Chemokines are secreted proteins with essential roles in leukocyte trafficking and cell migration during embryogenesis. CXCL14 displays a degree of evolutionary conservation unmatched by any other chemokine except for SDF1(CXCL12). However, its role during embryogenesis has not been studied. Here we describe the expression pattern of mouse and chicken CXCL14 during embryogenesis and compare it with that of SDF1. CXCL14 is widely expressed in embryonic ectoderm and shows a restricted and dynamic expression pattern in paraxial mesoderm, mesonephros, neural tube, and limbs. During limb development, CXCL14 marks a unique connective tissue subset that surrounds developing tendons. Comparison of CXCL14 and SDF1 reveals mostly non-overlapping or complementary expression patterns, suggesting an interactive regulation of developmental processes by these two chemokines. Our study identifies CXCL14 as a novel marker of tendon connective tissue and provides a conceptual framework for the coordinated action of two highly conserved chemokines in embryonic development.

The mitogen-activated protein kinase (MAPK) family comprises ERK, JNK, p38 and ERK5 (big-MAPK, BMK1). UV irradiation of squamous cell carcinoma cells induced up-regulation of gene expression of chemokine BRAK/CXCL14, stimulated p38 phosphorylation, and down-regulated the phosphorylation of ERK. Human p38 MAPKs exist in 4 isoforms: p38 alpha, beta, gamma and delta. The UV stimulation of p38 phosphorylation was not inhibited by the presence of SB203580 or PD169316, inhibitors of p38 alpha and beta, suggesting p38 phosphorylation was not dependent on these 2 isoforms and that p38 gamma and/or delta was responsible for the phosphorylation. In fact, inhibition of each of these 4 p38 isoforms by the introduction of short hairpin (sh) RNAs for respective isoforms revealed that only shRNA for p38 delta attenuated the UV-induced up-regulation of BRAK/CXCL14 gene expression. In addition, over-expression of p38 isoforms in the cells showed the association of p38 delta with ERK1 and 2, concomitant with down-regulation of ERK phosphorylation. The usage of p38 delta isoform by UV irradiation is not merely due to the abundance of this p38 isoform in the cells. Because serum deprivation of the cells also induced an increase in BRAK/CXCL14 gene expression, and in this case p38 alpha and/or beta isoform is responsible for up-regulation of BRAK/CXCL14 gene expression. Taken together, the data indicate that the respective stress-dependent action of p38 isoforms is responsible for the up-regulation of the gene expression of the chemokine BRAK/CXCL14.

Chemokines are a family of proteins originally identified for their activity promoting the recruitment of leukocytes to inflammatory sites. Recent evidence indicates that chemokines and their receptors may also regulate key developmental processes. In this paper we report the expression and regulation of the chemokine CXCL14 during Xenopus laevis embryogenesis. CXCL14 is first detected in several ectoderm derivatives, the dorsal aspect of the retina, the cement gland and the hatching gland. Later in development, additional domains of expression include the head mesenchyme and the medial ventral aspect of the otic vesicle. CXCL14 expression in the ectoderm is regulated by both Bmp and canonical Wnt signaling. In the hatching gland CXCL14 is co-expressed with the transcription factor Pax3. Using gain of function and knockdown approaches in whole embryos and animal explants we show that Pax3 is both necessary and sufficient for CXCL14 expression in this domain of the ectoderm.

CXCL14 is a CXC chemokine family that exhibits antimicrobial activity and contains an amphipathic cationic α-helical region in the C-terminus, a characteristic structure of antimicrobial peptides (AMPs). In this study, we designed three analogs of CXCL14CXCL14-C17) corresponding to the C-terminal α-helix of CXCL14, which displayed potential antimicrobial activity against a wide variety of gram-negative and gram-positive bacteria with minimum inhibitory concentrations of 4-16 μM without mammalian cell toxicity. Furthermore, two CXCL14-C17 analogs (CXCL14-C17-a1 and CXCL14-C17-a3) with improved cell selectivity were engineered by introducing Lys, Arg, or Trp in CXCL14-C17. Additionally, CXCL14-C17 analogs showed much greater synergistic effect (FICI: 0.3125-0.375) with chloramphenicol and ciprofloxacin against multidrug-resistant Pseudomonas aeruginosa (MDRPA) than LL-37 did (FICI: 0.75-1.125). CXCL14-C17 analogs were more active against antibiotic-resistant bacteria including methicillin-resistant Staphylococcus aureus (MRSA), MDRPA, and vancomycin-resistant Enterococcus faecium (VREF) than LL-37 and melittin. In particular, CXCL14-C17-a2 and CXCL14-C17-a3 completely inhibited the biofilm formation at sub-MIC and all of the peptides were able to eliminate pre-formed biofilm as well. Membrane depolarization, flow cytometry, sytox green uptake, ONPG hydrolysis and confocal microscopy revealed the possible target of the native peptide (CXCL14-C17) to likely be intracellular, and the amphipathic designed analogs targeted the bacterial membrane. CXCL14-C17 also showed DNA binding characteristic activity similar to buforin-2. Interestingly, CXCL14-C17-a2 and CXCL14-C17-a3 effectively inhibited the production and expression of nitric oxide (NO), tumor necrosis factor (TNF)-α, interleukin (IL)-6, and monocyte chemoattractant protein (MCP)-1 from lipopolysaccharide (LPS)-stimulated RAW264.7 cells, suggesting that these peptides could be promising anti-inflammatory and antimicrobial agents.

CXCL14 is a chemokine family member that is involved in various cellular responses in addition to immune cell activation. Although constitutive CXCL14 expression in normal epithelial cells may help protect against infection by activating immune systems, its expression in cancer cells has raised controversy regarding its possible role in tumorigenesis. However, the underlying mechanisms for this disparity remain unknown. Investigation of cellular CXCL14 binding properties might increase our understanding of the peptide's roles in tumorigenesis. In the present study, we found that CXCL14 binds to various cell types. Interestingly, binding to NCI-H460 cells was prevented by heparan sulfate and N-acetyl neuraminic acid. Next, we examined effect of CXCL14 binding in NCI-H460 and NCI-H23. CXCL14 enhanced proliferation and migration in NCI-H460 but had no effect on NCI-H23. A reporter gene assay with various transcription factor response elements revealed that only nuclear factor-κB (NF-κB) signaling was activated by CXCL14 in NCI-H460 cells, which was blocked by BAPTA-AM, TPCA-1, and brefeldin A. Exogenous expression of some glycoproteins such as syndecan-4, podoplanin, and CD43 in these cells enhanced CXCL14 binding and NF-κB activity. Collectively, these results demonstrate that CXCL14 binding to glycoproteins harboring heparan sulfate proteoglycans and sialic acids leads proliferation and migration of some cancer cells.

Long non-coding RNAs are involved with development and progression of cancer, and the advance of microarray technology allows the researchers to investigate the complete expression profile of lncRNA in various kinds of sample. We enrolled 5 male primary HCC cases with chronic HBV infection and the HCC and normal tissues have been obtained during the resection surgery. After total RNA extraction, the lncRNA microarray analysis was conducted to determine the lncRNA and mRNA expression signals. 612 lncRNAs and 1,064 mRNAs were significantly up-regulated in HCC tissue while 656 lncRNAs and 1,532 mRNAs were down-regulated in HCC tissues. Compared with normal tissues, XLOC_007433 (fold change: 12.80) and AC144449.1 (fold change: 27.20) were the most over- and under-expressed lncRNAs in HCC tissues. As for the mRNA, THBS4 (fold change:41.13) and CXCL14 (fold change: 58.03) were the most over- and under-expressed mRNAs in HCC tissues when comparing with their normal counterparts. In total, 4,552 pairs of lncRNA-mRNA were identified and the co-expression network was constructed. Moreover, the gene ontology enrichment analysis showed that the significantly different transcript between HCC and normal tissues were mainly associated with response to wounding, inflammatory response, protein hetrodimerization activity, response to stress which involved with biological process and molecular function. The pathway analysis suggested that the most significant pathways consisted of alcoholism, regulatory RNA pathways and RNA polymerase transcription. Several novel differentially expressed lncRNAs and mRNAs were identified in the present study.

OBJECTIVE

Recently, the microarray analysis has been an important tool used for studying the cancer type, biological mechanism, and diagnostic biomarkers. There are several machine-learning methods being used to construct the prognostic model based on the microarray data sets. However, most of these previous studies were focused on the supervised classification for predicting the clinical type of patients. In this study, we investigate whether or not the expression level of some significant genes identified can be used to predict the clinical metastases time of patients.

METHODS

We have used a regression method to remodel the data set of breast cancer published in 2002. Some significant genes were ranked and selected based on a wrapper method with 10-fold cross-validation procedure and the selected genes were used to fit the support vector regression (SVR) model. This method could model the relationship between the significant gene expression value and the clinical metastases time of breast cancer.

RESULTS

44 significant genes are selected for building the regression model and the corresponding cross-validated correlation coefficient obtained is 0.82 which is much superior to those reported previously by others using some different data sets. Moreover, there are two breast cancer related genes (the ligand 14 of the chemokine C-X-C motif (CXCL14) and estrogen receptor gene (ER)) selected in the gene set and one of them is never been included in the other data sets.

CONCLUSIONS

In this report, we have shown that the expression level of some significant genes identified could strongly correlate with the clinical metastases time of breast cancer patients. The 44 selected genes may be used as a benchmark to evaluate the risk of recurrence of breast cancer.

OBJECTIVE

To characterize the chemokine pattern in metastatic salivary adenoid cystic carcinoma (SACC).

METHODS

Real-time polymerase chain reaction (RT-PCR) was used to compare chemokine and chemokine receptor gene expression in two SACC cell lines: SACC-83 and SACC-LM (lung metastasis). Chemokines and receptor genes were then screened and their expression pattern characterized in human tissue samples of non-recurrent SACC and recurrent SACC with perineural invasion.

RESULTS

Expression of chemokine receptors C5AR1, CCR1, CCR3, CCR6, CCR7, CCR9, CCR10, CXCR4, CXCR6, CXCR7, CCRL1 and CCRL2 were higher in SACC-83 compared to SACC-LM. CCRL1, CCBP2, CMKLR1, XCR1 and CXCR2 and 6 chemokine genes (CCL13, CCL27, CXCL14, CMTM1, CMTM2, CKLF) were more highly expressed in tissues of patients without tumor recurrence/perineural invasion compared to those with tumor recurrence. CCRL1 (receptor), CCL27, CMTM1, CMTM2, and CKLF (chemokine) genes were more highly expressed in SACC-83 and human tissues of patients without tumor recurrence/perineural invasion.

CONCLUSIONS

CCRL1, CCL27, CMTM1, CMTM2 and CKLF may play important roles in the development of tumor metastases in SACC.

CXCL14, chemokine (C-X-C motif) ligand 14, is a novel highly conserved chemokine with unique features. Despite exhibiting the typical chemokine fold, it has a very short N-terminus of just two amino acid residues responsible for chemokine receptor activation. CXCL14 actively participates in homeostatic immune surveillance of skin and mucosae, is linked to metabolic disorders, fibrotic lung diseases, and possesses strong anti-angiogenic properties in early tumor development. In this work, we investigated the interaction of CXCL14 with various glycosaminoglycans (GAGs) by NMR spectroscopy, microscale thermophoresis, analytical heparin affinity chromatography, and in silico approaches to understand the molecular basis of GAG-binding. We observed different GAG binding modes specific for the GAG type used in the study. In particular, the CXCL14 epitope for heparin suggests a binding pose distinguishable from the ones of the other GAGs investigated (hyaluronic acid, chondroitin sulfate-A/C, -D, dermatan sulfate. This observation is also supported by computational methods that included molecular docking, molecular dynamics and free energy calculations. Based on our results, we suggest that distinct GAG sulfation patterns confer specificity beyond simple electrostatic interactions usually considered to represent the driving forces in protein-GAG interactions. The CXCL14-GAG system represents a promising approach to investigate the specificity of GAG protein interactions, which represents an important topic for developing the rational approaches to novel strategies in regenerative medicine.

To investigate the molecular mechanisms of lung cancer-induced bone metastasis, we established a bone-seeking subclone (HARA-B4) from a human squamous lung cancer cell line (HARA) using an in vivo selection method. We compared comprehensive gene expression profiles between HARA and HARA-B4, and identified the critical factors for the formation of bone metastasis using in vitro and in vivo assays. The number of bone metastatic colonies in the hind legs was significantly higher in HARA-B4-inoculated mice than in HARA-inoculated mice at 4 weeks after inoculation. In addition, visceral (adrenal) metastases were not found in HARA-B4-inoculated mice at autopsy, suggesting an increase in cancer cell tropism to bone in HARA-B4. Based on a comprehensive gene expression analysis, the expression level of CXC chemokine ligand 14 (CXCL14) was 5-fold greater in HARA-B4 than in HARA. Results of a soft agar colony formation assay showed that anchorage-independent growth ability was 4.5-fold higher with HARA-B4 than with HARA. The murine pre-osteoblast cell line MC3T3-E1 and the pre-osteoclast/macrophage cell line RAW264.7 migrated faster toward cultured HARA-B4 cells than toward HARA cells in a transwell cell migration assay. Interestingly, CXCL14 was shown to be involved in all events (enhancement of cancer cell tropism to the bone, anchorage-independent growth and/or recruitment of bone marrow cells) based on siRNA experiments in HARA-B4 cells. Furthermore, in clinical specimens of lung cancer-induced bone metastasis, expression of CXCL14 was observed in the tumor cells infiltrated in bone marrow in all specimens examined. CXCL14 was able to promote bone metastasis through enhancement of cancer cell tropism to the bone and/or recruitment of bone marrow cells around metastatic cancer cells.

Approximately, one-third to one-half of children with chronic granulomatous disease (CGD) develop gastrointestinal inflammation characteristic of idiopathic inflammatory bowel disease (IBD), usually Crohn's disease. We hypothesized that the overall IBD genetic risk, determined by IBD genetic risk score (GRS), might in part determine IBD development in CGD.

We reviewed medical records to establish IBD diagnoses in CGD subjects seen at NIAID. IBD risk single nucleotide polymorphism genotypes were determined using the Immunochip, and GRS were estimated by Mangrove.

Among 157 white patients with CGD, 55 were confirmed, 78 excluded, and 24 were uncertain for IBD. Two hundred one established, independent European IBD risk single nucleotide polymorphisms passed quality control. After sample quality control and removing non-IBD CGD patients with perianal disease, mean GRS for 40 unrelated patients with CGD-IBD was higher than 53 CGD non-IBD patients (in log2-scale 0.08 ± 1.62 versus -0.67 ± 1.64, P = 0.026) but lower than 239 IBD Genetics Consortium (IBDGC) young-onset Crohn's disease cases (0.76 ± 1.60, P = 0.025). GRS for non-IBD CGD was similar to 609 IBDGC controls (-0.69 ± 1.60, P = 0.95). Seven established IBD single nucleotide polymorphisms were nominally significant among CGD-IBD versus CGD non-IBD, including those near LACC1 (P = 0.005), CXCL14 (P = 0.007), and TNFSF15 (P = 0.016).

The weight of the common IBD risk alleles are significant determinants of IBD in CGD. However, IBD risk gene burden among CGD children with IBD is significantly lower than that in nonsyndromic pediatric Crohn's disease, congruent with the concept that defective superoxide production in CGD is also a major IBD risk factor. Individual IBD genes might interact with the CGD defect to cause IBD in CGD.

We reported previously that the forced expression of the chemokine BRAK, also called CXCL14 in head and neck squamous cell carcinoma (HNSCC) cells decreased the rate of tumor formation and size of tumor xenografts compared with mock-vector treated cells in athymic nude mice or in severe combined immunodeficiency mice. This suppression occurred even though the growth rates of these cells were the same under in vitro culture conditions, suggesting that a high expression level of the gene in tumor cells is important for the suppression of tumor establishment in vivo. The aim of this study was to determine whether CXCL14/BRAK transgenic mice show resistance to tumor cell xenografts or not. CXCL14/BRAK cDNA was introduced into male C57BL/6 J pronuclei, and 10 founder transgenic mice (Tg) were obtained. Two lines of mice expressed over 10 times higher CXCL14/BRAK protein levels (14 and 11 ng/ml plasma, respectively) than normal blood level (0.9 ng/ml plasma), without apparent abnormality. The sizes of Lewis lung carcinoma and B16 melanoma cell xenografts in Tg mice were significantly smaller than those in control wild-type mice, indicating that CXCL14/BRAK, first found as a suppressor of tumor progression of HNSCC, also suppresses the progression of a carcinoma of other tissue origin. Immunohistochemical studies showed that invasion of blood vessels into tumors was suppressed in tumor xenografts of CXCL14/BRAK Tg mice. These results indicate that CXCL14/BRAK suppressed tumor cell xenografts by functioning paracrine or endocrine fashion and that CXCL14/BRAK is a very promising molecular target for tumor suppression without side effects.

BRAK/CXCL14 is a chemokine that is expressed in many normal cells and tissues but is absent from or expressed at very low levels in transformed cells and cancerous tissues including head and neck squamous cell carcinoma (HNSCC). We reported previously that the forced expression of BRAK/CXCL14 in HNSCC cells decreased the rate of tumor formation and size of tumor xenografts in athymic nude mice and SCID mice, suggesting that expression level of the gene is important for tumor suppression. In order to study the regulatory mechanisms governing the expression of this gene, we determined the transcriptional start site and promoter motifs of the gene. The major transcriptional start site determined by 5'rapid amplification of cDNA end method was located 283 bp downstream of the first proposed site of the gene. Determination of luciferase activities of reporter gene constructs with various deletions or mutations showed that an atypical TATA-like sequence, TATTAA was essential for the transcription of the gene and that the AP-1 binding sequence and tandem GC box were necessary for stimulating the expression of the gene in human squamous epithelial cells. The human DNA region was highly homologous (95% base identity) to the mouse gene. In addition, okadaic acid, an inhibitor of serine/threonine phosphatases 1, 2A and 2B, stimulated TATTAA sequence and AP-1 binding-sequence dependent promoter activity as well as increased the level of BRAK/CXCL14 mRNA, indicating that these sequences are essential for the regulation of BRAK/CXCL14 gene expression in the cells.

BACKGROUND

In juvenile mammals, the epiphyses of long bones grow by chondrogenesis within the articular cartilage. A better understanding of the molecular mechanisms that regulate the growth of articular cartilage may give insight into the antecedents of joint disease, such as osteoarthritis.

METHODS

We used laser capture microdissection to isolate chondrocytes from the superficial, middle, and deep zones of growing tibial articular cartilage in the 1-wk-old mouse and then investigated expression patterns by microarray. To identify molecular markers for each zone of the growing articular cartilage, we found genes showing zone-specific expression and confirmed by real-time PCR and in situ hybridization.

RESULTS

Bioinformatic analyses implicated ephrin receptor signaling, Wnt signaling, and bone morphogenetic protein signaling in the spatial regulation of chondrocyte differentiation during growth. Molecular markers were identified for superficial (e.g., Cilp, Prg4), middle (Cxcl14, Tnn), and deep zones (Sfrp5, Frzb). Comparison between juvenile articular and growth plate cartilage revealed that the superficial-to-deep zone transition showed similarity with the hypertrophic-to-resting zone transition.

CONCLUSIONS

Laser capture microdissection combined with microarray analysis identified novel signaling pathways that are spatially regulated in growing mouse articular cartilage and revealed similarities between the molecular architecture of the growing articular cartilage and that of growth plate cartilage.

Ultra-small superparamagnetic iron oxide nanoparticles (USPIO-NPs) are widely used as clinical magnetic resonance imaging contrast agents for hepatic diseases diagnosis. USPIO-NPs often damage the hepatocytes and affect the function of liver but its mechanism of action remains unclear. In the present study, USPIO-NPs caused higher cytotoxicity and lactate dehydrogenase (LDH) leakage in hepatic L02 cells than SPIO-NPs. Subsequently, USPIO-NPs affected more genes' expression than SPIO-NPs analyzed through microarray and bioinformatics analysis. The affected genes were involved in several biological processes, including calcium ion homeostasis, inflammatory response-related leukocyte chemotaxis, and migration. In addition, the level of endoplasmic reticulum (ER) calcium ion was increased by USPIO-NPs. USPIO-NPs also upregulated the genes related to acute-phase inflammation, including IL1B, IL6, IL18, TNFSF12, TNFRSF12, SAA1, SAA2, JAK1, STAT5B, and CXCL14. Furthermore, interleukin-6 (IL-6) secretion was elevated by USPIO-NPs as detected using ELISA. On the other hand, USPIO-NPs changed the morphology of ER and triggered the ER stress and unfolded protein response PERK/ATF4 pathway. Furthermore, blocking ER stress with inhibitor or ATF4 small interfering RNA counteracted IL-6-related acute-phase inflammation and cytotoxicity caused by USPIO-NPs. Taken together, we found that the USPIO-NPs could trigger stronger IL-6-related acute-phase inflammation than SPIO-NPs in hepatocytes. We demonstrated, for the first time, that IL-6-related acute-phase inflammation caused by NPs was regulated by PERK/ATF4 signaling. The PERK/ATF4 pathway explored in this study could be a candidate for diagnostic and therapeutic target against NPs-induced liver injury and cytotoxicity, which would be helpful for USPIO-NPs medical application.

The aim of this study was to investigate the differential gene expression of cytokines in peripheral blood mononuclear cells (PBMCs) from patients with pulmonary embolism (PE) and controls. Twenty patients with PE and twenty control patients matched for gender and age with the PE group were recruited into the study. Human cDNA microarray analysis was used to detect differences in the expression of cytokine-associated genes between the two groups. In PE patients, the expression levels of the genes encoding IFNα5, IFNα6, IFNα8, IFNα14, IFNκ, IFNω1, IFNε1 and IFNγ were significantly lower compared with controls (P<0.05). The expression levels of the genes encoding IL1α, IL2, IL3, IL9, IL13, IL17β, IL19, IL22, IL23α, IL24, IL25 and IL31 were significantly lower (P<0.05), while IL10 and IL28A mRNA expression levels were higher in PE patients compared with controls (P<0.05). In PE patients, Cxcl1, Cxcl2, Cxcl6, Cxcl13 and Cxcl14 mRNAs were significantly upregulated (P<0.05), however, Cxcl10 mRNA was significantly downregulated (P<0.01). In PE patients, the mRNA expression levels of TNF superfamily members 1, 9 and 13, and TNF receptor superfamily members 1A, 1B, 9, 10B, 10C, 10D and 19L, were significantly upregulated (P<0.05), whereas TNF receptor superfamily members 11B, 19 and 25 were significantly downregulated compared with controls (P<0.05). The mRNA expression levels of granulocyte-macrophage colony‑stimulating factor, granulocyte colony-stimulating factor, erythropoietin, thrombopoietin and mast cell growth factor were significantly lower in PE patients compared with controls (P<0.05). In PE patients, the mRNA expression levels of a variety of cytokines were imbalanced and cellular immune function was downregulated compared with controls. Thus, PE patients may be more susceptible to infections caused by viruses, intracellular bacteria and parasites.

GBM tissues are comprised of not only tumor cells but also tumor-associated nontumor cells, such as stromal cells and immune cells, which dilute the purity of glioma cells and function in glioma biology. However, the roles of miRNAs in modulating glioma purity are not clarified. In total, 838 glioma samples with transcriptome data, including 537 RNAseq data from TCGA project and 301 microarray data from Chinese Glioma Genome Atlas (CGGA project), were recruited into our investigation. Tumor purity, molecular subtypes and IDH status were also available. R language was employed as the main tool for statistical analysis and graphical work. Screening miRNA profiling and paired TCGA samples' transcriptome data demonstrates that miR-17-5p expression harbors the most significant positive correlation with glioma purity among all miRNAs. CXCL14 shows robust negative correlation with miR-17-5p expression in TCGA and CGGA dataset. miR-17-5p directly targets CXCL14 and functions as a tumor-suppressor of GBM. CXCL14 showed lower expression in proneural subtype and may contribute as a potential marker for proneural subtype in glioma. Genes markedly correlated with CXCL14 are involved in essential functions associated with anti-tumor immune process. CXCL14 has a strong correlation with immune(T cells, Monocytic lineage and Neutrophils) and Fibroblasts within glioma environment. miR-17-5p and CXCL14 exhibited predictive values for high-grade glioma(HGG) patients: Higher miR-17-5p indicated significantly longer survival while lower CXCL14 indicated longer survival. Our results highlight the importance of the miR-17-5p-CXCL14 axis in regulating key steps of anti-tumor immune process and may serve as potential targets of immune treatments for gliomas.

The chemokine (C-X-C motif) ligand (CXCL) family plays an important role in inflammation. In order to understand the role of CXC chemokine family in carcinogenesis, this study explored a group of early gastric cancer (GC) patients, and assessed the level of CXC chemokine ligand (CXCL) in blood samples of patients representing systemic circulation and tumor microenvironment, detected the expression of CXC chemokine receptor (CXCR) in tumor tissues, and measured tumor infiltrating immune cell subsets. 69 patients with GC were included in a single center prospective study and were followed up for 6 years. The level of CXCL1-14 was determined by ELISA and the concentration gradient of chemokine was calculated. Western blot was used to detect the expression of CXCR1, CXCR2, CXCR3, and CXCR4 in tumor tissue. CXCL1-14 expression was inhibited by siRNA in HGC27 cells and then the migration ability of HGC27 cells was detected by cell scratch test. The results of this study showed that the chemokine concentrations of CXCL1, CXCL2, CXCL5, CXCL8, CXCL11, and CXCL13 in peripheral blood and tumor drainage blood of patients without recurrence after treatment were significantly lower than those before treatment. The concentrations of CXCL1, CXCL2, CXCL4, CXCL5, CXCL7, CXCL8, CXCL9, CXCL10, CXCL12, CXCL13, and CXCL14 in peripheral blood and tumor drainage blood were significantly higher than those in patients without recurrence. Patients with low expression of CXCR1 and CXCR3 had lower AFP (alpha fetoprotein), smaller tumor volume, and lower TNM tumor stage. Patients with lower expression of CXCR2 and CXCR4 had higher AFP (alpha fetoprotein) level, larger tumor volume, and higher TNM tumor stage. After down-regulation of CXCLs expression, the migration ability of most cell lines was significantly inhibited. This study suggests that CXCL chemokine family plays an important role in the pathogenesis of GC and can be used as a marker for the development of GC.

Reflux of gastroduodenal contents and consequent inflammatory responses are associated with the development of Barrett's oesophagus (BO) and the promotion of oesophageal adenocarcinoma (OAC). Deregulation of inflammatory processes is a hallmark of oesophageal cancer. In this study, we aimed to investigate (i) the transcriptional responses to deoxycholic acid (DCA) in cell lines representative of either end of the oesophageal cancer sequence, (ii) the expression of DCA-regulated genes in data charting oesophageal carcinogenesis and (iii) the impact of these genes on oesophageal inflammatory signalling. Gene expression microarrays were utilized to demonstrate differential transcriptional responses between squamous (HET-1A) and adenomatous (SKGT4) cell lines exposed to DCA. Differential basal and DCA-inducible expression of cytokines such as interleukin (IL) 8 was observed between both cell types. A cohort of DCA-regulated genes specific to each cell type was identified in microarray experimentation and subsequently validated. Cell type-specific genes included TRB3, CXCL14, GDF15 and LIF in HET-1A cells, with COX2-, ESM1-, URHF1- and IL1alpha-and IL1beta-specific expression in SKGT4 cells. Over 30% of the genes altered in BO and OAC were shown to be regulated by DCA utilizing an integrative genomic approach. One such gene, tribbles-homology-3 (TRB3) was induced specifically in HET-1A cells, absent in SKGT4 cells and decreased in BO samples in silico and in vivo. Inhibition and re-introduction of TRB3 in HET-1A and SKGT4 cells, respectively, demonstrated the ability of TRB3 to regulate inflammatory signalling through nuclear factor-kappaB. This study identifies mechanisms through which bile acids such as DCA, in conjunction with the loss of key signalling molecules, could regulate oesophageal metaplasticity.

The chemokine CXCL14 is a highly conserved, homeostatic chemokine that is constitutively expressed in skin epithelia. Responsible for immune cell recruitment and maturation, as well as impacting epithelial cell motility, CXCL14 contributes to the establishment of immune surveillance within normal epithelial layers. Furthermore, CXCL14 is critical to upregulating major histocompatibility complex class I expression on tumor cells. Given these important roles, CXCL14 is often dysregulated in several types of carcinomas including cervical, colorectal, endometrial, and head and neck cancers. Its disruption has been shown to limit critical antitumor immune regulation and is correlated to poor patient prognosis. However, other studies have found that in certain cancers, namely pancreatic and some breast cancers, overexpression of stromal CXCL14 correlates with poor patient survival due to increased invasiveness. Contributing to the ambiguity CXCL14 plays in cancer is that the native CXCL14 receptor remains uncharacterized, although several candidate receptors have been proposed. Despite the complexity of CXCL14 functions, it remains clear that this chemokine is a key regulatory factor in cancer and represents a potential target for future cancer immunotherapies.

Hypoxia is an integral component of the tumor microenvironment. Either as chronic or cycling hypoxia, it exerts a similar effect on cancer processes by activating hypoxia-inducible factor-1 (HIF-1) and nuclear factor (NF-κB), with cycling hypoxia showing a stronger proinflammatory influence. One of the systems affected by hypoxia is the CXC chemokine system. This paper reviews all available information on hypoxia-induced changes in the expression of all CXC chemokines (CXCL1, CXCL2, CXCL3, CXCL4, CXCL5, CXCL6, CXCL7, CXCL8 (IL-8), CXCL9, CXCL10, CXCL11, CXCL12 (SDF-1), CXCL13, CXCL14, CXCL15, CXCL16, CXCL17) as well as CXC chemokine receptors-CXCR1, CXCR2, CXCR3, CXCR4, CXCR5, CXCR6, CXCR7 and CXCR8. First, we present basic information on the effect of these chemoattractant cytokines on cancer processes. We then discuss the effect of hypoxia-induced changes on CXC chemokine expression on the angiogenesis, lymphangiogenesis and recruitment of various cells to the tumor niche, including myeloid-derived suppressor cells (MDSCs), tumor-associated macrophages (TAMs), tumor-associated neutrophils (TANs), regulatory T cells (T_regs) and tumor-infiltrating lymphocytes (TILs). Finally, the review summarizes data on the use of drugs targeting the CXC chemokine system in cancer therapies.

Keywords: CXC chemokine; HIF-1α; IL-8; NF-κB; SDF-1; cancer; cycling hypoxia; hypoxia; hypoxia-inducible factor; tumor.

The C-X-C motif ligand 14 (CXCL14) is a recently discovered chemokine that is highly conserved in vertebrates and expressed in various embryonic and adult tissues. CXCL14 signaling has been implicated to function as an antiangiogenic and anticancer agent in adults. However, its function during development is unknown. We previously identified novel expression of CXCL14 mRNA in various ocular tissues during development. Here, we show that CXCL14 protein is expressed in the anterior eye at a critical time during neurovascular development and in the retina during neurogenesis. We report that RCAS-mediated knockdown of CXCL14 causes severe neural defects in the eye including precocious and excessive innervation of the cornea and iris. Absence of CXCL14 results in the malformation of the neural retina and misprojection of the retinal ganglion neurons. The ocular neural defects may be due to loss of CXCL12 modulation since recombinant CXCL14 diminishes CXCL12-induced axon growth in vitro. Furthermore, we show that knockdown of CXCL14 causes neovascularization of the cornea. Altogether, our results show for the first time that CXCL14 plays a critical role in modulating neurogenesis and inhibiting ectopic vascularization of the cornea during ocular development.

The cellular prion protein (PrPC) is an evolutionary conserved protein abundantly expressed not only in the central nervous system but also peripherally including the immune system. A line of Norwegian dairy goats naturally devoid of PrPC (PRNP Ter/Ter) provides a novel model for studying PrPC physiology.

In order to explore putative roles for PrPC in acute inflammatory responses, we performed a lipopolysaccharide (LPS, Escherichia coli O26:B6) challenge of 16 goats (8 PRNP +/+ and 8 PRNP Ter/Ter) and included 10 saline-treated controls (5 of each PRNP genotype). Clinical examinations were performed continuously, and blood samples were collected throughout the trial. Genome-wide transcription profiles of the choroid plexus, which is at the blood-brain interface, and the hippocampus were analyzed by RNA sequencing, and the same tissues were histologically evaluated.

All LPS-treated goats displayed clinical signs of sickness behavior, which were of significantly (p < 0.01) longer duration in animals without PrPC. In the choroid plexus, a substantial alteration of the transcriptome and activation of Iba1-positive cells were observed. This response included genotype-dependent differential expression of several genes associated with the immune response, such as ISG15, CXCL12, CXCL14, and acute phase proteins, among others. Activation of cytokine-responsive genes was skewed towards a more profound type I interferon response, and a less obvious type II response, in PrPC-deficient goats. The magnitude of gene expression in response to LPS was smaller in the hippocampus than in the choroid plexus. Resting state expression profiles revealed a few differences between the PRNP genotypes.

Our data suggest that PrPC acts as a modulator of certain pathways of innate immunity signaling, particularly downstream of interferons, and probably contributes to protection of vulnerable tissues against inflammatory damage.

We recently reported that CXCL14 binds to CXCR4 with high affinity and inhibits CXCL12-mediated chemotaxis. Here we found that the C-terminal 51-77 amino acid residues of CXCL14 are responsible for CXCR4 binding. A disulfide dimer peptide of CXCL14(51-77) bound to CXCR4 with comparable affinity to full length CXCL14, and exhibited CXCL12 inhibitor activity. CXCR4 was efficiently internalized upon binding of dimeric CXCL14(51-77), thereby being reduced on the cell surface. Substitution of 5 amino acid residues in combination with the use of an oxime linker for dimerization increased the solubility and chemical stability of the dimeric CXCL14(51-77).