Cerebral malaria is a severe multifactorial condition associated with the interaction of high numbers of infected erythrocytes to human brain endothelium without invasion into the brain. The result is coma and seizures with death in more than 20% of cases. Because the brain endothelium is at the interface of these processes, we investigated the global gene responses of human brain endothelium after the interaction with Plasmodium falciparum-infected erythrocytes with either high- or low-binding phenotypes. The most significantly up-regulated transcripts were found in gene ontology groups comprising the immune response, apoptosis and antiapoptosis, inflammatory response, cell-cell signaling, and signal transduction and nuclear factor kappaB (NF-kappaB) activation cascade. The proinflammatory NF-kappaB pathway was central to the regulation of the P falciparum-modulated endothelium transcriptome. The proinflammatory molecules, for example, CCL20, CXCL1, CXCL2, IL-6, and IL-8, were increased more than 100-fold, suggesting an important role of blood-brain barrier (BBB) endothelium in the innate defense during P falciparum-infected erythrocyte (Pf-IRBC) sequestration. However, some of these diffusible molecules could have reversible effects on brain tissue and thus on neurologic function. The inflammatory pathways were validated by direct measurement of proteins in brain endothelial supernatants. This study delineates the strong inflammatory component of human brain endothelium contributing to cerebral malaria.

BACKGROUND

To investigate whether dendritic cell (DC) precursors, recruited by injection of chemokine ligand 3 (CCL3) and CCL20, induce anti-tumor immunity against gastric cancer induced by a DC vaccine expressing melanoma antigen gene-1 (MAGE-1) ex vivo and in vivo.

METHODS

B6 mice were injected with CCL3 and CCL20 via the tail vein. Freshly isolated F4/80-B220-CD11c+ cells cultured with cytokines were analyzed by phenotype analysis and mixed lymphocyte reaction (MLR). For adenoviral (Ad)-mediated gene transduction, cultured F4/80-B220-CD11c+ cells were incubated with Ad-MAGE-1. Vaccination of stimulated DC induced T lymphocytes. The killing effect of these T cells against gastric carcinoma cells was assayed by MTT. INF-gamma production was determined with an INF-gamma ELISA kit. In the solid tumor and metastases model, DC-based vaccines were used for immunization after challenge with MFC cells. Tumor size, survival of mice, and number of pulmonary metastatic foci were used to assess the therapeutic effect of DC vaccines.

RESULTS

F4/80-B220-CD11c+ cell numbers increased after CCL3 and CCL20 injection. Freshly isolated F4/80-B220-CD11c+ cells cultured with cytokines were phenotyically identical to typical DC and gained the capacity to stimulate allogeneic T cells. These DCs were transduced with Ad-MAGE-1, which were prepared for DC vaccines expressing tumor antigen. T lymphocytes stimulated by DCs transduced with Ad-MAGE-1 exhibited specific killing effects on gastric carcinoma cells and produced high levels of INF-gamma ex vivo. In vivo, tumor sizes of the experimental group were much smaller than both the positive control group and the negative control groups (P < 0.05). Kaplan-Meier survival curves showed that survival of the experimental group mice was significantly longer than the control groups (P < 0.05). In addition, MAGE-1-transduced DCs were also a therapeutic benefit on an established metastatic tumor, resulting in a tremendous decrease in the number of pulmonary metastatic foci.

CONCLUSIONS

CCL3 and CCL20-recruited DCs modified by adenovirus-trasnsduced, tumor-associated antigen, MAGE-1, can stimulate anti-tumor immunity specific to gastric cancer ex vivo and in vivo. This system may prove to be an efficient strategy for anti-tumor immunotherapy.

Lipocalin-2 (LCN2) is an antimicrobial protein and adipokine associated with insulin resistance, obesity and atherosclerotic disease. Psoriasis is a T-helper (Th)1/Th17-mediated, chronic inflammatory dermatosis related to metabolic syndromes and serum LCN2 levels are elevated in psoriatic patients. We examined the in vivo effects of LCN2 on topical imiquimod (IMQ)-induced psoriasiform skin in BALB/c mice and in vitro on human keratinocytes (KC). Clinically, i.p. injected LCN2 exacerbated erythema and scaling in IMQ-treated murine skin compared with phosphate-buffered saline injection alone, and it augmented interleukin (IL)-17A, IL-17F, IL-22, IL-23p19, IL-12p40, CCL20, tumor necrosis factor-α, chemokine (C-X-C motif) ligand (CXCL)1, CXCL2, DEFB4, DEFB14, LCN2 and S100A7 mRNA levels of IMQ-treated murine skin while it did not increase the mRNA levels of interferon-γ, IL-12p35 or CXCL10. LCN2 in synergy with IL-17 increased mRNA levels of CCL20, LCN2 and DEFB4A but not of CXCL10 in human KC in vitro. These results suggest that LCN2 enhances the expression of Th17 cytokines/chemokines and antimicrobial peptides in murine IMQ-treated psoriatic skin and KC. LCN2 may potentiate the development of psoriasis via the enhancement of Th17- and antimicrobial peptide-mediated inflammation.

Diabetic nephropathy (DN) is the leading cause of renal failure worldwide and its complications have become a public health problem. Inflammation, oxidative stress and fibrosis play central roles in the progression of DN that lead to renal failure. Potential deleterious effect of inflammation in early evolution of DN is not fully disclosed. Therefore, it is relevant to explore therapies that might modulate this process in order to reduce DN progression. We explored the beneficial effect of all-trans retinoic acid (ATRA) in early inflammation in glomeruli, proximal and distal tubules in streptozotocin (STZ)-induced diabetes. ATRA was administered (1 mg/kg daily by gavage) on days 3 to 21 after STZ administration. It was found that 21 days after STZ injection, diabetic rats exhibited proteinuria, increased natriuresis and loss of body weight. Besides, diabetes induced an increase in interleukins [IL-1β, IL-1α, IL-16, IL-13, IL-2; tumor necrosis factor alpha (TNF-α)] and transforming growth factor-beta 1 (TGF-β1), chemokines (CCL2, CCL20, CXCL5 and CXCL7), adhesion molecules (ICAM-1 and L-selectin) and growth factors (GM-CSF, VEGF, PDGF) in glomeruli and proximal tubules, whereas ATRA treatment remarkably ameliorated these alterations. To further explore the mechanisms through which ATRA decreased inflammatory response, the NF-κB/p65 signaling mediated by TLR4 was studied. We found that ATRA administration attenuates the TLR4/NF-κB inflammatory signaling and prevents NF-κB nuclear translocation in glomeruli and proximal tubules.

Background: Although immune modulation is a promising therapeutic avenue in coronavirus disease 2019 (COVID-19), the most relevant targets remain to be found. COVID-19 has peculiar characteristics and outcomes, suggesting a unique immunopathogenesis.

Methods: Thirty-six immunocompetent non-COVID-19 and 27 COVID-19 patients with severe pneumonia were prospectively enrolled in a single center, most requiring intensive care. Clinical and biological characteristics (including T cell phenotype and function and plasma concentrations of 30 cytokines) and outcomes were compared.

Results: At similar baseline respiratory severity, COVID-19 patients required mechanical ventilation for significantly longer than non-COVID-19 patients (15 [7-22] vs. 4 (0-15) days; p = 0.0049). COVID-19 patients had lower levels of most classical inflammatory cytokines (G-CSF, CCL20, IL-1β, IL-2, IL-6, IL-8, IL-15, TNF-α, TGF-β), but higher plasma concentrations of CXCL10, GM-CSF and CCL5, compared to non-COVID-19 patients. COVID-19 patients displayed similar T-cell exhaustion to non-COVID-19 patients, but with a more unbalanced inflammatory/anti-inflammatory cytokine response (IL-6/IL-10 and TNF-α/IL-10 ratios). Principal component analysis identified two main patterns, with a clear distinction between non-COVID-19 and COVID-19 patients. Multivariate regression analysis confirmed that GM-CSF, CXCL10 and IL-10 levels were independently associated with the duration of mechanical ventilation.

Conclusion: We identified a unique cytokine response, with higher plasma GM-CSF and CXCL10 in COVID-19 patients that were independently associated with the longer duration of mechanical ventilation. These cytokines could represent the dysregulated immune response in severe COVID-19, as well as promising therapeutic targets. ClinicalTrials.gov: NCT03505281.

Keywords: Acute respiratory distress syndrome; COVID-19; CXCL10; GM-CSF; Immune response; Mechanical ventilation; Pneumonia.

Psoriasis is a TNF-α/IL-23/IL-17A-mediated inflammatory skin disease that causes a significant socioeconomic burden in afflicted patients. IL-17A-producing immune cells, including Th17 cells, are crucial effector cells in the development of psoriasis. IL-17A stimulates epidermal keratinocytes to produce CCL20, which eventually recruits CCR6+ Th17 cells into the lesional skin. Thus, the CCL20/CCR6 axis works as a driving force that prepares an IL-17A-rich cutaneous milieu. In this review, we summarize the current research topics on the CCL20/CCR6 axis and the therapeutic intervention of this axis for psoriasis.

Objective: The transcription factor forkhead box protein O1 (FOXO1) is critical for regulating cytokine and chemokine secretion. However, its function in the tumor microenvironment (TME) remains largely unexplored. In this study, we characterized the prognostic value of FOXO1 and the interaction between tumor-derived FOXO1 and M2 macrophages in esophageal squamous cell carcinoma (ESCC). Methods: FOXO1 expression and macrophage infiltration in clinical samples and mouse models were quantified using quantitative real-time polymerase chain reaction (qRT-PCR) and immunohistochemistry staining. Western blotting, qRT-PCR, and enzyme-linked immunosorbent assay were used to evaluate chemokine ligand 20 (CCL20) and colony stimulating factor 1 (CSF-1) expression in FOXO1(+) and FOXO1(-) tumor cells. Macrophage phenotypes were determined using qRT-PCR, flow cytometry, and RNA sequencing. Transcriptional activity was measured using chromatin immunoprecipitation (ChIP)-qPCR. Tumor viability was investigated using XTT proliferation and foci formation assays. Results: FOXO1 upregulation in tumor tissues was found to drive the polarization of M0 macrophages and infiltration of M2 macrophages into the TME, resulting in worse prognosis in ESCC patients. CSF-1, a vital factor inducing M0-to-M2 polarization, was upregulated via a FOXO1-mediated mechanism. RNA sequencing results corroborated that the FOXO1-induced macrophages exhibited similar molecular signatures to the IL4-stimulated M2 macrophages. The transwell assays showed that FOXO1 promoted the migration of M2 macrophages via CCL20 secretion, which could be inhibited using an anti-CCL20 antibody. FOXO1(+) tumor-induced M2 macrophages promoted tumor proliferation via the FAK-PI3K-AKT pathway and the PI3K inhibitor could effectively impede the oncogenical process. Conclusions: FOXO1 facilitated M0-to-M2 polarization and the recruitment of M2 macrophages in the TME via the transcriptional modulation of CCL20 and CSF-1. Our data deciphered the FOXO1-dependent mechanism in M2 macrophage infiltration in the TME of ESCC, which has implications for the development of novel prognostic and therapeutic targets to optimize the current treatment against ESCC.

Keywords: FOXO1; M2 macrophage; cancer progression; esophageal squamous cell carcinoma.

Chemokine and its receptors play important roles in laryngeal cancer development and progression. CCR6 is the receptor of CCL20 chemokine, but its function in laryngeal cancer is not known. The aim of this study is to explore the roles of CCR6 and its regulation mechanism in laryngeal cancer. We found CCR6 expression was higher in laryngeal cancer tissues compared with their normal controls, so did in laryngeal cancer cells. Cellular function indicated that down-regulation of CCR6 in laryngeal cancer cells could suppress cell proliferation and metastasis. Further research showed that CCR6 could activate p38, which was related with the changes of microRNA (miRNA) profile in laryngeal cancer cells. We also found that CCR6 was positively associated with miR-20a-5p, miR-489 and negatively related to miR-29-3p, miR-632 and miR-1276 in laryngeal cancer tissues.

BACKGROUND

Migration of lymphocytes into and through the mucosal immune system depends upon adhesion molecules to attract circulating cells and chemokines to stimulate diapedesis into tissues. Decreased enteral stimulation significantly reduces mucosal addressin cellular adhesion molecule-1 (MAdCAM-1) levels, an adhesion molecule critical for homing of T and B cells to Peyer's patches (PP), which reduces PP and intestinal T and B cells. We studied the effect of type and route of nutrition on tissue specific chemokines in PP (CXCL-12, -13 and CCL-19, -20 and -21), small intestine (SI; CCL-20, -25 and -28) and lung (CXCL-12, CCL-28).

METHODS

Intravenously cannulated male Institute of Cancer Research (ICR) mice were randomized to chow or parenteral nutrition (PN) for 5 days. PP, SI, and lung chemokine mRNA levels were measured using real-time qRT-polymerase chain reaction, and analyzed semiquantitatively by the DeltaDeltaCt method. Protein levels were quantified using enzyme-linked immunosorbent assay (ELISA) techniques, and groups compared using Student's t-test.

RESULTS

PP CXCL13 protein significantly decreased, whereas CCL21 protein increased significantly in the parenterally fed group. Parenteral feeding significantly decreased SI CCL20 and CCL 25 protein levels. CCL28 decreased significantly in the SI and lung of intravenously fed animals. mRNA levels changed in the opposite direction (compared with protein) for all chemokines except CCL28.

CONCLUSIONS

Decreased enteral stimulation significantly alters key mucosal immune chemokine protein levels at multiple sites. In general, PN (and concomitant lack of enteral stimulation) results in decreased levels of chemokines that control lymphocyte migration within the mucosal immune system.

The follicle-associated epithelium (FAE) overlying the Peyer's patches and the microfold cells (M cells) within it are important sites of antigen transcytosis across the intestinal epithelium. Using a meta-analysis approach, we identified a transcriptional signature that distinguished the FAE from a large collection of mouse cells and tissues. A co-expressed cluster of 21 FAE-specific genes was identified, and the analysis of the transcription factor binding site motifs in their promoter regions indicated that these genes shared an underlying transcriptional programme. This cluster contained known FAE- (Anxa10, Ccl20, Psg18 and Ubd) and M-cell-specific (Gp2) genes, suggesting that the others were novel FAE-specific genes. Some of these novel candidate genes were expressed highly by the FAE and M cells (Calcb, Ces3b, Clca2 and Gjb2), and others only by the FAE (Ascl2, Cftr, Fgf15, Gpr133, Kcna1, Kcnj15, Mycl1, Pgap1 and Rps6kl). We also identified a subset of novel FAE-related genes that were induced in the intestinal epithelium after receptor activator of nuclear factor (NF)-κB ligand stimulation. These included Mfge8 which was specific to FAE enterocytes. This study provides new insight into the FAE transcriptome. Further characterization of the candidate genes identified here will aid the identification of novel regulators of cell function in the FAE.

While chemokines are clearly important in the generation of protective immunity, the role of individual chemokines in the control of bacterial infection is still poorly understood. In this study, we investigated the role of macrophage inflammatory protein (MIP)-3alpha/CCL20, a chemokine that attracts activated T and B lymphocytes and immature dendritic cells, in host responses to bacterial infection. CCL20 production was induced in subcutaneous tissue in the BALB/c mouse in response to Salmonella enteritidis, Staphylococcus aureus and zymosan, with S. enteritidis being the most potent. S. enteritidis induced CCL20 production in the spleen following either oral administration or injection into the peritoneal cavity. In contrast, no increase was observed in the Peyer's patches. In this model, following intraperitoneal injection, dose-dependent colonization of the spleen and Peyer's patches by S. enteritidis, expression of IFNgamma and IL-4, and production of antibodies against the S. enteritidis surface antigen SefA were observed. Prior treatment with neutralizing antibodies against CCL20 enhanced bacterial dissemination to the spleen and Peyer's patches and strongly biased the IFNgamma/IL-4 ratio towards a type 2 profile in the spleen, while the humoral response was unaffected. In contrast, treatment with neutralizing anti-MIP-1alpha/CCL3 antibodies enhanced the bacterial burden in the Peyer's patches but not in the spleen, had no significant effect on the cytokine ratio, but significantly inhibited anti-SefA production. Together, these results demonstrate an important role for CCL20 in the control of bacterial infection and more specifically in the regulation of cell-mediated immunity against intracellular bacteria such as S. enteritidis.

The aim of this study was to examine the effects of macrophage secretions on global gene expression in human preadipocytes using microarrays. Preadipocytes were cultured with unconditioned or conditioned medium from U937 macrophages, and gene expression examined with Agilent arrays (43,000 probes). 472 transcripts were differentially regulated (>2-fold difference; P<0.05) between preadipocytes in the conditioned medium compared to the unconditioned; 401 were upregulated and 71 downregulated. The upregulated transcripts were particularly linked to inflammation, including IL-1β, IL-6, and CCL20 (16.8-, 10.0-, and 8.9-fold increases, respectively) together with matrix metalloproteinases (MMP3, MMP9 and MMP12). Major pathways regulated by the conditioned medium were linked to inflammation, macrophage infiltration and lipid accumulation. Network analysis identified NFkB and IL-1β as central nodes in the upregulation of multiple inflammation-related genes. Treatment with an IL-1β neutralising antibody abolished the stimulation of IL-6 secretion by conditioned medium, indicating that IL-1β is a key regulator of preadipocyte IL-6 production. Macrophages evoke extensive changes in preadipocyte gene expression.

Cigarette smoke (CS) exposure is known to increase infection rates, but the mechanisms are not well understood. These studies tested the hypothesis that CS exposure would impair antimicrobial activity of apical conditioned media from human airway (BEAS-2B) cultures by reducing induction and release of the antimicrobial peptide CCL20. BEAS-2B cultures were exposed to CS extract and assayed for temporal and physical characteristics of release as well as for antimicrobial activity. E. coli were exposed to Beas-2B-conditioned media (BCM) and subsequent bacterial colonies were enumerated. In time course studies TLR-agonist-induced CCL20 transcription and release were rapid, of short duration and release was consistently targeted to the apical/luminal compartment. Cells treated with CS extract had diminished release of CCL20 under both constitutive and toll-like receptor (TLR) agonist stimulating conditions. Exposure of the cells to CS significantly reduced the antimicrobial activity in BCM and neutralizing antibodies to CCL20 brought antibacterial activity back to baseline levels demonstrating that antimicrobial activity in this culture system was primarily attributable to CCL20. These studies add to the understanding of CCL20 as a mucosal antimicrobial and improve insight into a likely mechanism linking infection to CS exposure.

Chemokines play an important role in the pathogenesis of non-small cell lung cancer (NSCLC). Although the deregulations of chemokines have been reported to be associated with the development and progression of many human cancers including lung cancer, polymorphisms of chemokine genes have not been examined with the survival of NSCLC. We systematically investigated associations of 23 common potentially functional SNPs in the key chemokine genes (CCL2, CCL5, CCL8, CCL20, CCL22, CXCL1, CXCL6, CXCL9 and CXCL12) with the survival of NSCLC in a case cohort of 568 NSCLC patients in a Chinese population. The results showed that variant genotypes of CCL2 rs3760396 and CCL8 rs3138035 were associated with a significantly decreased risk of death for NSCLC (dominant model: adjusted HR=0.65, 95% CI=0.48-0.89 for rs3760396; dominant model: adjusted HR=0.65, 95% CI=0.49-0.86 for rs3138035), while CXCL12 rs1804429 was associated with an increased risk of death for NSCLC (CC vs AA: adjusted HR=6.03, 95% CI=1.44-25.24). Further stepwise regression analysis suggested that only rs3138035, a SNP located at 5'-flanking region of CCL8, was an independently favorable factor for the prognosis of NSCLC and the protective effect was more evident in smokers (adjusted HR=0.61, 95% CI=0.42-0.87), patients with squamous cell cancer (adjusted HR=0.58, 95% CI=0.35-0.96), patients with early stage (adjusted HR=0.32, 95% CI=0.15-0.67) and patients treated with surgical operation (adjusted HR=0.47, 95% CI=0.31-0.71). In addition, the interaction analysis demonstrated that stage and surgical operation interacted with the genetic effect of rs3138035 in relation to NSCLC survival (adjusted P(interaction)=0.02 and 0.01, respectively). These findings suggest that CCL8 rs3138035 may be one of the candidate biomarkers for NSCLC survival and may modify death risk associated with stage and surgical operation. Larger studies incorporating functional evaluations are warranted to validate our findings.

Effector T cell migration through the tissue extracellular matrix (ECM) is an important step of the adaptive immune response and in the development of inflammatory diseases. However, the mechanisms involved in this process are still poorly understood. In this study, we addressed the role of a collagen receptor, the discoidin domain receptor 1 (DDR1), in the migration of Th17 cells. We showed that the vast majority of human Th17 cells express DDR1 and that silencing DDR1 or using the blocking recombinant receptor DDR1:Fc significantly reduced their motility and invasion in three-dimensional (3D) collagen. DDR1 promoted Th17 migration by activating RhoA/ROCK and MAPK/ERK signaling pathways. Interestingly, the RhoA/ROCK signaling module was required for MAPK/ERK activation. Finally, we showed that DDR1 is important for the recruitment of Th17 cells into the mouse dorsal air pouch containing the chemoattractant CCL20. Collectively, our results indicate that DDR1, via the activation of RhoA/ROCK/MAPK/ERK signaling axis, is a key pathway of effector T cell migration through collagen of perivascular tissues. As such, DDR1 can contribute to the development of Th17-dependent inflammatory diseases.

Endometrial Cancer is the most common female genital tract malignancy, its pathogenesis is complex, not yet fully described. To identify key genes of Endometrial Cancer we downloaded the gene chip GSE17025 from the Gene Expression Omnibus database. Differentially expressed genes (DEGs) were identified through the GEO2R analysis tool. Functional and pathway enrichment analysis were performed for DEGs using DAVID database. The network of protein–protein-interaction (PPI) was established by STRING website and visualized by Cytoscape. Then, functional and pathway enrichment analysis of DEGS were performed by DAVID database. A total of 1000 significant differences genes were obtained, contain 362 up-regulated genes and 638 down-regulated genes. PCDH10, SLC6A2, OGN, SFRP4, TRH, ANGPTL, FOSB are down-regulated genes. The gene of IGH, CCL20, ELF5, LTF, ASPM expression level in tumor patients are up-regulated. Biological function of enrichment include metabolism of xenobiotics by cytochrome P450, MAPK signaling pathway, Serotonergic synapse, Protein digestion and absorption, IL-17 signaling pathway, Chemokine signaling pathway, HIF-1 signaling pathway, p53 signaling pathway. All in all, the current study to determine endometrial differentially expressed genes and biological function, comprehensive analysis of intrauterine membrane carcinoma pathogenesis mechanism, and might be used as molecular targets and diagnostic biomarkers for the treatment of endometrial cancer.

BACKGROUND

Lupus nephritis is an autoimmune inflammatory disease and urgently needs effective anti-inflammation therapies. A20, tumor necrosis factor alpha induced protein 3 (TNFAIP3), is a key negative regulator of inflammation, however whether A20 can regulate lupus nephritis has not been clarified. This study aimed at investigating the potential therapeutic effect of A20 on renal inflammation in mouse pristine model oflupus.

RESULTS

Female BALB/c mice were intraperitoneally injected with pristine to establish lupus renal injury. The levels of serum IL-1β, IL-6 and autoantibodies and the degrees of renal injury and CCL2 and F4/80 levels were measured. The levels of the NF-κB and NLRP3 inflammasome activation in peritoneal macrophages were determined. We found that injection with pristine increased the levels of serum IL-1β, IL-6, autoantibodies and CCL20 and F4/80 expression in the kidney and induced renal injury, accompanied by enhancing the NF-κB and NLRP3 inflammasome activation in macrophages of mice. In contrast, treatment with Ad-A20, but not with Ad-control, significantly mitigated pristine-induced inflammatory responses and renal injury,and reduced the NF-κB and NLRP3 inflammasome activation in macrophages in mice.

CONCLUSIONS

Our data indicated that induction of A20 overexpression inhibited pristane induced lupus inflammation and renal injury in mice and may be a new therapeutic strategy for treatment of lupus nephritis.

Most gallbladder cancer (GBC) cases arise in the context of gallstones, which cause inflammation, but few gallstone patients develop GBC. We explored inflammation/immune-related markers measured in bile and serum in GBC cases compared to gallstone patients to better understand how inflammatory patterns in these two conditions differ. We measured 65 immune-related markers in serum and bile from 41 GBC cases and 127 gallstone patients from Shanghai, China, and calculated age- and sex-adjusted odds ratios (ORs) and 95% confidence intervals (95% CIs) for GBC versus gallstones. We then focused on the markers that were significantly elevated in bile and serum to replicate the findings in serum from 35 GBC cases and 31 gallstone controls from Chile. Comparing the highest versus lowest quantile, 15 markers (23%) were elevated in both serum and bile from GBC versus gallstone patients in the Shanghai study (p<0.05). The strongest OR was for CXCL8 (interleukin-8) in serum (96.8, 95% CI: 11.9-790.2). Of these 15 markers, 6 were also significantly elevated in serum from Chile (CCL20, C-reactive protein, CXCL8, CXCL10, resistin, serum amyloid A). Pooled ORs from Shanghai and Chile for these 6 markers ranged from 7.2 (95% CI: 2.8-18.4) for CXCL10 to 58.2 (95% CI: 12.4-273.0) for CXCL8. GBC is associated with inflammation above and beyond that generated by gallstones alone. This local inflammatory process is reflected systemically. Future longitudinal studies are needed to identify the key players in cancer development, which may guide translational efforts to identify individuals at high risk of developing GBC.

Effects of bacterial pathogens on the production of macrophage inflammatory protein 3alpha (MIP-3alpha) and MIP-3beta from human peripheral blood neutrophils were investigated. Neutrophils produced both chemokines by coincubation with either gram-positive or gram-negative bacteria. Neutrophils may initiate antigen-specific immune responses through the release of these chemokines that are capable of promoting selective recruitment of dendritic cells and T-cell subsets.

BACKGROUND

Graves' disease (GD) is a common autoimmune disease involving the thyroid gland. The altered balance of pro- and anti-inflammatory cytokines plays an important role in the pathogenesis of GD. Chemokine (C-C motif) ligand 20 (CCL20) is important for interleukin-17 (IL-17) signal activation and a potent chemoattractant for Th17 cells. Meanwhile, Osteopontin (OPN), a broadly expressed pleiotropic cytokine, has been implicated in GD through inducing Th1-involved response to enhance the production of proinflammatory cytokines and chemokines, but little is known about the role of OPN in regulating CCL20 and IL-17 signaling.

OBJECTIVE

This study sought to explore the possibility of CCL20 level as a biomarker for GD, as well as investigate the role of OPN in regulating CCL20 production.

METHODS

Fifty untreated GD patients, fifteen euthyroid GD patients, twelve TRAb-negative GD patients and thirty-five healthy control donors were recruited. OPN, CCL20 and other clinical GD diagnosis parameters were measured. CD4+T cells were isolated from peripheral blood mononuclear cells (PBMCs) using antibody-coated magnetic beads. Enzyme-linked immune-sorbent assay and quantitative polymerase chain reaction were used to determine CCL20 expression level.

RESULTS

We found that the plasma CCL20 level was enhanced in GD patients and decreased in euthyroid and TRAb-negative GD patients. In addition, CCL20 level correlated with GD clinical diagnostic parameters and plasma OPN level. Moreover, we demonstrated that recombinant OPN and plasma from untreated GD patients increased the expression of CCL20 in CD4+T cells, which could be blocked by OPN antibody. Furthermore, we found that the effect of OPN on CCL20 expression was mediated by β3 integrin receptor, IL-17, NF-κB and MAPK pathways.

CONCLUSIONS

These results demonstrated that CCL20 might serve as a biomarker for GD and suggested the possible role of OPN in induction of CCL20 expression.

In our previous study, we found that cycloheximide (CHX) induces hepatocellular necrosis as well as hepatocellular apoptosis. This article evaluates the role of Kupffer cells on cycloheximide-induced hepatic injury using gadolinium chloride (GdCl(3)) for the inhibition of Kupffer cells. One group of rats was treated with CHX (CHX group), and another was treated with GdCl(3) before being treated with the same dose of CHX (GdCl(3)/CHX group). The necrotic change in the GdCl(3)/CHX group was exacerbated under the induction of hepatocellular apoptosis by the CHX treatment. A substantial diminution of the number of ED1- or ED2-positive cells was demonstrated in the GdCl(3)/CHX group compared to the CHX group. In addition, the degree of decrease in ED2-positive cells was more apparent than that in ED1-positive cells. Increases in the mRNA levels of IL-10 and Stat3 were observed in the CHX group, but not in the GdCl(3)/CHX group. On the other hand, the hepatic mRNA levels of chemokines and adhesion molecules such as Ccl20, LOX-1, and E-selectin were significantly increased only in the GdCl(3)/CHX group. Thus, Kupffer cell inactivation by the GdCl(3) treatment leads to a loss of the capacity to produce IL-10, supposedly resulting in the enhancement of pro-inflammatory cytokine activities such as tumor necrosis factor (TNF) signaling. These events are suggested to be a factor of the inflammatory exacerbation in the livers of the GdCl(3)/CHX group. In conclusion, Kupffer cells may play a role in protecting hepatic necroinflammatory changes by releasing anti-inflammatory cytokines following the hepatocellular apoptosis resulting from CHX treatment.

Microarray analysis was used to investigate the effects of Bacillus anthracis lethal toxin (LT) on human neutrophil-like NB-4 cells to identify markers of intoxication. Genes down-regulated after a 2h LT exposure included those encoding chemokines and transcription factors. Significant decreases in the mRNA of interleukin-8, CCL20, CCL3 and CCL4, which are important chemoattractants for immune cells, were observed using real-time PCR (12.3, 4.0, 4.1 and 2.2-fold (p<0.05), respectively). The decreases were more pronounced at 4 and 8h and were LT-specific. Decreases in chemokine protein levels were evident after 24h and were sensitive to low concentrations of LT. Co-incubation with an anti-lethal factor mAb at 500 and 250ng/ml restored levels of interleukin-8 to 100% and 50%, respectively. The results reveal that LT suppresses the cellular immune response and that this assay is a useful tool for the analysis of toxin-neutralising antibody activity in a biologically relevant system.

The Ah receptor (AHR) is directly involved in the regulation of both innate and adaptive immunity. However, these activities are poorly understood at the level of gene regulation. The chemokine (c-c motif) ligand 20 (CCL20) plays a nonredundant role in the chemoattraction of C-C motif receptor 6 expressing cells (eg, T cells and others). A survey of promoter regions of chemokine genes revealed that there are several putative dioxin responsive elements in the mouse Ccl20 promoter. The addition of an AHR agonist along with lipopolysaccharide (LPS) to cultured primary peritoneal macrophages results in synergistic induction of both Ccl20 mRNA and protein, compared with each compound alone. Through the use of macrophage cultures derived from Ahr(-) (/) (-) and Ahr(nls/nls) mice, it was established that expression of the AHR and its ability to translocate into the nucleus are necessary for AHR ligand-mediated synergistic induction of Ccl20. Gel shift analysis determined that a potent tandem AHR binding site ~3.1 kb upstream from the transcriptional start site can efficiently bind the AHR/ARNT (aryl hydrocarbon receptor/AHR nuclear translocator) heterodimer upon activation with a number of AHR agonists. Furthermore, studies reveal that LPS increases AHR levels on the Ccl20 promoter while decreasing HDAC1 occupancy. The level of Ccl20 constitutive expression in the colon is greatly attenuated in Ahr(-) (/) (-) mice. These studies suggest that the presence of AHR ligands during localized inflammation may augment chemokine expression, thus participating in the overall response to pathogens.