Emedastine Inhibits Th1 and Th2 Cell Differentiation Mediated by Mast Cells.

We have previously clarified that emedastine, a second-generation antihistamine drug, inhibits T helper 1(Th1)/Th2 cell differentiation mediated by Langerhans cells (LCs). In addition, although we have recently found that mast cells also function as antigen-presenting cells (APCs) and induce Th1/Th2 cell differentiation, any influence of emedastine on this function remained unclear. Here we investigated the influence of emedastine on Th1/Th2 cell differentiation via mast cells. Mast cells were obtained by long-term culture of murine splenocytes in medium supplemented with tumor necrosis factor (TNF)-α. The mast cells were then incubated in the presence or absence of emedastine, and cultured with naïve CD4+ T cells in the presence of ovalbumin (OVA) peptide. Five days later, Th cells in the culture were stimulated with phorbol 12-myristate 13-acetate (PMA) and ionomycin, and Th1/Th2 cytokine production was examined by enzyme-linked immunosorbent assay. When mast cells treated with emedastine were used as APCs, production of interferon (IFN)-γ and interleukin (IL)-4 from activated Th cells was significantly suppressed. This suppression was associated with inhibition of CD86 expression on mast cells, and mast cells treated with emedastine were shown to obstruct the differentiation of both Th1 and Th2 cells by down-regulating their cell surface expression of CD86. The present data provide additional information that topical application of emedastine to the lesional skin of patients with atopic dermatitis (AD) would reduce not only LC- but also mast cell-mediated skin inflammation.

Combination Therapy with Betamethasone and Josamycin Demonstrates Superior Therapeutic Efficacy in an NC/Nga Mouse Model of Atopic Dermatitis.

We have previously demonstrated the excellent bactericidal activity of josamycin against Staphylococcus aureus isolated from patients with atopic dermatitis (AD), with therapeutic efficacy equal to that of betamethasone. The present study was designed to evaluate the effectiveness of combination therapy with betamethasone and josamycin for AD. Betametasone (0.1%) and josamycin (0.1%) were topically administered to NC/Nga mice with severe AD-like skin lesions. Skin severity scores, histological changes in skin lesions, and serum immunoglobulin E (IgE) levels were assessed as indicators of therapeutic efficacy. Topical treatment with both drugs suppressed the skin severity score to a greater degree than betamethasone alone. This was associated with a reduction of epidermal thickening, a reduced density of dermal cellular infiltration, a decreased mast cell count in the dermis, and a reduced serum IgE level. In addition, both drugs in combination markedly reduced the expression of interferon (IFN)-γ and interleukin (IL)-4 in auricular lymph node cells, as well as the S. aureus count on the lesioned skin. These results show that simultaneous topical application of both drugs can ameliorate severe AD-like skin lesions in NC/Nga mice. It is suggested that combination therapy with betamethasone and josamycin would be beneficial for control of severe AD lesions colonized by S. aureus by inhibiting the development of both T helper (Th) type 1 (Th1) and Th2 cells and also through elimination of superficially located S. aureus.

Effects of dual therapy with betamethasone and tetracycline in a NC/Nga mouse model of atopic dermatitis.

Purpose: Our previous study has demonstrated that tetracycline exerts excellent bactericidal activity against Staphylococcus aureus strains isolated from patients with atopic dermatitis (AD) while simultaneously inhibiting the development of T helper (Th) type 2 (Th2) cells. The present study was designed to evaluate the effectiveness of dual therapy with betamethasone and tetracycline for AD. Methods: Betametasone (0.1%) and tetracycline (3%) were topically administered to NC/Nga mice with AD-like skin lesions. Skin severity scores, histological changes to the lesioned skin, and serum IgE levels were assessed as indicators of therapeutic effectiveness. Results: Topical treatment with both drugs reduced the skin severity score more significantly than was the case with betamethasone alone or tetracycline alone. This was associated with a reduction in the degree of epidermal thickening, the density of cellular infiltration into the dermis, the mast cell count in the dermis and the serum IgE concentration. Furthermore, the degree of Th1/Th2 cell development in auricular lymph nodes and the S. aureus count on the lesioned skin were synergistically suppressed by simultaneous application of both drugs. Conclusion: The present results show that simultaneous topical application of betamethasone and tetracycline synergistically ameliorates AD-like skin lesions in NC/Nga mice. This suggests that dual therapy with betamethasone and tetracycline for AD lesions colonized by S. aureus might be one of the best options for inhibiting the development of both Th1 and Th2 cells and acting on superficially located S. aureus .

Influence of the Th1 Cytokine Environment on CCL5 Production from Langerhans Cells.

In the chronic skin lesions of atopic dermatitis (AD), T helper type 1 (Th1) cells appear in addition to Th2 cells, but the role played by Th1 cells in skin inflammation during the chronic phase remains unknown. Here we examined CCL5 production from Langerhans cells (LCs) in the Th1 cytokine environment. LCs were generated from mouse bone marrow cells, then stimulated with anti-CD40 antibody and the Th1 cytokine, interferon (IFN)-γ. Their CCL5 production was then measured. In addition, the LCs were incubated with naïve CD4+ T cells from a DO 11.10 TCR Tg mouse in the presence of ovalbumin peptide for 5 d, and their IFN-γ, interleukin-4 and CCL5 production was then measured. When LCs were stimulated with the anti-CD40 antibody in the presence of IFN-γ, significant levels of CCL5 production were confirmed. Furthermore, when LCs presented antigen to Th cells in the Th1 cytokine environment, significant levels of CCL5 production were induced. This CCL5 production was associated with IFN-γ activity and CD40L expression by Th cells in the culture. Our present data suggest that LCs augment CCL5 production by responding to IFN-γ while presenting antigen to Th cells, and that this augmentation of CCL5 production would likely contribute to infiltration of eosinophils and other Th1 cells into skin lesions, followed by expansion of chronic inflammation in the skin.

Effects of Josamycin on Scratching Behavior in NC/Nga Mice with Atopic Dermatitis-Like Skin Lesions.

Our previous study showed that chronic skin colonization by Staphylococcus aureus exacerbated atopic dermatitis (AD) and that control of such skin colonization using antibiotic ointment might relieve AD-related skin inflammation. However, the role of S. aureus colonization in the pruritus accompanying AD was not elucidated. The aim of the present study was to evaluate the effect of topically applied josamycin, a macrolide antibiotic, on the scratching behavior of NC/Nga mice with AD-like skin lesions. Josamycin (0.1%) was topically administered to NC/Nga mice with AD-like skin lesions induced by a mite antigen, Dermatophagoides farinae extract, and the therapeutic effects of josamycin were assessed by measurement of the skin severity score, S. aureus colonization, scratching count, and interleukin (IL)-31 mRNA expression in the skin lesions. Topical treatment with josamycin ointment significantly suppressed the increase of the skin severity score in NC/Nga mice. This suppressive effect was associated with decreases in the S. aureus count on the lesioned skin, scratching behavior of mice and IL-31 mRNA expression in the lesions. The present results show that the severity of AD-like skin inflammation in NC/Nga mice is correlated with the level of S. aureus colonization and subsequent IL-31 production in the skin. Therefore, topical application of josamycin to AD lesions colonized by S. aureus would be beneficial for control of AD by eliminating superficially located S. aureus and by suppressing the IL-31-induced scratching behavior.

Effects of anti-allergy drugs on Th1 cell and Th2 cell development mediated by Langerhans cells.

BACKGROUND: It is well known that Langerhans cells (LCs) work as the primary orchestrators in polarization towards T helper type 1 (Th1) or T helper type 2 (Th2) immune responses. In this study, we examined the effects of various anti-allergy drugs against the Th2 cell development by LCs. METHODS: The expression of cell surface molecules on LCs was investigated using reverse transcriptase polymerase chain reaction. The effects of anti-allergy drugs on T-cell immunoglobulin and mucin domain-containing protein (TIM)-4 expression in LCs were examined to predict whether they would inhibit Th2 cell development. Next, mice were primed via the hind footpad with ovalbumin (OVA)-pulsed LCs that had been treated with selected anti-allergy drugs. After 5 days, the cytokine response in the popliteal lymph nodes was investigated by enzyme-linked immunosorbent assay. The therapeutic effects of a selected drug on atopic dermatitis (AD) were assessed using AD-like skin lesions of NC/Nga mice. RESULTS: The first-generation histamine H1 receptorantagonists, cyproheptadine and promethazine, and the second-generation histamine H1 receptor antagonists, emedastine and loratadine, were selected as candidate inhibitors of Th2 cell development. As expected, OVA peptide-pulsed LCs that had been treated with each drug and injected into the hind footpads of mice inhibited Th2 cell development, as represented by down-regulation of interleukin (IL)-4 production. Furthermore, the LCs that had been treated withemedastine also inhibited Th1 cell development, as represented by down-regulation of interferon (IFN)-g production. This additional inhibition of Th1 cell development was accompanied by suppression of CD40 expression in LCs. Therefore, the therapeutic effect of emedastine on AD was examined. Topical application of emedastine significantly suppressed the increase in the skin severity score in NC/Nga mice with AD-like skin lesions. This suppressive effect was associated with a decrease in the production of IFN-g and IL-4 in auricular lymph node cells. CONCLUSIONS: These results suggest that topical application of emedastine to skin lesions of patients with AD may provide clinical benefits through the inhibition of both Th1 cell and Th2 cell development mediated by LCs.

Topical Application of Doxycycline Inhibits Th2 Cell Development Mediated by Langerhans Cells and Exerts a Therapeutic Effect on Atopic Dermatitis.

BACKGROUND: Langerhans cells (LCs) polarize the immune milieu towards a T helper type (Th) 1 or Th2 immune response. We investigated the effects of selected tetracyclines on Th cells development mediated by LCs, and their implications for the treatment of atopic dermatitis (AD). METHODS: Mice were primed with ovalbumin (OVA) peptide-pulsed LCs, which had been treated with each antibiotic, via the hind footpad. After 5 days, the Th1/Th2 cytokine response in the popliteal lymph nodes was investigated by enzyme-linked immunosorbent assay. The expression of cell surface molecules on LCs was investigated using reverse transcriptase polymerase chain reaction. The therapeutic effects of a selected antibiotic on AD-like skin lesions of NC/Nga mice were assessed in terms of the skin severity score, histological changes in the lesioned skin, the serum level of total IgE, and expression of Th1/Th2 cytokines in lymph nodes and skin lesions. RESULTS: Antibiotic-treated, OVA peptide-pulsed LCs inhibited development of Th2 cells but not Th1 cells. This was accompanied by suppression of T-cell immunoglobulin and mucin domain-containing protein (TIM)-4 expression in LCs. Doxycycline had the greatest activity against Staphylococcus aureus strains isolated from skin lesions of patients with AD, and a strong inhibitory effect on Th2 cell development. Doxycycline suppressed the increase in the skin severity score during the acute phase in NC/Nga mice similar to betamethasone. This suppressive effect was associated with a decrease in the serum IgE level and production of Th2 cytokines in auricular lymph node cells and skin lesions. CONCLUSION: Topical application of doxycycline to AD lesions would act on both superficial S. aureus colonization and epidermal LCs, thus possibly inhibiting the development of Th2 cells in vivo, with benefits for control of acute inflammation in AD.

Norfloxacin, a Fluoroquinolone Antibiotic, Inhibits Langerhans Cell-Mediated Th1 and Th2 Cell Development.

BACKGROUND: It is widely acknowledged that Langerhans cells (LCs) play a primary role in the polarization of T helper type 1 (Th1) or T helper type 2 (Th2) immune responses. Our aim was to find fluoroquinolone ("new quinolone") antibiotics that would inhibit LC-mediated Th2 cell development. METHODS: Expression of LC surface molecules was investigated using the reverse transcriptase polymerase chain reaction. The effects of fluoroquinolone antibiotics on T-cell immunoglobulin and mucin domain-containing protein (TIM)-4 expression in LCs were examined to predict whether they would inhibit Th2 cell development. Mice were primed via the hind footpad with ovalbumin (OVA) peptide-pulsed LCs that had been treated with a selected fluoroquinolone antibiotic, then 5 days later the cytokine response in popliteal lymph nodes was examined by enzyme-linked immunosorbent assay. RESULTS: Norfloxacin was selected as a candidate inhibitor of Th2 cell development. As expected, OVA peptide-pulsed LCs that had been treated with norfloxacin and injected into the hind footpads of mice inhibited Th2 cell development, as represented by down-regulation of interleukin (IL)-4 production, as well as Th1 cell development, as represented by down-regulation of interferon (IFN)- g production. This additional inhibition of Th1 cell development was accompanied by suppression of CD40 expression in LCs. In addition, Staphylococcus aureus strains isolated from skin lesions of patients with atopic dermatitis (AD) were more susceptible to norfloxacin than to gentamicin. Topical treatment with norfloxacin significantly suppressed the increase in the skin severity score in NC/Nga mice with AD-like skin lesions. This suppressive effect was associated with a decrease in the production of IFN-g and IL-4 in auricular lymph node cells. CONCLUSIONS: The present results show that topical application of norfloxacin inhibits the development of AD-like skin lesions in NC/Nga mice. This suggests that topical application of norfloxacin to AD lesions colonized with S. aureus would act on both superficial S. aureus and epidermal LCs, thus possibly inhibiting the development of Th1 and Th2 cells in vivo, and controlling the severity of AD.

Mast Cells Stimulated with Peptidoglycan from Staphylococcus aureus Augment the Development of Th1 Cells.

BACKGROUND: The skin of patients with atopic dermatitis (AD) is superficially colonized by Staphylococcus aureus. We have previously found that percutaneous permeation of peptidoglycan (PEG) from S. aureus increases the number of mast cells in the dermis, as seen in skin lesions of AD patients. The purpose of the present study was to clarify the influence of PEG on T helper type 1 (Th1)/ T helper type 2 (Th2) cell development mediated by mast cells. METHODS: Mast cells were induced by long-term culture of murine spleen cells in medium supplemented with tumor necrosis factor (TNF)- a. Ovalbumin (OVA) peptide-pulsed mast cells were incubated with naïve Th cells in the presence or absence of PEG. Five days later, Th cells in the culture were stimulated with phorbol 12-myristate 13-acetate (PMA) and ionomycin, and Th1/Th2 cytokine production was investigated by enzyme-linked immunosorbent assay. RESULTS: It was confirmed that the mast cells we obtained had surface expression of I-Ad, worked as antigen-presenting cells, and induced Th1 cell and Th2 cell development. The stimulation of mast cells with PEG enhanced the development of Th1 cells but not that of Th2 cells. The increase of Th1 cell development stimulated by PEG was associated with an increase in the expression of Notch ligand Delta 1 in the mast cells. Furthermore, treatment of mast cells with the macrolide antibiotic josamycin suppressed Th1 cell development and this was correlated with a reduction of both Delta 1 expression and interleukin (IL)-12 production in mast cells. CONCLUSIONS: Colonization of S. aureus on the lesioned skin of AD patients contributes to not only an increase in the number of mast cells but also Th1 cell development mediated by mast cells in the dermis and subsequent induction of chronic inflammation, which is characterized by up-regulation of the Th1 cytokine, interferon (IFN)- g. Therefore, application of josamycin to the lesional skin of AD patients may provide relief from chronic inflammation mediated by mast cells.

Topical Application of Josamycin Inhibits Development of Atopic Dermatitis-Like Skin Lesions in NC/Nga Mice.

BACKGROUND: Patients with atopic dermatitis (AD) have superficial skin colonization by Staphylococcus aureus and an increased number of T helper type 2 (Th2) cells in their peripheral blood. Our previous study showed that josamycin, a macrolide antibiotic, had excellent bactericidal activity against S. aureus strains isolated from AD patients and simultaneously inhibited Th1 and Th2 cell development mediated by Langerhans cells. The purpose of the present study was to evaluate the effect of topical application of josamycin on AD-like skin lesions in NC/Nga mice. METHODS: Josamycin (0.1%) was topically administered to NC/Nga mice with AD-like skin lesions induced by 2, 4, 6-trinitrochlorobenzene (TNCB). The therapeutic effects of josamycin were assessed by measurement of the skin severity scores, histological changes in the lesioned skin, serum levels of total IgE, and expression of interferon (IFN)-γ and interleukin (IL)-4 in lymph nodes and skin lesions. RESULTS: Topical treatment with josamycin significantly suppressed the increase in the skin severity score in NC/Nga mice. This suppressive effect was equal to that of betamethasone, and was associated with a decrease in the density of cellular infiltration into the dermis, the mast cell count in the dermis and the serum IgE level. Furthermore, topical application of josamycin reduced the expression of IFN-γ and IL-4 in auricular lymph node cells and the skin lesions. CONCLUSION: The present results show that topical application of josamycin inhibits the development of AD-like skin lesions in NC/Nga mice. This suggests that topical application of josamycin to AD lesions colonized by S. aureus would be beneficial for control of AD by acting on superficially located S. aureus and by inhibiting the development of Th1 and Th2 cells.This article is open to POST-PUBLICATION REVIEW. Registered readers (see "For Readers") may comment by clicking on ABSTRACT on the issue's contents page.

Betamethasone, but Not Tacrolimus, Suppresses the Development of Th2 Cells Mediated by Langerhans Cell-Like Dendritic Cells.

It is well known that Langerhans cells (LCs) work as the primary orchestrators in the polarization of the immune milieu towards a T helper type 1 (Th1) or T helper type 2 (Th2) response. In this study, we investigated the effects of tacrolimus and betamethasone, each used as topical applications in atopic dermatitis (AD), on Th2 cell development mediated by LCs. LC-like dendritic cells (LDCs) were generated from mouse bone marrow cells and used as substitutes for LCs. Mice were primed with ovalbumin (OVA) peptide-pulsed LDCs, which had been treated with tacrolimus or betamethasone, via the hind footpad. After 5 d, the cytokine response in the popliteal lymph nodes was investigated by enzyme-linked immunosorbent assay. The expression of cell surface molecules on LDCs was investigated via reverse transcriptase polymerase chain reaction. Administration of OVA peptide-pulsed LDCs, which had been treated with betamethasone, inhibited Th2 cell development, as represented by the down-regulation of interleukin-4 production, and also inhibited Th1 cell development, represented by the down-regulation of interferon-γ production. However, tacrolimus-treated LDCs did not induce such inhibition of the development of Th1 and Th2 cells. The inhibition of Th1 and Th2 cell development was associated with the suppression of CD40 and T-cell immunoglobulin, and mucin domain-containing protein (TIM)-4 expression, respectively, in LDCs. These results suggest that the topical application of betamethasone to skin lesions of patients with AD acts on epidermal LCs, and may inhibit the development of Th2 cells, thus being of benefit for the control of AD.

Effects of Macrolide Antibiotics on Th1 Cell and Th2 Cell Development Mediated by Langerhans Cells.

- Background: It is well known that Langerhans cells (LCs) work as the primary orchestrators in the polarization of the immune milieu towards a T helper type 1 (Th1) or a Th2 immune response. In this study, we investigated the effects of macrolide antibiotics on Th1 cell and Th2 cell development mediated by LCs. METHODS: LC-like dendritic cells (LDCs) were generated from mouse bone marrow cells and used as substitutes for LCs. Mice were primed with ovalbumin (OVA) peptide-pulsed LDCs, which had been treated with each macrolide antibiotic, via the hind footpad. After 5 days, the cytokine response in the popliteal lymph nodes was investigated by enzyme-linked immunosorbent assay. The expression of cell surface molecules on LDCs was investigated using reverse transcriptase polymerase chain reaction. RESULTS: Injection of OVA peptide-pulsed LDCs, which had been treated with josamycin or spiramycin, inhibited Th2 cell development as represented by down-regulation of interleukin (IL)-4 production as well as Th1 cell development as represented by down-regulation of interferon (IFN)-g production. This inhibition of Th1 cell and Th2 cell development was associated with suppression of CD86 and T-cell immunoglobulin and mucin domain-containing protein (TIM)-4 expression, respectively, in LDCs. Furthermore, Staphylococcus aureus strains isolated from skin lesions of patients with atopic dermatitis (AD) were more susceptible to josamycin than to spiramycin. CONCLUSIONS:   These results suggest that topical application of josamycin to AD lesions colonized with S. aureus would be beneficial for control of AD by acting on both superficial S. aureus and epidermal LCs, and inhibiting the development of Th2 cells.

CCL17 production by mouse langerhans cells stimulated with Staphylococcus aureus cell wall components.

Patients with atopic dermatitis (AD) show increased numbers of Th2 cells in their acute skin lesions and superficial skin colonization by Staphylococcus aureus. The purpose of this study was to clarify the effect of S. aureus cell wall components on Th2 chemokine production by murine Langerhans cells (LCs). Murine LCs were stimulated with peptidoglycan (PEG) and/or muramyldipeptide (MDP) for 24 or 48 h, and Th1 and Th2 chemokine production was investigated by reverse transcription polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA). PEG-stimulation of LCs induced production of the Th2 chemokine CCL17 and this was enhanced in the presence of interleukin (IL)-4. A low-molecular weight PEG fragment, MDP, did not induce CCL17 production by LCs. However, when LCs were stimulated with PEG in combination with MDP, PEG-induced CCL17 production was synergistically enhanced by MDP. Furthermore, PEG- and MDP-induced CCL17 production was enhanced to a greater extent in the presence of IL-4. These results suggest that S. aureus colonization in AD patients may enhance the Th2-prone immune response through upregulation of CCL17 production by LCs, which would occur as a result of simultaneous stimulation with PEG and MDP from S. aureus in a Th2 environment.

Peptidoglycan in combination with muramyldipeptide synergistically induces an interleukin-10-dependent T helper 2-dominant immune response.

In this study, peptidoglycan (PEG) from Staphylococcus aureus-stimulated, but not muramyldipeptide (MDP)-stimulated, Langerhans cells (LCs) induced a dose-dependent Th2-prone immune response. However, when LCs were stimulated with PEG in combination with MDP, the strength of Th2 immune responses was synergistically augmented by MDP. Furthermore, it was found that production of IL-10, but not of IL-12 p40, by PEG-stimulated LCs was also enhanced in the presence of MDP. These results suggest that MDP enhances Th2 cell development through up-regulation of IL-10 production from PEG-stimulated LCs, increase the importance of S. aureus colonization in patients with atopic dermatitis.

Peptidoglycan-induced T helper 2 immune response in mice involves interleukin-10 secretion from Langerhans cells.

Patients with atopic dermatitis (AD) have superficial skin colonization with Staphylococcus aureus and an increased number of T helper (Th)2 cells in their peripheral blood. The purpose of this study was to clarify the involvement of interleukin (IL)-10 secretion from Langerhans cells (LCs) in staphylococcal peptidoglycan (PEG)-induced Th2 immune responses in mice. Mice were primed with LCs pulsed with PEG (or LPS) and ovalbumin (OVA) and then given a booster OVA injection 2 days later in the hind footpad. Five days after the OVA injection, cytokine responses in the draining popliteal lymph nodes were investigated by RT-PCR and ELISA. Production of both IL-10 and IL-12 by cultured LCs was detected by ELISA. Administration of PEG- or LPS-stimulated LCs into the hind footpads of the mice induced Th2-prone and Th1-prone immune responses, respectively, as represented by expression of IL-4 and interferon-γ. In vitro experiments showed that PEG induced greater production of IL-12 p40 from LCs than did LPS, whereas LPS induced greater production of IL-12 p70 from LCs than did PEG. Furthermore, it was found that PEG-stimulated LCs induced greater production of IL-10 than did LPS-stimulated LCs, and that neutralization of IL-10 augmented IL-12 p70 production and inhibited Th2 development by PEG-stimulated LCs. These results suggest that PEG can induce Th2 development through down-regulation of IL-12 p70 production by LCs in an IL-10 production-dependent manner and would explain the role of S. aureus colonization in patients with AD.

Analysis of signaling pathways involved in peptidoglycan-induced RANTES production from murine Langerhans cells.

BACKGROUND: Our previous study showed that percutaneous application of peptidoglycan from Staphylococcus aureus induced eosinophil infiltration in murine skin through production of CCL5/RANTES (regulated upon activation in normal T cells expressed and secreted) from epidermal Langerhans cells. Although it is well known that peptidoglycan is an agonist of Toll-like receptor (TLR)-2, it is unclear whether other TLR agonists are able to induce RANTES production from Langerhans cells. METHODS: Langerhans cells were purified from murine epidermal cells by the panning method using anti-IA(d) monoclonal antibody. RANTES production by Langerhans cells was investigated by RT-PCR and ELISA. Analysis of the signaling pathways responsible for RANTES production by Langerhans cells was performed by ELISA using N-acetyl-L-cysteine, SP600125, SB203580 and PD98059, which are specific inhibitors of NF-kappaB activation, JNK, p38 MAPK and ERK, respectively, and was finally confirmed by Western blot analysis. RESULTS: Peptidoglycan, poly(I:C), lipopolysaccharide and CpG DNA, which signal through TLR-2, TLR-3, TLR-4 and TLR-9, respectively, were found to strongly induce RANTES production. Treatment with N-acetyl-L-cysteine inhibited all TLR agonist-induced RANTES production. However, treatment with SP600125 did not inhibit CpG DNA-induced RANTES production. Furthermore, treatment with SB203580 inhibited only peptidoglycan- and lipopolysaccharide-induced RANTES production and the inhibition was correlated with that of p38 MAPK phosphorylation. CONCLUSION: These results suggest that the signaling pathway of RANTES production from murine Langerhans cells induced by different TLR stimuli is not necessarily the same, and that inhibition of p38 MAPK may be a more specific therapeutic target for eosinophilic inflammation in patients with atopic dermatitis associated with S. aureus colonization.

Peptidoglycan from Staphylococcus aureus induces IL-4 production from murine spleen cells via an IL-18-dependent mechanism.

BACKGROUND: Atopic dermatitis (AD) is a chronic inflammatory skin disease with immunopathologic features that vary depending on the duration of the lesion. Skin lesions of AD patients show an increased number of Th2 cells in the dermis and superficial Staphylococcus aureus colonization. The purpose of this study was to predict the effects of peptido- glycan (PEG) from S. aureus on the induction of interleukin (IL)-4 production in AD patients. METHODS: PEG was applied to barrier-disrupted abdominal mouse skin every 5 days. Twenty days later, IL-4 production in the spleen was investigated by reverse transcription-polymerase chain reaction (RT-PCR). Spleen cells from normal mice were also treated in vitro with PEG and processed for IL-4 production by RT-PCR and enzyme-linked immunosorbent assay. RESULTS: IL-4 production was significantly increased in the spleen of PEG-treated mice compared with that of PBS-applied mice. In addition, in vitro experiments demonstrated that PEG was able to induce IL-4 production in murine spleen cells. Furthermore, IL-4 production was associated with IL-18 production, but not with IL-2 production from PEG-stimulated spleen cells, and IL-4 mRNA was expressed in CD4+ lymphocytes in the spleen cells. In in vivo experiments, percutaneous treatment with PEG induced mRNA expression not only for IL-4 but also for IL-18 in the spleen. CONCLUSION: These results suggest that PEG may influence the induction of a Th2-dominant cytokine response in AD patients through IL-4 production from CD4+ T cells stimulated with PEG-induced IL-18.

Effects of the macrolide antibiotic, midecamycin, on Staphylococcus aureus product-induced Th2 cytokine response in patients with atopic dermatitis.

In the present study, the effects of the macrolide antibiotic, midecamycin (MDM), on the Th2 cytokine response induced by the Staphylococcus aureus products, staphylococcal enterotoxin B (SEB), lipoteichoic acid (LTA), and peptidoglycan (PEG), was investigated in human peripheral blood mononuclear cells (PBMCs) from patients with atopic dermatitis (AD). MDM inhibited SEB-induced mRNA expression of the Th2 cytokines interleukin-4 (IL-4) and IL-5 in PBMCs from patients with AD. Furthermore, MDM also suppressed LTA-induced or PEG-induced IL-5 mRNA expression in these patients. Inhibition of mRNA expression by MDM correlated with the synthesis of cytokines in PBMCs, indicating that MDM controls Th2 cytokine production. In addition, S. aureus strains isolated from skin lesions of patients with AD were particularly susceptible to MDM compared with gentamicin, which is used widely in Japan as an antibiotic ointment combined with steroid for topical application in AD. These results suggest that topical administration of MDM might be beneficial in AD lesions infected with S. aureus.