BACKGROUND

Previous findings support the concept that IL-9 may play a significant role in mediating both pro-inflammatory and changes in airway responsiveness that characterizes the atopic asthmatic state. We previously demonstrated that human airway smooth muscle (ASM) cells express a functional IL-9R that mediate CCL11 expression. However, the signaling pathway governing this effect is not well understood.

RESULTS

In this study, we showed that IL-9 mediated CCL11 expression in ASM cells does not rely on STAT6 or STAT5 but on STAT3 pathway. IL-9 induced rapid STAT3 activation in primary ASM cells that was not observed in case of STAT6 or STAT5. STAT3 binding to CCL11 promoter was also observed in vivo upon IL-9 stimulation of ASM cells. Disruption of STAT3 activity with SH2 domain binding inhibitory peptide results in significant reduction of IL-9 mediated CCL11 promoter activity. DN STAT3beta over-expression in ASM cells, but not Ser 727 STAT3 or STAT6 DN, abolishes IL-9 mediated CCL11 promoter activity. Finally, STAT3 but not STAT6 silenced ASM cells showed significant reduction in IL-9 mediated CCL11 promoter activity and mRNA expression.

CONCLUSIONS

Taken together, our results indicate that IL-9 mediated CCL11 via STAT3 signalling pathway may play a crucial role in airway inflammatory responses.

BACKGROUND

The high affinity IgE receptor (FcepsilonRI) is a crucial structure for IgE-mediated allergic reactions. We have previously demonstrated that human airway smooth muscle (ASM) cells express the tetrameric (alphabetagamma2) FcepsilonRI, and its activation leads to marked transient increases in intracellular Ca(2+) concentration, release of Th-2 cytokines and eotaxin-1/CCL11. Therefore, it was of utmost importance to delineate the factors regulating the expression of FcepsilonRI in human (ASM) cells.

RESULTS

Incubation of human bronchial and tracheal smooth muscle (B/TSM) cells with TNF-alpha, IL-1beta or IL-4 resulted in a significant increase in FcepsilonRI-alpha chain mRNA expression (p<0.05); and TNF-alpha, IL-4 enhanced the FcepsilonRI-alpha protein expression compared to the unstimulated control at 24, 72 hrs after stimulation. Interestingly, among all other cytokines, only TNF-alpha upregulated the FcepsilonRI-gamma mRNA expression. FcepsilonRI-gamma protein expression remained unchanged despite the nature of stimulation. Of note, as a functional consequence of FcepsilonRI upregulation, TNF-alpha pre-sensitization of B/TSM potentially augmented the CC (eotaxin-1/CCL11 and RANTES/CCL5, but not TARC/CCL17) and CXC (IL-8/CXCL8, IP-10/CXCL10) chemokines release following IgE stimulation (p<0.05, n = 3). Furthermore, IgE sensitization of B/TSM cells significantly enhanced the transcription of selective CC and CXC chemokines at promoter level compared to control, which was abolished by Lentivirus-mediated silencing of Syk expression.

CONCLUSIONS

Our data depict a critical role of B/TSM in allergic airway inflammation via potentially novel mechanisms involving proinflammatory, Th2 cytokines and IgE/FcepsilonRI complex.

Chemokines have emerged as critical regulators of leukocyte function and as such represent attractive new targets for the therapy of allergic diseases. Recent studies have revealed important roles for the chemokine family in both the afferent and efferent limbs of the immune system, orchestrating and integrating innate and acquired immune responses. A subset of chemokines including eotaxin-1 (also called CCL11), eotaxin-2 (CCL24), eotaxin-3 (CCL26), MCP (monocyte chemoattractant protein)-3 (CCL7), MCP (monocyte chemoattractant protein)-4 (CCL13), TARC (thymus- and activation-regulated chemokine) (CCL17), and MDC (macrophage-derived chemokine) (CCL22) are highly expressed in allergic inflammation and are regulated by T helper type 2 cytokines. Receptors for these chemokines, including CCR3 (CC chemokine receptor 3), CCR4 (CC chemokine receptor 4) and CCR8 (CC chemokine receptor 8) are expressed on key leukocytes associated with allergic inflammation, such as T helper type 2 cells, eosinophils, mast cells and basophils, establishing a subset of chemokine/chemokine receptors potentially important in allergic inflammation. Recent data using inhibitory antibodies and chemokine antagonists support the concept that interfering with this subset of chemokines and their receptors represents a new approach to allergy immunotherapy.

Signaling of all-trans retinoic acid (ATRA) through nuclear retinoid acid (RA) receptors regulates several biological functions in airway epithelial cells, eosinophils, and immune cells, yet its impact on different in vivo aspects of pulmonary allergic reaction remains elusive. We compared the effect of a treatment with liposomally encapsulated ATRA (Lipo-ATRA) in a mouse model of ovalbumin (OVA)-induced T helper (Th) 2-type responses and airway remodeling. Daily intraperitoneal injections of 10 mg/kg Lipo-ATRA, at the time of each of the 2 systemic sensitizing injections, increased OVA-induced Immunoglobulin E synthesis, bronchoalveolar lavage (BAL) eosinophilia, and accumulation of IL-5, transforming-growth factor beta1, fibronectin, eotaxin/chemokine (C-C motif) ligand 11 (eotaxin/CCL11) and regulated upon activation, normal T expressed and secreted chemokine (C-C motif) ligand 5. In contrast, Lipo-ATRA, administered during each of the 4 intranasal OVA challenges, did not affect these variables. Regardless of the treatment regimen, Lipo-ATRA augmented mucin levels in BAL fluid and reduced lung total collagen content. In vitro incubation of mouse splenocytes or purified spleen cluster of differentiation (CD) 4-positive T lymphocytes, with ATRA, increased, respectively, OVA- and anti-CD 3 antibody-induced IL-4 and IL-5 production and inhibited IFNgamma release. These findings demonstrate that, when given during systemic sensitization, Lipo-ATRA exacerbates allergic immune and inflammatory responses, most likely by promoting Th2 development.

Enhanced disease is the term used to describe the aberrant Th2-skewed responses to naturally acquired human respiratory syncytial virus (hRSV) infection observed in individuals vaccinated with formalin-inactivated viral Ags. Here we explore this paradigm with pneumonia virus of mice (PVM), a pathogen that faithfully reproduces features of severe hRSV infection in a rodent host. We demonstrate that PVM infection in mice vaccinated with formalin-inactivated Ags from PVM-infected cells (PVM Ags) yields Th2-skewed hypersensitivity, analogous to that observed in response to hRSV. Specifically, we detect elevated levels of IL-4, IL-5, IL-13, and eosinophils in bronchoalveolar lavage fluid of PVM-infected mice that were vaccinated with PVM Ags, but not among mice vaccinated with formalin-inactivated Ags from uninfected cells (control Ags). Interestingly, infection in PVM Ag-vaccinated mice was associated with a approximately 10-fold reduction in lung virus titer and protection against weight loss when compared with infected mice vaccinated with control Ags, despite the absence of serum-neutralizing Abs. Given recent findings documenting a role for eosinophils in promoting clearance of hRSV in vivo, we explored the role of eosinophils in altering the pathogenesis of disease with eosinophil-deficient mice. We found that eosinophil deficiency had no impact on virus titer in PVM Ag-vaccinated mice, nor on weight loss or levels of CCL11 (eotaxin-1), IFN-gamma, IL-5, or IL-13 in bronchoalveolar lavage fluid. However, levels of both IL-4 and CCL3 (macrophage inflammatory protein-1alpha) in bronchoalveolar lavage fluid were markedly diminished in PVM Ag-vaccinated, PVM-infected eosinophil-deficient mice when compared with wild-type controls.

Inflammatory mediators, including cytokines, contribute to neuronal and axonal dysfunction and cell death. To examine the roles of cytokines in pathogenesis and regeneration in the central nervous system (CNS), we analyzed effects of cytokines on early gene regulation (6h) in neuronal cultures, employing gene arrays. Our hypothesis is that neuronal gene expression is differentially regulated in vitro by cytokine mixtures typical of Th1 and Th2 T cells and monocytes/macrophages (M/M). Th1 and M/M cytokines showed similar patterns for regulation of numerous pathways including cytokine-receptor interactions, MAP kinase, toll like receptors, apoptosis, PPAR signaling, cell adhesion molecules (CAMS), antigen processing, adipocytokine, and JAK-STAT signaling. M/M cytokines uniquely regulated genes in T cell, B cell and ECM receptor signaling pathways. Th2 cytokines had few effects on pathways regulated by Th1 and MM cytokines, but uniquely regulated genes related to neuroactive ligand-receptors and calcium. Th1 and MM cytokines markedly upregulated a wide array of cytokine-related genes. Notably, M/M cytokines uniquely upregulated G-CSF, GM-CSF, CXCL5 and lymphotactin (Xcl1). Th2 cytokines did not upregulate cytokine-related genes, with the exception of CCL11 and FMS-like tyrosine kinase 1, a VEGF receptor. In neuroactive ligand-receptor pathways, Th1 and M/M cytokines upregulated gene expression for tryptophan hydroxylase. Th1 cytokines upregulated gene expression for GABA A receptor, delta, while Th2 cytokines downregulated GABA A receptor, gamma 3. Significant changes occurred in several genes in the wnt and Notch signaling pathways, which are highly conserved and play critical roles in neuronal and glial differentiation. In the ubiquitin-proteasome pathway, proinflammatory cytokine mixtures induced upregulation of several genes, notably ubiquitin D (Ubd/FAT10), ubiquitin ligase and several proteasomal proteins. In agreement with microarray results, QRT-PCR showed marked upregulation of gene expression for Ubd with Th1 and M/M, for transglutaminase 2 with M/M, and for arginase 1 with Th2 cytokines. Expression of Ubd in the nervous system has not been previously reported. Both message and protein for Ubd are expressed in neurons, and upregulated by pro-inflammatory cytokines. Transglutaminase 2 has been implicated in neurodegenerative diseases, and proposed as a therapeutic target. Upregulation of arginase by Th2 cytokines could be potentially neuroprotective by decreasing NO generation and enhancing neurite outgrowth. Our analysis of changes in neuronal gene expression at the time of initial exposure to an abnormal cytokine milieu provides the opportunity to identify early changes that could be reversed to prevent later irreversible neuronal damage and death in multiple sclerosis and other CNS diseases.

Recent studies suggest that the chemokine eotaxin may participate in atherosclerosis. Threonine (T) for alanine (A) substitution at amino acid 23 in the eotaxin gene (CCL11) has been associated with risk of developing allergic-inflammatory disorders. However, no genetic-epidemiological data are available on the risk of cardiovascular disease associated with this polymorphism. Using DNA samples collected at baseline in a prospective cohort of 14,916 initially healthy American men, we evaluated the A23T polymorphism among 523 individuals who subsequently developed myocardial infarction (MI) and among 2092 individuals who remained free of reported cardiovascular disease over a mean follow-up period of 13.2 years. The T23 allele was significantly associated with risk of myocardial infarction (odds ratio (OR) in an age and smoking adjusted recessive model of inheritance, 1.86; 95% confidence interval (CI), 1.15-3.01; P = 0.012). This risk effect remained statistically significant in analyses further controlling for body mass index, history of hypertension, the presence of diabetes, and randomized treatment assignment (OR, 1.95; 95% CI, 1.19-3.18; P = 0.008). In this cohort, a T for A substitution at amino acid 23 in the eotaxin gene is associated with increased risk for incident myocardial infarction. If confirmed in other cohorts, these data support the emerging hypothesis that eotaxin participates in atherosclerosis.

BACKGROUND

Eosinophilic oesophagitis (EE) is a clinico-pathologically defined oesophageal disorder that is characterized by eosinophil migration into oesophageal tissues. There is growing support for EE being an allergic disease and for a contribution of T-helper type 2 (Th2)-associated cytokines in disease pathogenesis. The respiratory system has been shown to be critical in driving the development of EE in animal models. However, the mechanisms underlying the recruitment of eosinophils into the oesophagus remain unclear.

OBJECTIVE

We sought to investigate the influence of Th2-associated cytokines on the production of eosinophil-specific chemokines from the oesophagus directly.

METHODS

In order to eliminate the potential involvement of the lung, we utilized isolated oesophageal rings. These were treated in vitro with IL-4 or IL-13 and the expression and production of CCL11 and CCL24 were determined.

RESULTS

Our data demonstrate that IL-13 is a potent and direct inducer of both CCL11 and CCL24 production from the oesophagus, as is IL-4 also. The expression of CCL11 precedes CCL24 by several hours but then diminishes over time, as well as at high concentrations of IL-13. We demonstrate that there is an up-regulation of the inhibitory IL-13 receptor, IL-13Ralpha2 but that IL-13Ralpha1 remains unaltered. Oesophagus rings isolated from STAT6(-/-) mice were unable to produce CCL11 or CCL24 upon IL-13 treatment. Lastly, we demonstrate that oesophageal production of CCL11 and CCL24 upon IL-13 stimulation is sufficient to promote eosinophil migration.

CONCLUSIONS

IL-13 is capable of directly stimulating oesophageal tissue to produce eosinophil-attracting chemokines and drive eosinophil migration.

BACKGROUND

Inflammatory and immune alterations occur and may be relevant in patients with schizophrenia. Chemokines are a subgroup of cytokines that play a major role in the recruitment of determined subsets of leukocytes into tissues. To date no study has evaluated whether levels of chemokines are altered in patients with schizophrenia.

OBJECTIVE

To evaluate serum levels of CC and CXC chemokines of schizophrenic patients and age- and gender-matched controls.

METHODS

Forty male institutionalized schizophrenic patients (mean+/-SD age, 52.3+/-9.9) and 20 asymptomatic matched controls were recruited for this study. Severity of symptoms was assessed using BPRS, PANSS and AIMS. All patients were under typical antipsychotic treatment. Serum concentrations of chemokines were measured by ELISA.

RESULTS

There was no statistical difference in serum levels of CCL2, CCL3, CCL24, CXCL9 and CXCL10 between controls and patients. Serum levels of CCL11 were increased in schizophrenic patients when compared to controls. Serum levels of chemokines were not correlated with the length of disease or hospitalization and the severity of involuntary movements, positive and/or negative symptoms.

CONCLUSIONS

CCL11 is a ligand for CCR3, a receptor expressed preferentially on Th2 lymphocytes, mast cells and eosinophils. Higher serum levels of CCL11 in schizophrenia reinforce the view that this disease may be associated with a Th1/Th2 imbalance with a shift toward a Th2 immune response.

Ixodid ticks require comparatively large bloodmeals for their development and survival. Blood-feeding elicits signaling events in the host leading to wound healing responses (hemostasis, inflammation, and tissue repair) and immunity. Bioactive molecules present in tick saliva sabotage these host responses at several levels. One of them is neutralization of cellular communication by binding of specific saliva molecules to cytokines that have important roles in innate and adaptive immunity. Chemokines are a subset of cytokines having chemotactic activities. We show anti-chemokine activities in salivary gland extracts (SGE) of adult Rhipicephalus appendiculatus ticks against human chemokines CXCL8, CCL2, CCL3, CCL5, and CCL11. At comparable protein concentrations, male Ixodes ricinus SGE showed activity against all the chemokines; SGE of female I. ricinus had comparatively lower levels of activity against all the chemokines but no detectable activity against CCL5 and CCL11. However, when the equivalent of a single pair of salivary glands was tested, male I. ricinus showed little or no activity against CCL3 and CCL5. No fundamental differences in activity were observed against mouse compared with human chemokines. A comparison with previously published data for Dermacentor reticulatus and Amblyomma variegatum indicates that the level of anti-cytokine activity depends on the species, developmental stage (adult or nymph), and amount of SGE used, as well as on the number of days the tick has been feeding.

BACKGROUND

Although eosinophils have been detected in several human skin diseases in the vicinity of basophils, how eosinophils infiltrate the skin and the role of eosinophils in the development of skin inflammation have yet to be examined.

OBJECTIVE

Using murine irritant contact dermatitis (ICD) as a model, we sought to clarify the roles of eosinophils in ICD and the underlying mechanism of eosinophil infiltration of the skin.

METHODS

We induced croton oil-induced ICD in eosinophil-deficient ΔdblGATA mice with or without a reactive oxygen species (ROS) inhibitor. We performed cocultivation with fibroblasts and bone marrow-derived basophils and evaluated eosinophil migration using a chemotaxis assay.

RESULTS

ICD responses were significantly attenuated in the absence of eosinophils or by treatment with the ROS inhibitor. ROS was produced abundantly by eosinophils, and both basophils and eosinophils were detected in human and murine ICD skin lesions. In coculture experiments, basophils attracted eosinophils, especially in the presence of fibroblasts. Moreover, basophils produced IL-4 and TNF-α in contact with fibroblasts and promoted the expression of eotaxin/CCL11 from fibroblasts in vitro.

CONCLUSIONS

Eosinophils mediated the development of murine ICD, possibly through ROS production. Recruitment of eosinophils into the skin was induced by basophils in cooperation with fibroblasts. Our findings introduce the novel concept that basophils promote the recruitment of eosinophils into the skin through fibroblasts in the development of skin inflammation.

BACKGROUND

Oxidative Stress contributes to the pathogenesis of many diseases. The NRF2/KEAP1 axis is a key transcriptional regulator of the anti-oxidant response in cells. Nrf2 knockout mice have implicated this pathway in regulating inflammatory airway diseases such as asthma and COPD. To better understand the role the NRF2 pathway has on respiratory disease we have taken a novel approach to define NRF2 dependent gene expression in a relevant lung system.

METHODS

Normal human lung fibroblasts were transfected with siRNA specific for NRF2 or KEAP1. Gene expression changes were measured at 30 and 48 hours using a custom Affymetrix Gene array. Changes in Eotaxin-1 gene expression and protein secretion were further measured under various inflammatory conditions with siRNAs and pharmacological tools.

RESULTS

An anti-correlated gene set (inversely regulated by NRF2 and KEAP1 RNAi) that reflects specific NRF2 regulated genes was identified. Gene annotations show that NRF2-mediated oxidative stress response is the most significantly regulated pathway, followed by heme metabolism, metabolism of xenobiotics by Cytochrome P450 and O-glycan biosynthesis. Unexpectedly the key eosinophil chemokine Eotaxin-1/CCL11 was found to be up-regulated when NRF2 was inhibited and down-regulated when KEAP1 was inhibited. This transcriptional regulation leads to modulation of Eotaxin-1 secretion from human lung fibroblasts under basal and inflammatory conditions, and is specific to Eotaxin-1 as NRF2 or KEAP1 knockdown had no effect on the secretion of a set of other chemokines and cytokines. Furthermore, the known NRF2 small molecule activators CDDO and Sulphoraphane can also dose dependently inhibit Eotaxin-1 release from human lung fibroblasts.

CONCLUSIONS

These data uncover a previously unknown role for NRF2 in regulating Eotaxin-1 expression and further the mechanistic understanding of this pathway in modulating inflammatory lung disease.

BACKGROUND

Primary Sjögren's syndrome (pSS) is an autoimmune rheumatic disease mainly characterised by focal mononuclear cell infiltration in the salivary and lacrimal glands, and by the symptoms xerostomia and keratoconjunctivitis sicca. Germinal centre-like structures (GC) are found in the minor salivary glands of approximately 25% of patients. In this study, we aimed to assess genetic variations in pSS patients with GC-like formations (GC+) compared with patients without such formations (GC-).

METHODS

Minor salivary gland biopsies from Swedish and Norwegian pSS patients (n=320) were evaluated for GC-like formations, identifying 76 GC+ and 244 GC- patients. A panel of 1536 single-nucleotide polymorphisms (SNPs) in 107 genes was genotyped. Minor allele frequencies in GC+ and GC- patients were compared using Fisher's exact test, and associations were considered significant when p<4.7×10(-4) and suggestive when p<0.01.

RESULTS

In this case-only analysis, we identified two SNPs in CCL11 (eotaxin) associated with GC-like structures (p<4.7×10(-4), OR 0.45 and 0.41, respectively). A haplotype of the two minor alleles was associated with GC status with p=2.6×10(-4), OR 0.40. Suggestive associations (p<0.01) were found in SNPs in the B cell activation and/or GC-formation related genes AICDA, BANK1 and BCL2. Furthermore, SNPs in IL17A, ICA1, PKN1 and SNPs in the NF-κB pathway genes CARD8, IKBKE and TANK were found suggestively associated with GC-like structures.

CONCLUSIONS

Our findings suggest that genetic variations may explain why ectopic GC-like structures are present in some pSS patients, and support the hypothesis that GC+ and GC- patients represent distinct disease phenotypes.

Accumulating data have indicated a fundamental role of eosinophils in regulating adipose tissue homeostasis. Here, we performed whole-genome RNA sequencing of the small intestinal tract, which suggested the presence of impaired lipid metabolism in eosinophil-deficient ΔdblGATA mice. ΔdblGATA mice fed a high-fat diet (HFD) showed reduced body fat mass, impaired enlargement of adipocytes, decreased expression of adipogenic genes, and developed glucose intolerance. HFD induced accumulation of eosinophils in the perigonadal white adipose tissue. Concordantly, adipocyte-differentiated 3T3-L1 cells promoted the migration of eosinophils through the expression of CCL11 (eotaxin-1) and likely promoted their survival through the expression of interleukin (IL)-3, IL-5, and granulocyte-macrophage colony-stimulating factor. HFD-fed ΔdblGATA mice showed increased infiltration of macrophages, CD4+ T-cells, and B-cells, increased expression of interferon-γ, and decreased expression of IL-4 and IL-13 in white adipose tissue. Interferon-γ treatment significantly decreased lipid deposition in adipocyte-differentiated 3T3-L1 cells, while IL-4 treatment promoted lipid accumulation. Notably, HFD-fed ΔdblGATA mice showed increased lipid storage in the liver as compared with wild-type mice. We propose that obesity promotes the infiltration of eosinophils into adipose tissue that subsequently contribute to the metabolic homeostasis by promoting adipocyte maturation.

Sexual dimorphism in asthma links the estrogen and allergic immune responses. The function of estrogen was classically believed to be mediated through its nuclear receptors, i.e., estrogen receptors (ERs). However, recent studies established the important roles of G-protein-coupled estrogen receptor (GPER/GPR30) as a novel membrane receptor for estrogen. To date, the role of GPER in allergic inflammation is poorly understood. The purpose of this study was to examine whether GPER might affect the functions of eosinophils, which play an important role in the pathogenesis of asthma. Here, we demonstrated that GPER was expressed in purified human peripheral blood eosinophils both at the mRNA and protein levels. Although GPER agonist G-1 did not induce eosinophil chemotaxis or chemokinesis, preincubation with G-1 enhanced eotaxin (CCL11)-directed eosinophil chemotaxis. G-1 inhibited eosinophil spontaneous apoptosis and caspase-3 activities. The anti-apoptotic effect was not affected by the cAMP-phospodiesterase inhibitor rolipram or phosphoinositide 3-kinase inhibitors. In contrast to resting eosinophils, G-1 induced apoptosis and increased caspase-3 activities when eosinophils were co-stimulated with IL-5. No effect of G-1 was observed on eosinophil degranulation in terms of release of eosinophil-derived neurotoxin (EDN). The current study indicates the functional capacities of GPER on human eosinophils and also provides the previously unrecognized mechanisms of interaction between estrogen and allergic inflammation.

Animal models suggest that chemokines are important mediators in the pathophysiology of neuropathic pain. Indeed, these substances have been called "gliotransmitters," a term that illustrates the close interplay between glial cells and neurons in the context of neuroinflammation and pain. However, evidence in humans is scarce. The aim of the study was to determine a comprehensive cerebrospinal fluid (CSF) inflammatory profile of patients with neuropathic pain. Our hypothesis was that we would thereby find indications of a postulated on-going process of central neuroinflammation. Samples of CSF were collected from 2 cohorts of patients with neuropathic pain (n = 11 and n = 16, respectively) and healthy control subjects (n = 11). The samples were analyzed with a multiplex proximity extension assay in which 92 inflammation-related proteins were measured simultaneously (Proseek Multiplex Inflammation I; Olink Bioscience, Uppsala, Sweden). Univariate testing with control of false discovery rate, as well as orthogonal partial least squares discriminant analysis, were used for statistical analyses. Levels of chemokines CXCL6, CXCL10, CCL8, CCL11, CCL23 in CSF, as well as protein LAPTGF-beta-1, were significantly higher in both neuropathic pain cohorts compared with healthy controls, pointing to neuroinflammation in patients. These 6 proteins were also major results in a recent similar study in patients with fibromyalgia. The findings need to be confirmed in larger cohorts, and the question of causality remains to be settled. Because it has been suggested that prevalent comorbidities to chronic pain (eg, depression, anxiety, poor sleep, and tiredness) also are associated with neuroinflammation, it will be important to determine whether neuroinflammation is a common mediator.

Airway remodeling due to increased airway smooth muscle (ASM) mass, likely due to enhanced migration and proliferation, has been shown to be highly associated with decline in lung function in asthma. Thymic stromal lymphopoietin (TSLP) is an IL-7-like, pro-allergic cytokine that has been shown to be necessary and sufficient for the development of allergic asthma. Human ASM (HASM) cells express TSLP receptor (TSLPR), the activation of which leads to enhanced release of proinflammatory mediators such as IL-6, CCL11/eotaxin-1, and CXCL8/IL-8. We show here that TSLP induces HASM cell migration through STAT3 activation since lentiviral-shRNA inhibition of STAT3 abrogated the TSLP-induced cell migration. Moreover, TSLP induced multiple cytoskeleton changes in HASM cells such as actin polymerization, cell polarization, and activation of small GTPase Rac1. Collectively, our data suggest a pro-migratory function of TSLP in ASM remodeling and provides better rationale for targeting TSLP/TSLPR pathway for therapeutic approaches in allergic asthma.

Carcinoma associated fibroblasts (CAFs) play important roles in breast cancer development and progression. Recent studies show that microRNAs (miRNAs) are the main regulators in CAFs. MiR-29b is one of the significant down-regulated miRNAs in CAFs from the miRNA screening. The role of miR-29b in the interaction between CAFs and breast cancer is still unclear. In the present study, we investigated the effects of CAFs on breast cancer cell proliferation and metastasis regulated by miR-29b. We found that fibroblasts activated by co-cultured breast cancer cells produced higher levels of some chemokines like CCL11, CXCL14, which accelerated breast cancer cell growth and induced drug resistance and metastasis. Increased miR-29b expression in activated fibroblasts could suppress the activating p38-STAT1 signal pathway in breast cancer cells. We also found that the expression of CCL11 and CXCL14 could be regulated by miR-29b in CAFs. Our results illustrate that down-regulation of miR-29b in CAFs plays an important role in tumor stroma by activating p38-STAT1 in breast cancer cells. The study indicates that cancer cells and fibroblasts interaction promotes breast cancer cell growth, drug resistance, migration and invasion due to the lack of miR-29b expression in CAFs.

The chemokine eotaxin/CCL11 is an important mediator of leukocyte migration, but its effect on inflammatory cytokine signaling has not been explored. In this study, we find that CCL11 induces suppressor of cytokine signaling (SOCS)1 and SOCS3 expression in murine macrophages, human monocytes, and dendritic cells (DCs). We also discover that CCL11 inhibits GM-CSF-mediated STAT5 activation and IL-4-induced STAT6 activation in a range of hematopoietic cells. This blockade of cytokine signaling by CCL11 results in reduced differentiation and endocytic ability of DCs, implicating CCL11-induced SOCS as mediators of chemotactic inflammatory control. These findings demonstrate cross-talk between chemokine and cytokine responses, suggesting that myeloid cells tracking to the inflammatory site do not differentiate in the presence of this chemokine, revealing another role for SOCS in inflammatory regulation.

Determining the molecular events induced in the spleen during schistosome infection is an essential step in better understanding the immunopathogenesis of schistosomiasis and the mechanisms by which schistosomes modulate the host immune response. The present study defines the transcriptional and cellular events occurring in the murine spleen during the progression of Schistosoma japonicum infection. Additionally, we compared and contrasted these results with those we have previously reported for the liver. Microarray analysis combined with flow cytometry and histochemistry demonstrated that transcriptional changes occurring in the spleen were closely related to changes in cellular composition. Additionally, the presence of alternatively activated macrophages, as indicated by up-regulation of Chi3l3 and Chi3l4 and expansion of F4/80(+) macrophages, together with enhanced expression of the immunoregulatory genes ANXA1 and CAMP suggests the spleen may be an important site for the control of S. japonicum-induced immune responses. The most striking difference between the transcriptional profiles of the infected liver and spleen was the contrasting expression of chemokines and cell adhesion molecules. Lymphocyte chemokines, including the homeostatic chemokines CXCL13, CCL19 and CCL21, were significantly down-regulated in the spleen but up-regulated in the liver. Eosinophil (CCL11, CCL24), neutrophil (CXCL1) and monocyte (CXCL14, CCL12) chemokines and the cell adhesion molecules VCAM1, NCAM1, PECAM1 were up-regulated in the liver but unchanged in the spleen. Chemokines up-regulated in both organs were expressed at significantly higher levels in the liver. Co-ordinated expression of these genes probably contributes to the development of a chemotactic signalling gradient that promotes recruitment of effector cells to the liver, thereby facilitating the development of hepatic granulomas and fibrosis. Together these data provide, for the first time, a comprehensive overview of the molecular events occurring in the spleen during schistosomiasis and will substantially further our understanding of the local and systemic mechanisms driving the immunopathogenesis of this disease.

BACKGROUND

Respiratory syncytial virus (RSV) causes severe respiratory tract infections, which might have a role in the development of airway hyperreactivity. Mast cells are important effector cells in allergy, with sentinel cell roles in host defense. However, the role of mast cells in response to RSV infection is unknown.

OBJECTIVE

Human mast cell responses to RSV were investigated with a view to better understanding the role of mast cells in RSV-induced disease.

METHODS

Human cord blood-derived mast cells and the HMC-1 mast cell line were exposed to RSV or UV-inactivated RSV. Viral gene and protein expression were evaluated by using PCR and flow cytometry. The expression of interferon-stimulated genes and selected mediators were evaluated by using quantitative PCR and ELISA.

RESULTS

Human mast cells expressed multiple RSV genes after exposure to RSV, and a small percentage of mast cells supported RSV antigen protein expression. RSV induced mast cells to upregulate production of chemokines, including CCL4, CCL5, and CXCL10, as well as type I interferons, and interferon-stimulated gene expression. However, production of the granulocyte chemoattractants CXCL8 and CCL11 was not induced. Antibody blockade of the type I interferon receptor on human cord blood-derived mast cells reduced the RSV-mediated induction of CXCL10 and CCL4 but not CCL5. Leukotriene C4 production by mast cells was not enhanced by exposure to RSV.

CONCLUSIONS

Despite low levels of infection, human mast cells produce multiple chemokines in response to RSV through mechanisms that include responses to type I interferons. Such mast cell responses might enhance effector cell recruitment during RSV-induced disease.

Drug-induced liver injury (DILI) is a major health issue, as it remains difficult to predict which new drugs will cause injury and who will be susceptible to this disease. This is due in part to the lack of animal models and knowledge of susceptibility factors that predispose individuals to DILI. In this regard, liver eosinophilia has often been associated with DILI, although its role remains unclear. We decided to investigate this problem in a murine model of halothane-induced liver injury (HILI). When female Balb/cJ mice were administered halothane, eosinophils were detected by flow cytometry in the liver within 12 hours and increased thereafter proportionally to liver damage. Chemokines, eotaxin-1 (CCL11) and eotaxin-2 (CCL24), which are known to attract eosinophils, increased in response to halothane treatment. The severity of HILI was decreased significantly when the study was repeated in wildtype mice made deficient in eosinophils with a depleting antibody and in eosinophil lineage-ablated ΔdblGata(-/-) mice. Moreover, depletion of neutrophils by pretreating animals with Gr-1 antibody prior to halothane administration failed to reduce the severity of HILI at antibody concentrations that did not affect hepatic eosinophils. Immunohistochemical staining for the granule protein, major basic protein, revealed that eosinophils accumulated exclusively around areas of hepatocellular necrosis.

CONCLUSIONS

Our findings indicate that eosinophils have a pathologic role in HILI in mice and suggest that they may contribute similarly in many clinical cases of DILI.

In this review, we highlight experiments conducted in our laboratories that have elucidated functional roles for CD4+ T-helper type-2 lymphocytes (TH 2 cells), their associated cytokines, and eosinophils in the regulation of hallmark features of allergic asthma. Notably, we consider the complexity of type-2 responses and studies that have explored integrated signaling among classical TH 2 cytokines (IL-4, IL-5, and IL-13), which together with CCL11 (eotaxin-1) regulate critical aspects of eosinophil recruitment, allergic inflammation, and airway hyper-responsiveness (AHR). Among our most important findings, we have provided evidence that the initiation of TH 2 responses is regulated by airway epithelial cell-derived factors, including TRAIL and MID1, which promote TH 2 cell development via STAT6-dependent pathways. Further, we highlight studies demonstrating that microRNAs are key regulators of allergic inflammation and potential targets for anti-inflammatory therapy. On the background of TH 2 inflammation, we have demonstrated that innate immune cells (notably, airway macrophages) play essential roles in the generation of steroid-resistant inflammation and AHR secondary to allergen- and pathogen-induced exacerbations. Our work clearly indicates that understanding the diversity and spatiotemporal role of the inflammatory response and its interactions with resident airway cells is critical to advancing knowledge on asthma pathogenesis and the development of new therapeutic approaches.

BACKGROUND

There is a need for a readily available, non-invasive source of biomarkers that predict poor asthma control.

OBJECTIVE

We sought to determine if there is an association between the salivary inflammatory profile and disease control in children and adults with asthma.

METHODS

In this cross-sectional study, we collected demographic and clinical information from two independent populations at different sites, resulting in convenience samples of 58 pediatric and 122 adult urban asthmatics. Control was assessed by symptom questionnaire (children) and by Asthma Control Questionnaire and current exacerbation (adults). Saliva was collected in all subjects. We applied principal component analysis to a 10-plex panel of relevant inflammatory markers to characterize marker profiles and determined if profiles were associated with asthma control.

RESULTS

There were similar, strong correlations amongst biologically related markers in both populations: eosinophil-related: eotaxin-1/CCL11, RANTES/CCL5, and IL-5 (p<.001); myeloid/innate: IL-1β, IL-6, MCP-1/CCL2, and IL-8/CXCL8 (p<.001). The first three principal components captured ≥74% of variability across all ten analytes in both populations. In adults, the Principal Component 1 score, broadly reflective of all markers, but with greater weight given to myeloid/innate markers, was associated with Asthma Control Questionnaire score and exacerbation. The Principal Component 3 score, reflective of IP-10/CXCL10, was associated with current exacerbation. In children, the Principal Component 1, 2, and 3 scores were associated with recent asthma symptoms. The Principal Component 2 score, reflective of higher eosinophil markers, was inversely correlated with symptoms. The Principal Component 3 score was positively associated with all symptom outcomes.

CONCLUSIONS

The salivary inflammatory profile is associated with disease control in children and adults with asthma.