TARC Production Research Articles

Casuarinin suppresses TARC/CCL17 and MDC/CCL22 production via blockade of NF-κB and STAT1 activation in HaCaT cells

Casuarinin is a naturally occurring tannin that is isolated from the leaves of Hippophae rhamnoides. It has been shown to have anti-oxidant, anti-cancer, anti-viral, and anti-inflammatory activities. The aim of this study was to investigate the possible mechanism by which casuarinin inhibits TNF-α/IFN-γ-induced Th2 chemokines expression in the human keratinocytes cell line HaCaT. We found that casuarinin suppressed TNF-α/IFN-γ-induced expression of TARC and MDC mRNA and protein in HaCaT cells. Casuarinin significantly inhibited TNF-α/IFN-γ-induced activation of NF-κB, STAT1, and p38 MAPK. Furthermore, we observed that p38 MAPK contributes to inhibition of TNF-α/IFN-γ-induced TARC and MDC production by blocking NF-κB and STAT1 activation in HaCaT cells. Taken together, these results suggest that casuarinin may exert anti-inflammatory responses by suppressing TNF-α/IFN-γ-induced expression of TARC and MDC via blockage of p38 MAPK activation and subsequent activation of NF-κB and STAT1. We propose that it could therefore be used as a therapeutic agent against inflammatory skin diseases.

CCR4 Expression in Hodgkin Lymphoma

Abstract 2626In Hodgkin lymphoma (HL) a minority (<1%) of tumor cells is found in a reactive background. The tumor cells shape their environment by the production of several chemokines and cytokines. Production of the chemokine TARC will lead to the attraction of Th2 and Treg cells specifically to involved lymph nodes. Although Treg and Th2 cells are prominent in the infiltrate the expression of its receptor CCR4 is not very high in lymphocytes surrounding the tumor cells. We hypothesized that this is due to internalization of the receptor. We analyzed in which specific compartment of lymphocytes CCR4 is expressed and if CCR4 expression can be recovered when TARC and MDC levels are low.We analyzed 11 lymph node suspensions involved with HL and 11 with follicular hyperplasia (RLN) for expression of CD4, CD26, FoxP3 and CCR4 at t=0 and t=24 hours. We tested the effect of TARC and MDC on CCR4 expression and the mechanism of internalization on normal PBMCs.At t=0 the CD4+ T cell compartment was significantly lower for CD26 in HL (49% vs 68% in RLN, p<0.01) (Table 1). In the comparison between CD26+ and CD26- cells no difference was seen in CCR4 expression in HL but in RLN CCR4 in CD26- cells was significantly lower than in CD26+ cells (p<0.01).Recovery of CCR4 (in average %) after 24 hours was significantly lower in CD26+ cells and FoxP3+ cells in RLN (p<0.01 and p<0.001). Recovery of CCR4 as factor change is significantly higher in HL than in RLN for CCR4 as well as the 3 subpopulations (Table 2). Generally, internalization of CCR4 on lymphocytes is higher with MDC (54 to 90% internalization with 10 and 500ng/ml MDC) than TARC (28 to 48% with 10 and 1000ng/ml TARC) after 2 hours. Internalization occurs through the lipid raft/caveolae dependent pathway as filipin III could inhibit internalization completely and not by clathrin-mediated endocytosis since no effect of inhibition by sucrose was seen. Recovery of CCR4 back on the membrane is fast and by exocytosis since it can be disrupted by exocytosis inhibitor Brefeldin A.Table 1:Percentages subpopulations CD4+ T cells in Hodgkin lymphoma and reactive lymph node by flowcytometry on cell suspensionsHLRLNpCD264968<0.01CCR44137nsFoxP34.41.8nsCCR4/CD26+1925nsCCR4/CD26-2212nsCCR4/FoxP3+5560nsTable 2:Factor of CCR4 recovery after 24 hours in CD4+ subpopulations in Hodgkin lymphoma and reactive lymph node cell suspensionsHLRLNpCCR41.70.9<0.05CCR4 in CD26+1.30.8<0.05CCR4 in CD26-2.91.1<0.05CCR4 in FoxP3+1.60.8<0.05In conclusion, CD26 expression is significantly lower in HL than RLN. CCR4 expression is equally divided over CD26+ and CD26- cells in HL, but in RLN less is found in CD26-. Recovery of CCR4 expression after culture for 24 hours is lower in RLN than in HL. More CCR4 is recovered in HL especially in the CD26- CD4+ T cell population that is surrounding the tumor cells in HL. Disclosures:No relevant conflicts of interest to declare.

Quercetagetin, a component of premature Citrus unshiu (Swingle) Marcow., suppress the chemokines related with atopic dermatitis by regulating STAT1 signal

Atopic dermatitis (AD) is an ithchy and relapsing inflammatory skin disease. It was known that a predominant systemic Th2 dysbalance with increased IgE levels and eosinophilia is widely accepted in the pathogenesis of AD [1]. Thymus and activation-regulated chemokine (TARC/CCL17) and macrophage-derived chemokine (MDC/CCL22) are related with AD and are elevated in serum and lesional skin of AD patients [2, 3]. Citrus unshiu (CU) contains various flavonoids that have various bioactive effects [4]. In present study, we investigated the effect of a component of premature CU, quercetagetin, on the production of TARC and MDC in HaCaT human keratinocytes. As results, quercetagetin showed inhibitory activity on the protein production and mRNA expression of TARC and MDC in IFN-γ and TNF-α-stimulated HaCaT human keratinocytes. Also, quercetagetin inhibited the phosphorylation of STAT1, key transcription factor initiating IFN-γ signaling pathway, in a time- and dose-dependent manner. These results suggest that quercetagetin, a component of premature CU, may have an anti-atopic activity by inhibiting the inflammatory chemokines (TARC and MDC) via the STAT1 pathway.

TSLP: a novel potent proinflammatory mediator that activates myeloid dendritic cells to stimulate Th1 and Th17 activity in rheumatoid arthritis

BackgroundThymic stromal lymphopoietin (TSLP) is well known for its potent activation of myeloid dendritic cells (mDCs) resulting in Th2-mediated immune responses. TSLP signals cells via the interleukin (IL)-7 receptor-α chain...

Gene silencing of STAT6 with siRNA ameliorates contact hypersensitivity and allergic rhinitis

Silencing of genes using small interfering RNA (siRNA) is a recently developed strategy to regulate the synthesis of target molecules. Signal transducer and activator of transcription 6 (STAT6) is a nuclear transcription factor that mediates Th2-type immunity. To elucidate the therapeutic potential of using siRNA to inhibit STAT6 in allergic reactions, we determined the nucleotide sequences of siRNA specific for STAT6. The selected sequences of STAT6 siRNA specifically inhibited the generation of STAT6 synthesis in dermal fibroblasts and eotaxin (CCL11) production in response to IL-4/TNF-α in vitro. Local administration of STAT6 siRNA in vivo alleviated contact hypersensitivity responses to chemical haptens. This was accompanied by reduced local production of IL-4, IL-13, eotaxin (CCL11), TARC (CCL17) and MDC (CCL22). Similarly, consecutive intranasal instillation of STAT6 siRNA markedly inhibited inflammatory cellular infiltration of mucosal tissues in allergic rhinitis responses in association with reduced IL-4 and IL-5 production from regional lymph node cells. Immediate responses, such as sneezing and nasal rubbing behaviors, were also improved by STAT6 siRNA. Local administration of STAT6 siRNA is thus a promising therapeutic strategy for both Th2-mediated cutaneous diseases and allergic rhinitis.

Reactive oxygen species are involved in the IFN‐γ‐stimulated production of Th2 chemokines in HaCaT keratinocytes

The increased generation of reactive oxygen species (ROS) induces inflammation in different cell types. However, it is unclear whether ROS play an essential role in the production of thymus and activation-regulated chemokine (TARC/CCL17) and macrophage-derived chemokine (MDC/CCL22) in keratinocytes. Here, we investigated the function of ROS in the production of these two Th2 chemokines in interferon-gamma (IFN-γ)-treated HaCaT keratinocytes. We found that IFN-γ-induced production of both chemokines in parallel with the increased generation of intracellular ROS. A ROS scavenger, N-acetyl cysteine (NAC), significantly inhibited the IFN-γ-induced production of chemokines as well as the activation of I kappa-B (IκB)-nuclear factor-kappa B (NF-κB). Inhibitors of Janus family kinases (JAKs), p38 mitogen-activated kinase (MAPK), and NF-κB suppressed IFN-γ-induced production of TARC and MDC. NF-κB activation was inhibited by both inhibitors of JAKs and p38 MAPK. Importantly, IFN-γ-stimulated phosphorylation of p38 MAPK was significantly suppressed by JAKs inhibitors, but not significantly affected by NAC or L-buthionine sulfoximine (L-BSO). However, IFN-γ-stimulated activation of IκB and NF-κB was suppressed by NAC but enhanced by BSO. Furthermore, inhibition of p38 MAPK and JAKs did not affect ROS generation in IFN-γ-stimulated HaCaT cells. These results indicate that intracellular ROS and JAKs/p38 MAPK both contribute independently to IFN-γ-stimulated production of TARC and MDC in HaCaT keratinocytes, by increasing NF-κB activation.

Tannic acid and quercetin display a therapeutic effect on atopic dermatitis via suppression of angiogenesis and Th2 polarization. (97.7)

Abstract Atopic dermatitis (AD) is a chronic inflammatory skin disease that is characterized by inflammation, abnormal angiogenesis and elevated Th2 polarization. Tannic acid (TA) and quercetin are well known polypenolic compounds, have documented anti-inflammatory effect. However, little is known about the effects of TA and quercetin on atopic dermatitis and related mechanisms. The present study investigated whether TA and quercetin had a therapeutic effect on atopic dermatitis. In vitro, we found that TA and quercetins reduced pro-angiogenic factor and Th2-inducing cytokine in human keratinocytes. The inhibitory effect was higher in the combined application of the two agents than in the application of one agent alone. To investigate the effect of combined application with TA and quercetin in vivo, an AD-induced Nc/Nga mouse model was used. The group receiving TA and quercetin showed disease improvement with down-regulation of serum IgE, TARC production and angiogenesis level compare to non-treated group. Taken together, these data demonstrate that TA and quercetin have a therapeutic effect on AD by suppressing angiogenesis, Th2 polarization.

Suppressive activity of epinastine hydrochloride on TARC production from human peripheral blood CD4+ T cells in-vitro

Thymus- and activation-regulated chemokine (TARC) is an important molecule in the development and maintenance of allergic diseases. However, there is little information about the influence of anti-allergic agents on TARC production. The aim of this study is to examine the influence of epinastine hydrochloride, an H1-receptor antagonist, on TARC production from human peripheral blood CD4+ T cells using an in-vitro cell culture technique. CD4+ T cells prepared from healthy subjects were cultured in wells coated with a combination of OKT3 and anti-CD28 monoclonal antibody in the presence or absence of epinastine HCl for 24 h. The cells were also stimulated with interleukin (IL)-4 in a similar manner. Levels of TARC and IL-4 in culture supernatants were examined by ELISA. The addition of epinastine HCl exerted a dose-dependent suppressive effect on the production of both TARC and IL-4 from CD4+ T cells under co-stimulatory molecule stimulation. The minimum concentration of the agent showing a significant suppressive effect on TARC and IL-4 production was 5.0 microM and 2.5 microM, respectively. Epinastine HCl also suppressed the ability of cells to produce TARC in response to IL-4 stimulation, when the agent was added to cell cultures at more than 2.5 microM. It was concluded that this inhibitory action of epinastine HCl may be partially responsible for epinastine's attenuating effect on allergic diseases.

The adenylyl cyclase-cAMP system suppresses TARC/CCL17 and MDC/CCL22 production through p38 MAPK and NF-κB in HaCaT keratinocytes

Patients with atopic dermatitis (AD) have significantly reduced plasma cAMP levels, and the cAMP level is correlated with the immunopathogenesis of AD. The production of thymus and activation-regulated chemokine (TARC/CCL17) and macrophage-derived chemokine (MDC/CCL22) in keratinocytes is significantly enhanced in patients with AD. In the present study, we investigated the in vitro effects of the adenylyl cyclase-cAMP system on IFN-γ and TNF-α-stimulated production of TARC and MDC in human HaCaT keratinocytes. Both forskolin (a direct activator of adenylyl cyclase) and dibutyryl-cAMP (DBcAMP, a permeable analog of cAMP) suppressed production of TARC and MDC in parallel with the activation of NF-κB in IFN-γ and TNF-α-stimulated HaCaT cells. Moreover, inhibition of NF-κB suppressed TARC and MDC production induced by IFN-γ plus TNF-α. However, dideoxyforskolin, a forskolin derivative that does not activate cAMP, failed to suppress the secretion of these chemokines. An inhibitor of p38 MAPK suppressed the production of TARC and MDC in parallel to the activation of NF-κB in HaCaT cells. Of note, the IFN-γ plus TNF-α-stimulated activation of p38 MAPK was suppressed following incubation with forskolin or DBcAMP alone. These results indicate that the adenylyl cyclase-cAMP system has an inhibitory role in IFN-γ plus TNF-α-stimulated production of TARC and MDC in HaCaT keratinocytes by inhibiting NF-κB activation through p38 MAPK pathway, implying that the adenylyl cyclase-cAMP system could be a candidate therapeutic target of Th2-skewed skin inflammation such as AD.

Role of TARC/CCL17 and CTACK/CCL27 in the Pathogenesis of Atopic Dermatitis

TARC/CCL17 is designated as a Th2 type chemokine since it binds to CCR4. CTACK/CCL27 is a ligand for CCR10 and is selectively expressed in skin. Atopic dermatitis (AD) is a chronic inflammatory skin disease that is characterized by the predominant infiltration of Th2-type cells, especially in the acute phase. We have determined that serum levels of TARC and CTACK are elevated in AD patients and reflect the disease activity of AD. We investigated the regulation of production of TARC and CTACK from the human keratinocyte (KC) cell line, HaCaT cells. Production of both was upregulated after stimulation with inflammatory cytokines TNF-α/IFN-γ and TNF-α/IL-1β, respectively, and the upregulated production was downregulated by adding inhibitors for nuclear factor kappa B (NFκB) and p38. We have created transgenic (Tg) mice in which TARC or CTACK is overexpressed in KC. Interestingly, both TARC Tg mice and CTACK Tg mice show enhanced contact hypersensitivity to Th2, but not to Th1 stimuli. AD-like conditions such as increased number of mast cells and elevated levels of serum IgE were observed in both Tg mice by Th2-type stimuli. Thus, TARC and CTACK may participate in the pathogenesis of Th2-shifted skin diseases such as AD.

Vorinostat inhibits STAT6-mediated TH2 cytokine and TARC production and induces cell death in Hodgkin lymphoma cell lines

AbstractEpigenetic changes have been implicated in silencing several B-cell genes in Hodgkin and Reed-Sternberg cells (HRS) of Hodgkin lymphoma (HL), and this mechanism has been proposed to promote HRS survival and escape from immunosurveillance. However, the molecular and functional consequences of histone deacetylase (HDAC) inhibition in HL have not been previously described. In this study, we report that the HDAC inhibitor vorinostat induced p21 expression and decreased Bcl-xL levels causing cell-cycle arrest and apoptosis. Furthermore, vorinostat inhibited STAT6 phosphorylation and decreased its mRNA levels in a dose- and time-dependent manner, which was associated with a decrease in the expression and secretion of Thymus and Activation-Regulated Chemokine (TARC/CCL17) and interleukin (IL)–5 and an increase in IP-10 levels. Moreover, vorino-stat inhibited TARC secretion by dendritic cells that were activated by the thymic stromal lymphopoietin (TSLP). Collectively, these data suggest that pharmacologic HDAC inhibition in HL may induce favorable antitumor activity by a direct antiproliferative effect on HRS cells, and possibly by an immune mediated effect by altering cytokine and chemokines secretion in the microenvironment.

Regulatory effects of antihistamines on the responses to staphylococcal enterotoxin B of human monocyte-derived dendritic cells and CD4 + T cells

Antihistamines are widely used for the treatment of allergic diseases, such as urticaria and allergic rhinitis. They are also effective for the treatment of diseases in which T cells are mainly involved in the pathogenesis, such as atopic dermatitis (AD) and contact dermatitis. Dendritic cells (DCs) drive polarization of naive T cells into Th1 or Th2 subsets, and are also likely to be involved in AD pathogenesis. The aim of this study was to determine the effects of antihistamines on DCs and CD4+ T cells. Human monocyte-derived DCs (MoDCs) and autologous CD4+ T cells were obtained from healthy subjects, and cultured together or independently in the presence of antihistamines. As a stimulant, we used staphylococcal enterotoxin B or the combination of anti-CD3 monoclonal antibody (mAb) and anti-CD28 mAb. The concentrations of cytokines and chemokines in culture supernatants were measured by ELISA. The expression of surface molecules on MoDCs was measured by flow cytometry. Cell proliferation in the cocultures of MoDCs and CD4+ T cells (DC-T cocultures) was measured by a [(3)H] thymidine incorporation assay. Antihistamines inhibited the production of IFN-gamma, and enhanced the production of IL-4 in DC-T cocultures. Antihistamines inhibited the production of TNF-alpha, TARC, MDC, IP-10, and Mig from MoDCs. Epinastine, one of antihistamines, suppressed the expression of ICAM-1 (CD54) on MoDCs. Epinastine also inhibited the proliferation of CD4+ T cells cocultured with MoDCs. Our findings show that antihistamines regulate immune responses by affecting the interaction between DCs and CD4+ T cells, and further DCs and CD4+ T cells independently, which may partially contribute to the control of allergic diseases such as AD and contact dermatitis.

Suppressive effects of Houttuynia cordata Thunb (Saururaceae) extract on Th2 immune response

Ethnopharmacological relevance Houttuynia cordata Thunb (Saururaceae), known as ‘E-Sung-Cho’ in Korea, has been traditionally used for the treatment of herpes simplex, chronic sinusitis, and allergy. Aim of the study To investigate the inhibitory activity of Houttuynia cordata Thunb fractions (HcFs) on the T helper 2 (Th2) immune response, we evaluated the alternation of the release of Th2-type cytokines and chemokines such as interleukin (IL)-4 and IL-5, and thymus and activation-regulated chemokine (TARC/CCL17). Materials and methods Ethanol fraction was obtained from dried and powdered whole plants of Houttuynia cordata Thunb using ethanol. The residue was diluted with water and was then successively partitioned with n-hexane, EtOAc and BuOH. HcFs include ethanol, n-hexane, EtOAc, BuOH and water fractions. RT-PCR and ELISA were performed to measure mRNA and protein expression of cytokines. Results HcFs inhibited the expression of IL-4 and IL-5 in response to phorbol 12-myristate 13-acetate (PMA) and calcium ionophore (CaI) in Jurkat T cells and the human mast cell line, HMC-1. IL-4- and tumor necrosis factor-alpha (TNF-α)-induced TARC production was blocked by HcFs in skin fibroblast CCD-986sk cells, particularly by the ethanol extract of Hc. Stimulants included in PMA, phytohemagglutinin (PHA) and CaI, increased the mRNA level of CC chemokine receptor 4 (CCR4), a receptor of TARC, in Jurkat T cells, and the ethanol extract of HcF weakly blocked the increased mRNA level. However, the stimulants and ethanol extract had no effect on the CCR4 protein level. The ethanol extract inhibited TARC-induced migration, as well as basal migration of Jurkat T cells. Conclusions This study may show the usefulness of HcFs in the ethnopharmacological treatment of Th2-mediated or allergic inflammation, through the down-regulation of the production of Th2 cytokines and TARC, as well as cell migration.

Induction of TARC Production by Lipopolysaccharide and Interleukin-4 in Nasal Fibroblasts

Background: Th2 cell infiltration is a characteristic feature of allergic chronic sinusitis. However, the mechanisms that cause the predominance of Th2 cells in this disease have yet to be clarified. The airway is often exposed to not only Th2 cytokines but also bacterial products such as lipopolysaccharides (LPS). A CC chemokine, TARC, is a potent chemoattractant for Th2 cells. The objectives of this study were twofold. First, we examined whether nasal polyp fibroblasts were able to produce TARC when costimulated with LPS and IL-4. Second, we investigated whether there was any heterogeneity in TARC production among fibroblasts derived from different airway sites. Methods: Fibroblast lines were established from human biopsy tissue. The amount of TARC in the supernatants was measured by ELISA. The expression of TARC mRNA was quantitated by real-time PCR. Results: Combined stimulation with LPS and IL-4 significantly induced TARC production by nasal polyp fibroblasts in a dose- and time-dependent manner. This induction occurred in normal nasal fibroblasts, but not in normal lung fibroblasts. Conclusions: Via TARC production, nasal fibroblasts may play an important role in the recruitment of Th2 cells into the sinus mucosa as well as nasal polyps. The heterogeneity in TARC production may reflect functional differences between upper and lower airway fibroblasts.

Discovery of Biomarkers for Hodgkin Lymphoma Using Proteomics Technology.

Hodgkin lymphoma (HL) is characterized by a minority of neoplastic cells, the Hodgkin and Reed-Sternberg cells (HRS cells) and an inflammatory background. Abnormal cytokine and chemokine production may play an important role in the proliferation of HRS cells, the reactive background formation and the impaired immune response encountered in patients with HL. To detect new proteins that might be involved in the interaction between the HRS cells and the inflammatory background cells and that also might serve as useful biomarkers for HL, we used different proteomic techniques to study the proteome of HRS cells.Methods: The first step is to determine the secretome of HRS cells in vitro. Therefore, the cells needed to be cultured without serum. Hodgkin cell lines L428 and L1236 were washed extensively to deplete serum proteins and cultured in RPMI media without serum for 6–72 hours. The supernatant was collected, concentrated, and analyzed by Surface-Enhanced Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (SELDI-TOF-MS) to determine the protein composition and as a quality check. After culture process optimization, sample fractionation was carried out by SDS-PAGE. The whole lane was excised in 30–35 bands and trypsine digestion was performed on every band. Peptide mixtures were analyzed by LC-MS/MS for identification of the proteins.Results: We tested several culture approaches for the Hodgkin lymphoma cell lines for proteomics purpose. Serum-free media were found to be inappropriate because of the abundant presence of several proteins. We next tested efficiency of washing steps and culture characteristics of 5 Hodgkin lymphoma cell lines in RPMI without serum. Cell viability, TARC and MDC levels in the culture supernatant at different time points and also SELDI-TOF-MS spectra indicated that culturing for 6–24 hours gave the best results. For protein identification we started with analysis of small proteins from supernatant of the L428 cell line, which has the highest TARC production. Gel bands in the region between 6–14 kDa were isolated from SDS-PAGE and treated for protein identification. This revealed a total of 64 proteins identified with ≥95% confidence. Application of SecretomeP2.0 (Protein Eng.Des.Sel., 2004. TMHMM 2.0) software indicated the presence of 28 secreted proteins including TARC. A similar approach was used for the L1236 Hodgkin lymphoma cell line, but the whole mass range (3–188 kDa) was analyzed in this case. We identified a total of 215 different proteins, out of which 85 are secreted (24 proteins follow a classical pathway while the other 61 follow a non-classical pathway). The non-secreted proteins were localized to the ER lumen, mitochondrion, nucleus, and membrane. These proteins are probably derived from cells breaking up during culture or centrifugation.Conclusion: In this study we identified 85 secreted proteins in L1236 supernatant, and 28 secreted proteins in the low molecular mass range (6–14kDa) in L428 supernatant. Some of the secreted proteins, including TARC, macrophage migration inhibitory factor (MIF), CD44 and Activated Leukocyte Cell Adhesion Molecule (ALCAM), are known to be involved in inflammatory process, lymphocytes homing or cell adhesion. The most consistently observed proteins will be tested for potential use as biomarkers in the sera of HL patients.

Cytokines regulate the expression of the thymus and activation‐regulated chemokine (TARC; CCL17) in human skin fibroblast cells

Abstracts Allergic inflammation is thought to be a Th2 cell‐dominant immune response during which tissue‐resident fibroblasts produce chemokines which contribute to the recruitment of migratory leukocytes to sites of tissue injury. Thymus and activation‐regulated chemokine (TARC; CCL17) is a potent member of the CC chemokine family and a selective chemoattractant for Th2 cells. In order to study the regulatory profiles of TARC production by TNF‐α, IFN‐γ, and IL‐4 in human normal skin fibroblast, CCD‐986sk cell line was used. The expression of TARC protein was measured using ELISA, and mRNA level was detected by RT‐PCR. The combination of TNF‐α and IL‐4 induced a time‐ and dose‐dependent synergistic increase in the expression of TARC at both protein and mRNA levels in the cultured human skin fibroblasts. Exposure of the cells to single cytokine had no effect on TARC expression. The high concentration (100 ng/ml) and long incubation time (72 h) of IFN‐γ further enhanced the TARC production induced by TNF‐α/...

Effects of Corticosteroid on the Expression of Thymus and Activation-Regulated Chemokine in a Murine Model of Allergic Asthma

Background: Thymus and activation-regulated chemokine (TARC; CCL17) is a lymphocyte-directed CC chemokine that specifically attracts T-helper (Th) 2 cells positive for the CC chemokine receptor 4 (CCR4⁺). Corticosteroids reduce airway inflammation, as reflected by reduced numbers of eosinophils and T cells and reduced expression of cytokines. We investigated TARC production and the inhibitory effects of corticosteroids on TARC expression in a murine model of allergic asthma. Methods: BALB/c mice were sensitized by intraperitoneal injection of ovalbumin (OVA) with alum. Once daily for 1 week, mice received injections of dexamethasone or 0.2 ml saline (control), then 1 h later inhaled aerosolized 1% OVA for 30 min. Mice were killed 24 h after OVA challenge for bronchoalveolar lavage and lung tissue examination. Results: TARC was expressed mainly in the bronchial epithelial cells. Dexamethasone attenuated OVA-induced airway eosinophilia, lymphocyte infiltration, and airway hyperresponsiveness. Dexamethasone also decreased TARC production in the bronchoalveolar lavage fluid and decreased expression of TARC mRNA and TARC protein in lung tissue. Conclusions: The corticosteroid dexamethasone inhibits TARC production in a murine model of allergic asthma in vivo. The beneficial effect of corticosteroids in bronchial asthma is due in part to their direct inhibitory effects on TARC production.

The Effect of Suplatast Tosilate on TARC Production in Peripheral Blood Mononuclear Cells and TARC Plasma Levels

Thymus and activation-regulated chemokine (TARC/CCL17) is a highly specific ligand for CCR 4. TARC may contribute to the recruitment, activation, and development of Th2 polarized cells that express CCR4. These characteristics have led investigators to hypothesize that TARC is involved in the development of Th2 responses. Suplatast tosilate ((±)-[2-[4-(3-ethoxy-2-hydroxy-propoxy) phenylcarbamoyl] ethyl] dimethylsulfonium p-toluenesulfonate) is an anti-allergic agent that selectively suppresses the synthesis of Th2 cytokines. We examined the effect of suplatast tosilate on TARC production and CCR-4 expression in vitro . Furthermore, we attempted to clarify whether TARC production was suppressed after clinical administration of suplatast tosilate. Peripheral blood mononuclear cells (PBMCs) were isolated from patients with allergic rhinitis who tested positive to house dust. PBMCs were stimulated with mite antigen. TARC mRNA was detected by real time PCR. The amount of TARC was estimated using an ELISA kit. PBMCs expressing CCR-4 were sorted by flow cytometry. The plasma level of TARC was examined in patients with chronic allergic rhinitis before and after treatment with suplatast tosilate for 4 weeks. Suplatast tosilate significantly reduced TARC production by PBMCs. TARC mRNA was also suppressed in a concentration dependent manner. However, suplatast tosilate did not inhibit the expression of CCR-4 on PBMCs. The plasma level of TARC was significantly decreased in patients administered suplatast tosilate. Suplatast tosilate suppressed TARC production by PBMCs and decreased the plasma level of TARC in patients with chronic allergic rhinitis.

Interleukin‐4 and Dexamethasone Counterregulate Extracellular Matrix Remodelling and Phagocytosis in Type‐2 Macrophages

Alternatively activated macrophages (Mphi2) are induced by Th2 cytokines and by glucocorticoids (GC), and can be distinguished from classically activated effector macrophages (Mphi1) on the basis of their anti-inflammatory properties. In addition, Mphi2 are involved in Th2/Th1 skewing, enhance antigen uptake and processing and support tissue remodelling and healing. In order to elucidate the heterogeneity of Mphi2 population systematically, we analysed a number of genes involved in extracellular matrix (ECM) remodelling, inflammation and phagocytosis in Mphi2 populations generated with interleukin-4 (IL-4) or GC. Using real-time polymerase chain reaction, we demonstrated that the ECM component, tenascin-C, is stimulated by IL-4, whereas it is suppressed by dexamethasone. The ECM remodelling enzymes--MMP-1 and MMP-12--and tissue transglutaminase (TG) showed a similar regulation pattern. FXIIIa, another putative Mphi2-associated TG, was synergistically regulated by IL-4 and GC. Enzyme-linked immunosorbent assay analysis revealed that the production of Mphi2-associated chemokines, AMAC-1, MCP-4 or TARC, was induced by IL-4 and was modulated by GC. Phagocytosis of opsonized and non-opsonized particles was stimulated by GC, whereas IL-4 had only a modulatory effect, what may be partially explained by the expression pattern of hMARCO, a scavenger receptor for non-opsonized particles, that was strongly and selectively induced by GC. In conclusion, stimulation of Mphi with IL-4 and GC regulate antagonistically the expression of ECM remodelling-related molecules and phagocytosis of opsonized and non-opsonized particles.

Thymus and activation-regulated chemokine (TARC/CCL17) produced by mouse epidermal Langerhans cells is upregulated by TNF-alpha and IL-4 and downregulated by IFN-gamma.

Thymus and activation-regulated chemokine (TARC/CCL17) is a Th2-type chemokine and its receptor CC chemokine receptor 4 (CCR4) is preferentially expressed on Th2 cells. Langerhans cells (LC) are immature dendritic cells (DC) in the epidermis of the skin and play vital roles in immune response. In this study, we investigated TARC expression by murine freshly isolated LC and 48 h cultured (mature) LC, and the regulation of TARC production in cultured LC by various cytokines. Murine LC was prepared using a panning method from BALB/c mice. RT-PCR was performed using fresh and cultured LC to evaluate TARC mRNA levels. ELISA was carried out using supernatant of cultured LC to calculate secreted TARC protein levels. TARC mRNA was strongly upregulated during maturation of murine LC. TARC production by murine LC was upregulated by TNF-alpha and IL-4 and downregulated by IFN-gamma, dose-dependently. Th1 and Th2 cytokines reciprocally regulate the production of Th2-type chemokine TARC by murine LC. Th2 cytokine microenvironments in skin may increase TARC production by mature LC, providing attraction of Th2 cells in skin. This may be an amplification circuit in Th2-dominant inflammatory skin disease like atopic dermatitis.