The immunosuppressive properties of mesenchymal stromal cells (MSCs) have been clinically proven to be effective in treating graft-versus-host disease (GVHD). However, MSC therapy is limited by the need for laborious and expensive manufacturing processes that are fraught with batch-to-batch variability. Substitution of MSC therapy with key MSC-mediated immunomodulatory factors could be an option for GVHD treatment. Using a simulated in vitro model of the immunosuppressive effects of MSC on allogeneic graft reactions, a synergistic 2-factor combination (2FC) of CXCL5 and anti-CCL24 was identified from a panel of over 100 immunomodulatory factors as being superior to MSCs in the modulation of mixed lymphocyte reactions. This 2FC was superior to cyclosporine in ameliorating both moderate and severe GVHD while being equivalent to MSCs in moderate GVHD and superior to MSCs in severe GVHD. Its immunosuppressive efficacy could be further improved by extended treatment. Mechanistic studies revealed that in vitro the 2FC could only reduce the proliferation of Th 1 and Th 17, whereas in vivo CXCL5 acts in concert with anti-CCL24 antibody to reduce not only transplanted Th 1 and Th 17 but also cytotoxic T lymphocytes and natural killer cells to increase mouse immunosuppressive neutrophils without affecting human hematopoietic stem cell reconstitution. Concurrently, it reduced circulating human proinflammatory cytokines IFN-γ, IL-6, IL-17A, IL-8, macrophage inflammatory protein-1β, and monocyte chemoattractant protein-1. Both in vitro and in vivo data suggest that CXCL5 and anti-CCL24 antibody act in concert to ameliorate GVHD via suppression of Th 1 and Th 17 responses. We propose that this novel 2FC could substitute for MSC therapy in GVHD treatment.

BACKGROUND

B lymphocyte-induced maturation protein-1 (BLIMP1) is a transcriptional repressor, and plays a crucial role in the regulation of development and functions of various immune cells. Currently, there is limited understanding about the regulation of BLIMP1 expression in keratinocytes and crosstalk between EGFR and BLIMP1 in skin homeostasis.

OBJECTIVE

The aim of the study was to investigate the regulation and functional link between EGFR and BLIMP1 in human epidermal keratinocytes.

METHODS

Immunoblotting and Q-PCR were used to determine the molecular mechanism of BLIMP1 expression induced by EGFR in primary human epidermal keratinocytes (NHEK) and HaCaT cells. In functional assay, effects of BLIMP1 knockdown on EGFR-mediated cytokine production, differentiation, and migration in NHEK were evaluated by Q-PCR, ELISA, immunoblotting, and/or wound-healing assay.

RESULTS

EGFR activation by EGFR ligands could upregulate the protein and mRNA levels of BLIMP1 in NHEK and HaCaT cells. This effect was dependent on PKC, p38, and ERK activation. Additionally, the stability of BLIMP1 protein was under the control of the proteasome and lysosome degradation systems. EGF could also upregulate BLIMP1 expression in skin squamous cell carcinomas. In addition, BLIMP1 knockdown enhanced the EGFR-mediated IL8, CXCL5 and IL6 gene expression and keratinocyte migration, but reduced the EGFR-mediated suppression of differentiation marker K10.

CONCLUSIONS

Our findings shed new insights into the regulation of BLIMP1 expression by EGFR-mediated gene transcription and proteasome/lysosome-mediated degradation in keratinocytes. Functionally, BLIMP1 is a negative regulator of EGF-induced inflammation and migration in keratinocytes, and exerts a gene-specific regulation on keratinocyte differentiation.

<AbstractText>Blunt chest (thoracic) trauma (TxT) is known to contribute to the development of secondary pulmonary complications. Of these, acute lung injury (ALI) is common especially in multiply injured patients and might not only be due to the direct trauma itself, but seems to be caused by ongoing and multifactorial inflammatory changes. Nevertheless, the exact mechanisms and contributing factors of the development of ALI following blunt chest trauma are still elusive.</AbstractText><AbstractText>60 CL57BL/6N mice sustained either blunt chest trauma combined with laparotomy without further interventions or a double hit (DH) including TxT and cecal ligation puncture (CLP) after 24 h to induce ALI. Animals were killed either 6 or 24 h after the second procedure. Pulmonary expression of inflammatory mediators cxcl1, <em>cxcl5</em>, IL-1β and IL-6, neutrophil infiltration and lung tissue damage using the Lung Injury Score (LIS) were determined.</AbstractText><AbstractText>Next to a moderate increase in other inflammatory mediators, a significant increase in CXCL1, neutrophil infiltration and lung injury was observed early after TxT, which returned to baseline levels after 24 h. DH induced significantly increased gene expression of cxcl1, <em>cxcl5</em>, IL-1β and IL-6 after 6 h, which was followed by the postponed significant increase in the protein expression after 24 h compared to controls. Neutrophil infiltration was significantly enhanced 24 h after DH compared to all other groups, and exerted a slight decline after 24 h. LIS has shown a significant increase after both 6 and 24 h compared to both control groups as well the late TxT group.</AbstractText><AbstractText>Early observed lung injury with moderate inflammatory changes after blunt chest trauma recovered quickly, and therefore, may be caused by mechanical lung injury. In contrast, lung injury in the ALI group did not undergo recovery and is closely associated with significant changes of inflammatory mediators. This model may be used for further examinations of contributing factors and therapeutic strategies to prevent ALI.</AbstractText>

The aim of the present study was to identify key genes and signaling pathways associated with the pathogenesis of juvenile spondyloarthritis (JSA). The gene expression profile dataset GSE58667, including data from 15 human whole blood samples collected from 11 patients with JSA and four healthy controls, was analyzed to identify differentially expressed genes (DEGs) associated with disease characteristics. Additionally, Gene Ontology term and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses of the DEGs were performed. Protein‑protein, microRNA‑transcription factor and chemical‑gene interaction networks were constructed. A total of 326 DEGs, 196 upregulated and 130 downregulated, were identified. DEGs, including C‑X‑C motif chemokine ligand 5 (CXCL5), BCL2 interacting protein 3 like (BNIP3L), dual specificity phosphatase 5 (DUSP5) and tumor protein p53 (TP53) were enriched in functions associated with apoptosis, the cell cycle and immune responses. KEGG pathway enrichment analysis revealed that pathways associated with inflammation and the mitogen‑activated protein kinase 1 (MAPK) signaling pathway were the most enriched by DEGs. The results of the present study indicated that the MAPK signaling pathway and four genes, including CXCL5, BNIP3L, DUSP5 and TP53, may be implicated in the pathogenesis of JSA.

Lung epithelial cells are considered important sources of inflammatory molecules and extracellular matrix proteins that contribute to diseases such as asthma. Understanding the factors that stimulate epithelial cells may lead to new insights into controlling lung inflammation. This study sought to investigate the responsiveness of human lung epithelial cells to the TNF family molecules LIGHT and lymphotoxin αβ (LTαβ). Bronchial and alveolar epithelial cell lines, and primary human bronchial epithelial cells, were stimulated with LIGHT and LTαβ, and expression of inflammatory cytokines and chemokines and markers of epithelial-mesenchymal transition and fibrosis/remodeling was measured. LTβ receptor, the receptor shared by LIGHT and LTαβ, was constitutively expressed on all epithelial cells. Correspondingly, LIGHT and LTαβ strongly induced a limited but highly distinct set of inflammatory genes in all epithelial cells tested, namely the adhesion molecules ICAM-1 and VCAM-1; the chemokines CCL5, CCL20, CXCL1, CXCL3, CXCL5, and CXCL11; the cytokines IL-6, activin A and GM-CSF; and metalloproteinases matrix metalloproteinase-9 and a disintegrin and metalloproteinase domain-8. Importantly, induction of the majority of these inflammatory molecules was insensitive to the suppressive effects of the corticosteroid budesonide. LIGHT and LTαβ also moderately downregulated E-cadherin, a protein associated with maintaining epithelial integrity, but did not significantly drive production of extracellular matrix proteins or α-smooth muscle actin. Thus, LIGHT and LTαβ induce a distinct steroid-resistant inflammatory signature in airway epithelial cells via constitutively expressed LTβ receptor. These findings support our prior murine studies that suggested the receptors for LIGHT and LTαβ contribute to development of lung inflammation characteristic of asthma and idiopathic pulmonary fibrosis.

OBJECTIVE

The objective of the study was to investigate whether concentrations of chemokines in the umbilical cord blood of neonates are affected by delivery via cesarean section.

METHODS

Umbilical cord blood was obtained from 116 singleton late-preterm and full-term neonates without infections, born to healthy pregnant women. Concentrations of chemokines - MIP-1α (CCL3), MIP-1β 1 (CCL4), RANTES (CCL5), GRO-α (CXCL1) and ENA-78 (CXCL5) - were measured by ELISA. Logistic regression was used to investigate regression relationships between the occurrence of neonatal chemokines concentrations in umbilical cord blood and mode and time of delivery.

RESULTS

Concentrations of CXC chemokines in late-preterm neonates were the same as those in term neonates. RANTES concentrations in late-preterm cord blood were significantly lower than concentrations in term cord blood. Concentrations of the CC chemokine - RANTES (CCL5) - were noted to be lower in neonates born to cesarean section than in neonates born vaginally. Any anesthetic taken by the mothers during cesarean section did not affect CC chemokine production in the cord blood of full-term neonates. In a logistic regression model including gestational age as a variable, late-preterm delivery was associated with RANTES concentrations (OR = 3.8). After adjustment for variable mode of delivery in regression model, RANTES concentration (OR = 4.75).

CONCLUSIONS

Both late-preterm and cesarean delivery are essential risk factors of low RANTES (CCL5) concentrations in the umbilical cord blood.

BACKGROUND

The alveolar macrophage (AM) - first line of innate immune defence against pathogens and environmental irritants - constitutively expresses peroxisome-proliferator activated receptor γ (PPARγ). PPARγ ligand-induced activation keeps the AM quiescent, and thereby contributes to combat invaders and resolve inflammation by augmenting the phagocytosis of apoptotic neutrophils and inhibiting an excessive expression of inflammatory genes. Because of these presumed anti-inflammatory functions of PPARγ we tested the hypothesis, whether reduced functional receptor availability in mutant mice resulted in increased cellular and molecular inflammatory response during acute inflammation and/or in an impairment of its resolution.

METHODS

To address this hypothesis we examined the effects of a carbon-nanoparticle (CNP) lung challenge, as surrogate for non-infectious environmental irritants, in a murine model carrying a dominant-negative point mutation in the ligand-binding domain of PPARγ (P465L/wt). Animals were instilled intratracheally with Printex 90 CNPs and bronchoalveolar lavage (BAL) was gained 24 h or 72 h after instillation to investigate its cellular and protein composition.

RESULTS

Higher BAL cell numbers - due to higher macrophage counts - were found in mutants irrespective of treatment. Neutrophil numbers in contrast were slightly lower in mutants. Intratracheal CNP instillation resulted in a profound recruitment of inflammatory neutrophils into the alveolus, but genotype related differences at acute inflammation (24 h) and resolution (72 h) were not observed. There were no signs for increased alveolar-capillary membrane damage or necrotic cell death in mutants as determined by BAL protein and lactate-dehydrogenase content. Pro-inflammatory macrophage-derived cytokine osteopontin was higher, but galectin-3 lower in female mutants. CXCL5 and lipocalin-2 markers, attributed to epithelial cell stimulation did not differ.

CONCLUSIONS

Despite general genotype-related differences, we had to reject our hypothesis of an increased CNP induced lung inflammation and an impairment of its resolution in PPARγ defective mice. Although earlier studies showed ligand-induced activation of nuclear receptor PPARγ to promote resolution of lung inflammation, its reduced activity did not provide signs of resolution impairment in the settings investigated here.

Acute ethanol intoxication suppresses the host immune responses against Streptococcus pneumoniae. As interleukin 17 (IL-17) is a critical cytokine in host defense against extracellular pathogens, including S. pneumoniae, we hypothesized that ethanol impairs mucosal immunity against this pathogen by disrupting IL-17 production or IL-17 receptor (IL-17R) signaling. A chronic ethanol feeding model in simian immunodeficiency virus (SIV)-infected rhesus macaques and acute ethanol intoxication in a murine model were used. Transcriptome analysis of bronchial brushes in the nonhuman primate model showed downregulation of the expression of IL-17-regulated chemokines in ethanol-fed animals, a finding also replicated in the murine model. Surprisingly, recombinant CXCL1 and CXCL5 but not IL-17 or IL-23 plus IL-1β rescued bacterial burden in the ethanol group to control levels. Taken together, the results of this study suggest that ethanol impairs IL-17-mediated chemokine production in the lung. Thus, exogenous luminal restoration of IL-17-related chemokines, CXCL1 and CXCL5, improves host defenses against S. pneumoniae.

BACKGROUND

Phenotype of chronic rhinosinusitis (CRS) may be an important determining factor of the efficacy of anti-inflammatory treatments. Although both glucocorticoids and macrolide antibiotics have been recommended for the treatment of CRS, whether they have different anti-inflammatory functions for distinct phenotypic CRS has not been completely understood. The aim of this study is to compare the anti-inflammatory effects of clarithromycin and dexamethasone on sinonasal mucosal explants from different phenotypic CRS ex vivo.

METHODS

Ethmoid mucosal tissues from CRSsNP patients (n = 15), and polyp tissues from eosinophilic (n = 13) and non-eosinophilic (n = 12) CRSwNP patients were cultured in an ex vivo explant model with or without dexamethasone or clarithromycin treatment for 24 h. After culture, the production and/or expression of anti-inflammatory molecules, epithelial-derived cytokines, pro-inflammatory cytokines, T helper (Th)1, Th2 and Th17 cytokines, chemokines, dendritic cell relevant markers, pattern recognition receptors (PRRs), and tissue remodeling factors were detected in tissue explants or culture supernatants by RT-PCR or ELISA, respectively.

RESULTS

We found that both clarithromycin and dexamethasone up-regulated the production of anti-inflammatory mediators (Clara cell 10-kDa protein and interleukin (IL)-10), whereas down-regulated the production of Th2 response and eosinophilia promoting molecules (thymic stromal lymphopoietin, IL-25, IL-33, CD80, CD86, OX40 ligand, programmed cell death ligand 1, CCL17, CCL22, CCL11, CCL5, IL-5, IL-13, and eosinophilic cationic protein) and Th1 response and neutrophilia promoting molecules (CXCL8, CXCL5, CXCL10, CXCL9, interferon-γ, and IL-12), from sinonasal mucosa from distinct phenotypic CRS. In contrast, they had no effect on IL-17A production. The expression of PRRs (Toll-like receptors and melanoma differentiation-associated gene 5) was induced, and the production of tissue remodeling factors (transforming growth factor-β1, epidermal growth factor, basic fibroblast growth factor, platelet derived growth factor, vascular endothelial growth factor, and matrix metalloproteinase 9) was suppressed, in different phenotypic CRS by dexamethasone and clarithromycin in comparable extent.

CONCLUSIONS

Out of our expectation, our explant model study discovered herein that glucocorticoids and macrolides likely exerted similar regulatory actions on CRS and most of their effects did not vary by the phenotypes of CRS.

Angiogenesis as a pathological process in the eye can lead to blindness. In the cornea, suppression of angiogenesis by anti-VEGF treatment is only partially effective while steroids, although effective in treating inflammation and angiogenesis, have broad activity leading to undesirable side effects. In this study, genome-wide expression was investigated in a suture-induced corneal neovascularization model in rats, to investigate factors differentially targeted by dexamethasone and anti-Vegf. Topical treatment with either rat-specific anti-Vegf, dexamethasone, or normal goat IgG (sham) was given to sutured corneas for 48 hours, after which in vivo imaging, tissue processing for RNA microarray, and immunofluorescence were performed. Dexamethasone suppressed limbal vasodilation (P < 0.01) and genes in PI3K-Akt, focal adhesion, and chemokine signaling pathways more effectively than anti-Vegf. The most differentially expressed genes were confirmed by immunofluorescence, qRTPCR and Western blot. Strong suppression of Reg3g and the inflammatory chemokines Ccl2 and Cxcl5 and activation of classical complement pathway factors C1r, C1s, C2, and C3 occurred with dexamethasone treatment, effects absent with anti-Vegf treatment. The genome-wide results obtained in this study provide numerous potential targets for specific blockade of inflammation and angiogenesis in the cornea not addressed by anti-Vegf treatment, as possible alternatives to broad-acting immunosuppressive therapy.

OBJECTIVE

Iron-binding proteins H-ferritin (HF) and lactoferrin (LF), as well as chemokines, tumor necrosis factor (TNF)-alpha, and interferon (IFN)-gamma suppress hematopoietic progenitor cell (HPC) proliferation. Major histocompatibility complex (MHC) class II antigens have been associated with suppressive effects of HF and LF. Because the transcription factor class II transactivator (CIITA) regulates expression of MHC class II antigens, we evaluated influences of CIITA and MHC class II antigens on suppression of colony formation by murine bone marrow HPC in response to HF, LF, CC, and CXC chemokines, TNF-alpha, and IFN-gamma. We also evaluated hematopoiesis in mice deficient in both CIITA and MHC class II antigens (CIITA -/-), in mice deficient in MHC class II antigens but not in CIITA (MHC class II -/-), and in mice deficient in CIITA but not in MHC class II antigens (CIITA-IE).

METHODS

HF, LF, CCL3/MIP-1alpha, CXCL5/ENA-78, CXCL8/IL-8, CCL5/RANTES, TNF-alpha, and IFN-gamma were assessed for effects on colony formation by bone marrow HPC (colony-forming unit granulocyte-macrophage, burst-forming unit erythroid, and colony-forming unit multipotential) stimulated in vitro by combinations of growth factors including erythropoietin, stem cell factor, pokeweed mitogen mouse spleen cell conditioned medium, and hemin. Bone marrow cells were from CIITA -/-, MHC class II antigen -/-, CIITA-IE, and littermate control mice. We also evaluated cycling status (percent cells in S-phase) and absolute numbers of marrow and spleen HPC in these mice.

RESULTS

Multiple growth factor-stimulated colony formation by control bone marrow HPC was significantly suppressed by HF, LF, CCL3, CXCL5, CXCL8, TNF-alpha, and IFN-gamma, but not by CCL5. However, HPC from CIITA -/- and MHC class II antigen -/- mouse marrow was insensitive to inhibition by HF, LF, CCL3, CXCL5, CXCL8, and CCL5; these HPC were inhibited by TNF-alpha and IFN-gamma. Restoration of MHC class II expression in CIITA -/- (CIITA-IE) mice restored responsiveness of HPC to inhibition by HF, LF, CCL3, CXCL5, and CXCL8. Increased cycling of splenic HPC in CIITA -/- and MHC class II antigen -/-, compared to control and CIITA-IE, mice was noted.

CONCLUSIONS

Myelosuppressive effects of iron-binding proteins HF and LF and chemokines CCL3, CXCL5, and CXCL8 on mouse bone marrow HPC require expression of MHC class II antigens, and CIITA is involved in this responsiveness through its regulation of expression of MHC class II antigens.

OBJECTIVE

To study the impact of heparins on chemokines in decidualized human endometrial stromal cells (ESCs) in vitro.

METHODS

In vitro experiment.

METHODS

Research laboratory.

METHODS

Premenopausal women undergoing hysterectomy for benign reasons.

METHODS

ESCs were isolated from hysterectomy specimens, decidualized in vitro and incubated with unfractionated heparin and low-molecular-weight heparins (LMWHs) as well as tumor necrosis factor (TNF) α or thrombin with or without heparins.

METHODS

Chemokines CXCL1, CXCL5, CXCL8, CCL2, and CCL5 were measured with the use of ELISA, and CXCL5, CXCL8, CCL2, and CCL5 were detected with the use of real-time reverse-transcription polymerase chain reaction. Cell viability was determined with the use of a fluorometric assay.

RESULTS

TNF-α and thrombin stimulated distinct patterns of chemokines in ESCs. Unfractionated heparin and LMWHs attenuated the TNF-α-mediated induction of CXCL8 and enhanced CXCL5, CCL2, and CCL5. The stimulating effect of thrombin on CXCL8 could be inhibited by heparin, whereas heparin had no impact on thrombin-induced CXCL1 and CCL2. Nuclear factor of transcription κB signaling mediated the effects of TNF-α. The effects of thrombin were mediated via extracellular signal-regulated protein kinases 1/2.

CONCLUSIONS

Heparins have modulating effects on TNF-α- and thrombin-induced endometrial chemokines, which might have implications in the regulation of endometrial receptivity and early implantation.

Chemokines are components of the skin microenvironment, which enable immune cell chemotaxis. Traditionally, transcription factors involved in inflammatory signaling (eg, NFκB) are important mediators of chemokine expression. To what extent xenobiotics and their associated receptors control chemokine expression is poorly understood. The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor known to mediate physiological responses in the skin through the regulation of genes involved in xenobiotic metabolism, epidermal differentiation, and immunity. Here, we demonstrate that AHR activation within primary mouse keratinocytes regulates the expression of a neutrophil directing chemokine (C-X-C motif) ligand 5 (Cxcl5). AHR-mediated regulation of Cxcl5 is because of direct transcriptional activity upon treatment with AHR agonists such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Additionally, AHR mediates enhanced induction of Cxcl5 upon exposure to an agonist and the inflammatory cytokine interleukin 1 beta. This synergy is confined primarily to keratinocytes, as dermal fibroblasts did not achieve the same level of combinatorial induction. AHR-specific antagonists were able to reduce basal and induced levels of Cxcl5, demonstrating the potential for pharmacological intervention. Exposure of C57BL/6 J mice to ultraviolet (UV) light followed by topical treatment with the AHR agonist formylindolo(3,2-b)carbazole (FICZ) significantly induced Cxcl5 expression in skin compared with UV alone, and this response was absent in Ahr-/- mice. These results establish AHR as an important mediator of Cxcl5, with implications for the treatment of inflammatory skin diseases.

Originally recognized as a regulator of axon guidance in the nervous system, Semaphorin 4D (Sema4D, CD100) also participates in various immune responses and many immune-related diseases. However, whether Sema4D is involved in the pathogenesis of contact hypersensitivity (CHS) remains unclear. In this study, we explored the role of Sema4D in oxazolone-induced CHS using Sema4D knockout (KO) mice. We found that Sema4D KO mice developed attenuated CHS responses, as indicated by milder ear-swelling, lower expression of IL-1β, IL-6, CXCL2 and CXCL5, and decreased recruitment of neutrophils, CD8+ T cells and CD4+ T cells. CHS was impaired in the wide type (WT) mice reconstituted with bone marrow from Sema4D KO mice, indicating that deletion of Sema4D gene in hematopoietic cells played a key role in the alleviated CHS in Sema4D KO mice. CHS was also attenuated in the WT mice transferred with draining lymph nodes (dLNs) cells from oxazolone-sensitized Sema4D KO mice, and the activation and differentiation of hapten-specific CD8+ T cells were impaired in Sema4D KO mice. Furthermore, Sema4D KO mice expressed less IL-1β and CXCL2 than WT mice after oxazolone sensitization, and after transferred with dLNs cells from oxazolone-sensitized WT mice, naïve Sema4D KO mice showed attenuated CHS responses upon oxazolone challenge, indicating that the innate immune response of CHS in Sema4D KO mice was also abrogated. Taken together, our findings revealed for the first time that Sema4D positively regulated both the adaptive and innate immune responses in CHS.

Background. Bronchial smooth muscle cells (BSMC) are a major source of proinflammatory and proangiogenic cytokines and chemokines, including VEGF and CXC-chemokines. CXC-chemokines act primarily on neutrophils, mediating their recruitment to and activation at the site of inflammation. In humans, house-dust mite (HDM) allergens can cause asthmatic exacerbations and trigger an inflammatory response through protease-dependent mechanisms. Objective. We investigated the effect HDM extract on the release of pro-angiogenic and proinflammatory cytokines from BSMC. Methods. Human primary BSMC were stimulated with HDM extract in the absence or presence of fetal calf serum (FCS). Twenty angiogenic cytokines were detected by a specific antibody array and modified protein levels were confirmed by ELISA. Neutrophil migration was measured using a 96-well Boyden chamber. Results. ENA-78/CXCL5 protein levels in conditioned medium of BSMC stimulated with HDM extract were significantly reduced (n = 10, P < 0.05) but restored in the presence of 5% FCS. HDM extracts did not affect ENA-78/CXCL5 mRNA levels. Recombinant ENA-78/CXCL5 was degraded after incubation with HDM extracts (n = 7, P < 0.05) but restored after the addition of the serine protease AEBSF. Neutrophil migration towards recombinant ENA-78/CXCL5 was also reduced in the presence of HDM extract. Conclusion. HDM proteases degrade ENA-78/CXCL5. Thus exposure to HDM allergens may alter ENA-78/CXCL5 levels in the lungs and may affect angiogenesis and the inflammatory response in the airways of asthma patients.

OBJECTIVE

Major trauma initiates a systemic inflammatory response, which is characterized by systemic release of various chemokines. There is growing evidence for the extraordinary role of dendritic cells (DC) as professional antigen-presenting cells and activators of the immune response. Recently, the impact of severe trauma on DC transcriptomic activation was demonstrated. The purpose of the present study was to evaluate gene expression pattern in DC following multiple trauma to gain further understanding of the mechanisms of posttraumatic immune response.

METHODS

Ten patients with multiple injuries aged 20 to 46 years (mean 30 ± 9.2 years) were included in this study. The mean injury severity score (ISS) was 36 ± 10.4 points. Repeated blood samples were taken on the day of admission (day 0) and on five consecutive days (day 1 to day 5). Microarray analysis and RT-qPCR were performed in primary isolated DC.

RESULTS

A mean of 116,000 ± 21,466 DC with a purity of 96 ± 0,8 % were harvested. Gene expression of CCL5 and CXCL5 as well as TIMP1 and GUCY1B3 showed a significant increase within the first 4 days after trauma. The time-dependent increase of these genes correlated significantly with serum CRP concentration and the total number of DC but neither with age nor with injury severity.

CONCLUSIONS

Our study provides new data regarding temporal expression patterns of CCL5, CXCL5, TIMP1, and GUCY13B in multiple trauma. DC activation following trauma may follow a uniform pattern early after admission, eventually leading to cell recruitment.

BACKGROUND

Renal cell carcinoma (RCC) is one of the most common kidney cancers and is highly resistant to chemotherapy. We previously demonstrated that 5-aza-2'-deoxycytidine (DAC) could significantly increase the susceptibility of renal cell carcinoma (RCC) cells to paclitaxel (PTX) treatment in vitro, and showed the synergy of DAC and PTX against RCC. The purpose of this study is to investigated the gene transcriptional alteration and investigate possible molecular mechanism and pathways implicated in the synergy of DAC and PTX against RCC.

METHODS

cDNA microarray was performed and coupled with real-time PCR to identify critical genes in the synergistic mechanism of both agents against RCC cells. Various patterns of gene expression were observed by cluster analysis. IPA software was used to analyze possible biological pathways and to explore the inter-relationships between interesting network genes.

RESULTS

We found that lymphoid enhancer-binding factor 1 (LEF1), transforming growth factor β-induced (TGFBI), C-X-C motif ligand 5 (CXCL5) and myelocytomatosis viral related oncogene (c-myc) may play a pivotal role in the synergy of DAC and PTX. The PI3K/Akt pathway and other pathways associated with cyclins, DNA replication and cell cycle/mitotic regulation were also associated with the synergy of DAC and PTX against RCC.

CONCLUSIONS

The activation of PI3K/Akt-LEF1/β-catenin pathway could be suppressed synergistically by two agents and that PI3K/Akt-LEF1/β-catenin pathway is participated in the synergy of two agents.

OBJECTIVE

Microarray analyses of sinus mucosa in pediatric patients with chronic rhinosinusitis (CRS) have recently demonstrated increased messenger RNA expression of the inflammatory chemokines CXCL5 and CXCL13 and of the innate immune mediators β-defensin 1 (DEFB1), serum amyloid A2 (SAA2), and serpin B4. The objectives of this study were to determine whether these gene products were expressed at the protein level in pediatric sinus mucosa and to determine their localization.

METHODS

Immunohistochemical analysis was used to identify protein expression and cellular localization of CXCL5, CXCL13, DEFB1, SAA2, and serpin B4. Coimmunofluorescence staining of inflammatory cells was performed to further evaluate expression of CXCL5 and CXCL13.

METHODS

Pediatric tertiary care facility.

METHODS

Fifteen children with CRS who underwent endoscopic sinus surgery and 8 children who underwent craniofacial or neurosurgical procedures for abnormalities other than sinusitis.

METHODS

Protein expression and cellular localization of CXCL5, CXCL13, DEFB1, SAA2, and serpin B4 in pediatric sinus mucosa.

RESULTS

Ciliated and basal cells in the pseudostratified epithelium stained positively for the 5 mediators examined in both cohorts. Except for serpin B4, goblet cells did not stain for any mediators in either cohort. Glandular cells stained positively for all 5 mediators in both cohorts. Coimmunofluorescence staining of inflammatory cells showed that CXCL13 was expressed in macrophages, T and B cells but not in neutrophils. CXCL5 was detected only in T cells.

CONCLUSIONS

CXCL5, CXCL13, DEFB1, SAA2, and serpin B4 were expressed at the protein level in the sinus mucosa of controls and pediatric patients with CRS and exhibited cell-specific localization. These mediators, not typically associated with pediatric CRS, may be involved in the inflammatory response and mucus hypersecretion seen in pediatric CRS.

BACKGROUND

Tissue infiltration by neutrophils during acute inflammatory states causes substantial tissue injury. While the magnitude of tissue neutrophil accumulation in innate immune responses is profoundly greater in males than females, fundamental aspects of the molecular mechanisms underlying these sex differences remain largely unknown.

METHODS

We investigated sex differences in neutrophil stimulation and recruitment in ischemia/reperfusion (I/R; mesenteric or renal) or carrageenan pleurisy in rats or mice, as well as skin injury in human volunteers. The induction of potent chemoattractive mediators (chemokines) and neutrophil adhesion molecules were measured by real-time PCR, flow cytometry, and protein assays.

RESULTS

Mesenteric I/R in age-matched Wistar rats resulted in substantially more neutrophil accumulation and tissue injury at 2 h reperfusion in males than females. Using intravital microscopy, we show that the immediate (<30 min) neutrophil response to I/R is similar in males and females but that prolonged neutrophil recruitment occurs in males at sites local and distal to inflammatory insult partly due to an increase in circulating neutrophil populations with elevated surface expression of adhesion molecules. Sex differences in neutrophil kinetics were correlated with sustained induction of chemokine Cxcl5 in the tissue, circulation, and bone marrow of males but not females. Furthermore, blockade of Cxcl5 in males prior to ischemia resulted in neutrophil responses that were similar in magnitude to those in females. Conversely, administration of Cxcl5 to males in the absence of I/R was sufficient to increase levels of systemic neutrophils. Cxcl5 treatment of bone marrow neutrophils in vitro caused substantial induction of neutrophil-mobilizing cytokine granulocyte colony-stimulating factor (GCSF) and expression of β2 integrin that accounts for sexual dimorphism in circulating neutrophil populations in I/R. Moreover, male Cxcl5-stimulated bone marrow neutrophils had an increased capacity to adhere to β2 integrin ligand ICAM-1, implicating a greater sensitivity of male leukocytes to Cxcl5-mediated activation. Differential induction of Cxcl5 (human CXCL6) between the sexes was also evident in murine renal I/R, rat pleurisy, and human skin blisters and correlated with the magnitude of neutrophil accumulation in tissues.

CONCLUSIONS

Our study reveals that sex-specific induction of chemokine Cxcl5/CXCL6 contributes to sexual dimorphism in neutrophil recruitment in diverse acute inflammatory responses partly due to increased stimulation and trafficking of bone marrow neutrophils in males.

Resistance to erythropoietin (EPO) affects a significant number of anaemic patients with end-stage renal disease. Previous reports suggest that inflammation is one of the major independent predictors of EPO resistance, and the effects of EPO treatment on inflammatory mediators are not well established. The aim of this study was to investigate EPO-induced modification to gene expression in primary cultured leucocytes. Microarray experiments were performed on primed ex vivo peripheral blood mononuclear cells (PBMCs) and treated with human EPO-α. Data suggested that EPO-α modulated genes involved in cell movement and interaction in primed PBMCs. Of note, EPO-α exerts anti-inflammatory effects inhibiting the expression of pro-inflammatory cytokine IL-8 and its receptor CXCR2; by contrast, EPO-α increases expression of genes relating to promotion of inflammation encoding for IL-1β and CCL8, and induces de novo synthesis of IL-1α, CXCL1 and CXCL5 in primed cells. The reduction in MAPK p38-α activity is involved in modulating both IL-1β and IL-8 expression. Unlike the induction of MAPK, Erk1/2 activity leads to upregulation of IL-1β, but does not affect IL-8 expression and release. Furthermore, EPO-α treatment of primed cells induces the activation of caspase-1 upstream higher secretion of IL-1β, and this process is not dependent on caspase-8 activation. In conclusion, our findings highlight new potential molecules involved in EPO resistance and confirm the anti-inflammatory role for EPO, but also suggest a plausible in vivo scenario in which the positive correlation found between EPO resistance and elevated levels of some pro-inflammatory mediators is due to treatment with EPO itself.

BACKGROUND Acute respiratory distress syndrome (ARDS) is a common acute and severe disease in clinic. Recent studies indicated that Cxc chemokine ligand 5 (CXCL5), an inflammatory chemokine, was associated with tumorigenesis. The present study investigated the role of the CXCL5/Cxc chemokine receptor 2 (CXCR2) bio-axis in ARDS, and explored the underlying molecular mechanism. MATERIAL AND METHODS The pathological morphology of lung tissue and degree of pulmonary edema were assessed by hematoxylin-eosin staining and pulmonary edema score, respectively. Real-time PCR and Western blot analysis were performed to detect the expression levels of CXCL5, CXCR2, Matrix metalloproteinases 2 (MMP2), and Matrix metalloproteinases 9 (MMP9) in lung tissues. Enzyme-linked immunosorbent assay (ELISA) was performed to determine the expression levels of CXCL5 and inflammatory factors (IL-1ß, IL-6, TNF-alpha, and IL-10) in serum. RESULTS The results demonstrated that diffuse alveolar damage and pulmonary edema appeared in lipopolysaccharide (LPS)-induced ARDS and were positively correlated with the severity of ARDS. In addition, CXCL5 and its receptor CXCR2 were overexpressed by upregulation of MMP2 and MMP9 in lung tissues of ARDS. In addition, CXCL5 neutralizing antibody effectively alleviated inflammatory response, diffuse alveolar damage, and pulmonary edema, and decreased the expression levels of MMP2 and MMP9 compared to LPS-induced ARDS. CONCLUSIONS We found that CXCL5/CXCR2 accelerated the progression of ARDS, partly by upregulation of MMP2 and MMP9 in lung tissues with the release of inflammatory factors.

Non-alcoholic steatohepatitis (NASH) is characterized by hepatocellular steatosis with concomitant hepatic inflammation. Despite its pandemic proportions, no anti-NASH drugs have been approved yet. This is partially because drug development is decelerated due to the lack of adequate tools to assess the efficacy of potential new drug candidates. The present study describes the development and application of a new preclinical model for NASH using hepatic cells generated from human skin-derived precursors. Exposure of these cells to lipogenic (insulin, glucose, fatty acids) and pro-inflammatory factors (IL-1β, TNF-α, TGF-β) resulted in a characteristic NASH response, as indicated by intracellular lipid accumulation, modulation of NASH-specific gene expression, increased caspase-3/7 activity and the expression and/or secretion of inflammatory markers, including CCL2, CCL5, CCL7, CCL8, CXCL5, CXCL8, IL1a, IL6 and IL11. The human relevance of the proposed NASH model was verified by transcriptomics analyses that revealed commonly modulated genes and the identification of the same gene classes between the in vitro system and patients suffering from NASH. The application potential of this in vitro model was demonstrated by testing elafibranor, a promising anti-NASH compound currently under clinical phase III trial evaluation. Elafibranor attenuated in vitro key features of NASH, and dramatically lowered lipid load as well as the expression and secretion of inflammatory chemokines, which in vivo are responsible for the recruitment of immune cells. This reduction in inflammatory response was NFκB-mediated. In summary, this human-relevant, in vitro system proved to be a sensitive testing tool for the investigation of novel anti-NASH compounds.

Epidemiological and animal model studies suggest that sequelae of genital Chlamydia trachomatis infection are more often associated with second or subsequent infections than with initial infection. Further, in order to establish an acute or long-term persistent infection, C. trachomatis develops several strategies to circumvent host immune responses. Hence, resolution of the C. trachomatis infection may require modulation of host factors especially during persistent or chronic infection. Moreover, azithromycin treatment has been reported to possess anti-inflammatory properties but its mechanism of action is still not elucidated. Therefore, in order to better understand the effect of azithromycin in chronic conditions, our aim was to study changes in expression of key genes associated with inflammatory cytokines and receptors, mitogen-activated protein kinase (MAPK) signaling pathway, and apoptosis pathway before and after therapy with azithromycin in infertile women with recurrent C. trachomatis infection. Real-time polymerase chain reaction was performed to study inflammatory cytokines and receptors, MAPK signaling pathway, and apoptosis pathway before and after therapy with azithromycin in infertile women with recurrent C. trachomatis infection. Further, effect of azithromycin on activation of extracellular signal-regulated kinase was studied in epithelial cells by western blotting. Chemokine (C-C motif) ligand 2 (CCL2), CCL5, chemokine (C-X-C motif) ligand 1 (CXCL1), CXCL5, CXCL9, interleukin-1B (IL-1B), IL-8, baculoviral IAP repeat-containing 3 (BIRC3), myeloid cell leukemia sequence 1 (MCL1), and MAPK1 were downregualted after azithromycin treatment. In addition, phosphorylation of extracellular signal-regulated kinase was inhibited after azithromycin treatment in epithelial cells obtained from women with recurrent infection. Hence, our data suggest that azithromycin with its properties apart from antibacterial activity may contribute to its therapeutic potential in treatment of chronic recurrent infection in infertile women.