Oxidative stress plays a pivotal role in the pathogenesis of asthma. Aquaporin-3 (AQP3) is a small transmembrane water/glycerol channel that may facilitate the membrane uptake of hydrogen peroxide (H2O2). Here we report that AQP3 potentiates ovalbumin (OVA)-induced murine asthma by mediating both chemokine production from alveolar macrophages and T cell trafficking. AQP3 deficient (AQP3(-/-)) mice exhibited significantly reduced airway inflammation compared to wild-type mice. Adoptive transfer experiments showed reduced airway eosinophilic inflammation in mice receiving OVA-sensitized splenocytes from AQP3(-/-) mice compared with wild-type mice after OVA challenge, consistently with fewer CD4(+) T cells from AQP3(-/-) mice migrating to the lung than from wild-type mice. Additionally, in vivo and vitro experiments indicated that AQP3 induced the production of some chemokines such as CCL24 and CCL22 through regulating the amount of cellular H2O2 in M2 polarized alveolar macrophages. These results imply a critical role of AQP3 in asthma, and AQP3 may be a novel therapeutic target.

In the murine model, it was demonstrated that pro-inflammatory cytokines and chemokines are essential to the formation and modulation of Schistosoma-induced granulomatous inflammation. However, the relationship of these immune mediators and disease severity is hard to be established in naturally infected individuals. The current study evaluates the association between plasma concentrations of MIF, sTNF-R1, CCL3, CCL7 and CCL24 and schistosomiasis morbidity in Schistosoma mansoni-infected patients with a low parasite burden. For this propose, 97 S. mansoni-infected individuals were subjected to abdominal ultrasound analysis and clinical examination. Among them, 88 had plasma concentration of immune mediators estimated by ELISA assay. Multivariate linear regression models were used to evaluate the relationship between the plasma concentration of immune mediators and the variables investigated. Although most individuals presented low parasite burden, over 30% of them showed signs of fibrosis defined by ultrasound measurements and 2 patients had a severe form of schistosomiasis. No association between parasite burden and the plasma levels of chemokine/cytokines or disease severity was observed. There was a positive association between plasma concentration of CCL4, sTNF-R1, CCL3 and MIF with gall bladder thickness and/or with portal vein thickness that are liver fibrosis markers. In contrast, no association was found between CCL7 plasma concentrations with any of the schistosomiasis morbidity parameters evaluated. The data showed that CCL24, sTNFR1, MIF and CCL3 can be detected in plasma of S. mansoni-infected individuals and their concentration would be used as prognostic makers of Schistosoma-induced liver fibrosis, even in individuals with low parasite burden.

The majority of pregnancy loss in ruminants occurs during the preimplantation stage, which is thus the most critical period determining reproductive success. Here, we performed a comparative transcriptome study by sequencing total mRNA from corpus luteum (CL) collected during the preimplantation stage of pregnancy in Finnsheep, Texel and F1 crosses. A total of 21,287 genes were expressed in our data. Highly expressed autosomal genes in the CL were associated with biological processes such as progesterone formation (STAR, CYP11A1, and HSD3B1) and embryo implantation (e.g., TIMP1, TIMP2 and TCTP). Among the list of differentially expressed genes, sialic acid-binding immunoglobulin (Ig)-like lectins (SIGLEC3, SIGLEC14, SIGLEC8), ribosomal proteins (RPL17, RPL34, RPS3A, MRPS33) and chemokines (CCL5, CCL24, CXCL13, CXCL9) were upregulated in Finnsheep, while four multidrug resistance-associated proteins (MRPs) were upregulated in Texel ewes. A total of 17 known genes and two uncharacterized non-coding RNAs (ncRNAs) were differentially expressed in breed-wise comparisons owing to the flushing diet effect. The significantly upregulated TXNL1 gene indicated potential for embryonic diapause in Finnsheep and F1. Moreover, we report, for the first time in any species, several genes that are active in the CL during early pregnancy (including TXNL1, SIGLEC14, SIGLEC8, MRP4, and CA5A).

Chronic rhinosinusitis (CRS) is characterized by sustained mucosal inflammation, impaired mucociliary clearance, loss of cilia and epithelial barrier breakdown, and tissue remodeling. Certain glycosaminoglycans inhibit various inflammatory mediators, suppress bacterial growth, and provide important functions in mucosal tissue repair and mucociliary clearance. Herein, we evaluated the effects of a synthetic glycosaminoglycan, GM-1111, on the clinical signs and inflammatory tissue changes associated with CRS in mice. CRS was generated by repeated intranasal applications of Aspergillus fumigatus (A. fumigatus) extracts over 4 weeks. Mice were then intranasally administered GM-1111 (600 μg per dose, 5 times a week) or vehicle (phosphate buffered saline, PBS) for an additional 4 weeks while still being given A. fumigatus extracts to maintain a chronic inflammatory environment with acute exacerbations. Clinical signs indicative of sinonasal inflammation were recorded throughout the study. After 9 weeks, whole blood and sinonasal tissues were harvested for hematological, histological, and biochemical examination. The clinical signs, white blood cell counts, tissue markers of sinonasal inflammation, and histological changes caused by A. fumigatus extract administration were compared to the healthy (PBS vehicle) and GM-1111-treated groups (n = 12 per treatment group). Compared to vehicle-treated animals, animals treated with GM-1111 demonstrated significant reductions in clinical signs (p<0.05), degenerative tissue changes, goblet cell hyperplasia, inflammatory cell infiltration (p<0.01), innate immunity- (tlr2, tlr4, myd88, il1b, tnfa, il6, and il12) and adaptive immunity-associated (ccl11, ccl24, ccl5, il4, il5, and il13) cytokine gene expression (p<0.05 to p<0.0001) in sinonasal tissues, and serum IgE levels (p<0.01). Our data suggest that GM-1111 significantly reduces local and systemic effects of CRS-associated sinonasal inflammation.

Human chemokine receptor CCR3 (hCCR3) belongs to the G protein-coupled receptors (GPCRs) superfamily of membrane proteins and plays major roles in allergic diseases and angiogenesis. In order to study the structural and functional mechanism of hCCR3, it is essential to produce pure protein with biological functions on a milligram scale. Here we report the expression of hCCR3 gene in a tetracycline-inducible stable mammalian cell line. A cell clone with high hCCR3 expression was selected from 46 stably transfected cell clones and from this cell line pure hCCR3 on a milligram scale was obtained after two-step purification. Circular dichroism spectrum with a characteristic shape and magnitude for α-helix indicated proper folding of hCCR3 after purification. The biological activity of purified hCCR3 was verified by its high binding affinity with its endogenous ligands CCL11 and CCL24, with K D in the range of 10(-8) M to 10(-6) M.

EBI3 functions as the subunit of immune-regulatory cytokines, such as IL-27 and IL-35, by pairing with p28 and p35, respectively. We treated wild-type and EBI3-deficient mice with intratracheal administration of LPS and obtained bronchoalveolar lavage fluid (BALF) 24 h later. Although neutrophils were the predominant cells in BALF from both groups of mice, eosinophils were highly enriched and there was increased production of eosinophil-attracting chemokines CCL11 and CCL24 in BALF of EBI3-deficient mice. The bronchial epithelial cells and alveolar macrophages were the major producers of CCL11 and CCL24. Because no such increases in eosinophils were seen in BALF of p28/IL-27-deficient mice or WSX-1/IL-27Rα subunit-deficient mice upon intratracheal stimulation with LPS, we considered that the lack of IL-35 was responsible for the enhanced airway eosinophilia in EBI3-deficient mice. In vitro, IL-35 potently suppressed production of CCL11 and CCL24 by human lung epithelial cell lines treated with TNF-α and IL-1β. IL-35 also suppressed phosphorylation of STAT1 and STAT3 and induced suppressor of cytokine signaling 3. In vivo, rIL-35 dramatically reduced LPS-induced airway eosinophilia in EBI3-deficient mice, with concomitant reduction of CCL11 and CCL24, whereas neutralization of IL-35 significantly increased airway eosinophils in LPS-treated wild-type mice. Collectively, our results suggest that IL-35 negatively regulates airway eosinophilia, at least in part by reducing the production of CCL11 and CCL24.

Investigations revealed substantial parts accomplished by chemokines specifically eotaxins and their specific receptors. They are functionally involved in the modulation of the pathologic state of tissue inflammation which is as a result of allergic reactions. Chemokines as small proteins with approximately 8-10 kDa molecular weight are considered and fit in the bigger family of cytokines, containing basic heparin-binding polypeptide mediators. Chemokines actively interfere in the processes of selective, oriented leukocyte (including eosinophil) recruitment. As eminent from their name, more specifically, eotaxins are specialized for eosinophils' oriented locomotion toward allergic inflamed regions. To date, three members are defined for eotaxin subfamily as follows: eotaxin-1 (CCL11), eotaxin-2 (CCL24), and eotaxin-3 (CCL26), all of them bind to and activate CCR3 but have a low level of homology and appear to exhibit different physiological potentials. Allergy is described as a clinical state in which a pathologic hypersensitivity reaction is always initiated throughout an immunologic mechanism; similar to other immunologic reactions, an allergic reaction could also either be antibody or cell mediated. This type of allergic reactions occurs in all age groups and damages several different organs, having a significant impact on the emotional and social health of patients and their families and relatives. Concerning introductory comments introduced above, the authors of the present review attempted to collect and provide the latest evidences and information regarding the correlation between expression of eotaxin family members and allergy, in a wider extent, in two important allergic disorders: atopic asthma (AA) and atopic dermatitis (AD). Overall, concerning the most recent articles published within the database in the life sciences literature regarding the fundamental role(s) played by eotaxins in the pathogenesis of AA and AD, the authors of the current article propose that eotaxins (CCL11, CCL24, and CCL26) play key role(s) during symptomatic inflammatory responses raised in response to allergic crisis of these two clinical states.

Microbiota maintains host tissue homeostasis and influences tissue-resident macrophages. However, the mechanisms by which commensal bacteria in regulating the alveolar macrophages remain unclear. Here, by using an antibiotic-treated (Abt) mouse model, we found commensal bacteria depletion induced lower frequencies and numbers of alveolar macrophages. This effect was accompanied by the altered levels of genes involved in several biological pathways, including M2 macrophage polarization, as determined by gene expression analysis. Alveolar macrophages from the Abt mice had higher protein and gene levels of Arg1, CCL24, IL-13, IL-10, IL-6 and IL-1β, which could be recovered to normal levels by reconstructing commensal bacteria in the upper respiratory of Abt mice. Moreover, alveolar macrophages performed significant enhancement of M2 functions, especially CCL24 secretion, in the Abt mice challenged with B16/F10 melanoma. Adoptive transfer of normal alveolar macrophages or antibody neutralization of CCL24 significantly recovered the decrease of γδT17 cells and rescued the defect anti-tumor response of Abt mice, indicating the elevated amount of alveolar macrophage-derived CCL24 inhibited γδT cell mediated anti-tumor response. In conclusion, we demonstrated the ability of commensal bacteria to maintain alveolar macrophages with a low level of CCL24 production, which was necessary for the normal anti-tumor response in the lung.

The increasing number of eosinophils into bronchoaelvolar space is observed during noninfectious inflammatory lung diseases. Eotaxins (eotaxin-1/CCL11, eotaxin-2/CCL24, eotaxin-3/CCL26) are the strongest chemotactic agents for eosinophils. Inhibitors of phosphodiesterase 4 (PDE4), the enzyme decomposing cAMP, are anti-inflammatory agents which act through cAMP elevation and inhibit numerous steps of allergic inflammation. The effect of PDE4 inhibitors on eotaxin expression is not known in details. The aim of our study was to evaluate the influence of PDE4 inhibitors: rolipram and RO-20-1724 on expression of eotaxins in bronchial epithelial cell line BEAS-2B. Cells were preincubated with PDE4 inhibitors or dexamethasone for 1 hour and then stimulated with IL-4 or IL-13 alone or in combination with TNF-α. After 48 hours eotaxin protein level was measured by ELISA and mRNA level by real time PCR.

RESULTS

PDE4 inhibitors decreased CCL11 and CCL26 expression only in cultures co-stimulated with TNF-α. In cultures stimulated with IL-4 and TNF-α rolipram and RO-20-1724 diminished CCL11 mRNA expression by 34 and 37%, respectively, and CCL26 by 43 and 47%. In cultures stimulated with IL-13 and TNF-α rolipram and RO-20-1724 decreased expression of both eotaxins by about 50%. These results were confirmed at the protein level. The effect of PDE4 inhibitors on eotaxin expression in BEAS-2B cells, in our experimental conditions, depends on TNF-α contribution.

Calprotectin is a heterodimer of the proteins S100A8 and S100A9 and is an abundant innate immune protein associated with inflammation. In humans, calprotectin transcription and protein abundance are associated with asthma and disease severity. However, mechanistic studies in experimental asthma models have been inconclusive, identifying both protective and pathogenic effects of calprotectin. To clarify the role of calprotectin in asthma, calprotectin-deficient S100A9^-/- and wild-type C57BL/6 mice were compared in a murine model of allergic airway inflammation. Mice were intranasally challenged with extracts of the clinically relevant allergen Alternaria alternata (Alt Ext) or PBS every third day over 9 days. On day 10, bronchoalveolar lavage fluid and lung tissue homogenates were harvested and allergic airway inflammation was assessed Alt Ext challenge induced release of S100A8/S100A9 to the alveolar space and increased protein expression in the alveolar epithelium of wild-type mice. Compared to wild-type mice, S100A9^-/- mice displayed significantly enhanced allergic airway inflammation, including production of IL-13, CCL11, CCL24, serum IgE, eosinophil recruitment, and airway resistance and elastance. In response to Alt Ext, S100A9^-/- mice accumulated significantly more IL-13⁺IL-5⁺CD4⁺ Th2 cells. S100A9^-/- mice also accumulated a significantly lower proportion of CD4⁺ T regulatory cells in the lung that had significantly lower expression of CD25. Calprotectin enhanced wild-type T regulatory cell suppressive activity in vitro. Therefore, this study identifies a role for the innate immune protein S100A9 in protection from CD4⁺ Th2 hyperinflammation in response to Alt Ext. This protection is mediated, at least in part, by CD4⁺ T regulatory cell function.

The aim of this study was to investigate the modulation of plasma CXCL10, CCL20, CCL22, CCL2, CCL17 and CCL24 levels in HIV-positive patients grouped according to extreme phenotypes of progression to AIDS, and at different stages of HIV infection. HIV-positive individuals with extreme phenotypes of AIDS progression (n=58) at different clinical stages (chronic individuals, both pre-HAART and under-HAART) and HIV-negative controls (n=20) were evaluated. Additionally, HIV-positive individuals that initiated HAART with >350CD4+T-cells/mm3 were compared with those who initiated treatment with <350CD4+T-cells/mm3. Plasma levels of six chemokines were quantified by a Luminex assay. Higher CXCL10 levels were observed in individuals immediately before their CD4+T-cell levels were indicative for HAART (pre-HAART), independently of their progressor status, i.e. slow (SPs) or rapid progressors (RPs). SPs pre-HAART showed higher CXCL10 levels compared to elite controllers and RPs under HAART (pc=0.009 and pc=0.007, respectively). CXCL10 levels were higher in SPs HAART CD4<350 (initiated HAART with <350 CD4+T-cells) when compared with SPs HAART CD4>350 (initiated HAART with >350 CD4+T-cells) (1096 vs. 360.33pg/mL, p=0.0101). Normalisation of CXCL10 levels seems to depend on the CD4+T-cell nadir at HAART initiation. CCL20 levels were higher in chronic SPs, SPs pre-HAART, SPs HAART and RPs HAART compared with the HIV-negative controls, indicating persistent CCL20 expression. In conclusion, our results indicate that CXCL10 levels are a hallmark in the clinical evolution of HIV infection. However, our results must be verified in a study evaluating a larger number of AIDS progressors.

IL-13 and IL-4 are potent mediators of type 2-associated inflammation such as those found in atopic dermatitis (AD). IL-4 shares overlapping biological functions with IL-13, a finding that is mainly explained by their ability to signal via the type 2 IL-4 receptor (R), which is composed of IL-4Rα in association with IL-13Rα1. Nonetheless, the role of the type 2 IL-4R in AD remains to be clearly defined. Induction of two distinct models of experimental AD in Il13ra1^-/- mice, which lack the type 2 IL-4R, revealed that dermatitis, including ear and epidermal thickening, was dependent on type 2 IL-4R signaling. Expression of TNF-α was dependent on the type 2 IL-4R, whereas induction of IL-4, IgE, CCL24, and skin eosinophilia was dependent on the type 1 IL-4R. Neutralization of IL-4, IL-13, and TNF-α as well as studies in bone marrow-chimeric mice revealed that dermatitis, TNF-α, CXCL1, and CCL11 expression were exclusively mediated by IL-13 signaling via the type 2 IL-4R expressed by nonhematopoietic cells. Conversely, induction of IL-4, CCL24, and eosinophilia was dependent on IL-4 signaling via the type 1 IL-4R expressed by hematopoietic cells. Last, we pharmacologically targeted IL-13Rα1 and established a proof of concept for therapeutic targeting of this pathway in AD. Our data provide mechanistic insight into the differential roles of IL-4, IL-13, and their receptor components in allergic skin and highlight type 2 IL-4R as a potential therapeutic target in AD and other allergic diseases such as asthma and eosinophilic esophagitis.

Colorectal cancer (CRC) is one of the most common malignancies in the world, with a global incidence of almost 2 million new cases every year. Despite the availability of many diagnostic tests, including laboratory tests and molecular diagnostics, an increasing number of new cases is observed. Thus, it is very important to search new markers that would show high diagnostic sensitivity and specificity in the detection of colorectal cancer in early stages of the disease. Eotaxins are proteins that belong to the cytokine group-small molecules with a variety of applications. Their main role is the activation of basophils and eosinophils involved in inflammatory processes. Therefore, we performed an extensive search of the literature pertaining to our investigation via the MEDLINE/PubMed database. On the basis of available literature, we can assume that eotaxins accumulate in cancer cells in the course of CRC. This leads to a decrease in the chemotaxis of eosinophils, which are effector immune cells with anti-tumor activity. This may explain a decrease in their number as a defense mechanism of cancer cells against their destruction and may be useful when attempting anti-tumor therapy with the use of chemokines.

Keywords: CCL11; CCL24; CCL26; CCR3; CRC.

BACKGROUND

Macrophages can differentiate into pro-inflammatory (M1) or anti-inflammatory (M2) phenotypes upon exposure to a pathogen or a cytokine microenvironment. However, M1/M2 macrophage polarization in polymicrobial sepsis has not been fully characterized.

METHODS

The polarity of peritoneal exudate (PE) cells from mice that had undergone cecal ligation and puncture (CLP) and the response of those cells to lipopolysaccharide (LPS) in terms of cytokine and chemokine expression were examined.

RESULTS

PE cells from CLP mice demonstrated a shift toward the M2 phenotype in terms of marker enzyme expression. In addition, the CLP-derived PE cells showed apparent unresponsiveness to LPS stimulation with regard to expression of pro-inflammatory cytokines such as TNF-α, while the expression of anti-inflammatory cytokines such as IL-10 was induced. Nevertheless, the CLP-PE cells failed to express M2 chemokines including chemokine (C-C motif) ligand 17 (CCL17), CCL22, and CCL24, all of which are important for T cell recruitment.

CONCLUSIONS

The results suggested that a shift of naïve monocytes/macrophages to the M2 phenotype, along with the lack of M2 chemokine expression in septic monocytes/macrophages, might be responsible for immunosuppression after sepsis.

Bupleurum chinense is distributed in East Asia and has been used as a traditional herbal medicine for more than a thousand years. Though B. chinense has been reported to have immunomodulatory, anti-inflammatory, anti-oxidant, hepato-protective, antipyretic, analgesic and antifibrotic effects, its specific effect on allergic rhinitis disease has not been clarified. In this study, we investigated the anti-allergic and anti-inflammation effects of B. chinense extract (BCE) in an ovalbumin (OVA)-induced allergic rhinitis (AR) mouse model. Oral administration of BCE in a dose-independent manner regulated the balance of Th1/Th2/Treg cell differentiation in AR mice. Accordingly, BCE attenuated the expression of Th2-related cytokines such as IL-4, IL-5 and IL-13 in nasal lavage fluid (NALF) and nasal tissue and up-regulated the secretion of Th1/Treg cells including IL-10, IL-12 and IFN- <mml:math>γ</mml:math> . Also, BCE inhibited the formation and migration of eosinophils to the nasal mucosa and NALF, as well as suppressed <em>CCL24</em>, an eosinophil-specific chemoattractant in NALF. The levels of anti-OVA specific IgE and anti-OVA specific IgG1 were decreased, and as a result, the allergic response was attenuated by BCE via inhibiting mast cells accumulation in nasal mucosa and serum histamine release. The nasal allergy symptoms, nasal mucosal swelling, epithelial barrier disruption and mucus hyperplasia were obviously ameliorated. These results suggest that BCE may have therapeutic potential for treating allergic rhinitis through modulating the accumulation and activation of important leukocytes in the immune system such as Th1, Th2, Treg, eosinophils and mast cells.

Chemokines (CKs) and chemokine receptors (CKR) promote leukocyte recruitment into cardiac tissue infected by the Trypanosoma cruzi. This study investigated the long-term treatment with subantimicrobial doses of doxycycline (Dox) in association, or not, with benznidazole (Bz) on the expression of CK and CKR in cardiac tissue. Thirty mongrel dogs were infected, or not, with the Berenice-78 strain of T. cruzi and grouped according their treatments: (i) two months after infection, Dox (50 mg/kg) 2x/day for 12 months; (ii) nine months after infection, Bz (3,5 mg/kg) 2x/day for 60 days; (iii) Dox + Bz; and (iv) vehicle. After 14 months of infection, hearts were excised and processed for qPCR analysis of Th1 (CCL2, CCL3, CCL4, CCL5, CXCL9, and CXCL11), Th2 (CCL1, CCL17, CCL24, and CCL26), Th17 (CCL20) CKs, Th1 (CCR5, CCR6, and CXCR3), and Th2/Th17 (CCR3, CCR4, and CCR8) CKR, as well as IL-17. T. cruzi infection increases CCL1, CCL2, CCL4, CCL5, CCL17, CXCL10, and CCR5 expression in the heart. Dox, Bz, or Dox + Bz treatments cause a reversal of CK and CKR and reduce the expression of CCL20, IL-17, CCR6, and CXCR3. Our data reveal an immune modulatory effect of Dox with Bz, during the chronic phase of infection suggesting a promising therapy for cardiac protection.

Although elevated blood or sputum eosinophils are present in many patients with chronic obstructive pulmonary disease (COPD), uncertainties remain regarding the anatomical distribution pattern of lung-infiltrating eosinophils. Basophils have remained virtually unexplored in COPD. This study mapped tissue-infiltrating eosinophils, basophils, and eosinophil-promoting immune mechanisms in COPD-affected lungs.Surgical lung tissue and biopsies from major anatomical compartments were obtained from COPD patients with severity grades GOLD I-IV; never-smokers/smokers served as controls. Automated immunohistochemistry and in-situ hybridisation identified immune cells, the type 2 immunity marker GATA3, and eotaxins (CCL11, CCL24).Eosinophils and basophils were present in all anatomical compartments of COPD-affected lungs and increased significantly in very severe COPD. The eosinophilia was strikingly patchy, and focal eosinophil-rich microenvironments were spatially linked with GATA3⁺ cells, including Th₂ lymphocytes and type 2 innate lymphoid cells. A similarly localised and IL-33/ST2-dependent eosinophilia was demonstrated in influenza-infected mice. Both mice and patients displayed spatially confined eotaxin signatures with CCL11⁺ fibroblasts and CCL24⁺ macrophages.In addition to identifying tissue basophilia as a novel feature of advanced COPD, the identification of spatially confined eosinophil-rich type 2 microenvironments represents a novel type of heterogeneity in the immunopathology of COPD that will likely have implications for personalised treatment.

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.

OBJECTIVE

The expression of cytokine-associated genes in dendritic cells (DCs) derived from umbilical cord blood (UCB) and adult peripheral blood (APB) was comprehensively compared in order to elucidate the difference in DC function between newborns and adults.

METHODS

Immature DCs were obtained from UCB and APB of healthy human donors. Several cytokines were added to generate mature DCs. Gene expression was compared using cDNA microarray containing 553 cytokine-associated genes. Eleven genes with differential expression were selected and determined their expression levels in DCs by quantitative real-time RT-PCR.

RESULTS

The expression of the Th1 response-related genes (IL-12B and IL-18) and chemokine genes (CXCL9, CXCL13, CCL18 and CCL24) was significantly lower in UCB-DCs than in APB-DC in both maturation states. On the other hand, calgranulins A and B, which are speculated to induce immune tolerance, showed higher expression in UCB-DCs. The expression of cell cycle-related genes (CDC2 and cyclin B1) was significantly higher in UCB-DCs than in APB-DCs, and immature UCB-DCs proliferated more rapidly than immature APB-DCs.

CONCLUSIONS

The expression of genes related to immune responses was significantly different between UCB- and APB-DCs, which may cause a decreased DC-mediated immunity and an increased susceptibility to infection in newborns.

Targeting chemokine signaling is an attractive avenue for the treatment of inflammatory disorders. Tyrosine sulfation is an important post-translational modification (PTM) that enhances chemokine-receptor binding and is also utilized by a number of pathogenic organisms to improve binding affinity of immune-suppressive chemokine binding proteins (CKBPs). Here we report the display selection of tyrosine-sulfated cyclic peptides using a reprogrammed genetic code to discover high-affinity ligands for the chemokine CCL11 (eotaxin-1). The selected cyclic sulfopeptides possess high affinity for the target chemokine (as well as one or more of the related family members CCL2, CCL7 and CCL24) and inhibit CCL11 activation of CC chemokine receptor 3 (CCR3). This work demonstrates the utility in exploiting native PTMs as binding motifs for the generation of new leads for medicinal chemistry.

The process of spatial rearrangement of cells of the inner cell mass (ICM) that are destined to become hypoblast is not well understood. The observation that the chemokine (C-C motif) ligand 24 (CCL24) and several other genes involved in chemokine signaling are expressed more in the ICM than in the trophectoderm of the bovine embryo resulted in the hypothesis that CCL24 participates in spatial organization of the ICM. Temporally, expression of CCL24 in the bovine embryo occurs coincidently with blastocyst formation: transcript abundance was low until the late morula stage, peaked in the blastocyst at Day 7 of development and declined by Day 9. Treatment of embryos with two separate antagonists of C-C motif chemokine receptor 3 (the prototypical receptor for CCL24) decreased the percent of GATA6+ cells (hypoblast precursors) that were located in the outside of the ICM. Similarly, injection of zygotes with a CCL24-specific morpholino decreased the percent of GATA6+ cells in the outside of the ICM. In conclusion, CCL24 assists in spatial arrangement of the ICM in the bovine embryo. This experiment points to new functions of chemokine signaling in the bovine embryo and is consistent with the idea that cell migration is involved in the spatial organization of hypoblast cells in the blastocyst.

Background: Eosinophils develop from CD34⁺ progenitor cells in the bone marrow under the influence of interleukin (IL)-5. Several cell types produce IL-5, including type 2 innate lymphoid cells (ILC2s). The alarmin cytokine IL-33 is known to activate ILC2s in mucosal tissues, but little is known about IL-33-responsive ILC2s in the bone marrow in allergen-induced airway inflammation. Methods: Wild type (WT) and Rag1 deficient (Rag1 ^-/-) mice, which lack mature T and B cells, received intranasal doses of papain to induce acute allergic inflammation. In some experiments, mice were pre-treated with anti-IL-5 prior to the papain challenge. Furthermore, recombinant IL-33 was administered to WT mice, Rag1 ^-/- mice, lymphocyte deficient mice (Rag2 ^-/- Il2rg ^-/-) and to ex vivo whole bone marrow cultures. Bone marrow eosinophils and ILC2s were analyzed by flow cytometry. Eosinophil count was assessed by differential cell count and secreted IL-5 from bone marrow cells by ELISA. Results: Intranasal administration of papain or IL-33 increased the number of mature eosinophils in the bone marrow despite the absence of adaptive immune cells in Rag1 ^-/- mice. In parallel, an increased number of eosinophils was observed in the airways together with elevated levels of Eotaxin-2/CCL24. Bone marrow ILC2s were increased after papain or IL-33 administration, whereas ILC2s was found to be increased at baseline in Rag1 ^-/- mice compared to WT mice. An upregulation of the IL-33 receptor (ST2) expression on bone marrow ILC2s was observed after papain challenge in both Rag1 ^-/- and WT mice which correlated to increased number of bone marrow eosinophilia. Furthermore, an increased number of ST2⁺ mature eosinophils in the bone marrow was observed after papain challenge, which was further dependent on IL-5. In addition, bone marrow-derived ILC2s from both mouse strains produced large amounts of IL-5 ex vivo after IL-33 stimulation of whole bone marrow cultures. In contrast, IL-33-induced bone marrow and airway eosinophilia were abolished in the absence of ILC2s in Rag2 ^-/- Il2rg ^-/- mice and no production of IL-5 was detected in IL-33-stimulated bone marrow cultures. Conclusion: These findings establish bone marrow ILC2s and the IL-33/ST2 axis as promising targets for modulation of uncontrolled IL-5-dependent eosinophilic diseases including asthma.

Keywords: ILC2; allergy; asthma; bone marrow; eosinophilia; interleukin-33; interleukin-5; papain.

The underlying inflammation present in chronic airway diseases is orchestrated by increased expression of CC chemokines that selectively recruit leukocyte populations into the pulmonary system. Human CCL26 signals through CC chemokine receptor 3 (CCR3), is dramatically upregulated in challenged asthmatics, and stimulates recruitment of eosinophils (EOSs) and other leukocytes. CCL26 participates in regulation of its receptor CCR3 and modulates expression of a variety of chemokines in alveolar type II cells. Utilizing the A549 alveolar type II epithelial cell culture model, we carried out studies to test the hypothesis that CCL26-siRNA treatment of these cells would ameliorate Th2-driven release of the eotaxins and other CCR3 ligands that would, in turn, decrease recruitment and activation of EOSs. Results demonstrate that CCL26-siRNA treatments decreased interleukin-4-induced CCL26 and CCL24 expression by >70%. CCL26-directed small-interfering RNA (siRNA) treatments significantly decreased release of CCL5 (RANTES), CCL15 (MIP-1δ), CCL8 (MCP-2), and CCL13 (MCP-4). In bioactivity assays it was shown that EOS migration and activation were reduced up to 80% and 90%, respectively, when exposed to supernatants of CCL26-siRNA-treated cells. These results provide evidence that CCL26 may be an appropriate target for development of new therapeutic agents designed to alleviate the underlying inflammation associated with chronic diseases of the airways.

Paracoccidioidomycosis (PCM) is the most common systemic mycosis in Latin America. A major problem in the management of PCM is to determine the best time to discontinue therapy due to the high relapse rate among patients. Soluble TNF receptors (sTNF-R) levels and chemokines are associated with disease activity in several infectious, inflammatory and autoimmune disorders. The aim of the present work was to evaluate levels of sTNF-R1, sTNF-R2 and chemokines in serum of patients with adult type of PCM, before and after antifungal therapy, and to correlate those levels to disease activity. Concentrations of sTNF-R1, sTNF-R2 and CXCL9 were higher in untreated patients and decreased progressively with treatment. The serum marker with the best accuracy to discriminate PCM cases from controls was sTNF-R2. sTNF-R1 did not drop to control levels before 36 months of treatment. CCL2 and CCL3 levels were low at baseline in PCM patients, raised significantly after 12 months of treatment and diminished thereafter. CCL24 levels were higher after 36 months of antifungal therapy in PCM patients. CCL11 levels were not statistically different from control subjects. sTNF-R1, sTNF-R2 and CXCL9 may be useful as laboratory parameters to assess disease activity in PCM patients.