Mouse Model Of Skin Inflammation Research Articles

Heme oxygenase-1-mediated anti-inflammatory effects of tussilagonone on macrophages and 12-O-tetradecanoylphorbol-13-acetate-induced skin inflammation in mice

The dried flower buds of Tussilago farfara L. have been used in traditional medicine, mainly as an antitussive in the treatment of cough and other respiratory problems. In the present study, we investigated the anti-inflammatory signaling pathway via the upregulation of heme oxygenase-1 (HO-1) in response to tussilagonone (TGN), a sesquiterpene compound isolated from T. farfara. TGN induced HO-1 expression and nuclear factor-E2-related factor 2 (Nrf2) activation in RAW 264.7 cells. Nuclear translocation of Nrf2 by TGN also increased in a time- and dose-dependent manner, indicating that TGN induced HO-1 via the Nrf2 pathway. Consistent with the notion that HO-1 has anti-inflammatory properties, TGN suppressed inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) expression and reduced the mRNA expression of proinflammatory cytokines, as well as nitric oxide (NO) and prostaglandin E2 (PGE2) production in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. TGN inhibited the phosphorylation and degradation of inhibitory κB-α (IκB-α) and the nuclear translocation of nuclear factor (NF)-κB. However, a specific inhibitor of HO-1 reversed the TGN-mediated suppression of NO production and knockdown of HO-1 by small interfering RNA abrogated inhibitory effects of TGN on iNOS and COX-2 protein expression and NF-κB nuclear translocation. Furthermore, TGN reduced iNOS and COX-2 expression in a 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced skin inflammation mouse model. Taken together, these findings suggest an important role for TGN-induced HO-1 activation in regulating inflammatory responses. Moreover, TGN is a potent therapeutic candidate for targeting the crosstalk between Nrf2/HO-1 and the NF-κB signaling pathway in the prevention or treatment of inflammation-associated diseases.

Imiquimod Treatment Causes Systemic Disease in Mice Resembling Generalized Pustular Psoriasis in an IL-1 and IL-36 Dependent Manner.

Generalized pustular psoriasis (GPP) is a severe form of psoriasis that can be caused by missense mutations in the interleukin-36 (IL-36) receptor antagonist. In addition to neutrophil rich skin inflammation, GPP patients typically also experience anorexia, fever, malaise, and pain. The imiquimod-induced skin inflammation mouse model has rapidly become a popular way to study plaque psoriasis, which typically does not involve symptoms of systemic disease. In this model, neutrophil recruitment to the skin is dependent upon the inflammatory mediators IL-1, via its receptor IL-1R1, and IL-36α. Unexpectedly, we observed that mice also exhibited signs of anorexia (weight loss and decreased food intake), general malaise (decreased activity and loss of interest in building nests), and pain (nose bulging and hunched posture). A scoring system allowing quantitative comparisons of test groups was developed. Female mice were found to develop more severe disease than male mice. Furthermore, mice deficient in both IL-1R1 and IL-36α are nearly disease-free, while mice lacking only one of these inflammatory mediators have less severe disease than wild type mice. Hence, the imiquimod-induced skin inflammation mouse model recapitulates not only plaque psoriasis, but also the more severe symptoms, that is, anorexia, malaise, and pain, seen in GPP.

Imiquimod-induced psoriasis-like skin inflammation is suppressed by BET bromodomain inhibitor in mice through RORC/IL-17A pathway modulation

Psoriasis is one of the most common skin disorders characterized by erythematous plaques that result from hyperproliferative keratinocytes and infiltration of inflammatory leukocytes into dermis and epidermis. Recent studies suggest that IL-23/IL-17A/IL-22 cytokine axis plays an important role in the pathogenesis of psoriasis. The small molecule bromodomain and extraterminal domain (BET) inhibitors, that disrupt interaction of BET proteins with acetylated histones have recently demonstrated efficacy in various models of inflammation through suppression of several pathways, one of them being synthesis of IL-17A/IL-22 which primarily depends on transcription factor, retinoic acid receptor-related orphan receptor C (RORC). However, the efficacy and mechanistic aspect of a BET inhibitor in mouse model of skin inflammation has not been explored previously. Therefore, this study investigated the role of BET inhibitor, JQ-1 in mouse model of psoriasis-like inflammation. Mice were topically applied imiquimod (IMQ) to develop psoriasis-like inflammation on the shaved back and ear followed by assessment of skin inflammation (myeloperoxidase activity, ear thickness, and histopathology), RORC and its signature cytokines (IL-17A/IL-22). JQ-1 suppressed IMQ-induced skin inflammation as reflected by a decrease in ear thickness/myeloperoxidase activity, and RORC/IL-17A/IL-22 expression. Additionally, a RORα/γ agonist SR1078 was utilized to investigate the role of RORC in BET-mediated skin inflammation. SR1078 reversed the protective effect of JQ-1 on skin inflammation at both histological and molecular levels in the IMQ model. The current study suggests that BET bromodomains are involved in psoriasis-like inflammation through induction of RORC/IL-17A pathway. Therefore, inhibition of BET bromodomains may provide a new therapy against skin inflammation.

Protective effect of small molecule analogues of the Acanthocheilonema viteae secreted product ES-62 on oxazolone-induced ear inflammation

ES-62 is the major secreted protein of the rodent filarial nematode Acanthocheilonema viteae. The molecule contains covalently attached phosphorylcholine (PC) residues, which confer anti-inflammatory properties on ES-62, underpinning the idea that drugs based on this active moiety may have therapeutic potential in human diseases associated with aberrant inflammation. Here we demonstrate that two synthetic small molecule analogues (SMAs) of ES-62 termed SMA 11a and SMA 12b are protective in the oxazolone-induced acute allergic contact dermatitis mouse model of skin inflammation, as measured by a significant reduction in ear inflammation following their administration before oxazolone sensitisation and before oxazolone challenge. Furthermore, it was found that when tested, 12b was effective at reducing ear swelling even when first administered before challenge. Histological analysis of the ears showed elevated cellular infiltration and collagen deposition in oxazolone-treated mice both of which were reduced by treatment with the two SMAs. Likewise, the oxazolone-induced increase in IFNγ mRNA in the ears was reduced but no effect on other cytokines investigated was observed. Finally, no influence on the mast cell populations in the ear was observed.

Antibody-mediated delivery of interleukin 4 to the neo-vasculature reduces chronic skin inflammation

BackgroundThe antibody-mediated delivery of cytokines (“immunocytokines”) to sites of pathological angiogenesis represents an attractive strategy for the development of innovative biopharmaceuticals, capable of modulating the activity of the immune system in cancer and in chronic inflammatory conditions. ObjectiveRecombinant IL4 has previously been shown to be therapeutically active in patients with psoriasis. The antibody-mediated delivery of this cytokine to sites of chronic skin inflammatory conditions should lead to an improved potency and selectivity, compared to non-targeted IL4. MethodsThe therapeutic activity of F8-IL4, a fusion protein of the F8 antibody (specific to the alternatively-spliced EDA domain of fibronectin) with murine IL4, was investigated in three immunocompetent mouse models of skin inflammation: two induced by the TLR7/8 ligand imiquimod (in Balb/c and C57BL/6) and one mediated by the over-expression of VEGF-A. ResultsThe EDA domain of fibronectin, a marker for angiogenesis, is expressed in the inflamed skin in all three models and F8-IL4 selectively localized to inflamed skin lesions following intravenous administration. The F8-IL4 fusion protein mediated a therapeutic benefit, which was superior to the one of a non-targeted version of IL4 and led to increased levels of key regulatory cytokines (including IL5, IL10, IL13, and IL27) in the inflamed skin, while IL2 levels were not affected in all treatment groups. A murine version of etanercept and a murine anti-IL17 antibody were used as positive control in the therapy experiments. ConclusionSkin inflammatory lesions can be selectively targeted using anti-EDA antibody–cytokine fusion proteins and the pharmacodelivery of IL4 confers a therapeutic benefit by shifting the cytokine balance.

Repositioning drugs for inflammatory disease - fishing for new anti-inflammatory agents.

Inflammation is an important and appropriate host response to infection or injury. However, dysregulation of this response, with resulting persistent or inappropriate inflammation, underlies a broad range of pathological processes, from inflammatory dermatoses to type 2 diabetes and cancer. As such, identifying new drugs to suppress inflammation is an area of intense interest. Despite notable successes, there still exists an unmet need for new effective therapeutic approaches to treat inflammation. Traditional drug discovery, including structure-based drug design, have largely fallen short of satisfying this unmet need. With faster development times and reduced safety and pharmacokinetic uncertainty, drug repositioning – the process of finding new uses for existing drugs – is emerging as an alternative strategy to traditional drug design that promises an improved risk-reward trade-off. Using a zebrafish in vivo neutrophil migration assay, we undertook a drug repositioning screen to identify unknown anti-inflammatory activities for known drugs. By interrogating a library of 1280 approved drugs for their ability to suppress the recruitment of neutrophils to tail fin injury, we identified a number of drugs with significant anti-inflammatory activity that have not previously been characterized as general anti-inflammatories. Importantly, we reveal that the ten most potent repositioned drugs from our zebrafish screen displayed conserved anti-inflammatory activity in a mouse model of skin inflammation (atopic dermatitis). This study provides compelling evidence that exploiting the zebrafish as an in vivo drug repositioning platform holds promise as a strategy to reveal new anti-inflammatory activities for existing drugs.

In Vitro and in Vivo Anti-inflammatory Effects of a Novel 4,6-Bis ((E)-4-hydroxy-3-methoxystyryl)-1-phenethylpyrimidine-2(1H)-thione

Inflammation plays a critical defensive role in the human body. However, uncontrolled or aberrant inflammatory responses contribute to various acute and chronic diseases. The Nrf2-ARE pathway plays a pivotal role in the regulation of inflammatory markers, such as inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). On the basis of this concept, we synthesized a novel anti-inflammatory 4,6-bis ((E)-4-hydroxy-3-methoxystyryl)-1-phenethylpyrimidine-2(1H)-thione (HPT), and in vitro experiments using HepG2-C8 ARE-luciferase-transfected cells demonstrated the induction of Nrf2-ARE activity. In lipopolysaccharide (LPS)-induced RAW 264.7 cells, HPT treatment reduced the production of nitric oxide (NO) as well as the protein and mRNA expression levels of COX-2 and iNOS, in a dose-dependent manner. In addition, HPT suppressed the mRNA expression of inflammatory cytokines, including tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, and IL-6. In LPS-induced macrophages, HPT inhibited COX-2 and iNOS by blocking the activation of p38 and c-Jun NH2-terminal kinase (JNK) but not extracellular signal-regulated kinase (ERK1/2). Furthermore, an in vivo anti-inflammatory study was performed using a TPA-induced skin inflammation mouse model, and the results showed that HPT reduced TPA-induced inflammation and attenuated the expression of COX-2 and iNOS in TPA-induced mouse skin tissue. Thus, HPT demonstrated anti-inflammatory activity both in LPS-induced RAW 264.7 cells and TPA-stimulated mouse skin and may therefore serve as a potential anti-inflammatory agent.

S100A8-S100A9 Protein Complex Mediates Psoriasis by Regulating the Expression of Complement Factor C3

Psoriasis is a common heterogeneous inflammatory skin disease with a complex pathophysiology and limited treatment options. Here we performed proteomic analyses of human psoriatic epidermis and found S100A8-S100A9, also called calprotectin, as the most upregulated proteins, followed by the complement component C3. Both S100A8-S100A9 and C3 are specifically expressed in lesional psoriatic skin. S100A9 is shown here to function as a chromatin component modulating C3 expression in mouse and human cells by binding to a region upstream of the C3 start site. When S100A9 was genetically deleted in mouse models of skin inflammation, the psoriasis-like skin disease and inflammation were strongly attenuated, with a mild immune infiltrate and decreased amounts of C3. In addition, inhibition of C3 in the mouse model strongly reduced the inflammatory skin disease. Thus, S100A8-S100A9 can regulate C3 at the nuclear level and present potential new therapeutic targets for psoriasis.

IL-1R1 Signaling Facilitates Munro’s Microabscess Formation in Psoriasiform Imiquimod-Induced Skin Inflammation

Munro’s microabscesses contain polymorphonuclear leukocytes and form specifically in the epidermis of psoriasis patients. The mechanism whereby the neutrophils are recruited into the epidermis is poorly understood. Using a combination of human and mouse primary keratinocyte cell cultures and the imiquimod-induced psoriasis-like mouse model of skin inflammation we explored the role of interleukin-1 (IL-1) signaling in microabscess formation. In vitro imiquimod stimulated production of IL-1α and neutrophil recruiting chemokines. Imiquimod activated chemokine expression was dependent upon adenosine signaling and independent of IL-1α and IL-1 receptor type 1 (IL-1R1); nevertheless, IL-1α could enhance chemokine expression initiated by imiquimod. Topical application of imiquimod in vivo led to epidermal microabscess formation, acanthosis and increased IL-1α and chemokine expression in the skin of wild type mice. However, in IL-1R1 deficient mice these responses were either absent or dramatically reduced. These results demonstrate that IL-1α and IL-1R1 signaling is essential for microabscess formation, neutrophil recruiting chemokine expression and acanthosis in psoriasis-like skin inflammation induced by imiquimod.

Role of SERPINB3, SERPINB4, and their mouse homolog Serpinb3a in allergen-induced cutaneous inflammation. (P3347)

Abstract The serine protease inhibitors SERPINB3 and SERPINB4 have been implicated in allergic disorders. We have recently demonstrated a role for the serine protease inhibitor Serpinb3a (the mouse homolog of SERPINB3 and SERPINB4) in a mouse model of asthma. Elevated levels of SERPINB3 and SERPINB4 are detected in the serum of patients with atopic dermatitis (AD) and they have been proposed as possible biomarkers for AD. Serine proteases are critical for epidermal barrier homeostasis and aberrant expression and/or activity of serine proteases have been associated with AD in human studies. We also observe a significant induction in the expression of Serpinb3a in a mouse model of cutaneous inflammation that has many similarities to AD. However the role of Serpinb3a in the skin is unknown. Using our mouse model of skin inflammation, we observed attenuation in trans-epidermal water loss, skin inflammation scores, decreased levels of total IgE, and pro-inflammatory markers like S100A8 in mice lacking Serpinb3a following exposure to Aspergillus fumigatus extract. In humans, we have identified a single nucleotide polymorphism (SNP) in the locus that encodes SERPINB3 and SERPINB4 that is associated with trans-epidermal water loss (p-val=0.031). In conclusion, we demonstrate that SERPINB3 and SERPINB4 contribute to cutaneous inflammation and work is ongoing to elucidate the role of this protein in regulating the skin barrier.

Apoptotic and necroptotic cell death in cutaneous inflammation.

Epidermal keratinocytes provide an essential structural and immunological barrier forming the first line of defense against potentially pathogenic microorganims. Mechanisms regulating barrier integrity and innate immune responses in the epidermis are important for the maintenance of skin immune homeostasis and the pathogenesis of inflammatory skin diseases. Cell death, and in particular, apoptosis, has been suggested to play a key role in numerous skin inflammatory diseases. Supporting these reports, studies in mouse models have emphasized the role of increased keratinocyte apoptosis in cutaneous inflammation. Necrosis has long been considered as a passive form of cell death, but recent reports have unraveled the molecular regulation of necrosis. Programmed necrosis, also termed necroptosis, has been recently implicated in mouse models of skin inflammation. In this review, we discuss the respective roles of apoptotic or necrotic cell death of epidermal keratinocytes in mouse models of cutaneous inflammation and in the physiopathology of human inflammatory dermatoses.

Hiding under the skin: Interleukin-17–producing γδ T cells go under the skin?

Despite the irrefutable role of inflammation in psoriasis, a complete knowledge of what immune cells and cytokines are involved during initiation and progression of this skin disease is lacking. Moreover, the complexities of the immune cell network and potential differences between mice and humans have led to translational failures. It is therefore important that we acquire in-depth understanding of what inflammatory players, of the many involved, are crucial, if we wish to develop effective therapies. In 'Bedside to Bench', James Krueger discusses how a subset of T cells, TH17 cells, which release interleukin-17 in humans, seem to be essential for pathogenesis of psoriasis. The interplay between interleukin-17 and other cytokines that may potentially be involved in psoriasis also needs further investigation. Additionally, there are open questions as to what subset of T cells, other than TH17, also produce interleukin-17 and when. In 'Bench to Bedside', Burkhard Becher and Stanislav Pantelyushin examine this issue by looking at a mouse model of skin inflammation that resembles psoriasis in humans. A class of skin-invading innate immune cells called γδ T cells was shown to drive skin inflammation in this model, particularly during the early stages of the disease, suggesting that innate immunity plays an important part in the initiation of psoriasis.

Humanized Mouse Model of Skin Inflammation Is Characterized by Disturbed Keratinocyte Differentiation and Influx of IL-17A Producing T Cells

Humanized mouse models offer a challenging possibility to study human cell function in vivo. In the huPBL-SCID-huSkin allograft model human skin is transplanted onto immunodeficient mice and allowed to heal. Thereafter allogeneic human peripheral blood mononuclear cells are infused intra peritoneally to induce T cell mediated inflammation and microvessel destruction of the human skin. This model has great potential for in vivo study of human immune cells in (skin) inflammatory processes and for preclinical screening of systemically administered immunomodulating agents. Here we studied the inflammatory skin response of human keratinocytes and human T cells and the concomitant systemic human T cell response.As new findings in the inflamed human skin of the huPBL-SCID-huSkin model we here identified: 1. Parameters of dermal pathology that enable precise quantification of the local skin inflammatory response exemplified by acanthosis, increased expression of human β-defensin-2, Elafin, K16, Ki67 and reduced expression of K10 by microscopy and immunohistochemistry. 2. Induction of human cytokines and chemokines using quantitative real-time PCR. 3. Influx of inflammation associated IL-17A-producing human CD4+ and CD8+ T cells as well as immunoregulatory CD4+Foxp3+ cells using immunohistochemistry and -fluorescence, suggesting that active immune regulation is taking place locally in the inflamed skin. 4. Systemic responses that revealed activated and proliferating human CD4+ and CD8+ T cells that acquired homing marker expression of CD62L and CLA. Finally, we demonstrated the value of the newly identified parameters by showing significant changes upon systemic treatment with the T cell inhibitory agents cyclosporine-A and rapamycin.In summary, here we equipped the huPBL-SCID-huSkin humanized mouse model with relevant tools not only to quantify the inflammatory dermal response, but also to monitor the peripheral immune status. This combined approach will gain our understanding of the dermal immunopathology in humans and benefit the development of novel therapeutics for controlling inflammatory skin diseases.

Topical cis-urocanic acid attenuates oedema and erythema in acute and subacute skin inflammation in the mouse

cis-Urocanic acid (cis-UCA) is an endogenous immunosuppressive molecule of the epidermis. We investigated the effects of topical cis-UCA creams (2·5% and 5%) in acute and subacute mouse models of skin inflammation. Acute skin irritation was induced by applying dimethyl sulphoxide (DMSO) on the earlobe of CD-1 mice. Topical cis-UCA, hydrocortisone (1%) or tacrolimus (0·1%) were applied 10 min later. In another model, subacute inflammation was provoked and maintained by three applications of 12-O-tetradecanoylphorbol-13-acetate (TPA) on the ears of NMRI mice on days 1, 2 and 4. The test products were applied topically twice a day during 6 days. In the acute DMSO model, cis-UCA creams suppressed ear swelling at 1 h significantly more efficiently than hydrocortisone (P < 0·01) and tacrolimus (P < 0·001). Ear swelling was significantly inhibited by cis-UCA (P < 0·001) in the subacute TPA model as well. The 5% cream also decreased erythema, whereas tacrolimus enhanced skin reddening. Treatments with cis-UCA did not affect TPA-induced infiltration of neutrophils to the skin. In contrast to hydrocortisone, cis-UCA did not reduce epidermal thickness. The results suggest that cis-UCA - unlike hydrocortisone and tacrolimus - is efficient in both acute and subacute skin inflammation, attenuating skin oedema and erythema. Topical drug therapy with cis-UCA may provide a safe and effective drug treatment modality in inflammatory skin disorders.

Periostin promotes chronic allergic inflammation in response to Th2 cytokines

Allergic inflammation triggered by exposure of an allergen frequently leads to the onset of chronic inflammatory diseases such as atopic dermatitis (AD) and bronchial asthma. The mechanisms underlying chronicity in allergic inflammation remain unresolved. Periostin, a recently characterized matricellular protein, interacts with several cell surface integrin molecules, providing signals for tissue development and remodeling. Here we show that periostin is a critical mediator for the amplification and persistence of allergic inflammation using a mouse model of skin inflammation. Th2 cytokines IL-4 and IL-13 stimulated fibroblasts to produce periostin, which interacted with αv integrin, a functional periostin receptor on keratinocytes, inducing production of proinflammatory cytokines, which consequently accelerated Th2-type immune responses. Accordingly, inhibition of periostin or αv integrin prevented the development or progression of allergen-induced skin inflammation. Thus, periostin sets up a vicious circle that links Th2-type immune responses to keratinocyte activation and plays a critical role in the amplification and chronicity of allergic skin inflammation.

Abstract 4243: Potential protective role of phytoalexins derived from soybean by biotic elicitor on inflammatory mechanism

Abstract Given the preventive effect of soy intake against several chronic diseases, this study was conducted to investigate the inhibitory activity of phytoalexins glyceollins derived from soybean isoflavones by treating with biotic elicitor against LPS-induced inflammation in RAW 264.7 cell culture system. Our data showed that glyceollins effectively inhibited NO production, IL-6 release, and COX-2 expression induced by LPS. In particular, glyceollins suppressed the LPS-induced phosphorylation of NF-B p65, suggesting that the compounds inhibit the production of NO and transcriptional activation of COX-2 by regulating NF-B activity. In another experiment we found that glyceollins enhanced the expression of heme oxygenase 1 and repressed the phosphorylation of ERK1/2 and p38 but not JNK in LPS-treated RAW 264.7 cells. Glyceollins also reduced TPA-induced skin inflammation in mouse model, confirming the anti-inflammatory activity of glyceollins in vivo system as well as cell culture system. Taken together, glyceollins merit further study as potential therapeutic agents for inflammatory disorders. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4243. doi:10.1158/1538-7445.AM2011-4243

Identification of the Rage-dependent gene regulatory network in a mouse model of skin inflammation

BackgroundIn the past, molecular mechanisms that drive the initiation of an inflammatory response have been studied intensively. However, corresponding mechanisms that sustain the expression of inflammatory response genes and hence contribute to the establishment of chronic disorders remain poorly understood. Recently, we provided genetic evidence that signaling via the receptor for advanced glycation end products (Rage) drives the strength and maintenance of an inflammatory reaction. In order to decipher the mode of Rage function on gene transcription levels during inflammation, we applied global gene expression profiling on time-resolved samples of mouse back skin, which had been treated with the phorbol ester TPA, a potent inducer of skin inflammation.ResultsRanking of TPA-regulated genes according to their time average mean and peak expression and superimposition of data sets from wild-type (wt) and Rage-deficient mice revealed that Rage signaling is not essential for initial changes in TPA-induced transcription, but absolutely required for sustained alterations in transcript levels. Next, we used a data set of differentially expressed genes between TPA-treated wt and Rage-deficient skin and performed computational analysis of their proximal promoter regions. We found a highly significant enrichment for several transcription factor binding sites (TFBS) leading to the prediction that corresponding transcription factors, such as Sp1, Tcfap2, E2f, Myc and Egr, are regulated by Rage signaling. Accordingly, we could confirm aberrant expression and regulation of members of the E2f protein family in epidermal keratinocytes of Rage-deficient mice.ConclusionsIn summary, our data support the model that engagement of Rage converts a transient cellular stimulation into sustained cellular dysfunction and highlight a novel role of the Rb-E2f pathway in Rage-dependent inflammation during pathological conditions.

Potential use of iontophoresis for transdermal delivery of NF-κB decoy oligonucleotides

Topical application of nuclear factor-κB (NF-κB) decoy appears to provide a novel therapeutic potency in the treatment of inflammation and atopic dermatitis. However, it is difficult to deliver NF-κB decoy oligonucleotides (ODN) into the skin by conventional methods based on passive diffusion because of its hydrophilicity and high molecular weight. In this study, we evaluated the in vitro transdermal delivery of fluorescein isothiocyanate (FITC)-NF-κB decoy ODN using a pulse depolarization (PDP) iontophoresis. In vitro iontophoretic experiments were performed on isolated C57BL/6 mice skin using a horizontal diffusion cell. The apparent flux values of FITC-NF-κB decoy ODN were enhanced with increasing the current density and NF-κB decoy ODN concentration by iontophoresis. Accumulation of FITC-NF-κB decoy ODN was observed at the epidermis and upper dermis by iontophoresis. In mouse model of skin inflammation, iontophoretic delivery of NF-κB decoy ODN significantly reduced the increase in ear thickness caused by phorbol ester as well as the protein and mRNA expression levels of tumor necrosis factor-α (TNF-α) in the mice ears. These results suggest that iontophoresis is a useful and promising enhancement technique for transdermal delivery of NF-κB decoy ODN.

Identification of TSLP-dependent mediators in a calcipotriol (Dovonex®) driven mouse model of skin inflammation (140.16)

Abstract The epithelial-derived cytokine thymic stromal lymphopoietin (TSLP) is sufficient to induce asthma or atopic dermatitis (AD)-like phenotypes when selectively overexpressed in transgenic mice. Although T and B cells have been reported to be dispensable for the TSLP-induced inflammation in these models, little is known about the downstream pathways or mediators involved in the inflammatory response driven by TSLP. Topical treatment with vitamin D3 (1α,25-(OH)2D3) or MC903, the low-calcemic analog of vitamin D3, has been shown to induce TSLP expression in epidermal keratinocytes. This induction resulted in an AD-like phenotype similar to that seen in TSLP transgenic mice. 0.005% calcipotriol (Dovonex®) is a synthetic, low-calcemic vitamin D3 analog which is FDA-approved for the treatment of psoriasis. Following topical application of Dovonex® onto mouse ears, TSLP mRNA expression increased over time. Concomitant increases in CCL17/TARC, CCL22/MDC, and IL-6 mRNA were also observed. These increases were dependent on TSLP as an antibody that blocked TSLP signaling abrogated the response. Similarly, a variety of TSLP-dependent protein analytes were increased in the skin following application of Dovonex®. Together these data identify TSLP-dependent mediators that may play a role in the development of skin inflammatory diseases such as atopic dermatitis.

318 Identification of TSLP-dependent mediators in a calcipotriol mouse model of skin inflammation

318 Identification of TSLP-dependent mediators in a calcipotriol mouse model of skin inflammation