Interleukin-8 (IL-8) is a pro-inflammatory cytokine which exerts its effects via binding to 2 receptors, CXCR1 and CXCR2 and is known to promote angiogenesis, mitogenesis and motogenesis in cancer. IL-8 is over expressed in endometrial carcinoma, but the expression of CXCR1 and CXCR2 in endometrial carcinoma has not been previously investigated. The aim of this study was to determine the expression of IL-8 receptors in endometrial carcinoma. The expression of CXCR1 and CXCR2 was studied in endometrial carcinomas and normal endometrium by immunohistochemistry in 101 tumours. IL-8 and IL-8 receptor expression was also studied by Real-time quantitative PCR (RT-qPCR) in 17 tumours in comparison to normal endometrium. The expression profile was correlated to the clinico-pathological features of the tumours. Immunohistochemistry showed CXCR1 and CXCR2 were expressed in all cases of endometrial carcinoma, with CXCR1 showing stronger expression. There was a statistically significant correlation between CXCR2 staining intensity and tumour grade (P=0.012) and disease free survival (P=0.015) independently. On RT-qPCR, 14/17, 15/17 and 16/17 tumours showed significant increase in IL-8, CXCR1 and CXCR2 expression levels in comparison to normal endometrium, with median fold increase of 42-fold, 51-fold and 27-fold, respectively. This is the first report of the expression of IL-8 receptors in endometrial carcinoma and the results show an association between IL-8 and IL-8 receptors and the pathogenesis of endometrial carcinoma, and represent potential prognostic biomarkers and therapeutic targets.

Fabry disease (FD) is an X-linked lysosome storage disease that results in the accumulation of globotriaosylceramide (Gb3) throughout the body leading to irreversible target organ damage. As the role of secondary mediators (inflammatory molecules) and their mechanisms has not been fully elucidated, we focused on the interleukin (IL)-6 system in adult FD patients and in matched healthy subjects. To obtain insights into the complex regulation of IL-6 actions, we used a novel approach that integrates information from plasma and exosomes of FD patients (n = 20) and of healthy controls (n = 15). Soluble IL-6 receptor (sIL-6R) levels were measured in plasma with the ELISA method, and membrane-bound IL-6R was quantified in plasma and urinary exosomes using flow cytometry. In FD patients, the levels of soluble IL-6R in plasma were higher than in control subjects (28.0 ± 5.4 ng/mL vs. 18.9 ± 5.4 ng/mL, p < 0.0001); they were also higher in FD subjects with the classical form as compared to those with the late-onset form of the disease (36.0 ± 11.4 ng/mL vs. 26.1 ± 4.5 ng/mL, p < 0.0001). The percentage of urinary exosomes positive for IL-6R was slightly lower in FD (97 ± 1 vs. 100 ± 0% of events positive for IL-6R, p < 0.05); plasma IL-6 levels were not increased. These results suggest a potential role of IL-6 in triggering the inflammatory response in FD. As in FD patients only the levels of sIL-6Rs are consistently higher than in healthy controls, the IL-6 pathogenic signal seems to prevail over the homeostatic one, suggesting a potential mechanism causing multi-systemic damage in FD.

Since recent work has identified an apoptotic pathway in sympathetic neurons that is mediated by autocrine interleukin-1 (IL-1), we investigated whether cultured sympathetic neurons possess functional IL-1 receptors. Cultured sympathetic neurons express levels of IL-1RI and IL-1RAcP mRNAs consistent with signal transduction. Neurons stimulated with IL-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 IL-1, including a member of the reticulon family. We conclude that IL-1 stimulates a potential component of a neuronal secretory pathway.

Interleukin (IL)-17A, a pro-inflammatory cytokine that is linked to the pathology of several inflammatory diseases, has been shown to be upregulated in early human tendinopathy and to mediate inflammatory and tissue remodelling events. However, it remains unclear which cells in tendons can respond to IL-17A, and how IL-17A, and its family members IL-17F and IL-17AF, can affect intracellular signalling activation and mRNA expression in healthy and diseased tendon-derived fibroblasts. Using well-phenotyped human tendon samples, we show that IL-17A and its receptors IL-17RA and IL-17RC are present in healthy hamstring, and tendinopathic and torn supraspinatus tendon tissue. Next, we investigated the effects of IL-17A, IL-17F, or IL-17AF on cultured patient-derived healthy and diseased tendon-derived fibroblasts. In these experiments, IL-17A treatment significantly upregulated IL6, MMP3, and PDPN mRNA expression in diseased tendon-derived fibroblasts. IL-17AF treatment induced moderate increases in these target genes, while little change was observed with IL-17F. These trends were reflected in the activation of intracellular signalling proteins p38 and NF- κ B p65, which were significantly increased by IL-17A, modestly increased by IL-17AF, and not increased by IL-17F. In combination with TNF-α, all three IL-17 cytokines induced IL6 and MMP3 mRNA expression to similar levels. Therefore, this study confirms that healthy and diseased tendon-derived fibroblasts are responsive to IL-17 cytokines and that IL-17A induces the most profound intracellular signalling activation and mRNA expression of inflammatory genes, followed by IL-17AF, and finally IL-17F. The ability of IL-17 cytokines to induce a direct response and activate diverse pro-inflammatory signalling pathways through synergy with other inflammatory mediators suggests a role for IL-17 family members as amplifiers of tendon inflammation and as potential therapeutic targets in tendinopathy.

Chronic pain is a major comorbidity of chronic inflammatory diseases. Here, we report that the cytokine IL-1β, which is abundantly produced during multiple sclerosis (MS), arthritis (RA), and osteoarthritis (OA) both in humans and in animal models, drives pain associated with these diseases. We found that the type 1 IL-1 receptor (IL-1R1) is highly expressed in the mouse and human by a subpopulation of TRPV1+ dorsal root ganglion neurons specialized in detecting painful stimuli, termed nociceptors. Strikingly, deletion of the Il1r1 gene specifically in TRPV1+ nociceptors prevented the development of mechanical allodynia without affecting clinical signs and disease progression in mice with experimental autoimmune encephalomyelitis and K/BxN serum transfer-induced RA. Conditional restoration of IL-1R1 expression in nociceptors of IL-1R1-knockout mice induced pain behavior but did not affect joint damage in monosodium iodoacetate-induced OA. Collectively, these data reveal that neuronal IL-1R1 signaling mediates pain, uncovering the potential benefit of anti-IL-1 therapies for pain management in patients with chronic inflammatory diseases.

Myelin was previously shown to possess neurotransmitter and cytokine receptors that trigger well-defined signaling mechanisms within the multilamellar structure. The present study reveals the presence of an interleukin-2 (IL-2) receptor in isolated mouse CNS myelin that responds to recombinant mouse IL-2 by activating diacylglycerol kinase (DAGK) and phosphoinositide 3-kinase (PI3K); additional evidence suggests participation by protein tyrosine kinase. Activation of myelin DAGK by IL-2 occurred in brain stem tissue mince and was blocked by chelerythrin chloride, indicating an essential role for myelin-localized protein kinase C. Two inhibitors of PI3K, wortmannin and LY294002, blocked endogenous PI3K as well as that enhanced by IL-2. Activation of PI3K by IL-2 was also blocked by tyrphostin A25, a selective inhibitor of PTK, suggesting activation of the latter by IL-2 is upstream to PI3K activation. This reaction resulted in tyrosine phosphorylation of a protein tentatively identified as the p85 subunit of PI3K. Developmental changes were noted in that receptor density and signaling activity were robust during the period of rapid myelination and declined rapidly thereafter.

Decreased thyroidal hormone production is found during lipopolysaccharide (LPS)-induced endotoxic shock in animals as well as in critically ill patients. Here we studied the role of the thyroid hormone receptors (TRs) in activation of STAT3, NF-κB and ERK, which play a key role in the response to inflammatory cytokines during sepsis. TR knockout mice showed down-regulation of hepatic inflammatory mediators, including interleukin 6 (IL-6) in response to LPS. Paradoxically, STAT3 and ERK activity were higher, suggesting that TRs could act as endogenous repressors of these pathways. Furthermore, hyperthyroidism increased cytokine production and mortality in response to LPS, despite decreasing hepatic STAT3 and ERK activity. This suggested that TRs could directly repress the response of the cells to inflammatory mediators. Indeed, we found that the thyroid hormone T3 suppresses IL-6 signalling in macrophages and hepatocarcinoma cells, inhibiting STAT3 activation. Consequently, the hormone strongly antagonizes IL-6-stimulated gene transcription, reducing STAT3 recruitment and histone acetylation at IL-6 target promoters. In conclusion, TRs are potent regulators of inflammatory responses and immune homeostasis during sepsis. Reduced responses to IL-6 should serve as a negative feedback mechanism for preventing deleterious effects of excessive hormone signaling during infections.

Soluble interleukin 2 receptor (SIL-2R) and interleukin 1-B (IL-1B) levels in peripheral blood were assayed by enzyme-linked immunoabsorbent method in 25 patients with Behçet's disease and in 20 healthy controls. Eighteen patients suffered from active Behçet's disease and 6 patients had severe disease manifestations. SIL-2R levels were followed for a period of 16 months and were correlated with disease activity. In all the patients with Behçet's disease the levels of SIL-2R and IL-1B were significantly higher than their control, 958 +/- 712 vs. 404 +/- 119 u/ml (P < 0.001) for SIL-2R and 639 +/- 741 vs. 59 +/- 53 pg/ml (P < 0.001) for IL-1B. No correlation was found between disease activity and severity and the levels of SIL-2R and IL-1B. It is suggested that levels of SIL-2R and IL-1B are elevated in patients with Behçet's disease but do not serve as disease activity markers.

Inflammatory response and cytokine activation are markedly stimulated in skeletal muscle during various conditions. Interleukin-6 (IL-6), a pro-inflammatory cytokine, has pleiotropic effects on skeletal muscle. Adenosine, released by all cell types, binds to a class of G protein-coupled receptors to induce various skeletal muscle effects. The aim of this work was to investigate whether activation of adenosine receptors, particularly adenosine A2B receptors, could stimulate IL-6 gene expression in rat L6 skeletal muscle cells.

Sickness responses to lipopolysaccharide (LPS) were examined in mice with deletion of the interleukin (IL)-1 type 1 receptor (IL-1R1). IL-1R1 knockout (KO) mice displayed intact anorexia and HPA-axis activation to intraperitoneally injected LPS (anorexia: 10 or 120µg/kg; HPA-axis: 120µg/kg), but showed attenuated but not extinguished fever (120µg/kg). Brain PGE2 synthesis was attenuated, but Cox-2 induction remained intact. Neither the tumor necrosis factor-α (TNFα) inhibitor etanercept nor the IL-6 receptor antibody tocilizumab abolished the LPS induced fever in IL-1R1 KO mice. Deletion of IL-1R1 specifically in brain endothelial cells attenuated the LPS induced fever, but only during the late, 3rd phase of fever, whereas deletion of IL-1R1 on neural cells or on peripheral nerves had little or no effect on the febrile response. We conclude that while IL-1 signaling is not critical for LPS induced anorexia or stress hormone release, IL-1R1, expressed on brain endothelial cells, contributes to the febrile response to LPS. However, also in the absence of IL-1R1, LPS evokes a febrile response, although this is attenuated. This remaining fever seems not to be mediated by IL-6 receptors or TNFα, but by some yet unidentified pyrogenic factor.

Interleukin-6 (IL-6) is a cytokine implicated in proinflammatory as well as regenerative processes and acts via receptor complexes consisting of the ubiquitously expressed, signal-transducing receptor gp130 and the IL-6 receptor (IL-6R). The IL-6R is expressed only on hepatocytes and subsets of leukocytes, where it mediates specificity of the receptor complex to IL-6 as the subunit gp130 is shared with all other members of the IL-6 cytokine family such as IL-11 or IL-27. The amount of IL-6R at the cell surface thus determines the responsiveness of the cell to the cytokine and might therefore be decisive in the development of inflammatory disorders. However, how the expression levels of IL-6R and gp130 at the cell surface are controlled is largely unknown. Here, we show that IL-6R and gp130 are constitutively internalized independent of IL-6. This process depends on dynamin and clathrin and is temporally controlled by motifs within the intracellular region of gp130 and IL-6R. IL-6 binding and internalization of the receptors is a prerequisite for activation of the Jak/STAT signaling cascade. Targeting of gp130, but not of the IL-6R, to the lysosome for degradation depends on stimulation with IL-6. Furthermore, we show that after internalization and activation of signaling, both the IL-6R and gp130 are recycled back to the cell surface, a process that is enhanced by IL-6. These data reveal an important function of IL-6 beyond the pure activation of signaling.

Microglia sense their environment using an array of membrane receptors. While P2Y12 receptors are known to play a key role in targeting directed motility of microglial processes to sites of damage where ATP/ADP is released, little is known about the role of P2Y13 , which transcriptome data suggest is the second most expressed neurotransmitter receptor in microglia. We show that, in patch-clamp recordings in acute brain slices from mice lacking P2Y13 receptors, the THIK-1 K+ current density evoked by ADP activating P2Y12 receptors was increased by ~50%. This increase suggested that the P2Y12 -dependent chemotaxis response should be potentiated; however, the time needed for P2Y12 -mediated convergence of microglial processes onto an ADP-filled pipette or to a laser ablation was longer in the P2Y13 KO. Anatomical analysis showed that the density of microglia was unchanged, but that they were less ramified with a shorter process length in the P2Y13 KO. Thus, chemotactic processes had to grow further and so arrived later at the target, and brain surveillance was reduced by ~30% in the knock-out. Blocking P2Y12 receptors in brain slices from P2Y13 KO mice did not affect surveillance, demonstrating that tonic activation of these high-affinity receptors is not needed for surveillance. Strikingly, baseline interleukin-1β release was increased fivefold while release evoked by LPS and ATP was not affected in the P2Y13 KO, and microglia in intact P2Y13 KO brains were not detectably activated. Thus, P2Y13 receptors play a role different from that of their close relative P2Y12 in regulating microglial morphology and function.

The spread of cancer from organ to organ (metastasis) is responsible for the vast majority of cancer deaths; however, most current anti-cancer drugs are designed to arrest or reverse tumor growth without directly addressing disease spread. It was recently discovered that tumor cell-secreted interleukin-6 (IL-6) and interleukin-8 (IL-8) synergize to enhance cancer metastasis in a cell-density dependent manner, and blockade of the IL-6 and IL-8 receptors (IL-6R and IL-8R) with a novel bispecific antibody, BS1, significantly reduced metastatic burden in multiple preclinical mouse models of cancer. Bispecific antibodies (BsAbs), which combine two different antigen-binding sites into one molecule, are a promising modality for drug development due to their enhanced avidity and dual targeting effects. However, while BsAbs have tremendous therapeutic potential, elucidating the mechanisms underlying their binding and inhibition will be critical for maximizing the efficacy of new BsAb treatments. Here, we describe a quantitative, computational model of the BS1 BsAb, exhibiting how modeling multivalent binding provides key insights into antibody affinity and avidity effects and can guide therapeutic design. We present detailed simulations of the monovalent and bivalent binding interactions between different antibody constructs and the IL-6 and IL-8 receptors to establish how antibody properties and system conditions impact the formation of binary (antibody-receptor) and ternary (receptor-antibody-receptor) complexes. Model results demonstrate how the balance of these complex types drives receptor inhibition, providing important and generalizable predictions for effective therapeutic design.

Soluble interleukin 2 receptors (sIL-2R) exert a potential role in immunoregulation. We investigated the in vitro effects of sIL-2R on several interleukin 2 (IL-2)-dependent cellular events. Cytotoxicity of human rIL-2-stimulated PBMC against K562 and Daudi was correlated inversely to the concentration of sIL-2R in the culture medium during rIL-2 stimulation. sIL-2R concentrations higher than 4.0 pM produced a significant loss of cytotoxicity (P < 0.01). The effect of different sIL-2R concentrations added to cultured human PBMC on secondary sIL-2R production was tested by ELISA. Secondary sIL-2R production was abrogated by high initial sIL-2R dosages whereas low initial dosages were followed by a continuing production of secondary sIL-2R after five days of culture. Proliferation of the IL-2-dependent mouse cell line CTLL-2-was suppressed by sIL-2R added to the culture medium in a dose-dependent way. The neutralizing capacity of sIL-2R strongly depended on the initial number of CTLL set in per proliferation assay. In contrast, variation of rIL-2-concentration had no significant effect on reduction of proliferation by sIL-2R. Furthermore, preincubation of sIL-2R with rIL-2 did not enhance growth suppression. These last findings indicate that there is at least no functional interaction between sIL-2R and free IL-2, whereas an interaction of sIL-2R with the membrane-bound receptor for IL-2 seems possible.

Interleukin-10 (IL-10) is a cytokine whose main biological function is to suppress the immune response by induction of a signal(s) leading to inhibition of synthesis of a number of cytokines and their cellular receptors. Signal transduction is initiated upon formation of a ternary complex of IL-10 with two of its receptor chains, IL-10R1 and IL-10R2, expressed on the cell membrane. The affinity of IL-10R1 toward IL-10 is very high, which allowed determination of the crystal structure of IL-10 complexed with the extracellular/soluble domain of IL-10R1, while the affinity of IL-10R2 toward either IL-10 or IL-10/sIL-10R1 complex is quite low. This so far has prevented any attempts to obtain structural information about the ternary complex of IL-10 with its receptor chains.

The interleukin (IL) -1 family members play an important role in regulating inflammatory responses and their functions are mediated by a group of receptors consisting of immunoglobulin and Toll/IL-1 receptor (TIR) domains. In humans, 10 IL-1Rs are found. In this study, 5 IL-1 receptors including IL-1R3/IL-1RAcP, IL-1R8/SIGIRR, IL-1R9a/IL-1RAcPL1a, IL-1R9b/IL-1RAcPL1b and IL-1R10/IL-1RAcPL2 were identified in grass carp (Ctenopharyngodon idella). Phylogenetic analysis reveals that the IL-1R9a/IL-1RAcPL1a and IL-1R9b/IL-1RAcPL1b share significantly high sequence similarity and are believed to have been duplicated from the same gene prior to the radiation of teleosts. Further, these two receptors closely relate to the IL-1R10/IL-1RAcPL2, suggesting that they may have evolved from a common ancestor. The IL-1R3/IL-1RAcP, IL-1R9a/IL-1RAcPL1a, IL-1R9b/IL-1RAcPL1b and IL-1R10/IL-1RAcPL2 are highly expressed in the brain. Stimulation of primary spleen leucocytes by LPS and intraperitoneal injection of fish with poly (I:C) or bacterial infection results in significant increases of IL-1R3/IL-1RAcP expression. Interestingly, the IL-1R8/SIGIRR and IL-1R10/IL-1RAcPL2 showed similar expression patterns.

Interleukin (IL)-4 and -13 are structurally and functionally related cytokines sharing common receptor subunits. They regulate immune responses and, moreover, are involved in the pathogenesis of a variety of human neoplasms. Three different receptors have been described for IL-4, but only IL-4 receptor type II (IL-4Rα/IL-13Rα1) is expressed in solid tumors. While IL-13 can also bind to three different receptors, IL-13 receptor type I (IL-4Rα/IL-13Rα1/IL-13Rα2) and type II (IL-4Rα/IL-13Rα1) are expressed in solid tumors. After receptor binding, IL-4 and IL-13 can mediate tumor cell proliferation, survival, and metastasis in gastric or colon cancer. This review summarizes the results about the role of IL-4/IL-13 and their receptors in gastric and colon cancer.

Interleukin-6 (IL-6) is one of the most pleiotropic cytokines due to its importance in both innate and adaptive immune responses and other physiological processes. In this study, we identified the zebrafish (Danio rerio) IL-6 homologue by investigating the synteny between the human (Homo sapiens), the fugu (Takifugu rubripes) and the zebrafish genome. Although zebrafish IL-6 showed a low sequence homology with other IL-6 sequences in other species, it presented a high structural similarity to human IL-6. We also analysed IL-6 expression in several different tissues, along with analysis of the expression of the genes that form the IL-6 receptor complex, IL-6R and gp130. After treatment with bacterial or viral stimuli, zebrafish IL-6 expression was modulated in a manner similar to that of other proinflammatory molecules, such as IL-1β and TNF-α. The expression of IL-6, IL-6R and gp130 was also studied during the ontogeny of zebrafish larvae using quantitative PCR and in situ hybridisation. Our results indicated that the transcripts were detected very early, increased during the first week of life and were predominantly expressed in the head, epidermis and neuromasts of the anterior and posterior lateral line system, suggesting their involvement in the normal development of these tissues.

Using human osteoblastic SaM-1 cells, we investigated the effects of lysophosphatidic acid (LPA) on the production of interleukin (IL)-6 and IL-8, molecules which are capable of stimulating the development of osteoclasts from their haematopoietic precursors, and examined the signal transduction systems involved in their effect on these cells. These human osteoblasts constitutively expressed endothelial differentiation genes (Edg)-2 and Edg-4, which are LPA receptors. LPA increased gene and protein expression of IL-6 and IL-8 in SaM-1 cells. The expression of IL-6 and IL-8 mRNAs was maximal at 1-3h, and the increase in IL-6 and IL-8 synthesis in response to lysophosphatidic acid (1-10 microM) occurred in a concentration-dependent manner. These increases were blocked by Ki16425, an Edg-2/7 antagonist. In addition, LPA caused an increase in the intracellular Ca(2+) concentration ([Ca(2+)](i)), which was inhibited by pretreatment with Ki16425 or 2-aminoethoxy-diphenylborate (2-APB), an inositol 1,4,5-triphosphate (IP(3)) receptor (IP(3)R) blocker. The pretreatment of SaM-1 cells with U-73122, a phospholipase C (PLC) inhibitor, and 2-APB also inhibited the increase in IL-6 and IL-8 synthesis in response to LPA. These findings suggest that extracellular LPA-induced IL-6 and IL-8 synthesis occurred through Edg-2 (LPA(1) receptor) and the activation of PLC and IP(3)-mediated intracellular calcium release in SaM-1 cells.

Interleukin-33 (IL-33) induces T helper type 2 (Th2) cytokine production and eosinophilia independently of acquired immunity, leading to innate immunity-mediated allergic inflammation. Allergy-related innate myeloid cells such as eosinophils, basophils and mast cells express the IL-33 receptor (IL-33R), but it is still unknown how IL-33 regulates allergic inflammation involving these cells and their progenitors. Here, we revealed that the functional IL-33R was expressed on eosinophil progenitors (EoPs), basophil progenitors (BaPs) and mast cell progenitors (MCPs). In the presence of IL-33, these progenitors did not expand, but produced a high amount of Th2 and pro-inflammatory cytokines such as IL-9, IL-13, IL-1β and IL-6. The amount of cytokines produced by these progenitors was greater than that by mature cells. In vivo, IL-33 stimulated the expansion of EoPs, but it was dependent upon the elevated serum IL-5 that is presumably derived from type 2 innate lymphoid cells that express functional IL-33R. These data collectively suggest that EoPs, BaPs and MCPs are not only the sources of allergy-related granulocytes, but can also be sources of allergy-related cytokines in IL-33-induced inflammation. Because such progenitors can differentiate into mature granulocytes at the site of inflammation, they are potential therapeutic targets in IL-33-related allergic diseases.