Thymus And Activation-regulated Chemokine Production Research Articles

Poly(I:C) synergizes with Th2 cytokines to induce TARC/CCL17 in middle ear fibroblasts established from mucosa of otitis media with effusion

Conclusion: These results suggest that middle ear fibroblasts contribute to the recruitment of Th2 cells into the middle ear by producing thymus and activation-regulated chemokine (TARC). Objectives: Intractable otitis media is more common in atopic subjects and asthmatics than in the otherwise normal population. Although type 2 T helper (Th2) cytokines play crucial roles in the middle ear of these populations, the mechanism underlying the predominance of Th2 cytokines has yet to be clarified. TARC has been known to facilitate recruitment of Th2 polarized cells, resulting in high levels of Th2 cytokines in the middle ear. We investigated whether middle ear-derived fibroblasts produce TARC when stimulated with poly(I:C) and Th2 cytokines (IL-4, IL-13). Materials and methods: Fibroblast lines were established from middle ear mucosa. TARC mRNA expression was evaluated by real-time RT-PCR. The amount of TARC in the culture supernatants was measured by ELISA. Results: Poly(I:C) induced only TARC gene expression in middle ear-derived fibroblasts. Combined stimulation with poly(I:C) and Th2 cytokine (IL-4, IL-13) synergistically induced TARC production by the cultured middle ear-derived fibroblasts. This response was dose and time dependent.

Combined Stimulation of Nasal Polyp Fibroblasts With Poly IC, Interleukin 4, and Tumor Necrosis Factor α Potently Induces Production of Thymus- and Activation-Regulated Chemokine

To examine the effects of cytokines and poly IC on the expression of thymus- and activation-regulated chemokine (TARC), a potent chemoattractant for helper T-cell type 2 (T(H)2) cells, in nasal polyp fibroblasts. Quantitative reverse transcription-polymerase chain reaction (RT-PCR) analysis. Academic research. Primary fibroblast lines were established from human nasal polyp biopsy tissue specimens (n = 5) removed at polypectomy. The expression of TARC messenger RNA (mRNA) was evaluated by real-time RT-PCR. The amount of TARC in the supernatants was measured by enzyme-linked immunosorbent assay. Combined stimulation with interleukin 4 (IL-4) and tumor necrosis factor alpha (TNF-alpha) or with poly IC and IL-4 induced TARC production. Combined exposure of cells to poly IC, IL-4, and TNF-alpha resulted in substantial amounts of TARC release into the culture medium. Quantitative RT-PCR analysis revealed that simultaneous stimulation with those 3 compounds induced a tremendous increase in the amount of TARC mRNA in the nasal polyp fibroblasts. Nasal polyp fibroblasts contribute to T(H)2 cell infiltration and RNA virus-induced exacerbation of T(H)2-type airway inflammatory conditions such as allergic chronic sinusitis.

Toll-like receptor 2, 3, 4, 5 ligands and interleukin-4 synergistically induce TARC production in nasal polyp fibroblasts

Objective Although type 2 T helper (Th2) cytokines such as IL-4 and IL-5 play a crucial role in the pathogenesis of chronic sinusitis with allergy, the mechanism underlying the predominance of Th2 cytokines has yet to be clarified. Thymus and activation-regulated chemokine (TARC) has been known to facilitate the recruitment of Th2 polarized cells, resulting in high levels of Th2 cytokines in the sinus mucosa as well as nasal polyps. The nasal and sinus cavities are ideal sites for studying the interplay between microbial Toll-like receptor (TLR) ligands and chemokines. We investigated whether nasal polyp fibroblasts produce TARC when stimulated with the breakdown products of microorganisms (TLR ligands) and a Th2 cytokine (IL-4). Methods Fibroblast lines were established from nasal polyp tissues. The expression of TARC mRNA was evaluated by real-time RT-PCR. The amount of TARC in the supernatants was measured by ELISA. Results Combined stimulation with TLR 2, 3, 4, 5 ligands and IL-4 induced TARC gene expression and protein production in the cultured nasal polyp fibroblasts. This response was time-dependent. Conclusions These results suggest that nasal polyp fibroblasts contribute to innate immunity and may play an important role in the recruitment of Th2 cells into nasal polyps through the production of TARC.

Inhibition of Dermatitis Development by Sopungsan in Nc/Nga Mice.

Sopungsan (SS) is a traditional Korean decoction used for the treatment of dermatitis. The aim of this study is to confirm whether or not SS has a preventive effect on the development of atopic dermatitis in dinitrochlorobenzene-applied Nc/Nga mice. SS was administered orally to Nc/Nga mice, which led to the remarkable suppression of the development of dermatitis, as determined by a histological examination and the serum IgE levels. Moreover, SS inhibited the production of thymus- and activation-regulated chemokine (TARC) and its mRNA expression in a keratinocyte cell line, HaCaT, which had been stimulated with tumor necrosis factor-α (TNF-α) and interferon-γ (IFN-γ). Activation of the nuclear factor-κB (NF-κB) or activator protein-1 (AP-1) is one of key steps in the signaling pathways mediating induction of TARC. In this study, SS selectively suppressed NF-κB activation which may be essential for TARC expression in TNF-α/IFN-γ treated keratinocytes. The inhibitory effect of SS on NF-κB activation and TARC production might be associated with the anti-dermatitic effects of SS.

Circulating levels of both Th1 and Th2 chemokines are elevated in patients with sarcoidosis

In sarcoidosis, the T helper type 1 (Th1) response tends to predominate at affected disease sites; however, whether Th1/Th2 polarization occurs in the peripheral circulation is unknown. Fifty-two patients with sarcoidosis and 21 healthy volunteers were investigated. The concentrations of interferon-inducible protein 10 (IP-10)/CXCL10 and thymus- and activation-regulated chemokine (TARC)/CCL17 in the serum, bronchoalveolar lavage fluid (BALF) and culture supernatant were measured by an enzyme-linked immunosorbent assay. The circulating CXCR3+ CD4+ T cells and CCR4+ CD4+ T cells were assessed by flow cytometry. The CXCR3- or CCR4-positive ratios among CD4+ T cells were both higher in sarcoidosis than in healthy volunteers. The serum levels of both IP-10 and TARC of the patients with sarcoidosis were significantly higher than those of the healthy volunteers. In patients with sarcoidosis, a larger amount of IP-10 was generated by the BALF cells, whereas IP-10 production by peripheral blood mononuclear cells did not increase in comparison to the control subjects. The TARC levels produced by peripheral blood mononuclear cells of sarcoidosis patients were significantly higher than those of the controls, while no difference existed between the 2 groups regarding TARC production by BALF cells. IP-10 is mainly produced at the lung and TARC in the peripheral circulation in sarcoidosis patients. Both IP-10 and TARC cooperatively play a role in the pathogenesis of sarcoidosis.

Respiratory Syncytial Virus Synergizes with Th2 Cytokines to Induce Optimal Levels of TARC/CCL17

Respiratory syncytial virus (RSV) is a ubiquitous virus that preferentially infects airway epithelial cells, causing asthma exacerbations and severe disease in immunocompromised hosts. Acute RSV infection induces inflammation in the lung. Thymus- and activation-regulated chemokine (TARC) recruits Th2 cells to sites of inflammation. We found that acute RSV infection of BALB/c mice increased TARC production in the lung. Immunization of BALB/c mice with individual RSV proteins can lead to the development of Th1- or Th2-biased T cell responses in the lung after RSV infection. We primed animals with a recombinant vaccinia virus expressing either the RSV fusion (F) protein or the RSV attachment (G) protein, inducing Th1- and Th2-biased pulmonary memory T cell responses, respectively. After RSV infection, TARC production significantly increased in the vaccinia virus G-primed animals only. These data suggest a positive feedback loop for TARC production between RSV infection and Th2 cytokines. RSV-infected lung epithelial cells cultured with IL-4 or IL-13 demonstrated a marked increase in the production of TARC. The synergistic effect of RSV and IL-4/IL-13 on TARC production reflected differential induction of NF kappa B and STAT6 by the two stimuli (both are in the TARC promoter). These findings demonstrate that RSV induces a chemokine TARC that has the potential to recruit Th2 cells to the lung.

Down-Regulation of E-Cadherin in Human Bronchial Epithelial Cells Leads to Epidermal Growth Factor Receptor-Dependent Th2 Cell-Promoting Activity

Airway epithelial cells are well-known producers of thymus- and activation-regulated chemokine (TARC), a Th2 cell-attracting chemokine that may play an important role in the development of allergic airway inflammation. However, the mechanism responsible for up-regulation of TARC in allergy is still unknown. In the asthmatic airways, loss of expression of the cell-cell contact molecule E-cadherin and reduced epithelial barrier function has been observed, which may be the result of an inadequate repair response. Because E-cadherin also suppressed multiple signaling pathways, we studied whether disruption of E-cadherin-mediated cell contact may contribute to increased proallergic activity of epithelial cells, e.g., production of the chemokine TARC. We down-regulated E-cadherin in bronchial epithelial cells by small interference RNA and studied effects on electrical resistance, signaling pathways, and TARC expression (by electric cell-substrate impedance sensing, immunodetection, immunofluorescent staining, and real-time PCR). Small interference RNA silencing of E-cadherin resulted in loss of E-cadherin-mediated junctions, enhanced phosphorylation of epidermal growth factor receptor (EGFR), and the downstream targets MEK/ERK-1/2 and p38 MAPK, finally resulting in up-regulation of TARC as well as thymic stromal lymphopoietin expression. The use of specific inhibitors revealed that the effect on TARC is mediated by EGFR-dependent activation of the MAPK pathways. In contrast to TARC, expression of the Th1/Treg cell-attracting chemokine RANTES was unaffected by E-cadherin down-regulation. In summary, we show that loss of E-cadherin-mediated epithelial cell-cell contact by damaging stimuli, e.g., allergens, may result in reduced suppression of EGFR-dependent signaling pathways and subsequent induction of Th2 cell-attracting molecule TARC. Thus, disruption of intercellular epithelial contacts may specifically promote Th2 cell recruitment in allergic asthma.

Sopungyangjae‐Tang Inhibits Development of Dermatitis in Nc/Nga Mice

Sopungyangjae-Tang (SYT) is a traditional Korean decoction used for the treatment of dermatitis. The aim of this study was to confirm whether or not SYT has a preventive effect on the development of atopic dermatitis in dinitrochlorobenzene-applied Nc/Nga mice. SYT was administered orally to Nc/Nga mice, which led to the remarkable suppression of the development of dermatitis, as determined by a histological examination and the serum IgE levels. Moreover, SYT inhibited the production of thymus- and activation-regulated chemokine (TARC) and its mRNA expression in a keratinocyte cell line, HaCaT, which had been stimulated with tumor necrosis factor-α (TNF-α) and interferon-γ (IFN-γ). Activation of the nuclear factor-κB (NF-κB) or activator protein-1 (AP-1) is one of the key steps in the signaling pathways mediating induction of TARC. In this study, SYT selectively suppressed NF-κB activation, which may be essential for TARC expression in TNF-α/IFN-γ treated keratinocytes. The inhibitory effect of SYT on NF-κB activation and TARC production might be associated with the anti-dermatitic effects of SYT.

Der p, IL-4, and TGF-β Cooperatively Induce EGFR-Dependent TARC Expression in Airway Epithelium

Thymus and Activation-Regulated Chemokine (TARC) may be critical in Th2 cell recruitment in allergic inflammation; however, the mechanisms of allergen-induced TARC release are unclear. Since airway epithelium is the first line of defense to inhaled allergens, we questioned whether house dust mite allergen (Der p) can induce TARC expression in bronchial epithelial cells, how this is regulated at the molecular level, and if micro-environmental cytokines augment this effect. We examined the effects of Der p and the cytokines IL-4 and TGF-beta on TARC expression in 16HBE cells and primary bronchial asthma epithelium. Real-time PCR and immunofluorescence demonstrated that Der p induces TARC expression in bronchial epithelium. Supernatants from Der p-stimulated 16HBE cells were able to induce TARC-dependent T cell trafficking. IL-4 and TGF-beta cooperatively enhanced Der p-induced TARC expression in 16HBE cells. Specific inhibitors, immunodetection, and gel-shifts revealed that these effects are mediated by phosphorylation of the epidermal growth factor receptor (EGFR), mitogen-activated protein kinase (MAPK) signaling and subsequent nuclear factor (NF)-kappaB activation. A Disintegrin And Metalloproteinase (ADAM), a family of proteins involved in shedding of various growth factors, was shown to be responsible for EGFR activation. The increase in TARC production by direct interaction of Der p with the bronchial epithelium may be an important initial step in the generation of allergic inflammation, which is further potentiated by micro-environmental cytokines. Interference with ADAM or EGFR activity may be a novel promising target to prevent TARC release and subsequent allergic inflammation.

Inhibition of T helper 2 chemokine production by narrowband ultraviolet B in cultured keratinocytes

Narrowband ultraviolet B (NB-UVB) has recently been used for the treatment of various skin disorders. Its effects on the production of cytokines and chemokines by keratinocytes are unknown. To investigate the effect of NB-UVB on production of chemokines and proinflammatory cytokines by keratinocytes in comparison with broadband (BB)-UVB. Normal human epidermal keratinocytes (or the human keratinocyte cell line HaCaT in some experiments) at semiconfluency were irradiated with NB-UVB at 10, 100, 500 or 1000 mJ cm(-2) or BB-UVB at 10 or 100 mJ cm(-2). The cultures were maintained in the presence or absence of interferon (IFN)-gamma at 200 U mL(-1). The 72-h culture supernatants were analysed by enzyme-linked immunosorbent assay to quantify T helper (Th)1 chemokines (IFN-inducible protein 10 and monokine induced by IFN-gamma), Th2 chemokines [macrophage-derived chemokine (MDC) and thymus and activation-regulated chemokine (TARC)] and proinflammatory cytokines [interleukin (IL)-1alpha and tumour necrosis factor (TNF)-alpha]. The expression of mRNA for these molecules was simultaneously assessed by reverse transcriptase-polymerase chain reaction. The culture supernatants were also tested for their chemotactic activity for Th1 and Th2 cells. The two UVB sources were compared on the basis of their minimal erythemal doses and clinically used doses. Although both NB-UVB and BB-UVB increased the production of IL-1alpha and TNF-alpha, the augmentative effect of NB-UVB was less than that of BB-UVB. Both wavelength ranges of UVB enhanced or had no effect on Th1 chemokine production, but suppressed the production of Th2 chemokines MDC and TARC. This was confirmed by chemotactic assay, which showed decreased chemotactic activity for Th2 cells by the culture supernatants from NB-UVB-irradiated keratinocytes. NB-UVB reduces the production of Th2 chemokines without excess production of proinflammatory cytokines, suggesting its therapeutic effectiveness on Th2-mediated skin disorders as well as its relative safety in clinical usage.

TARC and sensitive skin

TARC and sensitive skin

Cultured human endometrial epithelial cells produce thymus and activation-regulated chemokine with stimulation of interleukin-4 and interleukin-13

To evaluate the effects of T-helper (Th)1 and Th2 cytokines on the production of thymus and activation-regulated chemokine (TARC) and macrophage-derived chemokine (MDC) by cultured endometrial epithelial cells (EEC) and endometrial stromal cells (ESC). The effects of interleukin (IL)-2, IL-4, IL-5, IL-10, IL-12, IL-13, interferon-gamma (IFN-gamma), and tumor necrosis factor-beta (TNF-beta) on the production of TARC and MDC were investigated. Research laboratory at a medical school. Fifteen endometrial specimens in the mid-late secretory phase were used. The EEC and ESC were incubated for 24 hours with recombinant human IL-2, IL-4, IL-5, IL-10, IL-12, IL-13, IFN-gamma, and TNF-beta. The concentrations of TARC and MDC in the culture media were measured using ELISA. Small amounts of TARC were detected in the culture medium of nonstimulated EEC. The increase in levels of TARC in the culture media of EEC paralleled the addition of increasing amounts of IL-4 and IL-13. Other cytokines, however, did not affect the production of TARC by EEC. Production of TARC by ESC was not detected under either nonstimulated or cytokine-stimulated conditions. Production of MDC was not detected in the culture media of EEC and ESC. These results suggest that IL-4 and IL-13 secreted from the embryo during the implantation period may selectively up-regulate the production of TARC by EEC. The controlled production of TARC in the endometrium may contribute to the modulation of the immune reaction by the regulation of Th2 lymphocyte trafficking and functions.

17β-Estradiol (E2) modulates cytokine and chemokine expression in human monocyte-derived dendritic cells

AbstractThe effects of estrogen on the immune system are still largely unknown. We have investigated the effect of 17β-estradiol (E2) on human monocyte-derived immature dendritic cells (iDCs). Short-term culture in E2had no effect on iDC survival or the expression of cell surface markers. However, E2treatment significantly increased the secretion of interleukin 6 (IL-6) in iDCs and also increased secretion of osteoprotegerin (OPG) by DCs. Furthermore, E2significantly increased secretion of the inflammatory chemokines IL-8 and monocyte chemoattractant protein 1 (MCP-1) by iDCs, but not the production of the constitutive chemokines thymus and activation-regulated chemokine (TARC) and macrophage-derived chemokine (MDC). However, after E2pretreatment the lipopolysaccharide (LPS)–induced production of MCP-1, TARC, and MDC by DCs was clearly enhanced. Moreover, mature DCs pretreated with E2stimulated T cells better than control cells. Finally, we found that E2provides an essential signal for migration of mature DCs toward CCL19/macrophage inflammatory protein 3β (MIP3β). In summary, E2may affect DC regulation of T-cell and B-cell responses, as well as help to sustain inflammatory responses. This may explain, in part, the reason serum levels of estrogen correlate with the severity of certain autoimmune diseases.

Enhanced TARC production by dust-mite allergens and its modulation by immunosuppressive drugs in PBMCs from patients with atopic dermatitis

Background : Thymus and activation regulated chemokine (TARC) is a CC chemokine that attracts CCR4 + T cells. We reported previously that TARC is an important chemokine that defines Th2 imbalance in the pathogenesis of atopic dermatitis (AD). Objectives : This study was undertaken to clarify TARC producing cells in peripheral blood mononuclear cells (PBMCs), the regulation of dust mite-allergen clude extract (DME) and different immunosuppressive drugs (Tacrolimus (FK506), cyclosporine (CsA), dexamethasone (Dex)) on TARC production by peripheral PBMCs from AD patients in vitro. Methods : Monocyte derived dendritic cells (MoDCs) were generated from and TARC mRNA levels were examined and comapared with those from T cells in PBMCs from AD patients. PBMCs were cultured with or without DME and/or immunosuppressive drugs (Tacrolimus, CsA, Dex) for 7 days and TARC levels were measured. Results : PBMCs from AD patients which were cultured with DME stimulation for 7 days showed significantly higher levels of TARC production than those from healthy controls. RT–PCR demonstrated that TARC mRNA was expressed in CD4 + T cells, CD8 + T cells and MoDCs. Tacrolimus, CsA and Dex individually suppressed TARC production by PBMCs from AD patients which were co-cultured with DME for 7 days. Gel shift analysis revealed differential inhibitory effects of these immunosuppressive drugs on NFκB activity in PBMCs from AD patients. Conclusion : Our data demonstrate that TARC producing cells are MoDCs, T cells as well as epidermal keratinocytes in AD. We suggest that MoDCs might regulate the immune responses by attracting T cells and CD25 + T cells in the pathogenesis of AD. We also showed the important role of DME on TARC production and the inhibitory effect of the immunosuppressive drugs on TARC production by PBMCs from AD patients, that can regulate ongoing immune responses in the pathogenesis of AD.

Suppressive Activity of Fexofenadine Hydrochloride on Thymus- and Activation-Regulated Chemokine Production from Human Peripheral Blood Leukocytes in Response to Antigenic Stimulation in vitro

Background: Thymus- and activation-regulated chemokine (TARC) is accepted as being an important molecule in the development and maintenance of allergic diseases. However, there is little information about the influence of antiallergic agents on TARC production after allergen challenge. The aim of this study is to examine the influence of fexofenadine hydrochloride (FEX), an H₁-receptor antagonist, on TARC production from human peripheral blood leukocytes (PBL) using an in vitro cell culture technique. Methods: PBL prepared from donors with pollinosis were cultured with either Japanese cedar pollen allergen, Cry j 1, or interleukin (IL)-4 in the presence of various doses of FEX for 6 days. Levels of TARC and the T cell cytokines IL-4 and interferon (IFN)-γ in culture supernatants were examined by ELISA. Results: FEX did not affect PBL proliferation induced by Cry j 1 stimulation, even when 500 ng/ml of the agent, twice the therapeutic blood levels, was added to cell cultures as assessed by measuring ³H-thymidine incorporation into DNA. On the other hand, FEX at 250 ng/ml (but not 125 ng/ml), similar to therapeutic blood levels, significantly inhibited the ability of PBL to produce IL-4 (but not IFN-γ), which was enhanced by Cry j 1 stimulation. FEX at concentrations of more than 250 ng/ml also exerted a suppressive effect on TARC production from PBL in response to Cry j 1 and IL-4 stimulation in vitro. Conclusion: This inhibitory action of FEX may be partially responsible for the attenuating effect of the agent on allergic diseases.

Lack of evidence for TARC/CCL17 production by normal human keratinocytes in vitro

Background: thymus and activation-regulated chemokine (TARC)/CCL17 is a CC chemokine that selectively attracts Th2-type lymphocytes. Immunohistochemical analyses have revealed that TARC is expressed in the epidermal keratinocytes of atopic dermatitis (AD), suggesting TARC involvement in the pathogenesis of the disease. However, keratinocyte TARC production has been described only in the transformed keratinocyte cell line HaCaT. Objective: to examine TARC production in normal human epidermal keratinocytes (NHEK) in vitro. Methods: the expression of TARC mRNA and protein were examined in NHEK and HaCaT cells stimulated with various cytokines. Results: stimulation with inflammatory cytokines, including interleukin (IL)-1, IL-4, IL-6, IL-10, interferon (IFN)-α, IFN-β, IFN-γ, and tumor necrosis factor (TNF)-α failed to induce TARC mRNA expression in NHEK. However, stimulation with IFN-γ and TNF-α together enhanced expression slightly. ELISA analysis failed to detect TARC protein in NHEK culture supernatant, even following stimulation with IFN-γ and TNF-α. In contrast, HaCaT cells produced TARC protein even without stimulation of cytokines. Conclusion: these results indicate that production of TARC by HaCaT cells is a phenomenon specific to the cell line and the observation on TARC in HaCaT cells can not be generalized. NHEK do not produce TARC protein in vitro.

Production of TARC and MDC by naive T cells in asthmatic patients.

The helper (Th)2 cell-attracting chemokines thymus and activation-regulated chemokine (TARC) and macrophage-derived chemokine (MDC) are ligands for the chemokine receptor CCR4. A number of cellular sources of TARC and MDC have been identified, including not only macrophages, dendritic cells, and natural killer cells, but also bronchial epithelial cells. Recent studies report that TARC and MDC may serve as pivotal chemokines for the development of Th2-dominated experimental allergen-induced asthma. This study was designed to assess TARC and MDC production by CD4+ T cells, including naive T cells and memory/effector T cells, purified from peripheral blood mononuclear cells in patients with asthma. Asthmatic subjects included in this study had mild asthmatic symptoms, positive skin test responses to house dust mite allergen, and elevated level of Dermatophagoides farinae immunoglobulin E in the sera. CD4+ T cells--CD45RA+ CD4+ T cells--as naive T cells and CD45RO+ CD4+ T cells--as memory/effector T cells--were purified by negative selection from peripheral blood mononuclear cells obtained from asthmatic patients (n = 6) and healthy controls (n = 6). These cells and established Th1/Th2 cell lines were then cultured in the presence of both anti-CD3 and -CD28 antibodies. After 48 hr of incubation, concentrations of TARC, MDC, interleukin (IL)-4, IL-5, and interferon-gamma in the supernatants were measured by enzyme-linked immunoadsorbent assay. Reverse transcriptase-polymerase chain reaction was performed to analyze mRNA expression of TARC and MDC. Our results clearly showed that TARC and MDC were produced by activated CD45RA+ CD4+ T cells rather than by activated CD45RO+ CD4+ T cells, and the levels of these chemokines in the asthmatic patients were higher than those in the healthy controls. Furthermore, these chemokines production by Th2 cell lines were greater than those by Th1 cell lines, but the level were smaller than those by naive T cells. Our studies suggest that TARC and MDC are produced by naive T cells rather than by memory/effector T cells, including Th2 cells, in asthmatic patients, and these chemokines were produced at modest levels in any T-cell populations from healthy controls. Taken together, naive T cells in asthma have a peculiar function to produce TRAC and MDC, which contribute to local migration of Th2 cells into lung and lymphoid tissues, along with a function as precursor for memory/effector T cell. This novel function of naive T cells may be implicated in the development of asthma.

Suplatast tosilate inhibits thymus- and activation-regulated chemokine production by antigen-specific human Th2 cells.

Suplatast tosilate is an anti-allergic agent that suppresses cytokine production by human Th2 cells. We investigated the effects of suplatast tosilate on the production of thymus- and activation-regulated chemokine (TARC) by T cells from allergic patients with asthma. Purified protein derivative (PPD)-specific Th1 cell lines and Dermatophagoides farinae (Der f)-specific Th2 cell lines were established from nine patients with house dust mite-allergic asthma. The effects of suplatast tosilate on mRNA expression of TARC and protein production of TARC from antigen-specific Th1 or Th2 cell lines were investigated after stimulation with relevant antigens or phytohemagglutinin (PHA). In addition, the effects of IL-4, IL-10, and IFN-gamma on TARC production by Der f-specific Th2 cell lines in the presence or absence of suplatast tosilate were studied. Although PPD-specific Th1 cell lines did not produce TARC after stimulation with PPD antigen or PHA, stimulation of Der f-specific Th2 cell lines with Der f antigen or PHA increased production of TARC. Suplatast tosilate significantly and dose-dependently inhibited production of TARC by Der f-specific Th2 cell lines stimulated with either Der f antigen (76.5% inhibition at 100 microg/mL, P < 0.01) or PHA (81.9% inhibition at 100 microg/mL, P < 0.01). TARC production by Der f-specific Th2 cell lines was significantly increased only by activation with IL-4 but not with IL-10 or IFN-gamma; this increase in TARC production was significantly inhibited by suplatast tosilate (97.5% inhibition at 100 microg/mL, P < 0.01). Suplatast tosilate inhibits TARC production by human Th2 cells. Therefore, this agent inhibits both Th2 cytokine and Th2 chemokine and may be a useful anti-allergic agent.

Synergistic Effect of TNF-α and IL-4 on the Expression of Thymus- and Activation-Regulated Chemokine in Human Corneal Fibroblasts

Chemokine production by resident fibroblasts contributes to the recruitment of migratory leukocytes to sites of tissue injury. The effects of cytokines on the expression of thymus- and activation-regulated chemokine (TARC), a potent chemoattractant for Th2 cells, were examined in cultured human corneal fibroblasts. The culture supernatants of cells incubated for 24 h in the absence or presence of either TNF-α (10 ng/ml) or IL-4 (10 ng/ml) alone did not contain TARC detectable by ELISA. However, exposure of cells to both cytokines resulted in marked release of TARC into the culture medium in a time- and dose-dependent manner. Similarly, quantitative RT-PCR analysis revealed that the two cytokines induced a synergistic increase in the amount of TARC mRNA in the cultured fibroblasts. This synergistic effect of TNF-α and IL-4 on TARC production by fibroblasts may contribute to the Th2 cell infiltration and consequent tissue damage associated with allergic inflammation.

A Th2 Chemokine, TARC, Produced by Keratinocytes May Recruit CLA+CCR4+ Lymphocytes into Lesional Atopic Dermatitis Skin

Atopic dermatitis is an inflammatory skin disease in which the inflammation is characterized by the influx of lymphocytes into the dermis. It is generally believed that atopic dermatitis is a Th2-type disease, i.e., the T lymphocytes produce interleukin-4, interleukin-5, interleukin-10, and interleukin-13, although it has become evident in recent years that the cytokine profile in the skin changes during the course of the disease towards a Th1-Th2 mixed cytokine profile (interferon-gamma, tumor necrosis factor alpha, and interleukin-2). The lymphocytes that home into the skin express cutaneous lymphocyte-associated antigen, and it has recently been shown that most of the lymphocytes in this population express the chemokine receptor CCR4. CCR4 is the receptor for the CC chemokine TARC (thymus and activation regulated chemokine), and this chemokine is expressed predominantly by keratinocytes in the basal layer of the epidermis of lesional atopic dermatitis skin in mice. In humans, however, it was shown to be expressed in the endothelial cells of the dermis. We have examined the peripheral blood mononuclear cells of atopic dermatitis patients for the expression of cutaneous lymphocyte-associated antigen and CCR4 and compared them with peripheral blood mononuclear cells from normal controls. We found that the proportion of CLA+CCR4+ lymphocytes is upregulated in atopic dermatitis patients. In addition we have examined skin biopsies of lesional and non-lesional skin from atopic dermatitis patients and found that the keratinocytes, but not the endothelial cells, produce TARC in the lesional but not in the nonlesional skin. To gain insight in the stimulatory mechanisms for TARC production in keratinocytes, as previously observed in mice, we cultured HaCaT cells and found that interferon-gamma and tumor necrosis factor alpha work synergistically to induce TARC production. These observations suggest that the induction of TARC production in keratinocytes plays an important role in the late phase skin invasion by CCR4+CLA+ Th2-type lymphocytes in atopic dermatitis.