent CD4+ T-cell Th1 polarization. An adjuvant that emulates the mechanism by which the BCG vaccine stimulates neonatal IL-12 p35 and p40 production could improve vaccine strategies at birth for protection against intracellular pathogens and toxins.

Association of the Precursor of Interleukin-1β and Peritoneal Inflammation-Role in Pathogenesis of Endometriosis.

The most important proinflammatory cytokine is interleukin (IL)-1β, however its precursor, prointerleukin-1β (proIL-1β), can also potentiate inflammatory state. The aim of this study was to explore the involvement of proIL-1β in pathogenesis of endometriosis. For this purpose, we evaluated concentrations of proIL-1β, IL-1β, and soluble IL-1 receptor type 2 (sIL-1R2) in peritoneal fluid (PF) and macrophage culture medium of women with endometriosis. PF from 55 women with and without endometriosis was collected during laparoscopy. Peritoneal macrophages were cultured in basal and stimulated with lipopolysaccharide (LPS) conditions. Concentrations of cytokines were measured with enzyme-linked immunosorbent assays (ELISA). PF proIL-1β and IL-1β levels in endometriosis women were higher than in the control. Higher basal and stimulated macrophage secretion of cytokines in endometriosis patients than in the control was observed. However, in endometriosis, there was a higher level of proIL-1β than for the mature molecule. Additionally, lower PF and macrophages culture medium sIL-1R2 levels were observed in women with endometriosis. Abnormal proIL-1β concentration in PF and higher macrophage secretion can escalate peritoneal inflammation and endometriosis formation. The results are presented as a total IL-1β, which is a sum of proIL-1βand IL-1β, and we believe that it reflects the actual cytokine production. The imbalance among all studied cytokines in endometriosis may be linked with an ability to transform acute inflammation to the chronic inflammation.

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Objectives Endothelial dysfunction is thought to be one of the mechanism involving in pre-eclampsia manifestation. Likewise, one of the inflammatory mediators involved in this process is interleukin (IL)-6, a mediator synthesized by mononuclear phagocytes, endothelial cells and fibroblasts in response to inflammatory stimuli. IL-6 has been described increased in women with pre-eclampsia. Thus, we aimed to investigate the association between endothelial dysfunction (by flow-mediated dilatation) and IL-6 levels in women with pre-eclampsia. Methods Following ethical approval, a written consent was obtained from 21 women diagnosed with pre-eclampsia. Endothelial function was evaluated by brachial artery flow-mediated dilatation. IL-6 was quantified using MagPlexTH-C - microspheres system, in maternal plasma. Pre-eclamptic groups was divided into altered (n = 9) and normal (n = 12) endothelial function, considering a cutoff point of 10%, below or above, respectively. The Mann–Whitney U-tests was carried out to compare the IL-6 values between pre-eclamptic groups. Correlations between the parameters were tested with a Spearman’s Rank correlation tests. The null hypothesis was rejected when p Results Higher levels of IL-6 was observed in altered flow-mediated dilatation test group in relation to normal endothelial group (median [IQR]): 4.34 pg/mL [3.38; 6.22] and 2.56 pg/mL [1.57; 3.67]; p = 0.028, respectively. When an analysis was made between flow-mediated dilatation levels and IL-6, a negative correlation was found (r = −0.514, p = 0.017). Conclusions As expected in patients with pre-eclampsia, a high inflammatory response showed a low flow-mediated dilatation. The significant correlation between a potential clinical prediction method and a well-known inflammatory cytokine support the importance of working not only with one potential biomarker to predict pre-eclampsia, but also, with a group of molecules and/or techniques, in order to increase the chance to diagnose the disease as early in pregnancy as possible – before signals and symptoms. Disclosures M.R. Hentschke: None. M.C. Vieira: None. E.V. da Cunha Filho: None. J. Guaragna: None. C.E. Poli de Figueiredo: None. B.E. Pinheiro da Costa: None.

T cell Suppression In Vitro During Toxoplasma gondii Infection is the Result of IL‐2 Competition Between Tregs and T cells Leading to Death of Proliferating T cells

A reduced proliferation to T cell mitogens is observed in vitro in murine cells isolated during the acute phase of Toxoplasma gondii infection. Foxp3(+) regulatory T cells (Tregs) mediate this suppression, which is interleukin (IL)-2 dependent. In this work, we analysed the mechanism of this Treg-mediated suppression. We found that removal of antigen-presenting cells (APC) from spleen cells from infected mice did not modify suppression but further elimination of Tregs led to a restored proliferation, demonstrating that Tregs mediate suppression in the absence of APC. Production of IL-2 by T cells from infected animals was abolished but partially restored when Tregs were removed. However, IL-2 levels and T cell proliferation were restored when Tregs and T cells were separated by transwells, indicating that Tregs require close proximity with T cells to induce suppression. Tregs from infected mice were able to reduce proliferation of CTLL-2 cells in the classical IL-2 bioassay, strongly suggesting that Tregs compete with T cells for IL-2. We found that T cells from infected mice died after a few rounds of division in vitro, but addition of recombinant IL-2 or removal of Tregs abolished this effect. Our results showed that suppression of T cell proliferation during acute Toxoplasma gondii infection is the result of death of proliferating T cells by Treg-mediated IL-2 competition. Thus, immunosuppression is due to death of proliferating T cells as a consequence of low IL-2 availability.

Interleukin-8 Responses of Multi-Layer Gingival Epithelia to Subgingival Biofilms: Role of the “Red Complex” Species

Periodontitis is an infectious inflammatory disease that results in the destruction of the tooth-supporting (periodontal) tissues. The Gram-negative anaerobic species Porphyromonas gingivalis, Tannerella forsythia and Treponema denticola, (also known as the “red complex” species) are highly associated with subgingival biofilms at periodontitis-affected sites. A major chemokine produced by the gingival epithelium in response to biofilm challenge, is interleukin (IL)-8. The aim of this in vitro study was to investigate the relative effect of the “red complex” species as constituents of subgingival biofilms, on the regulation of IL-8 by gingival epithelia. Multi-layered organotypic human gingival epithelial cultures were challenged with a 10-species in vitro subgingival biofilm model, or its 7-species variant, excluding the “red complex”. IL-8 gene expression and secretion analyses were performed by qPCR and ELISA, respectively. After 3 h, both biofilms up-regulated IL-8 gene expression, but the presence of the “red complex” resulted in 3-fold greater response. IL-8 secretion was also up-regulated by both biofilms, with no differences between them. After 24 h, the 10-species biofilm reduced IL-8 secretion to 50% of the control, but this was not affected when the “red complex” was absent. In conclusion, as part of biofilms, “red complex” species differentially regulate IL-8 in gingival epithelia, potentially affecting the chemotactic responses of the tissue.

Impact of lead and mercuric ions on the interleukin-2-dependent proliferation and survival of T cells

Mercury and lead are widespread in the environment, causing chronic exposure of a large population to low concentrations of these metals. While several studies demonstrated that low levels of both metals affect the immune system, little is known about underlying molecular mechanisms. The objective of this study was to investigate the impact of mercuric (Hg(2+)) and lead (Pb(2+)) ions on T cells. Up to 100 μM Pb(NO(3))(2) had no effect on cellular viability and proliferation. In contrast, HgCl(2) caused a concentration-dependent decline of viable leukocytes and especially of activated T cells. Additionally, Hg(2+) induced reactive oxygen species (ROS) generation accompanied by the loss of mitochondrial transmembrane potential, measured by Dihydrorhodamine and Rhodamine-123, respectively. The antioxidant N-acetylcysteine partially reversed the toxic effects of Hg(2+), pointing to an involvement of ROS. The major cytokine controlling T-cell survival and proliferation is interleukin (IL)-2. Hg(2+) had no effect on the secretion of IL-2, but on IL-2 mediated signal transduction pathways, reducing phosphorylation of the downstream kinases ERK1/2 and AKT. Moreover, Hg(2+) led to an arrest of the cells in the S phase of the cell cycle. Taken together, these data fit a model in which Hg(2+) disrupts mitochondria, and the resulting release of ROS inhibits IL-2-dependent signal transduction, reducing proliferation and survival of T cells.

Synergistic Reversal of Type 1 Diabetes in NOD Mice With Anti-CD3 and Interleukin-1 Blockade

Inflammatory cytokines are involved in autoimmune diabetes: among the most prominent is interleukin (IL)-1β. We postulated that blockade of IL-1β would modulate the effects of anti-CD3 monoclonal antibody (mAb) in treating diabetes in NOD mice. To test this, we treated hyperglycemic NOD mice with F(ab′)2 fragments of anti-CD3 mAb with or without IL-1 receptor antagonist (IL-1RA), or anti–IL-1β mAb. We studied the reversal of diabetes and effects of treatment on the immune system. Mice that received a combination of anti-CD3 mAb with IL-1RA showed a more rapid rate of remission of diabetes than mice treated with anti-CD3 mAb or IL-1RA alone. Combination-treated mice had increased IL-5, IL-4, and interferon (IFN)-γ levels in circulation. There were reduced pathogenic NOD-relevant V7 peptide-V7+ T cells in the pancreatic lymph nodes. Their splenocytes secreted more IL-10, had increased arginase expression in macrophages and dendritic cells, and had delayed adoptive transfer of diabetes. After 1 month, there were increased concentrations of IgG1 isotype antibodies and reduced intrapancreatic expression of IFN-γ, IL-6, and IL-17 despite normal splenocyte cytokine secretion. These studies indicate that the combination of anti-CD3 mAb with IL-1RA is synergistic in reversal of diabetes through a combination of mechanisms. The combination causes persistent remission from islet inflammation.

Accurate and Simple Measurement of the Pro-inflammatory Cytokine IL-1&amp;beta; using a Whole Blood Stimulation Assay

Inflammatory processes resulting from the secretion of soluble mediators by immune cells, lead to various manifestations in skin, joints and other tissues as well as altered cytokine homeostasis. The innate immune system plays a crucial role in recognizing pathogens and other endogenous danger stimuli. One of the major cytokines released by innate immune cells is Interleukin (IL)-1. Therefore, we utilize a whole blood stimulation assay in order to measure the secretion of inflammatory cytokines and specifically of the pro-inflammatory cytokine IL-1β(1, 2, 3). Patients with genetic dysfunctions of the innate immune system causing autoinflammatory syndromes show an exaggerated release of mature IL-1β upon stimulation with LPS alone. In order to evaluate the innate immune component of patients who present with inflammatory-associated pathologies, we use a specific immunoassay to detect cellular immune responses to pathogen-associated molecular patterns (PAMPs), such as the gram-negative bacterial endotoxin, lipopolysaccharide (LPS). These PAMPs are recognized by pathogen recognition receptors (PRRs), which are found on the cells of the innate immune system (4, 5, 6, 7). A primary signal, LPS, in conjunction with a secondary signal, ATP, is necessary for the activation of the inflammasome, a multiprotein complex that processes pro-IL-1β to its mature, bioactive form (4, 5, 6, 8, 9, 10). The whole blood assay requires minimal sample manipulation to assess cytokine production when compared to other methods that require labor intensive isolation and culturing of specific cell populations. This method differs from other whole blood stimulation assays; rather than diluting samples with a ratio of RPMI media, we perform a white blood cell count directly from diluted whole blood and therefore, stimulate a known number of white blood cells in culture (2). The results of this particular whole blood assay demonstrate a novel technique useful in elucidating patient cohorts presenting with autoinflammatory pathophysiologies.

Lipopolysaccharide (LPS)-induced Interferon (IFN)-gamma Production by Decidual Mononuclear Cells (DMNC) is Interleukin (IL)-2 and IL-12 Dependent

Th1-shifted immune response is believed to be harmful for successful pregnancy because of activation of maternal cytotoxic T lymphocytes and natural killer cells. However, its effects on Toll-like receptor (TLR)-mediated innate immune response are so far unknown and this study has been undertaken to address the issue. decidual tissues were obtained from 16 pregnant women undergoing elective termination during the first trimester pregnancy for socioeconomic reasons. Decidual Mononuclear Cells (DMNC) were stimulated with suboptimal doses of IL-2 and IL-12 with/without LPS, considered to be a TLR4 ligand, for 48 hr. Productions of IFN-γ and tumor necrosis factor (TNF)-α in culture supernatant were measured with ELISA. (i) IFN-γ production was induced with LPS alone which was strongly up-regulated in the presence of IL-2 and IL-12. (ii) TNF-α was also induced by LPS but was not affected by the presence of IL-2 and IL-12. IL-2 and IL-12 up-regulated the production of IFN-γ in DMNC through increasing their susceptibility to LPS. TNF-α production is independent of such a mechanism.

Mast Cell Protease 6 Is Required for Allograft Tolerance

It has been shown that mast cells (MC) are absolutely required for transplant acceptance. However, only a few of the numerous mediators produced by MC have been proposed as potential mechanisms for the observed immunosuppression. The role of proteases in acquired immune tolerance as such has not yet been addressed. In this study, we have shown the requirement for MC protease 6 (MCP6), an MC-specific tryptase, to establish tolerance toward an allogeneic skin graft. The substrate for MCP6 is interleukin (IL)-6, cytokine generally considered to indicate transplant rejection. Herein we have shown an inverse correlation between MCP6 and IL-6. High expression of MCP6 is accompanied by low levels of IL-6 when the allograft is accepted, whereas low expression of MCP6 in combination with high levels of IL-6 are observed in rejecting grafts. Moreover, tolerance toward an allogeneic graft cannot be induced in MCP6 −/− mice. Rejection observed in these mice was comparable to that of MC-deficient hosts; it is T-cell mediated. These findings suggest that MCP6 actively depletes the local environment of IL-6 to maintain tolerance.