BACKGROUND

This study compares inflammation-related biomarkers with established cardiometabolic risk factors in the prediction of incident type 2 diabetes and incident coronary events in a prospective case-cohort study within the population-based MONICA/KORA Augsburg cohort.

RESULTS

Analyses for type 2 diabetes are based on 436 individuals with and 1410 individuals without incident diabetes. Analyses for coronary events are based on 314 individuals with and 1659 individuals without incident coronary events. Mean follow-up times were almost 11 years. Areas under the receiver-operating characteristic curve (AUC), changes in Akaike's information criterion (ΔAIC), integrated discrimination improvement (IDI) and net reclassification index (NRI) were calculated for different models. A basic model consisting of age, sex and survey predicted type 2 diabetes with an AUC of 0.690. Addition of 13 inflammation-related biomarkers (CRP, IL-6, IL-18, MIF, MCP-1/CCL2, IL-8/CXCL8, IP-10/CXCL10, adiponectin, leptin, RANTES/CCL5, TGF-β1, sE-selectin, sICAM-1; all measured in nonfasting serum) increased the AUC to 0.801, whereas addition of cardiometabolic risk factors (BMI, systolic blood pressure, ratio total/HDL-cholesterol, smoking, alcohol, physical activity, parental diabetes) increased the AUC to 0.803 (ΔAUC [95% CI] 0.111 [0.092-0.149] and 0.113 [0.093-0.149], respectively, compared to the basic model). The combination of all inflammation-related biomarkers and cardiometabolic risk factors yielded a further increase in AUC to 0.847 (ΔAUC [95% CI] 0.044 [0.028-0.066] compared to the cardiometabolic risk model). Corresponding AUCs for incident coronary events were 0.807, 0.825 (ΔAUC [95% CI] 0.018 [0.013-0.038] compared to the basic model), 0.845 (ΔAUC [95% CI] 0.038 [0.028-0.059] compared to the basic model) and 0.851 (ΔAUC [95% CI] 0.006 [0.003-0.021] compared to the cardiometabolic risk model), respectively.

CONCLUSIONS

Inclusion of multiple inflammation-related biomarkers into a basic model and into a model including cardiometabolic risk factors significantly improved the prediction of type 2 diabetes and coronary events, although the improvement was less pronounced for the latter endpoint.

Bronchiolar epithelial cells are the prime targets for influenza A virus infection. It still remains to be clarified which signals are generated from these cells to initiate an immune response. Among chemokines, viral infection of primary lung epithelial cells triggered exclusively the release of CXCL8/interleukin-8 (IL-8), which contrasts with our previous observation that influenza A virus induced in monocytes the expression of mononuclear-leukocyte-attracting chemokines and even suppressed the production of neutrophil-attracting chemokines. Therefore, we speculated that it may be advantageous for respiratory epithelial cells to release primarily neutrophil-attracting CXCL8/IL-8 since neutrophils rapidly remove necrotic debris and are the first line of defense against bacterial superinfections. This concept has also been supported by our finding that influenza A virus infection led to necrosis of lung epithelial cells. This is in striking contrast to previous studies where influenza A virus infection induced apoptosis in monocytes and epithelial cells from origins other than the lung. Thus, the cell type instead of the virus determines which death pathway will be followed. In addition to the release of CXCL8/IL-8, we obtained a massive release of macrophage migration inhibitory factor (MIF) from virus-infected lung cells. However, whereas the CXCL8/IL-8 secretion was accompanied by induced gene activation, the transcription rate of MIF remained unchanged during the infection course and the virus-induced MIF release was predominantly a discharge from intracellular stores, suggesting that MIF is passively released upon cell death. Despite virus induced necrosis, the passively liberated MIF remained bioactive. Considering the well-established immunostimulatory effects of MIF on different leukocyte subsets, is its very likely that enhanced levels of MIF may contribute to the host immune response during the acute phase of influenza A virus infection in humans.

Multiple Sclerosis (MS) is a chronic inflammatory disease of the CNS which is characterized by large mononuclear cell infiltration and significant demyelination. CXCL8 is a chemo-attractant for both neutrophils and monocytes and triggers their firm adhesion to endothelium. In this study, we demonstrate that serum CXCL8 and CXCL8 secretion from PBMCs are significantly higher in untreated MS patients compared to controls and are significantly reduced in MS patients receiving interferon-beta1a therapy. We suggest that CXCL8 may serve as a marker of monocyte activity in MS and may play a role in monocyte recruitment to the CNS.

The Kaposi's sarcoma herpesvirus encodes a G-protein-coupled chemokine receptor termed KSHV-GPCR. Expression of this constitutively active GPCR leads to cell transformation and vascular overgrowth characteristic of Kaposi's sarcoma. Previously, we have shown that CXCR2, the closest human homolog, is similarly able to transform cells if continuously stimulated or constitutively activated by amino-acid exchange D138V of the DRY sequence. Here, we demonstrate that STAT3 activation is a prerequisite for transformation in KSHV-GPCR and CXCR2 transfected NIH 3T3 cells. In KSHV-GPCR and D138V transfected cells, STAT-3 is constitutively phosphorylated on Tyr705. In CXCR2 transfected NIH 3T3 cells and human microvascular endothelial cells (HMEC), which express the CXCR2 constitutively, STAT3 is phosphorylated upon stimulation with IL-8 (CXCL8). Focus formation in NIH 3T3 cells transfected with the KSHV-GPCR, CXCR2, or the D138V mutant, was blocked by the specific JAK2 inhibitor AG490. Typical functions of the CXCR2 including actin stress fiber formation, haptotaxis, and the angiogenic response in HMEC shown by tube formation in Matrigel were blocked by AG490. These data suggest that the transforming capacity and migratory responses that are involved in tumor development, metastasis, and angiogenesis in KSHV or CXCR2-expressing cells is at least partially mediated through a JAK2-STAT3 dependent pathway.

Eosinophils are multifunctional leukocytes involved in various inflammatory processes, as well as tissue remodeling and immunoregulation. During inflammation and infection, injured cells and damaged tissues release uric acid and monosodium urate (MSU) crystals as important endogenous danger signals. Uric acid is also implicated in the immunogenic effects of an authentic Th2 adjuvant, aluminum hydroxide. Eosinophils often localize at sites of Th2-type chronic inflammation; therefore, we hypothesized that eosinophils may react to endogenous danger signals. We found that human eosinophils migrate toward soluble uric acid and MSU crystals in a gradient-dependent manner. Eosinophils incubated with MSU crystals, but not those incubated with uric acid solution, produced elevated levels of IL-6 and IL-8/CXCL8. Other cytokines and chemokines, including IL-1beta, IL-10, IL-17, IFN-gamma, CCL2, CCL3, CCL4, TNF-alpha, G-CSF, GM-CSF, fibroblast growth factor, vascular endothelial growth factor, and TGF-beta, were also produced by eosinophils incubated with MSU crystals. Eosinophils exposed to MSU crystals rapidly (i.e., within 1 min of exposure) released ATP into the extracellular milieu. Importantly, this autocrine ATP was necessary for eosinophils to produce cytokines in response to MSU crystals, and P2 nucleotide receptors, in particular P2Y(2), are likely involved in this positive feedback loop. Finally, at higher concentrations, MSU crystals promoted P2R-dependent release of a granule protein (eosinophil-derived neurotoxin) and cell death. Thus, human eosinophils may respond to particulate damage-associated endogenous danger signals. These responses by eosinophils to tissue damage may explain the self-perpetuating nature of chronic inflammation in certain human diseases, such as asthma.

An important checkpoint in the progression of melanoma is the metastasis to lymph nodes. Here, to investigate the role of lymph node NK cells in disease progression, we analyze frequency, phenotype and functions of NK cells from tumour-infiltrated (TILN) and tumour-free ipsilateral lymph nodes (TFLN) of the same patients. We show an expansion of CD56(dim)CD57(dim)CD69 + CCR7 +KIR+ NK cells in TILN. TILN NK cells display robust cytotoxic activity against autologous melanoma cells. In the blood of metastatic melanoma patients, the frequency of NK cells expressing the receptors for CXCL8 receptor is increased compared with healthy subjects, and blood NK cells also express the receptors for CCL2 and IL-6. These factors are produced in high amount in TILN and in vitro switch the phenotype of blood NK cells from healthy donors to the phenotype associated with TILN. Our data suggest that the microenvironment of TILN generates and/or recruits a particularly effective NK cell subset.

BACKGROUND

Malaria during pregnancy is characterized by the sequestration of malaria-infected red blood cells (iRBC) in the intervillous spaces of the placenta, often accompanied by the infiltration of maternal mononuclear cells, causing substantial maternal and foetal/infant morbidity. The iRBC bind to receptors expressed by the syncytiotrophoblast (ST). How ST responds to this interaction remains poorly understood. Because it is known that ST is immunoactive and can respond to infectious agents, the consequences of this ST-iRBC interaction should be investigated.

METHODS

An in vitro system was used to assess the biochemical and immunological changes induced in ST by ST-adherent iRBCs. Changes in ST mitogen-activated protein kinase (MAPK) activation were assessed by immunoblotting and mRNA expression levels of selected cytokine and chemokines in primary ST bound by iRBC were determined using real-time, reverse transcription PCR. In addition, secreted cytokine and chemokine proteins were assayed by standard ELISA, and chemotaxis of PBMC was assessed using a two-chamber assay system.

RESULTS

Following iRBC/ST interaction, ST C-Jun N-terminal kinase 1 (JNK1) was activated and modest increases in the mRNA expression of TGF-beta and IL-8/CXCL8 were observed. In addition, this interaction increased secretion of MIF and MIP-1alpha/CCL3 by ST and induced migration of PBMC towards iRBC-stimulated ST.

CONCLUSIONS

Results from this study provide the first evidence that ST participates in shaping the local immunological milieu and in the recruitment of maternal immune cells to the maternal blood space during placental malaria infection.

BACKGROUND

The effectiveness of systemic treatment of psoriasis with fumaric acid esters has been proven, but their mode of action at the cellular and molecular level has not yet been fully elucidated.

OBJECTIVE

To study the effect of dimethylfumarate (DMF) on the production of the chemokines CXCL1, CXCL8, CXCL9, CXCL10 and CXCL11, formerly known as GROalpha, interleukin-8, Mig, IP-10 and IP-9/I-TAC, respectively, in human keratinocytes and peripheral blood mononuclear cells (PBMC).

METHODS

Cultured keratinocytes were stimulated with interferon (IFN) -gamma to produce CXCL9, CXCL10 and CXCL11 and with phorbol myristate acetate to produce CXCL1 and CXCL8 in the absence and presence of DMF (5, 15 and 45 micromol L(-1)). PBMC were stimulated with either IFN-gamma to produce CXCL9 and CXCL10 or lipopolysaccharide to produce CXCL8, in the absence and presence of DMF (5, 15 and 45 micromol L(-1)). RNA preparations from isolated keratinocytes were analysed by Northern blotting; protein production by keratinocytes and PBMC was monitored by an enzyme-linked immunosorbent assay.

RESULTS

Northern blot analysis on isolated keratinocyte RNA preparations showed a dose-dependent inhibition of CXCL1, CXCL8, CXCL9, CXCL10 and CXCL11 transcription by DMF. At 45 micromol L(-1) the inhibition was almost complete. In addition, keratinocytes and PBMC showed in the presence of DMF a dose-dependent inhibition of CXCL8, CXCL9 and CXCL10 protein production.

CONCLUSIONS

These results show the ability of DMF to inhibit the production of chemokines that may be critically involved in the development and perpetuation of psoriatic lesions. This might explain, at least in part, the beneficial effects of treatment with fumaric acid esters in psoriasis patients.

Tumors are dynamic organs, in which active processes of cell motility affect disease course by regulating the composition of cells at the tumor site. While sub-populations of tumor-promoting leukocytes are recruited inward and endothelial cell migration stands in the basis of vascular branching throughout the tumor, cancer cells make their way out of the primary site towards specific metastatic sites. This review describes the independent and cross-regulatory roles of inflammatory chemokines and of the inflammatory cytokine tumor necrosis factor α (TNFα) in determining cell motility processes that eventually have profound effects on tumor growth and metastasis. First, the effects of inflammatory chemokines such as CCL2 (MCP-1), CCL5 (RANTES) and CXCL8 (IL-8) are described, regulating the inward flow of leukocyte sub-populations with pro-tumoral activities, such as tumor-associated macrophages (TAM), myeloid-derived suppressor cells (MDSC), tumor-associated neutrophils (TAN), Th17 cells and Tregs. Then, the ability of inflammatory chemokines to induce endothelial cell migration, sprouting and tube formation is discussed, with its implications on tumor angiogenesis. This part is followed by an in depth description of the manners by which TNFα potentiates the above activities of the inflammatory chemokines, alongside with its ability to directly induce migratory processes in the tumor cells thus promoting metastasis. Note worthy is the ability of TNFα to induce in the tumor cells the important process of epithelial-to-mesenchymal transition (EMT). Emphasis is given to the ability of TNFα to establish an inflammatory network with the chemokines, and in parallel to form a cell re-modeling network together with transforming growth factor β (TGFβ). The review concludes by discussing the implications of such networks on disease course, and on the future design of therapeutic measures in cancer.

BACKGROUND

Chronic inflammation and oxidative stress are key features of chronic obstructive pulmonary disease (COPD). Oxidative stress enhances COPD inflammation under the control of the pro-inflammatory redox-sensitive transcription factor nuclear factor-kappaB (NF-κB). Histone acetylation plays a critical role in chronic inflammation and bromodomain and extra terminal (BET) proteins act as "readers" of acetylated histones. Therefore, we examined the role of BET proteins in particular Brd2 and Brd4 and their inhibitors (JQ1 and PFI-1) in oxidative stress- enhanced inflammation in human bronchial epithelial cells.

METHODS

Human primary epithelial (NHBE) cells and BEAS-2B cell lines were stimulated with IL-1β (inflammatory stimulus) in the presence or absence of H2O2 (oxidative stress) and the effect of pre-treatment with bromodomain inhibitors (JQ1 and PFI-1) was investigated. Pro-inflammatory mediators (CXCL8 and IL-6) were measured by ELISA and transcripts by RT-PCR. H3 and H4 acetylation and recruitment of p65 and Brd4 to the native IL-8 and IL-6 promoters was investigated using chromatin immunoprecipitation (ChIP). The impact of Brd2 and Brd4 siRNA knockdown on inflammatory mediators was also investigated.

RESULTS

H2O2 enhanced IL1β-induced IL-6 and CXCL8 expression in NHBE and BEAS-2B cells whereas H2O2 alone did not have any affect. H3 acetylation at the IL-6 and IL-8 promoters was associated with recruitment of p65 and Brd4 proteins. Although p65 acetylation was increased this was not directly targeted by Brd4. The BET inhibitors JQ1 and PFI-1 significantly reduced IL-6 and CXCL8 expression whereas no effect was seen with the inactive enantiomer JQ1(-). Brd4, but not Brd2, knockdown markedly reduced IL-6 and CXCL8 release. JQ1 also inhibited p65 and Brd4 recruitment to the IL-6 and IL-8 promoters.

CONCLUSIONS

Oxidative stress enhanced IL1β-induced IL-6 and CXCL8 expression was significantly reduced by Brd4 inhibition. Brd4 plays an important role in the regulation of inflammatory genes and provides a potential novel anti-inflammatory target.

BACKGROUND

This review presents the current in vitro and in vivo animal and human research on the roles of IL-8 in ocular inflammatory diseases.

METHODS

Data sources were a literature review using Pub Med, Medline, and ISI databases (from 1990 to 2011). Search items included interleukine-8 (IL-8), CXCL8, chemokines, cytokines, alone or in combination with the, serum, aqueous, vitreous, eye, ocular, ocular tissues, ophthalmic, and review.

RESULTS

IL-8 may be involved in primary or secondary ocular inflammations. Ocular effects of IL-8 differ based on the source of the secretion and site of the action. The most important effects of IL-8 in the eyes are angiogenic activities and induction of ocular inflammation.

CONCLUSIONS

IL-8 plays important roles in ocular inflammation and angiogenesis in conjunctiva, cornea, iris, retina, and orbit. Anti-IL-8 targeted immunotherapy has been introduced as an important treatment modality, provided that IL-8 signal blocking takes place in desired areas and tissues.

BACKGROUND

Leucocyte activating chemokines such as CCL2, CCL3, and CXCL8 together with proinflammatory IFNgamma, TNFalpha and downmodulatory IL10 play a central role in the restriction of M. tuberculosis infections, but is unclear whether these markers are indicative of tuberculosis disease severity.

METHODS

We investigated live M. tuberculosis- and M. bovis BCG-induced peripheral blood mononuclear cell responses in patients with tuberculosis (TB) and healthy endemic controls (ECs, n = 36). TB patients comprised pulmonary (PTB, n = 34) and extrapulmonary groups, subdivided into those with less severe localized extrapulmonary TB (L-ETB, n = 16) or severe disseminated ETB (D-ETB, n = 16). Secretion of CCL2, IFNgamma, IL10 and CCL3, and mRNA expression of CCL2, TNFalpha, CCL3 and CXCL8 were determined.

RESULTS

M. tuberculosis- and BCG-induced CCL2 secretion was significantly increased in both PTB and D-ETB (p<0.05, p<0.01) as compared with L-ETB patients. CCL2 secretion in response to M. tuberculosis was significantly greater than to BCG in the PTB and D-ETB groups. M. tuberculosis-induced CCL2 mRNA transcription was greater in PTB than L-ETB (p = 0.023), while CCL2 was reduced in L-ETB as compared with D-ETB (p = 0.005) patients. M. tuberculosis-induced IFNgamma was greater in L-ETB than PTB (p = 0.04), while BCG-induced IFNgamma was greater in L-ETB as compared with D-ETB patients (p = 0.036). TNFalpha mRNA expression was raised in PTB as compared with L-ETB group in response to M. tuberculosis (p = 0.02) and BCG (p = 0.03). Mycobacterium-induced CCL3 and CXCL8 was comparable between TB groups.

CONCLUSIONS

The increased CCL2 and TNFalpha in PTB patients may support effective leucocyte recruitment and M. tuberculosis localization. CCL2 alone is associated with severity of TB, possibly due to increased systemic inflammation found in severe disseminated TB or due to increased monocyte infiltration to lung parenchyma in pulmonary disease.

IL-31 is a novel T(h) type 2 cytokine that can induce pruritus and dermatitis in mice resembling human atopic dermatitis (AD). Eosinophil infiltration in skin lesions is a predominant pathological feature of AD. In the present study, we investigated the effects of IL-31 on the activation of human eosinophils and epidermal keratinocytes. Eosinophils and keratinocytes were cultured either together or separately in the presence or absence of IL-31 stimulation. IL-31 could significantly induce the release of pro-inflammatory cytokines IL-1beta, IL-6 and AD-related chemokines CXCL1, CXCL8, CCL2 and CCL18 from eosinophils, via functional cell surface IL-31 receptor. Such induction was further enhanced upon the co-culture of eosinophils and keratinocytes, in which eosinophils were the main source for releasing pro-inflammatory cytokines and chemokines. The presence of transwell inserts in co-culture system demonstrated that the direct interaction between eosinophils and keratinocytes was required for IL-31-induced cytokine and chemokine release. Cell surface expression of adhesion molecule CD18 on eosinophils and intercellular adhesion molecule-1 on keratinocytes was up-regulated in the co-culture, and levels were further enhanced upon IL-31 stimulation. The interaction between eosinophils and keratinocytes under IL-31 stimulation was differentially mediated through intracellular mitogen-activated protein kinases, nuclear factor-kappaB and phosphatidylinositol 3-kinase-Akt pathways. The above findings suggest a crucial immunopathological role of IL-31 in AD through activation of eosinophils-keratinocytes system.

Systemic lupus erythematosus (SLE) is a systemic autoimmune disease associated with aberrant activation of T and B lymphocytes for the production of inflammatory cytokines and autoreactive antibodies. Animal studies of SLE have indicated that Toll-like receptors (TLR) are important in the pathogenesis of murine lupus. In the present clinical study, differential protein expressions of TLR-1-9 of monocytes and different lymphocyte subsets from patients with SLE and normal control subjects were determined by flow cytometry. Results showed that the expression of intracellular TLRs (TLR-3, -8, -9) and extracellular TLRs (TLR-1, -2, -4, -5, -6) were elevated in monocytes, CD4(+) T lymphocytes, CD8(+) T lymphocytes and B lymphocytes of SLE patients compared to control subjects (all P < 0.001). Moreover, cell surface expression of TLR-4 on CD4(+) T lymphocytes and CD8(+) T lymphocytes, and TLR-6 on B lymphocytes, were correlated positively with SLE disease activity index (SLEDAI) (TLR-4 on CD4(+) T lymphocytes and CD8(+) T lymphocytes: r = 0.536, P = 0.04; r = 0.713, P = 0.003; TLR-6 in B lymphocytes: r = 0.572, P = 0.026). In concordance with the above results, there is an observable increased relative induction (%) of inflammatory cytokine interleukin (IL)-1beta, IL-6, IL-10 and IL-12, chemokines CCL2, CXCL8, CCL5 and CXCL10 from peripheral blood mononuclear cells (PBMC) upon differential stimulation by PolyIC (TLR-3 ligand), lipopolysaccharide (TLR-4 ligand), peptidoglycan (TLR-2 ligand), flagellin (TLR-5 ligand), R837 (TLR-7 ligand) and CpG DNA (TLR-9 ligand) in SLE patients compared to controls. These results suggest that the innate immune response for extracellular pathogens and self-originated DNA plays immunopathological roles via TLR activation in SLE.

We have recently shown that several proinflammatory chemokines can be stored in secretory granules of endothelial cells (ECs). Subsequent regulated exocytosis of such chemokines may then enable rapid recruitment of leukocytes to inflammatory sites. Although IL-8/CXCL8 and eotaxin-3/CCL26 are sorted to the rod-shaped Weibel-Palade body (WPB), we found that GROalpha/CXCL1 and MCP-1/CCL2 reside in small granules that, similarly to the WPB, respond to secretagogue stimuli. In the present study, we report that GROalpha and MCP-1 colocalized in 50- to 100-nm granules, which occur throughout the cytoplasm and at the cell cortex. Immunofluorescence confocal microscopy revealed no colocalization with multimerin or tissue plasminogen activator, i.e., proteins that are released from small granules of ECs by regulated exocytosis. Moreover, the GROalpha/MCP-1-containing granules were Rab27-negative, contrasting the Rab27-positive, WPB. The secretagogues PMA, histamine, and forskolin triggered distinct dose and time-dependent responses of GROalpha release. Furthermore, GROalpha release was more sensitive than IL-8 release to inhibitors and activators of PKA and PKC but not to an activator of Epac, a cAMP-regulated GTPase exchange factor, indicating that GROalpha release is regulated by molecular adaptors different from those regulating exocytosis of the WPB. On the basis of these findings, we designated the GROalpha/MCP-1-containing compartment the type 2 granule of regulated secretion in ECs, considering the WPB the type 1 compartment. In conclusion, we propose that the GROalpha/MCP-1-containing type 2 granule shows preferential responsiveness to important mediators of EC activation, pointing to the existence of selective agonists that would allow differential release of selected chemokines.

Interaction between chemokines and heparan sulfate (HS) is essential for leukocyte recruitment during inflammation. Previous studies have shown that a non-HS-binding mutant form of the inflammatory chemokine CCL7 can block inflammation produced by wild-type chemokines. This study examined the anti-inflammatory mechanism of a non-HS-binding mutant of the homeostatic chemokine CXCL12. Initial experiments demonstrated that mutant CXCL12 was an effective CXCR4 agonist. However, this mutant chemokine failed to promote transendothelial migration in vitro and inhibited the haptotactic response to wild-type CCL7, CXCL12, and CXCL8, and naturally occurring chemoattractants in synovial fluid from the rheumatoid synovium, including CCL2, CCL7, and CXCL8. Notably, intravenous administration of mutant CXCL12 also inhibited the recruitment of leukocytes to murine air pouches filled with wild-type CXCL12. Following intravenous administration, wild-type CXCL12 was cleared from the circulation rapidly, while the mutant chemokine persisted for >24 h. Chronic exposure to mutant CXCL12 in the circulation reduced leukocyte-surface expression of CXCR4, reduced the chemotactic response of these cells to CXCL12, and inhibited normal chemokine-mediated induction of adhesion between the alpha4beta1 integrin, VLA-4, and VCAM-1. These data demonstrate that systemic administration of non-HS-binding variants of CXCL12 can mediate a powerful anti-inflammatory effect through chemokine receptor desensitization.

Epidermal growth factor (EGF) and their receptor (EGFR) play an important role in the development of cancer proliferation, and metastasis, although the mechanism remains unclear. The present study aimed at investigating the role of EGF-EGFR signalling pathway in the development of human hepatocellular carcinoma (HCC) inflammatory environment. Gene profiles of inflammatory cytokines from HCC were measured. Cell bio-behaviours of HCC with low or high metastasis were detected by the live cell monitoring system. Cell proliferation was measured by CCK8. The protein level of CXCL5 and CXCL8 was measured by ELISA. The phosphorylation of PI3K, ERK, MAPK was measured by western blot. EGF significantly induced cell proliferation in HepG2 cells, but not in HCCLM3 cells. EGF prompted the cell movement in both HepG2 and HCCLM3 and regulated the production of CXCL5 and CXCL8 from HCC, which were inhibited by EGFR inhibitor, Erk inhibitor (U0126), or PI3K inhibitors (BEZ-235 and SHBM1009). HCC proliferation, metastasis and production of inflammatory cytokines were regulated via EGF-EGFR signal pathways. CXCL5 could interact with CXCL8, possibly by CXCR2 or the cross-talk between CXCR2 and EGFR. EGF-EGFR signaling pathway can be the potential target of therapies for HCC.

IL-21 is a cytokine known to mediate its biological action via the IL-21R, composed of a specific chain, IL-21Ralpha, and the common gamma-chain (CD132). Recent data suggest that IL-21 possesses proinflammatory properties. However, there is no clear evidence that IL-21 induces inflammation in vivo and, curiously, the interaction between IL-21 and neutrophils has never been investigated, despite the fact that these cells express CD132 and respond to other CD132-dependent cytokines involved in inflammatory disorders. Using the murine air pouch model, we found that IL-21 induced inflammation in vivo, based on recruitment of neutrophil and monocyte populations. In contrast to LPS, administration of IL-21 into the air pouch did not significantly increase the concentration of IL-6, CCL5, CCL3, and CXCL2. We demonstrated that HL-60 cells expressed IL-21Ralpha, which is down-regulated during their differentiation toward neutrophils, and that IL-21Ralpha is not detected in neutrophils. Concomitant with this, IL-21 induced Erk-1/2 phosphorylation in HL-60 cells, but not in neutrophils. To eliminate the possibility that IL-21 could activate neutrophils even in the absence of IL-21Ralpha, we demonstrated that IL-21 did not modulate several neutrophil functions. IL-21-induced Erk-1/2 phosphorylation was not associated with proliferation or differentiation of HL-60 toward neutrophils, monocytes, or macrophages. IL-21Ralpha was detected in human monocytes and monocyte-derived macrophages, but IL-21 increased CXCL8 production only in monocyte-derived macrophages. We conclude that IL-21 is a proinflammatory cytokine, but not a neutrophil agonist. We propose that IL-21 attracts neutrophils indirectly in vivo via a mechanism independent of IL-6, CCL3, CCL5, and CXCL2 production.

Recent data indicate a role for airway epithelial necroptosis, a regulated form of necrosis, and the associated release of damage-associated molecular patterns (DAMPs) in the development of chronic obstructive pulmonary disease (COPD). DAMPs can activate pattern recognition receptors (PRRs), triggering innate immune responses. We hypothesized that cigarette smoke (CS)-induced epithelial necroptosis and DAMP release initiate airway inflammation in COPD. Human bronchial epithelial BEAS-2B cells were exposed to cigarette smoke extract (CSE), and necrotic cell death (membrane integrity by propidium iodide staining) and DAMP release (i.e., double-stranded DNA, high-mobility group box 1, heat shock protein 70, mitochondrial DNA, ATP) were analyzed. Subsequently, BEAS-2B cells were exposed to DAMP-containing supernatant of CS-induced necrotic cells, and the release of proinflammatory mediators [C-X-C motif ligand 8 (CXCL-8), IL-6] was evaluated. Furthermore, mice were exposed to CS in the presence and absence of the necroptosis inhibitor necrostatin-1, and levels of DAMPs and inflammatory cell numbers were determined in bronchoalveolar lavage fluid. CSE induced a significant increase in the percentage of necrotic cells and DAMP release in BEAS-2B cells. Stimulation of BEAS-2B cells with supernatant of CS-induced necrotic cells induced a significant increase in the release of CXCL8 and IL-6, in a myeloid differentiation primary response gene 88-dependent fashion. In mice, exposure of CS increased the levels of DAMPs and numbers of neutrophils in bronchoalveolar lavage fluid, which was statistically reduced upon treatment with necrostatin-1. Together, we showed that CS exposure induces necrosis of bronchial epithelial cells and subsequent DAMP release in vitro, inducing the production of proinflammatory cytokines. In vivo, CS exposure induces neutrophilic airway inflammation that is sensitive to necroptosis inhibition.

Psychiatric disorders are highly prevalent and disabling conditions of increasing public health relevance. Much recent research has focused on the role of cytokines in the pathophysiology of psychiatric disorders; however, the related family of immune proteins designated chemokines has been relatively neglected. Chemokines were originally identified as having chemotactic function on immune cells; however, recent evidence has begun to elucidate novel, brain-specific functions of these proteins of relevance to the mechanisms of psychiatric disorders. A systematic review of both human and animal literature in the PubMed and Google Scholar databases was undertaken. After application of all inclusion and exclusion criteria, 157 references were remained for the review. Some early mechanistic evidence does associate select chemokines with the neurobiological processes, including neurogenesis, modulation of the neuroinflammatory response, regulation of the hypothalamus-pituitary-adrenal axis, and modulation of neurotransmitter systems. This early evidence however does not clearly demonstrate any specificity for a certain psychiatric disorder, but is primarily relevant to mechanisms which are shared across disorders. Notable exceptions include CCL11 that has recently been shown to impair hippocampal function in aging - of distinct relevance to Alzheimer's disease and depression in the elderly, and pre-natal exposure to CXCL8 that may disrupt early neurodevelopmental periods predisposing to schizophrenia. Pro-inflammatory chemokines, such as CCL2, CCL7, CCL8, CCL12, and CCL13, have been shown to drive chemotaxis of pro-inflammatory cells to the inflamed or injured CNS. Likewise, CX3CL has been implicated in promoting glial cells activation, pro-inflammatory cytokines secretion, expression of ICAM-1, and recruitment of CD4+ T-cells into the CNS during neuroinflammatory processes. With further translational research, chemokines may present novel diagnostic and/or therapeutic targets in psychiatric disorders.

There is accumulating evidence indicating that endothelial factors are involved in the pathogenesis of GVHD. We have recently shown that steroid-refractory, but not sensitive, GVHD is characterized by higher pretransplantation serum levels of angiopoetin-2 (ANG2), a hormone mediating endothelial vulnerability. To evaluate whether endothelial vulnerability is a risk factor for GVHD per se or becomes important only when noticeable GVHD is established, we measured ANG2 along with additional serum markers of endothelial stress, including soluble thrombomodulin (sTM), IL-8 (CXCL8), and hepatocyte growth factor (HGF), in patients with no, low-grade, or severe GVHD. Patients with refractory GVHD exhibited elevated serum levels of ANG2, sTM, HGF, and IL-8 posttransplantation compared with patients with sensitive GVHD and patients without GVHD. Pretransplantation ANG2 was the only growth factor correlated with the risk of refractoriness and mortality, and then only within the subset of patients who developed grade III-IV GVHD. In contrast, ANG2 was not predictive of GVHD or nonrelapse mortality (NRM) in patients with no GVHD or low-grade GVHD. These findings provide evidence that endothelial function plays an important role in the pathogenesis of steroid refractoriness in ongoing GVHD; however, endothelial vulnerability does not predict incidence of GVHD.

Chemokines are increasingly recognised as playing a role in depression. Here we meta-analyse the data on concentrations of all chemokines in patients diagnosed with a major depression versus healthy controls. We included studies which utilised Diagnostic and Statistical Manual (DSM)-IV diagnostic criteria for major depression, participants free from major medical conditions, studies with healthy controls, and unstimulated measurements of chemokines. We only included chemokines which had ≥3 studies performed. Two chemokines and 15 studies in total met criteria for this meta-analysis; 8 for Monocyte Chemotactic Protein (MCP)-1/CCL2 (n=747), and 7 for Interleukin (IL)-8/CXCL8 (n=560). There were significantly higher concentrations of CCL2/MCP-1 in depressed subjects compared with control subjects - overall mean difference of 36.43pg/mL (95% CI: 2.43 to 70.42). There was significant heterogeneity across these studies (I2=98.5%). The estimates of mean difference between the control and depression groups did not remain significant when the trim-and-fill procedure was used to correct for publication bias. There was no significant difference in concentrations of IL-8/CXCL8 in depressed subjects compared with control subjects. Significant heterogeneity was found across these studies (I2=96.7%). The estimates of mean difference between the control and depression groups remained non-significant when the trim-and-fill procedure was used to correct for publication bias. This meta-analysis reports significantly heterogeneity in this field among studies. There are higher concentrations of the chemokine MCP-1/CCL2 in depressed subjects compared with control subjects, and no differences for IL-8/CXCL8. More high quality research and consistent methodologies are needed in this important area of enquiry.

The dynamics of neutrophil chemotaxis under competing chemoattractant gradients was studied using a microfluidic platform. This microfluidic platform, which establishes a stable and dynamic gradient of chemoattractants across a cell culture chamber, enabled the investigation of human neutrophil migration patterns in the presences of four different chemoattractants (leukotriene B(4), chemokine C-X-C motif ligands 2 and 8, and fMLP) and competing gradients of all pairwise combinations. The migration patterns for individual cells were tracked and quantitatively analyzed, and the results suggest a hierarchy among these chemoattractants of fMLP>> CXCL8>> CXCL2>> leukotriene B(4). In all conditions, over 60% of neutrophils exposed to a competing gradient move toward the stronger signal though the weaker chemoattractant still influences neutrophil motility. These results yield insight about how each chemoattractant contributes to overall neutrophil chemotaxis within complex physiological environments.

Cxcl8 is a pro-inflammatory chemokine, best known for its role in neutrophil chemotaxis. Signalling through its receptors, Cxcr1 and Cxcr2, is induced by inflammatory stimuli evoked by microbial, chemical or environmental stress, and hormonal signals. While it is recognised that Cxcl8 signalling is active in the gut mucosa, this is not as well understood as its role in leukocyte trafficking. Here, we report the characterisation of genes encoding the zebrafish Cxcl8, Cxcr1 and Cxcr2. By a combination of genomic, expression and functional analyses, we show that the Cxcl8 signalling pathway is conserved in zebrafish. As in humans, cxcl8 is expressed in zebrafish leukocytes. Transcripts were also detected in intestinal epithelial cells, and this expression is upregulated under inflammatory conditions caused by bacterial or chemical insult. Expression of cxcr1 and cxcr2 is robust within the developing gut. This work provides a model for the study of Cxcl8 signalling during gut inflammation.