Angiogenesis in ischemic organs is modulated by immune cells. Systemic neovascularization of the ischemic lung requires macrophages, with chemokines playing a central role in new vessel growth. Because regulatory T (Treg) cells modulate tumor-induced neovascularization, we questioned whether this CD4(+) lymphocyte subset impacts blood vessel growth during ischemia. In a model of left lung ischemia, an increase in CD4(+) CD25(+) forkhead homeobox protein-3 (Foxp3)(+) cells was observed 3-5 days after the onset of ischemia in wild-type C57Bl/6 mice. Using transgenic mice where Foxp3(+) Treg cells can be depleted with diphtheria toxin (DT; Foxp3(DTR)), we unexpectedly found that Foxp3(+) Treg depletion led to markedly reduced lung angiogenesis (90% reduction from Foxp3(gfp) controls). Adoptive transfer studies using CD4(+) CD25(+) splenocytes from congenic CD45.1 mice into Foxp3(+) Treg-depleted mice showed an almost complete recovery of the angiogenic phenotype (80% of Foxp3(gfp) controls). A survey of lung gene expression of angiogenic (lipopolysaccharide-induced CXC chemokine [LIX], IL-6, IL-17) and angiostatic (IFN-γ, transforming growth factor-β, IL-10) cytokines showed Treg-dependent differences only in LIX (CXCL5) and IL-6. Protein confirmation demonstrated a significant reduction in LIX in Treg-deficient mice compared with controls 5 days after the onset of ischemia. Phenotyping other inflammatory cells in the lung by multicolor flow cytometry demonstrated a significantly reduced number of macrophages (major histocombatibility complex class II [MHCII](int), CD11C(+)) in Treg-deficient lungs compared with Treg-sufficient lungs. Treg cells are essential for maximal systemic angiogenesis after pulmonary ischemia. One likely mechanism responsible for the decrease in angiogenesis in Treg-depleted mice was the decline in the essential CXC chemokine, LIX.

UNASSIGNED

CXCL5 is a member of the CXC-type chemokine family, which has been found to play important roles in tumorigenesis and cancer progression. Recent studies have demonstrated that CXCL5 could serve as a potential prognostic biomarker for cancer patients. However, the prognostic value of CXCL5 is still controversial.

UNASSIGNED

We systematically searched PubMed, Embase and Web of Science to obtain all relevant articles investigating the prognostic significance of CXCL5 expression in cancer patients. Hazards ratios (HR) with corresponding 95% confidence intervals (CI) were pooled to estimate the association between CXCL5 expression levels with survival of cancer patients.

UNASSIGNED

A total of 15 eligible studies including 19 cohorts and 5070 patients were enrolled in the current meta-analysis. Our results demonstrated that elevated expression level of CXCL5 was significantly associated with poor overall survival (OS) (pooled HR 1.70; 95% CI 1.36-2.12), progression-free survival (pooled HR 1.65; 95% CI 1.09-2.49) and recurrence-free survival (pooled HR 1.49; 95% CI 1.15-1.93) in cancer patients. However, high or low expression of CXCL5 made no difference in predicting the disease-free survival (pooled HR 0.63; 95% CI 0.11-3.49) of cancer patients. Furthermore, we found that high CXCL5 expression was associated with reduced OS in intrahepatic cholangiocarcinoma (HR 1.91; 95% CI 1.31-2.78) and hepatocellular carcinoma (HR 1.87; 95% CI 1.55-2.27). However, there was no significant association between expression level of CXCL5 with the OS in lung cancer (HR 1.25; 95% CI 0.79-1.99) and colorectal cancer (HR 1.16; 95% CI 0.32-4.22, p = 0.826) in current meta-analysis.

UNASSIGNED

In conclusion, our meta-analysis suggested that elevated CXCL5 expression might be an adverse prognostic marker for cancer patients, which could help the clinical decision making process.

BACKGROUND

We previously found plasma levels of CD40 ligand (CD40L), chemokine (C-X-C motif) ligand 5 (CXCL5), chemokine (C-C motif) ligand 5 (CCL5) and epidermal growth factor (EGF) to be low in aplastic anemia (AA) patients and to be correlated with platelet count.

OBJECTIVE

To study the association of CD40L, CXCL5, CCL5 and EGF with platelets.

METHODS

We measured cytokines in the plasma of immune thrombocytopenic purpura (ITP) and AA patients using the Luminex assay and confirmed the results in a mouse model and in vitro experiments.

RESULTS

Both ITP and AA showed similarly low levels of CD40L, CXCL5, CCL5 and EGF, compared with healthy controls. In ITP, levels of these proteins were significantly greater in patients with higher platelet counts than in those with lower platelet counts. In a murine thrombocytopenia model, levels of CD40L, CXCL5, CCL5 and EGF decreased with platelet count after immune-mediated destruction, while the cytokine levels increased when the platelet count recovered. In vitro, concentrations of these cytokines in the supernatants of platelet suspensions were proportional to platelet numbers, and levels in sera prepared by simple blood coagulation were equivalent to those in platelet-rich plasma-converted sera. mRNA expression for CXCL5, CCL5 and EGF was higher in platelets than in megakaryocytes, peripheral blood mononuclear cells, granulocytes and non-megakaryocytic bone marrow cells.

CONCLUSIONS

Plasma CD40L, CXCL5, CCL5 and EGF are mainly platelet-derived, suggesting a role of platelets in immune responses and inflammation. Measurement of CD40L, CXCL5, CCL5 and EGF in human blood allowed testable inferences concerning physiology and pathophysiology in quantitative platelet disorders.

Although lungs are densely innervated by the peripheral nervous system (PNS), the role of the PNS in the progression of lung cancer is unknown. In this study, we report that mouse adult Schwann cells (SC), the principal glial cells of the PNS, can regulate the motility of lung cancer cells in vitro and the formation of metastases in vivo SCs promoted epithelial-to-mesenchymal transition (EMT) and the motility of two lung cancer cell lines by increasing expression of Snail and Twist in tumor cells; blocking of Snail and Twist expression abolished SC-induced motility of tumor cells. SC-derived CXCL5 was responsible for EMT in lung cancer cells, as the inhibition of CXCL5 or its receptor CXCR2 reduced SC-induced expression of Snail and Twist and reduced motility in tumor cells. CXCL5/CXCR2 binding activated the PI3K/AKT/GSK-3β/Snail-Twist signaling pathway in lung cancer cells, and the PI3K inhibitor blocked CXCL5-dependent phosphorylation of AKT and GSK-3β, reduced expression of Snail/Twist, and limited tumor cell invasiveness. SC conditioning of tumor cells prior to their injection into mice significantly increased the formation of metastases in the regional lymph nodes. In summary, SCs can regulate the CXCL5/CXCR2/PI3K/AKT/GSK-3β/Snail-Twist pathway to promote EMT, invasiveness, and metastatic potential of lung cancer cells. Our results reveal a new role of the PNS in the functional organization of the tumor microenvironment and tumor progression.Significance: This study increases our understanding of how nerves and, in particular, specific glial cells, Schwann cells, in the peripheral nervous system, may help promote tumor growth and metastasis. Cancer Res; 78(20); 5927-39. ©2018 AACR.

In order to evaluate probiotic strains applicable for the beneficial immunomodulation of the porcine gut (immunobiotics), we previously developed a porcine intestinal epitheliocyte cell line (PIE cells). Here, transcriptomic studies using PIE cells were performed considering that this information would be valuable for understanding the mechanisms involved in the protective activity of the immunobiotic strain Lactobacillus jensenii TL2937 against intestinal inflammatory damage in pigs. In addition, those studies would provide criteria for selecting biomarkers for the screening of new immunobiotic strains. We performed microarray analysis to investigate the transcriptomic response of PIE cells to the challenge with heat-stable enterotoxigenic Escherichia coli (ETEC) pathogen-associated molecular patterns (PAMPs) and, the changes induced by L. jensenii TL2937 in that response. The approach allowed us to obtain a global overview of the immune genes involved in the response of PIE cells to heat-stable ETEC PAMPs. We observed that L. jensenii TL2937 differently modulated gene expression in ETEC PAMPs-challenged PIE cells. Microarray and RT-PCR analysis indicated that the most remarkable changes in PIE cells transcriptomic profile after heat-stable ETEC PAMPs challenge were observed in chemokines, adhesion molecules, complement and coagulation cascades factors. In addition, an anti-inflammatory effect triggered by TL2937 strain in PIE cells was clearly demonstrated. The decrease in the expression of chemokines (CCL8, CXCL5, CXCL9, CXCL10, and CXCL11), complement (C1R, C1S, C3, and CFB), and coagulation factors (F3) by L. jensenii TL2937 supports our previous reports on the immunoregulatory effect of this strain. These results provided clues for the better understanding of the mechanism underlying host-immunobiotic interaction in the porcine host. The comprehensive transcriptomic profiles of PIE cells provided by our analyses successfully identified a group of genes, which could be used as prospective biomarkers for the screening and evaluation of new anti-inflammatory immunobiotics for the prevention of inflammatory intestinal disorders in pigs.

Paget's disease of bone (PDB) is a chronic focal skeletal disorder that affects 2-3% of the population over 55 years of age. PDB is marked by highly localized areas of bone turnover with increased osteoclast activity. Evidence suggests a functional role for measles virus nucleocapsid protein (MVNP) in the pathogenesis of PDB. In the present study, we identified elevated levels (≈ 180-fold) of CXCL5 mRNA expression in bone marrow cells from patients with PDB compared with that in normal subjects. In addition, CXCL5 levels are increased (five-fold) in serum samples from patients with PDB. Furthermore, MVNP transduction in human bone marrow monocytes significantly increased CXCL5 mRNA expression. Real-time PCR analysis showed that CXCL5 stimulation increased (6.8-fold) RANKL mRNA expression in normal human bone marrow-derived stromal (SAKA-T) cells. Moreover, CXCL5 increased (5.2-fold) CXCR1 receptor expression in these cells. We further showed that CXCL5 treatment elevated the expression levels of phospho-ERK1/2 and phospho-p38. CXCL5 also significantly increased phosphorylation of CREB (cAMP response element-binding) in bone marrow stromal/preosteoblast cells. Chromatin immuneprecipitation (ChIP) assay confirmed phospho-CREB binding to RANKL gene promoter region. Further, the suppression of p-CREB expression by the inhibitors of ERK1/2, p38 and PKA significantly decreased CXCL5 stimulation of hRANKL gene promoter activity. Thus, our results suggest that CREB is a downstream effector of CXCL5 signaling and that increased levels of CXCL5 contribute to enhanced levels of RANKL expression in PDB.

BACKGROUND

Destrin (DSTN) is a member of the ADF/cofilin family of proteins and is an important regulator of actin dynamics. The primary function of destrin is to depolymerize filamentous actin into its monomeric form and promote filament severing. While progress has been made in understanding the biochemical functions of the ADF/cofilin proteins, the study of an animal model for cells deficient for DSTN provides an opportunity to investigate the physiological processes regulated by proper actin dynamics in vivo. A spontaneous mouse mutant, corneal disease 1(corn1), is deficient for DSTN, which causes epithelial hyperproliferation and neovascularization in the cornea. Dstn(corn1) mice exhibit an actin dynamics defect in the cornea as evidenced by the formation of actin stress fibers in the epithelial cells. Previously, we observed a significant infiltration of leukocytes into the cornea of Dstn(corn1) mice as well as the upregulation of proinflammatory molecules. In this study, we sought to characterize this inflammatory condition and explore the physiological mechanism through which a loss of Dstn function leads to inflammation.

RESULTS

Through immunofluorescent analyses, we observed a significant recruitment of neutrophils and macrophages to the Dstn(corn1) cornea, demonstrating that the innate immune system is spontaneously activated in this mutant. The inflammatory chemokine, CXCL5, was ectopically expressed in the corneal epithelial cells of Dstn(corn1) mice, and targeting of the receptor for this chemokine inhibited neutrophil recruitment. An inflammatory reaction was not observed in the cornea of allelic mutant strain, Dstn(corn1-2J), which has a milder defect in actin dynamics in the corneal epithelial cells.

CONCLUSIONS

This study shows that severe defects in actin dynamics lead to an autoinflammatory condition that is mediated by the expression of CXC chemokines.

BACKGROUND

Surgery for trachomatous trichiasis (TT) is a key component of the SAFE Strategy for trachoma control. Unfortunately, recurrent TT following surgery is common, probably due to various surgical and disease factors. To develop strategies to reduce recurrence rates it is necessary to understand its pathological basis. In this study we investigated the relationship between recurrent trichiasis and the expression of various cytokines and fibrogenic genes during a two-year follow-up period.

RESULTS

Individuals undergoing surgery for TT were examined at baseline (pre-operative), 6, 12, 18 and 24 months. Conjunctival swab samples were collected from the tarsal conjunctiva for RNA isolation on each occasion. Individuals who developed recurrent TT with at least 3 lashes touching the eye on one or more occasion were designated "cases" and an equal number of "controls" were randomly selected from those without recurrent TT, frequency matched for age and baseline TT severity. The expression of the following genes was measured by quantitative RT-PCR: S100A7, IL1B, CXCL5, TNFA, NOS2A, CTGF, MMP7, MMP9 and MMP12. Thirteen hundred individuals were enrolled and underwent surgery. By two years 122 had developed recurrent TT with at least 3 lashes touching the eye. Recurrent TT was consistently associated across multiple time points with about a 2-fold increase in S100A7 expression (p = 0.008). Clinically visible conjunctival inflammation was associated with increased S100A7, IL1B, CXCL5, MMP9 and MMP12 expression.

CONCLUSIONS

Increased S100A7 expression was associated with trachomatous conjunctival scarring and may be linked to the pathophysiology of recurrent TT. S100A7 expression could be a potential biomarker for this disease process. As part of the epithelial innate immune response S100A7 has multiple actions, potentially contributing to a chronic pro-inflammatory response, which may lead to ongoing tissue damage and increased scarring.

With new frontiers of pharmaceutical therapies focusing on tumor growth and angiogenesis, understanding the interaction between immune system and tumor microenvironment has become ever more important. Chemokines and chemokine receptors appear to play an integral role in tumor characteristics. Evidence suggests CXCR4, CXCL5, CXCR7, and stromal derived factor-1 appear to be crucial in survival, growth, and metastasis of renal cell carcinoma. As the role of chemokines in renal cancer is becoming more evident, further research will lead to a better understanding of tumor biology and the development of new therapeutic targets to help improve survival.

Protease-activated receptors (PARs) are G-protein-coupled receptors with an active role in host defense. The two most highly expressed members of the PAR family in gingival epithelial cells (GECs) are PAR1 and PAR2. The major virulence factors of periodontal pathogen Porphyromonas gingivalis are its proteases which can activate PAR2. However, little is known about the function of PARs in GECs when they are activated by their endogenous agonist enzymes. The purpose of this study was to characterize how the expression of innate immune markers is modulated when PAR1 and PAR2 are activated by their agonist enzymes, thrombin and trypsin, respectively. Here, we report that activation of PAR1 and PAR2 induces cell proliferation at low concentration. Activation of PAR via proteolytic activity of thrombin and trypsin induces expression of CXCL5/ENA-78 and CCL20/MIP3alpha in a concentration-dependent manner. Induction of CXCL5 via PAR1 was inhibited in the presence of PAR1 cleavage blocking antibodies and by PAR1 siRNA. The induction of CXCL5 and CCL20 via PAR2 was inhibited by PAR2 siRNA. These findings indicate an active role in innate immune responses by PAR1 and PAR2 in GECs. Modulation of innate immunity by PARs may contribute to co-ordinated and balanced immunosurveillance in GECs.

Hemocyanins induce a potent Th1-dominant immune response with beneficial clinical outcomes when used as a carrier/adjuvant in vaccines and nonspecific immunostimulant in cancer. However, the mechanisms by which hemocyanins trigger innate immune responses, leading to beneficial adaptive immune responses, are unknown. This response is triggered by a proinflammatory signal from various components, of which macrophages are an essential part. To understand how these proteins influence macrophage response, we investigated the effects of mollusks hemocyanins with varying structural and immunological properties, including hemocyanins from Concholepas concholepas, Fissurella latimarginata, and Megathura crenulata (keyhole limpet hemocyanin), on cultures of peritoneal macrophages. Hemocyanins were phagocytosed and slowly processed. Analysis of this process showed differential gene expression along with protein levels of proinflammatory markers, including IL-1β, IL-6, IL-12p40, and TNF-α. An extended expression analysis of 84 cytokines during a 24-h period showed a robust proinflammatory response for F. latimarginata hemocyanin in comparison with keyhole limpet hemocyanin and C. concholepas hemocyanin, which was characterized by an increase in the transcript levels of M1 cytokines involved in leukocyte recruitment. These cytokine genes included chemokines (Cxcl1, Cxcl3, Cxcl5, Ccl2, and Ccl3), ILs (Il1b and Ifng), growth factors (Csf2 and Csf3), and TNF family members (Cd40lg). The protein levels of certain cytokines were increased. However, every hemocyanin maintains downregulated key M2 cytokine genes, including Il4 and Il5 Collectively, our data demonstrate that hemocyanins are able to trigger the release of proinflammatory factors with different patterns of cytokine expression, suggesting differential signaling pathways and transcriptional network mechanisms that lead to the activation of M1-polarized macrophages.

Prostate cancer primarily metastasizes to bone, and the interaction of cancer cells with bone cells results in a local activation of bone formation and/or bone resorption. Since the cellular and molecular mechanisms underlying the development of these tumor-induced osteoblastic or osteolytic lesions are still poorly understood, we have compared the effects of two prostate cancer cell lines, osteoblastic MDA-PCa-2b cells and osteolytic PC-3 cells, on bone-forming osteoblasts. Using Affymetrix Gene Chip hybridization followed by qRT-PCR confirmation we were able to identify specific genes, including Smpd3 and Dmp1, whose expression is significantly reduced upon treatment with PC-3-conditioned medium. Moreover, we observed that PC-3-conditioned medium led to a marked induction of several cytokine genes, including Cxcl5, Cxcl12 and Tnfsf11, the latter one encoding for the osteoclast differentiation factor Rankl. Likewise, when we analyzed the effects of MDA-PCa-2b- and PC-3-conditioned medium on signal transduction in osteoblasts we did not only observe opposite effects on the canonical Wnt signalling pathway, but also a specific induction of Erk and Stat phosphorylation by PC-3-conditioned medium. Most importantly, the induction of Cxcl5, Cxcl12 and Tnfsf11 in osteoblasts by PC-3-conditioned medium was abrogated by the Stat3/5 inhibitor piceatannol, whereas the selective blockade of Stat1 and Erk activation had no effect. Together with the finding, that activated Stat3 in osteoblasts was detectable in bone biopsies from patients with osteolytic metastases, our data suggest that the Stat3/5-dependent activation of cytokine expression in osteoblasts may have a significant impact on cancer cell migration and proliferation, but also on osteoclast activation.

Inflammation is an important feature of the pathophysiological response to ischaemic stroke. The ischaemic brain-invading leukocytes, neutrophils in particular, contribute to the exacerbation of tissue injury in stroke. Chemokines are a growing family of proteins performing chemotactic activity on selective leukocyte subpopulations. Chemokines are broadly divided into two major subfamilies on the basis of the arrangement of the two N-terminal cysteine residues, CXC and CC, depending on whether the first two cysteine residues have an amino acid between them (CXC) or are adjacent (CC). CXC chemokines possessing, close to the N terminus, the amino acid sequence glutamic acid-leucine-arginine (ELR motif) specifically act on neutrophils. CXCL5 is one of the ELR-expressing CXC chemokines and is a potent neutrophil attractant and activator. The objective of the study was to detect CXCL5 levels in the cerebrospinal fluid (CSF) and sera of stroke patients and to investigate the relation between these levels and the volume of brain computed tomography (CT) hypodense areas representing early ischaemic lesions. A total of 23 ischaemic stroke patients were studied. CSF and blood sampling and brain CT were performed within the first 24 hours of stroke. The control group consisted of 15 patients with tension headache. CXCL5 levels were determined by the ELISA method. CSF CXCL5 levels in stroke patients were significantly higher in comparison with the control group (38.2 +/- 18.4 pg/ml vs. 18.7 +/- 8.2 pg/ml; p < 0.001). No significant differences in serum CXCL5 levels were found between the stroke patients and the control group. CSF CXCL5 levels correlated positively with the volume of early brain CT hypodense areas (p < 0.0001). The results suggest that CXCL5 may play a role in the inflammatory reaction during the early phase of ischaemic stroke.

OBJECTIVE

CXCL5 (chemokine [C-X-C motif] ligand 5, also known as epithelial neutrophil-activating peptide 78 [ENA78]) belongs to the CXC chemokine family and has been shown to have promitotic effects on hepatocytes. The aim of our study was to assess CXCL5 plasma levels in patients with chronic liver disease.

METHODS

CXCL5 plasma levels were measured in 111 patients with chronic liver disease and 98 healthy controls. The gene expression of CXCL5 and its main receptor, CXC receptor-2, were also determined in liver biopsies from 46 patients.

RESULTS

CXCL5 levels were correlated with clinical presentation, laboratory parameters, and liver histology. Plasma CXCL5 levels in patients with liver cirrhosis were lower than those in healthy controls, and correlated with hepatic biosynthetic capacity, Child-Pugh and model for end-stage liver disease scores. Patients with hepatic necroinflammation and fibrosis on liver histology showed lower plasma CXCL5 levels. In patients with typical clinical complications of cirrhosis, CXCL5 levels were found to be decreased. Intrahepatically, CXCL5 expression was increased in patients with advanced fibrosis and cirrhosis. The isolation of different cellular compartments from mouse livers suggested that hepatic stellate cells and sinusoidal endothelial cells are the main sources of hepatic CXCL5.

CONCLUSIONS

Plasma CXCL5 levels are lower in patients with chronic liver disease, suggesting that CXCL5 might be involved in the pathogenesis of chronic liver disease. CXCL5 could serve as an additional biomarker for hepatic necroinflammation and fibrosis.

CXC chemokines are thought to play an important role at sites of inflammation. Because ELR(+) CXC chemokines are expressed in different types of tonsillitis we investigated the role of the surface/crypt epithelium of human tonsils in producing ELR(+) CXC chemokines: interleukin (IL)-8 (CXCL8), ENA-78 (CXCL5), GRO-alpha (CXCL1) and GCP-2 (CXCL6). Tonsillar tissue was obtained from patients undergoing tonsillectomy and chemokine expression was investigated by means of immunohistochemistry. A549 cells were used as a model to study kinetics of chemokine expression in epithelial cells. Cells were stimulated with tumour necrosis factor (TNF)-alpha, lipopolysaccharide (LPS) and supernatants derived from aerobic/anaerobic Staphylococcus aureus strains. Chemokine expression was measured by quantitative reverse transcription-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA). We observed epithelial expression of IL-8, GRO-alpha and GCP-2 in different types of tonsillitis, whereas ENA-78 was rarely expressed. In A549 cells abundant expression of ENA-78 was detected. IL-8 and GCP-2 are expressed in an acute type of tonsillitis whereas GRO-alpha was frequently detectable both in chronically and acutely inflamed tonsils. ENA-78 does not seem to play a pivotal role in tonsillitis in vivo.

The heat shock (HS) response, a phylogenetically conserved ubiquitous response to stress, is generally characterized by the induced expression of heat shock protein (HSP) genes. Our earlier studies showed that the stress-activated transcription factor, heat shock factor-1 (HSF1), activated at febrile range or HS temperatures also modified expression of non-HSP genes including cytokine and chemokine genes. We also showed by in silico analysis that 28 among 29 human and mouse CXC chemokine genes had multiple putative heat shock response elements (HSEs) present in their gene promoters. To further determine whether these potential HSEs were functional and bound HSF1, we analyzed the recruitment of HSF1 to promoters of 5 human CXC chemokine genes (CXCL-1, 2, 3, 5 and 8) by chromatin immunoprecipitation (ChIP) assay and analyzed the effect of HS exposure on tumor necrosis factor-α (TNFα)-induced expression of these genes in human lung epithelial-like A549 cells. HSF1 ChIP analysis showed that HSF1 was recruited to all but one of these CXC chemokine genes (CXCL-3) and HS caused a significant increase in recruitment of HSF1 to one or multiple HSEs present in the promoters of CXCL-1, 2, 5 and 8 genes. However, the effect of HS exposure on expression of these genes showed a variable gene-specific effect. For example, CXCL8 expression was markedly enhanced (p<0.05) whereas CXCL5 expression was significantly repressed (p<0.05) in cells exposed to HS coincident with TNFα stimulation. In contrast, expression of CXCL1 and CXCL2, despite HSF1 recruitment to their promoters, was not affected by HS exposure. Our results indicate that some, if not all, putative HSEs present in the CXC chemokine gene promoters are functional and recruit HSF1 in vivo but the effects on gene expression are variable and gene specific. We speculate, the physical proximity and interactions of other transcription factors and co-regulators with HSF1 could be critical to determining the effects of HS on the expression of these genes.

Human immunodeficiency virus-1 (HIV-1)-infected monocytes/macrophages and microglia release increased levels of proinflammatory cytokines and chemokines, including ELR+ (containing glutamic acid-leucine-arginine motif) chemokines. To investigate the role of HIV-1 infection on chemokine regulation, monocyte-derived macrophages (MDMs) from normal donors were infected with HIV-1 and the expression of chemokines and their downstream biological functions were evaluated. Among the tested chemokines, CXCL5 was upregulated significantly both at the mRNA and protein level in the HIV-1-infected MDMs compared with mock-infected cultures. Upregulation of CXCL5 in the HIV-1-infected MDMs is, in part, regulated by increased interleukin-1β (IL-1β) production and phosphorylation of ERK1/2. Functional analyses indicate that HIV-1-induced overexpression of CXCL5 has enhanced the ability to attract neutrophils, as observed by chemotaxis assay. However, exposure of NT2, SH-SY5Y cells, and primary neurons to HIV-1-infected MDM supernatants resulted in cell death that was not rescued by anti-CXCL5 antibody suggesting that CXCL5 does not have direct effect on neuronal death. Together, these results suggest that the increased level of CXCL5 in tissue compartments, including the central nervous system of HIV-1-infected individuals might alter the inflammatory response through the infiltration of neutrophils into tissue compartment, thus causing secondary effects on resident cells.

OBJECTIVE

To assess the relationship between serum chemokine CXC ligand 5 (CXCL5) and intima-media thickness (IMT) of the common carotid artery, a marker of preclinical atherosclerosis.

METHODS

We measured the IMT (mean of three segments of both carotid arteries by ultrasonography), serum CXCL5, fasting plasma glucose, serum insulin, serum urea, serum uric acid, HbA1c, serum CRP, adiponectin and lipid profile of 730 Chinese type 2 diabetic participants older than 30 years in a cross-sectional community-based study performed in downtown Shanghai.

RESULTS

Serum CXCL5 correlated with carotid IMT (r = 0·112, P = 0·019) after adjustment for age, gender, serum CRP and HbA1c. Multivariate linear regression analysis demonstrated that age (P = 2·30 × 10(-7) ), gender (P = 2·70 × 10(-2) ), systolic blood pressure (P = 9·26 × 10(-3) ), serum CRP (P = 2·12 × 10(-2) ), low-density lipoprotein cholesterol (P = 2·40 × 10(-2) ), CXCL5 (P = 5·27 × 10(-3) ) and high-density lipoprotein cholesterol (P = 3·80 × 10(-3) ) were independently associated with carotid IMT.

CONCLUSIONS

Serum CXCL5 is an important determinant of carotid artery IMT in patients with type 2 diabetes. The correlation is independent of inflammation and glucose control.

It is well established that maladaptive innate immune responses to sterile tissue injury represent a fundamental mechanism of disease pathogenesis. In the context of cardiac ischemia reperfusion injury, neutrophils enter inflamed heart tissue, where they play an important role in potentiating tissue damage and contributing to contractile dysfunction. The precise mechanisms that govern how neutrophils are recruited to and enter the injured heart are incompletely understood. Using a model of cardiac transplant-mediated ischemia reperfusion injury and intravital 2-photon imaging of beating mouse hearts, we determined that tissue-resident CCR2+ monocyte-derived macrophages are essential mediators of neutrophil recruitment into ischemic myocardial tissue. Our studies revealed that neutrophil extravasation is mediated by a TLR9/MyD88/CXCL5 pathway. Intravital 2-photon imaging demonstrated that CXCL2 and CXCL5 play critical and nonredundant roles in guiding neutrophil adhesion and crawling, respectively. Together, these findings uncover a specific role for a tissue-resident monocyte-derived macrophage subset in sterile tissue inflammation and support the evolving concept that macrophage ontogeny is an important determinant of function. Furthermore, our results provide the framework for targeting of cell-specific signaling pathways in myocardial ischemia reperfusion injury.

Cell lines derived from the small intestine that reflect authentic properties of the originating intestinal epithelium are of high value for studies on mucosal immunology and host microbial homeostasis. A novel immortalization procedure was applied to generate continuously proliferating cell lines from murine E19 embryonic small intestinal tissue. The obtained cell lines form a tight and polarized epithelial cell layer, display characteristic tight junction, microvilli and surface protein expression and generate increasing transepithelial electrical resistance during in vitro culture. Significant up-regulation of Cxcl2 and Cxcl5 chemokine expression upon exposure to defined microbial innate immune stimuli and endogenous cytokines is observed. Cell lines were also generated from a transgenic interferon reporter (Mx2-Luciferase) mouse, allowing reporter technology-based quantification of the cellular response to type I and III interferon. Thus, the newly created cell lines mimic properties of the natural epithelium and can be used for diverse studies including testing of the absorption of drug candidates. The reproducibility of the method to create such cell lines from wild type and transgenic mice provides a new tool to study molecular and cellular processes of the epithelial barrier.

The cyclin D1 gene encodes the regulatory subunit of a holoenzyme that drives cell autonomous cell cycle progression and proliferation. Herein we show cyclin D1 abundance is increased >30-fold in the stromal fibroblasts of patients with invasive breast cancer, associated with poor outcome. Cyclin D1 transformed hTERT human fibroblast to a cancer-associated fibroblast phenotype. Stromal fibroblast expression of cyclin D1 (cyclin D1Stroma) in vivo, enhanced breast epithelial cancer tumor growth, restrained apoptosis, and increased autophagy. Cyclin D1Stroma had profound effects on the breast tumor microenvironment increasing the recruitment of F4/80+ and CD11b+ macrophages and increasing angiogenesis. Cyclin D1Stroma induced secretion of factors that promoted expansion of stem cells (breast stem-like cells, embryonic stem cells and bone marrow derived stem cells). Cyclin D1Stroma resulted in increased secretion of proinflammatory cytokines (CCL2, CCL7, CCL11, CXCL1, CXCL5, CXCL9, CXCL12), CSF (CSF1, GM-CSF1) and osteopontin (OPN) (30-fold). OPN was induced by cyclin D1 in fibroblasts, breast epithelial cells and in the murine transgenic mammary gland and OPN was sufficient to induce stem cell expansion. These results demonstrate that cyclin D1Stroma drives tumor microenvironment heterocellular signaling, promoting several key hallmarks of cancer.

BACKGROUND

The observation that angiogenesis, the process of new blood vessel formation, in healthy prostate and early prostate cancer is androgen-dependent gave rise to significant questions on how hypervascularization and increased angiogenesis is also achieved at the molecular level in advanced androgen-independent prostate cancer. The exact paracrine molecular network that is hardwired into the proteome of the endothelial and cancer subpopulations participating in this process remains partially understood.

METHODS

Here, we interrogated the signaling pathways and the molecular functional signatures across the proteome of endothelial cells after interacting with various secretomes produced by androgen-dependent and -independent prostate cancer cells.

RESULTS

We found the significant overexpression (P < 0.05) of prominent markers of angiogenesis, such as vonWillebrand factor (vWF) (∼ 2.5-fold) and CD31 (∼ 2-fold) in HUVECs stimulated with conditioned media from the androgen-independent prostate cancer cell line PC3. By mining the proteome of PC3 conditioned media, we discovered a signature of chemokine CXC motif ligands (i.e., CXCL3, CXCL5, CXCL6 and CXCL8) that could potentially coordinate increased angiogenesis in androgen-independent prostate cancer and verified their increased expression (P < 0.05) in both in vitro and xenograft models of androgen-independence.

CONCLUSIONS

Our findings form the basis for understanding the regulation of crucial metastatic phenomena during the transition of androgen-dependent prostate cancer into the highly aggressive, androgen-independent state and provide further insight on potential therapeutic targets of cancer-related angiogenesis.

Pemphigus vulgaris (PV) and bullous pemphigoid (BP) are autoimmune blistering diseases, and substantial numbers of CD163+ tissue-associated macrophages (TAMs) are detected in both diseases. PV and BP possess different subsets of helper T cells, suggesting that the cytokine profiles of PV and BP might be different. The purpose of this study was to investigate the microenvironment of lesional skin and serum of PV and BP patients, focusing on the immunomodulatory factors related to TAMs, such as periostin (POSTN), chemokines, cytokines and matrix metalloproteinases (MMPs). We first performed immunohistological staining of POSTN in PV and BP lesions. POSTN was prominent in the superficial dermis in both PV and BP lesions. Next, to validate the activation of CD163+ TAMs in PV and BP patients, we examined the serum levels of soluble (s)CD163. The serum sCD163 levels in PV and BP patients are significantly higher than in healthy controls. To further elucidate the molecular mechanisms of the effects of POSTN on CD163+ TAMs in PV and BP, we examined chemokines, MMPs and cytokines selected by DNA microarray database. The serum CXCL5 levels from PV patients are significantly higher than those in BP patients and healthy controls. The IL-36γ expression on infiltrating macrophages was prominent only in the lesional skin of PV, while the MMP12 deposition was detected in both PV and BP lesions. Our results shed light on the novel pathogenesis of PV through CD163+ TAMs.

Epithelial neutrophil-activating peptide-78 (CXCL5), a member of the subgroup of CXC-type chemokine family, is an inflammatory factor involved in the progression of lung cancer, but the underlying mechanism remains unclear. In this study, we investigated the effects of CXCL5 on proliferation and migration in non-small cell lung cancer (NSCLC) using tissue microarrays from NSCLC patients and H460 cells transfected with a CXCL5-interfered lentivirus vector or stimulated with recombinant CXCL5. We observed that the expression of CXCL5 was significantly higher in lung cancer cell lines, and high CXCL5 was associated with high chemokine (C-X-C motif) receptor 2 expression and was significantly associated with poor differentiation. The high expression of CXCL5 was associated with poor NSCLC prognosis and was an independent predictive factor. Furthermore, downregulation of CXCL5 in H460 cells significantly reduced proliferation and migration. Recombinant CXCL5 promoted H460 cell proliferation and movement by activating MAPK/ERK1/2 and PI3K/AKT signaling. Our study elucidates the important role of CXCL5 in the progression and prognosis of NSCLC. These findings suggested that CXCL5 might be a potential biomarker and novel therapeutic target for lung cancer.