Interleukin-4 is a critical cytokine in contact sensitivity.

This study demonstrates an essential role for interleukin-4 (IL-4) in the delayed hypersensitivity reaction, as illustrated by contact sensitivity (CS) to trinitrochlorobenzene (TNCB). Injection of mice with monoclonal antibody to IL-4, but not with control antibody, reduced CS after active immunization by 75%, as judged by ear swelling. The histological alterations of CS were also reduced. IL-4 was essential to the effector stage, as inhibition of its production or action blocked the passive transfer of CS. In particular, treatment of immune lymph node cells with antisense oligonucleotide to IL-4 inhibited the systemic transfer of CS. Transfer was also inhibited by monoclonal antibody to IL-4 given to the recipient. The present results indicate that IL-4 is an essential cytokine at the effector stage of the CS reaction.     

Interleukin-27 as a negative regulator of human neutrophil function

Interleukin-27 (IL-27) is a novel cytokine of the IL-6/12 family with a broad range of immune regulation properties, which has been considered as a potential therapeutic agent for immune diseases and cancers. However, little is known about the effect of IL-27 on human neutrophils before its clinical administration. In this study, we investigated the effects of IL-27 on human neutrophil functions including adhesion, reactive oxygen species (ROS)/cytotoxic granule components production, inflammatory cytokines production, major histocompatibility complex (MHC) molecules expression and neutrophils' survival. We showed that IL-27 receptor complex, WSX-1/TCCR and gp130, is constitutively expressed on human neutrophils. In vitro, IL-27 suppressed neutrophil adhesion in response to fMLP, which might depend on the down-regulation of Mac-1. IL-27 also suppressed lipopolysaccharide-induced ROS production and attenuated cytotoxic granule components production in the cytoplasm of human neutrophils. In addition, IL-27 enhanced the production of IL-1β but not TNF-α from neutrophils. However, IL-27 failed to regulate the expression of MHC molecules and the survival of human neutrophils. In conclusion, our data demonstrate that IL-27 mainly down-modulates human neutrophil function, which might extend our understanding of the role of IL-27 in the innate immune response.     

Interleukin-8 induces neutrophil transendothelial migration.

Interleukin-8 (IL-8) is a potent neutrophil chemotactic stimulant. We have used chemically synthesized IL-8 to investigate its role in human neutrophil adhesion and transendothelial migration. IL-8 enhanced the adhesiveness of human neutrophils to plastic, and to both unstimulated and tumour necrosis factor (TNF)-stimulated endothelial monolayers in vitro. Using a two-compartment model separated by a confluent endothelial monolayer, we have shown that IL-8 chemotactic stimulation induced transmigration across the monolayer of up to 87.4 +/- 2.1% of added neutrophils (compared to random unstimulated transmigration of 2.2 +/- 0.7%), while chemokinetic stimulation led to transmigration of 21 +/- 3.8% of neutrophils. Preincubation of endothelium with TNF also induced transmigration in this model, and was additive when combined with an IL-8 chemotactic stimulus. Endothelial permeability was increased at maximal rates of chemotactic transmigration, which may correlate with increased permeability of vessels at inflammatory sites in vivo. The property of IL-8 to stimulate movement of neutrophils across endothelial monolayers in vitro supports the concept of a central role for this molecule in the accumulation of neutrophils at inflammatory lesions in vivo.     

6-trans-leukotriene B4 is a neutrophil chemotaxin in the guinea pig dermis.

The products of the 5- and 12-lipoxygenase (5-LO, 12-LO) pathways of arachidonic acid metabolism are implicated as proinflammatory mediators in a number of disease states. 12(R)-hydroxyeicosatetraenoic acid [12(R)-HETE] is present in large quantities in human psoriatic lesional skin and can be further metabolized by 5-LO to 5(S), 12(R)-dihydroxy-(6E,8Z,10E,14Z)-eicosatetraenoic acid (6-trans-LTB4). Furthermore, leukotriene B4 (LTB4) and the sulfidopeptide leukotrienes (LTC4, LTD4) can be transformed to 6-trans-LTB4. When injected into the guinea pig dermis, 6-trans-LTB4 (1.0, 10.0, 20.0 micrograms/intradermal site) caused a significant (P less than 0.02) infiltration of polymorphonuclear leukocytes (PMN) at 4 hr as assessed by histology and the levels of the PMN marker enzyme myeloperoxidase. 6-trans-LTB4 is a more potent PMN chemoattractant than 12(R)-HETE in the guinea pig dermis but is far less potent than LTB4. Pharmacological interdiction of leukotriene production or receptor binding should take into account the proinflammatory activity of 6-trans-LTB4.     

PLUNC is a secreted product of neutrophil granules

Airway epithelia and neutrophils are frequently recruited to release host defense factors in response to a variety of pulmonary pathogens. One abundant product of airway epithelia is palate, lung, nasal epithelium clone (PLUNC), a proposed innate immune protein expressed in submucosal glands and surface airway epithelia. In this study, we report the expression of PLUNC in human neutrophils, a previously unrecognized source of this protein. Immunoblots performed on polymorphonuclear cell (PMN) lysates and PMN subcellular fractions indicated that PLUNC was present in the specific granules of the neutrophil. Furthermore, secretion assays demonstrated that PLUNC protein was released by neutrophils upon stimulation with secretogogues, including formyl methionyl leucyl phenylalanine and the calcium ionophore A23187. Although recombinant PLUNC protein failed to exhibit antibacterial activity in our studies, its storage and secretion by a professional phagocytic cell support the hypothesis that PLUNC participates in an aspect of the inflammatory response that contributes to host defense. These studies suggest that PLUNC expression is less restricted than previously believed, and highlight new avenues of research for the study of PLUNC function.     

Interleukin-4 (IL-4) and IL-13 suppress excessive neutrophil infiltration and hepatocyte damage during acute murine schistosomiasis japonica

Due to the importance of neutrophils and proinflammatory cytokines in schistosomal liver damage, we analyzed the mechanisms underlying neutrophil and proinflammatory responses in murine schistosomiasis japonica. We found that granulomatous inflammation around parasite eggs in the liver was greater in Schistosoma japonicum-infected IL-4-/- IL-13-/- (double-knockout [DKO]) mice than in infected wild-type (WT) mice at 6 weeks, but not at 8 weeks, postinfection, suggesting the importance of Th2 responses in these typical hepatic lesions. Infected DKO mice also showed increased neutrophil infiltration accompanying more severe pathology, as shown by the enhanced necrosis of hepatocytes. This was not likely due to a Th1/Th2 imbalance, because there was no detectable increase in gamma interferon (IFN-γ) production in these DKO mice. mRNA expression of interleukin-17A (IL-17A), proinflammatory cytokines, and the neutrophil chemoattractant CXCL2 in liver was higher in infected DKO mice than in WT mice. However, in IL-4-/- IL-13-/- IL-17A-/- (triple-knockout [TKO]) mice, the absence of IL-17A was associated with only marginal differences in schistosomal liver damage, suggesting that IL-17A is only partially responsible for neutrophil-driven hepatic damage. Furthermore, the expression of mRNAs encoding proinflammatory cytokines was not under the control of IL-17A in TKO mice. These findings indicate that IL-4 and IL-13 suppress excessive neutrophil recruitment, proinflammatory cytokine production, and hepatic damage during the acute stage of S. japonicum infection, suggesting that neutrophils and proinflammatory cytokines are mainly responsible for hepatocyte damage during acute murine schistosomiasis japonica. However, neutrophil induction and the production of proinflammatory cytokines were not due solely to IL-17A.     

Platelet-activating factor (PAF-acether), an activator of neutrophil functions

The effect of totally synthetic PAF-acether (1-O-octadecyl-2-O-acetyl-sn-glyceryl-3-phosphorylcholine), 2-lyso PAF-acether (1-O-octadecyl-sn-glyceryl-3-phosphorylcholine) and lyso-phosphatidylcholine on enzyme release and superoxide production from human polymorphonuclear neutrophils (PMN were studied. PMN (2×10⁶ ml^–1) were incubated at 37°C with various concentrations of phospholipids in the absence of cytochalasin B. At 10^–7 M, PAF-acether induced superoxide production and -glucuronidase, acid phosphatase and lysosyme release, but not that of cytoplasmic lactic dehydrogenase. In the same condition 2-lyso PAF-acether and lyso-phosphatidylcholine were ineffective. In the presence of phagocytic stimuli PAF-acether enhanced in the range from 10^–7 M to 10^–10 M the enzyme release and only at 10^–7 M the superoxide production. Thus, the capacity of PAF-acether to stimulate PMN, as well as platelet function, indicates a prominent role for this lipid mediator in inflammatory processes.     

Interleukin-8 (IL-8) is a major neutrophil chemotaxin from human alveolar macrophages stimulated with staphylococcal enterotoxin A (SEA)

Since Staphylococcus aureus is an important human pathogen, and infection of the lungs in characterized by neutrophil infiltration we studied the role of a staphylococcal toxin, enterotoxin A (SEA) on the synthesis and secretion of IL-8 by human alveolar macrophages. As SEA concentration was increased, the IL-8 accumulation in the macrophage conditioned medium increased. The concentration of mRNA encoding IL-8 was also elevated in the macrophage in response to increases in SEA concentration. Although the monocytic cell line U937 was able to respond to SEA and secrete IL-8, treatment with PMA prior to SEA stimulation increased the IL-8 accumulation around fifty fold indicating that maturation of the undifferentiated cell to a more macrophage-like cell facilitated IL-8 accumulation. Stimulating human alveolar macrophages with high concentrations of SEA caused an increase in IL-1 accumulation. However, when the cells were incubated with SEA in the presence of IL-1 receptor antagonist, there was no decrease in IL-8 accumulation. Addition of a neutralizing anti-IL-8 monoclonal antibody to the culture medium of SEA-stimulated macrophages significantly reduced the neutrophil chemotactic activity of the medium. These studies showed that IL-8 is a major neutrophil chemotaxin from human alveolar macrophages stimulated with SEA.accepted by I. Ahnfelt-Rønne     

Interleukin-4 and listeriosis

Summary: Experimental infection of mice with Listeria monocytogenes (L. monocytogenes) has served as an appropriate model for analyzing Thl nil driven immune responses. Generally, Th2 responses are absent and IL-4 is not detectable. Here, we describe experimental settings under which IL-4 is detectable in listeriosis. Our data suggest that IL-4 is rapidly produced after infection. This prompt IL-4 burst seems to stimulate chemokine responses and, therefore, may participate in the regulation of the early antilisterial host response. Soon thereafter, lL-4 production wanes. At least partially this seems to be caused by downregulation of IL-4–producing CD4+ NK1+ TCRαβ^int lymphocytes by IL-12. In the absence of IFN-γ responsiveness, IL-4 production is demonstrable during acquired immunity against L. monocytogenes, and this elevated IL-4 production apparently contributes to disease exacerbation. In conclusion, the data are consistent with a detrimental role of IL-4 in listeriosis and active control of IL-4 synthesis by the antilisterial immune response. The rapid, but transient, IL-4 burst in listeriosis probably contributes to host defense without impairing development of the acquired T-cell response because of its shortness.     

Interleukin-6 mediates neutrophil mobilization from bone marrow in pulmonary hypertension

Myeloid cells, such as neutrophils, are produced in the bone marrow in high quantities and are important in the pathogenesis of vascular diseases such as pulmonary hypertension (PH). Although neutrophil recruitment into sites of inflammation has been well studied, the mechanisms of neutrophil egress from the bone marrow are not well understood. Using computational flow cytometry, we observed increased neutrophils in the lungs of patients and mice with PH. Moreover, we found elevated levels of IL-6 in the blood and lungs of patients and mice with PH. We observed that transgenic mice overexpressing Il-6 in the lungs displayed elevated neutrophil egress from the bone marrow and exaggerated neutrophil recruitment to the lungs, resulting in exacerbated pulmonary vascular remodeling, and dysfunctional hemodynamics. Mechanistically, we found that IL-6-induced neutrophil egress from the bone marrow was dependent on interferon regulatory factor 4 (IRF-4)-mediated CX₃CR1 expression in neutrophils. Consequently, Cx₃cr1 genetic deficiency in hematopoietic cells in Il-6-transgenic mice significantly reduced neutrophil egress from bone marrow and decreased neutrophil counts in the lungs, thus ameliorating pulmonary remodeling and hemodynamics. In summary, these findings define a novel mechanism of IL-6-induced neutrophil egress from the bone marrow and reveal a new therapeutic target to curtail neutrophil-mediated inflammation in pulmonary vascular disease.     

Interleukin-4 and Interleukin-4 Receptor Alpha Gene Polymorphisms in Recurrent Aphthous Stomatitis

ABSTRACT

Background: Recurrent aphthous stomatitis (RAS) is a common oral condition with a major impact on the quality of life. The condition is thought to be due to the overexpression of T helper-1(Th1)-related cytokines. Since interleukin-4 (IL-4) and its receptor (IL-4Rα) are antagonistic to Th-1 pathways, polymorphisms in their genes may also be involved in the pathogenesis of aphthous stomatitis.

Methods: Sixty-four patients diagnosed with minor RAS and 141 (age- and sex-matched) healthy controls were assessed for 3 single-nucleotide polymorphisms (SNPs) within the promoter region of the IL-4 gene (?1098G/T, ?590C/T, and ?33C/T), and 1 SNP in IL-4Rα gene (+1902 A/G).

Results: No significant differences were detected between the patient and the control group regarding IL-4 allele frequencies. However, the patient group demonstrated a higher frequency of IL-4 ?590 CC genotype and a lower rate of IL-4 ?590 TC genotype.

The TCT, GTT, GCT, and GTC haplotypes of the IL-4 gene (?1098, ?590, ?33) were significantly more frequent in the patients and the GCC, and TTT haplotypes were more common in healthy controls. No significant differences were found in IL-4Rα gene polymorphism between the 2 groups.

Conclusions: Certain polymorphisms of IL4 gene could predispose individuals to RAS.     

Intact proteoglycan is a polyclonal activator of murine B-lymphocytes

Destruction of articular cartilage is the hallmark of both inflammatory and degenerative arthritides. Since degradation of cartilage results in the release of proteoglycan (PG) monomers and fragments into the synovial fluid, the present study was initiated to determine whether hyaline cartilage PG can induce a cellular immune response. Nonimmune spleen cells obtained from A/St and C57Bl/6 mice were cultured with a crude extract of bovine nasal cartilage, a PG aggregate fraction, PG monomer or degraded PG monomer for varying time periods. Only intact PG monomer induced a proliferative response which peaked at day 2. The responding cell was a B-lymphocyte which did not require T-helper cell activity. Our results suggest that intact PG monomer is a polyclonal activator of B-lymphocytes.     

Interleukin-10 is an unequivocal Th2 parameter in the rat, whereas interleukin-4 is not

Exposure of Wistar rats to the immunotoxic compounds hexachlorobenzene (HCB), bis(tri-n-butyltin)oxide, and benzo(a)pyrene was previously found to affect mRNA expression of interleukin (IL)-2, IL-2R alpha-chain, and interferon (IFN)-gamma, the prototypic Th1 cytokine. In contrast, the mRNA expression of IL-4, the prototypic Th2 cytokine, was unaffected. This latter finding suggested that the IL-4 mRNA expression may not be an unequivocal parameter for Th2 responses in the rat. In order to obtain such a parameter the present study was performed, consisting of two types of experiments. Expression and production of IL-4 as well as IL-10, a second Th2 cytokine, were measured. First, Lewis (Th1 prone) and Brown Norway (BN; Th2 prone) rats were exposed to HCB. Exposure was previously found to increase the serum immunoglobulin (Ig)E levels, an IL-4-dependent response, in BN but not Lewis rats, and in Lewis rats to aggravate experimental allergic encephalomyelitis (EAE), severity being inversely related to IL-10 levels. Secondly, BN rats were infected with Trichinella spiralis, an infection previously found to induce IL-4 production. HCB exposure did not affect IL-4 mRNA expression in either strain, while IL-4 production was decreased in Lewis and unaffected in BN rats. In Lewis rats both the mRNA expression and the production of IL-10 were decreased. The T. spiralis infection induced IL-4 and IL-10 mRNA expression, as well as IL-10 production. In contrast, the IL-4 production was strongly reduced. Thus, both the IL-10 mRNA expression and production correlated with the EAE development and T. spiralis infection. In HCB exposed Lewis rats and T. spiralis infected BN rats the IL-4 mRNA expression correlated with IgE levels and T. spiralis infection, respectively, whereas the IL-4 production lacked correlation in all cases. Collectively, these results suggest that IL-10 is an unequivocal Th2 parameter in the rat, whereas IL-4 is not.     

Interleukin-8 is a major neutrophil chemotactic factor derived from cultured human gingival fibroblasts stimulated with interleukin-1 beta or tumor necrosis factor alpha.

Inflammatory mediators produced by cells in the gingiva have been implicated in the initiation and progression of periodontal disease, a common infectious disease. In this study, we examined the biological activity of neutrophil chemotactic factors and the kinetics of expression of interleukin-8 (IL-8) mRNA derived from normal gingival fibroblasts in response to inflammatory mediators in an in vitro model. Gingival fibroblasts stimulated by either recombinant human interleukin-1 beta or recombinant human tumor necrosis factor alpha produced neutrophil chemotactic factors after 4 h, whereas expression of cell-derived IL-8 mRNA was detected within 1 h after stimulation. Furthermore, in a neutralization assay, rabbit anti-recombinant human IL-8 antiserum inhibited neutrophil chemotactic activity to basal levels. These results provide evidence that gingival fibroblasts synthesize potent chemotactic factors such as IL-8 in the presence of the inflammatory mediators interleukin-1 beta and tumor necrosis factor alpha. The activity of these factors may contribute to neutrophil-mediated processes in the pathogenesis of periodontal disease.     

Microbicidal protein psoriasin is a multifunctional modulator of neutrophil activation

As effector cells in host defence, neutrophils actively destroy invading microorganisms via a potent antimicrobial arsenal composed of oxidants and antimicrobial peptides. Psoriasin, an Escherichia coli-cidal antimicrobial protein, has been found to be overexpressed in psoriasis, a skin disease characterized by infiltration of neutrophils. In addition to its microbicidal activities and chemotaxis of neutrophils reported previously, we hypothesized that psoriasin might regulate other neutrophil functions such as cytokine and chemokine production, reactive oxygen species generation, and release of antimicrobial peptides. In the current study, we demonstrate that psoriasin activates neutrophils to produce a range of cytokines and chemokines including interleukin-6 (IL-6), IL-8/CXCL8, tumour necrosis factor-alpha, macrophage inflammatory protein-1alpha (MIP-1alpha)/CCL3, MIP-1beta/CCL4 and MIP-3alpha/CCL20. Furthermore, psoriasin induces phosphorylation of mitogen-activated protein kinase p38 and extracellular signal-regulated kinase (ERK), but not c-Jun N-terminal kinase (JNK), both of which are required for the production of cytokines and chemokines as evidenced by the inhibitory effects of p38 and ERK inhibitors on psoriasin-mediated neutrophil activation. Moreover, psoriasin stimulates the generation of reactive oxygen species from neutrophils, most likely via nicotinamide adenine dinucleotide phosphate oxidase activation. Finally, we demonstrate that psoriasin enhances messenger RNA expression of alpha-defensins, termed human neutrophil peptides (HNP) 1 to 3, and induces their extracellular release. Besides its antimicrobial properties, therefore, psoriasin may contribute to innate immunity through enhancing neutrophil host defence functions at sites of inflammation or infection.     

Interleukin-8 is a prognostic indicator in human hilar cholangiocarcinoma

Interleukin-8 (IL-8), matrix metalloproteinase-9 (MMP-9) and neovascularization have been implicated to be associated with biological processes, especially cancer progression. However, few studies have investigated the role of IL-8 in human hilar cholangiocarcinoma. In this study we detected the expression of IL-8 combined with MMP-9 and microvessel density (MVD) in hilar cholangiocarcinoma to evaluate their clinicopathological significance and prognostic value. A total of 62 patients with hilar cholangiocarcinoma who underwent curative surgery were enrolled in this study. The expression of IL-8, MMP-9 and MVD were examined immunohistochemically. The correlation of IL-8 with MMP-9 expression, MVD, clinicopathological features and survival time of patients were then analyzed. Expression of IL-8 was observed in 56.5% tumors, which was related to advanced TNM stage (P = 0.026) and tumor recurrence (P = 0.018). IL-8 had a positive correlation with MMP-9 expression and MVD. Furthermore, patients with high IL-8 expression had a significantly shorter overall survival than those with low IL-8 expression (P = 0.01). Multivariate analysis confirmed IL-8 as an independent prognostic factor (P = 0.005). In conclusion, IL-8 expression significantly correlated with MMP-9 expression and MVD, and IL-8 was a valuable prognostic factor for human hilar cholangiocarcinoma.