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Background/Aim: Mesenchymal stem cells (MSCs) have been suggested as an alternative therapeutic option in atopic dermatitis. Palatine tonsils are lymphoepithelial tissue located around the oropharynx and have been proposed as one of the important alternative sources of MSCs. The purpose of this study was to evaluate the protective and therapeutic effects of tonsil-derived MSCs (TMSCs) in a 2,4-dinitrofluorobenzene (DNFB)-induced mouse model of atopic dermatitis (AD). Materials and Methods: The effect of TMSCs was evaluated in 20 C57BL/6J mice that were randomly divided into four groups (normal, DNFB-PBS, DNFB-TMSC7, and DNFB-TMSC16 group). TMSCs were subcutaneously injected into DNFB-sensitized mice on day 7 (DNFB-TMSC7 group) and day 16 (DNFB-TMSC16 group). Several parameters of inflammation were assessed. Results: Subcutaneously injected TMSCs significantly improved the inflammatory symptoms in a DNFB-induced AD model mice, particularly showing therapeutic effects rather than protective effects. TMSC treatment inhibited T-cell-mediated inflammatory responses by decreasing the levels of IL-6, IL-1β, TNF-α (Th1 cell marker), IL-4 (Th2 cell marker), and B-cell-mediated serum IgE. In contrast, TMSCs enhanced the anti-inflammatory cytokine TGF-β. Conclusion: In vitro and in vivo results suggest that TMSC treatment improved inflammatory skin lesions in the DNFB-induced AD mice model via immunomodulatory effects of the TMSCs. TMSCs inhibit T-cell and B-cell mediated responses, and enhance the anti-inflammatory responses. [ABSTRACT FROM AUTHOR]

The glutathione (GSH) conjugates, S-(1,2-dichlorovinyl)-glutathione (DCVG) and S-(1,2-dichlorovinyl)-L-cysteine (DCVC), have been implicated in kidney toxicity and kidney cancer from trichloroethylene (TCE) exposure. Considerable differences in blood and tissue levels of DCVG and DCVC have been reported, depending on whether HPLC/UV (High Performance Liquid Chromatography-Ultraviolet) or HPLC/MS (HPLC-Mass Spectrometry) was used. A side-by-side comparison of analytical results with HPLC/UV and HPLC/MS/MS (High Performance Liquid Chromatography-Tandem Mass Spectrometry) detection was undertaken to quantitatively compare estimates for DCVG and DCVG using rat and human tissues. For the HPLC method, DCVG and DCVC were initially derivatized with fluorodinitrobenzene (DNP). The results from the HPLC/UV method showed that derivatized-DCVC eluted at the solvent front and could not be quantified. Derivatized-DCVG, however, was quantified but significant interference was observed in all four control tissues (rat blood, liver, kidney; and human blood), resulting in average spike recoveries of 222–22,990%. In contrast, direct analysis of spiked tissues by HPLC/MS/MS resulted in recoveries of 82–127% and 89–117% for DCVG and DCVC, respectively. These differences in analytical results were further confirmed in tissues from TCE-treated rats, e.g. , DCVG levels in rat liver were 18,000 times higher by HPLC/UV as compared to HPLC/MS/MS. Fraction collection of the derivatized-DCVG peak (obtained with the HPLC-UV method), followed by peak identification via an HPLC/UV/Q-TOF/MS/MS method, identified DNP-derivatized endogenous glutamate as the primary interfering substance that contributed to and exaggerated recoveries of DCVG. Thus, estimates of DCVG based on the HPLC/UV methods are not reliable; they will over-estimate the formation of the GSH conjugates of TCE and will artifactually exaggerate the potential cancer risk in humans from TCE exposure. Therefore, it is recommended that any characterization of cancer risks from TCE exposure attributable to the GSH conjugates of TCE rely on results obtained with the more specific and reliable HPLC/MS/MS method. [ABSTRACT FROM AUTHOR]

The aim of this study is to determine whether AST2017-01 which consists of Rumex crispus and Cordyceps militaris would improve atopic dermatitis (AD). We analyzed anti-AD effects of AST2017-01 and chrysophanol, a bioactive compound of AST2017-01, using a 2,4-dinitrofluorobenzene-induced AD murine model. AST2017-01 and chrysophanol relieved clinical severity in AD-like skin lesions and significantly decreased scratching behavior. The thickness of epidermis and infiltration of inflammatory cells in AD-like skin lesions were reduced by AST2017-01 or chrysophanol. AST2017-01 and chrysophanol significantly suppressed the levels of histamine, immunoglobulin E, thymic stromal lymphopoietin (TSLP), interleukin (IL)-4, IL-6, and tumor necrosis factor-α in serum of AD mice. The protein levels of TSLP, intercellular adhesion molecule-1, and macrophage inflammatory protein 2 were significantly inhibited in the skin lesions. The mRNA expressions of TSLP, thymus and activation-regulated chemokine/CCL17, and C-C chemokine receptor 3 were inhibited in the skin lesions by AST2017-01 or chrysophanol. In addition, AST2017-01 and chrysophanol significantly suppressed the expressions and activities of caspase-1 in the skin lesions. Taken together, these results suggest that AST2017-01 has beneficial effects on AD and may be used as a health functional food in AD. [ABSTRACT FROM AUTHOR]

Abstract: Background: The interactive aggravation of pruritus and depression is well‐known, but an appropriate experimental model that could mimic this behavioral phenomenon is still lacking. Thus, a systematic animal behavioral investigation was carried out in this study. This will promote the research and treatment of pruritus and depression. Methods: The 2,4‐dinitrofluorobenzene (DNFB)‐induced chronic itch model was established to measure the depression index by forced swimming test (FST), tail suspension test (TST), and splash test (ST). The chronic unpredicted mild stress (CUMS)‐induced depression model was established to measure spontaneous itch and acute histamine or chloroquine‐induced itch behaviors. A depression and itch combining model was also established to measure the scratching and depression behaviors. The motor function of DNFB mice was analyzed by the rotarod test. Results: The scratching number, the immobility time in the FST and TST, and the grooming number in the ST test were all significantly increased in the chronic itch model. Mice receiving CUMS treatment showed significantly increased spontaneous scratching number, immobility time in the FST and TST tests, and grooming number in the ST. The combined model showed increased immobility time in FST and TST tests and increased grooming number in ST comparing to the depression model, and showed increased scratching number comparing to the chronic itch model. After histamine (His) or chloroquine (CQ) injection, the scratching numbers of CUMS mice were all significantly increased compared to those of His‐ and CQ‐control, respectively. Anti‐depression drug ketamine could significantly inhibit the depression‐like behaviors of CUMS mice, and simultaneously stopped the promoting effect on His‐induced acute itch. Conclusions: This study established an appropriate cross aggravation experimental mode and demonstrated that there is cross aggravation between pruritus and depression. The illumination of related mechanisms underlying this cross aggravation effect will provide theoretical basis for the prevention and treatment of depression and pruritus. [ABSTRACT FROM AUTHOR]

Background: Although immunosuppressants for therapy of atopic dermatitis (AD) are still being sought, proteasome inhibitors are also potential candidates for the treatment of AD. Proteasome inhibitors exert various effects by blocking proteasomal degradation and help regulate processes such as apoptosis induction via caspase-9, cell cycle progression via cyclins, NF-κB inactivation via IκB, and downregulation of antigen cross-presentation. The cells targeted by proteasome inhibitors are therefore activated cells undergoing proliferation or differentiation, and antigen-presenting cells carrying out protein degradation. Objectives: This study investigated the therapeutic effects and side effects of a proteasome inhibitor, MG132, on the treatment of AD. Methods: AD-like disease in NC/Nga mice housed under specific pathogen-free conditions was induced by repeated application of 2,4-dinitrofluorobenzene (DNFB). Disease progression was evaluated by inflammation score, histopathology, and serum IgE level, and the effects of systemic MG132 administration were investigated. The percentages and absolute numbers for each population of Th1, Th2, and Th17 cells in the axillary lymph nodes were analyzed by flow cytometry. Results: DNFB application increased the expression of a unique major histocompatibility complex class I mutant molecule D/Ldm7 in dendritic cells (DCs), and increased Th1 and Th17 cells in NC/Nga mice. In vivo MG132 administration to NC/Nga mice with DNFB-induced dermatitis reduced Th17 cells but maintained the level of Th1 cells, resulting in the alleviation of dermatitis lesions by decreasing both serum IgE hyperproduction and mast cell migration. To understand the mechanisms maintaining Th1 cell levels following in vivo MG132-administration, we focused on the role of proteasomes regulating D/Ldm7 expression. Interestingly, 20S proteasome activity was higher in NC/Nga DCs than in BALB/c DCs. In vitro MG132 administration partially increased D/Ldm7 expression in a dose-dependent manner during DC maturation, and induced IFN-γ production from autoreactive CD8+ T cells but not from CD4+ T cells following coculturing with D/Ldm7-upregulated DCs. Conclusion: Although MG132 administration temporarily alleviated AD pathogenesis in NC/Nga mice, prolonged MG132 treatment may result in immunopathogenesis leading to chronic AD due to its side effect of maintaining Th1 levels via autoreactive CD8+ T cells. [ABSTRACT FROM AUTHOR]

The study aimed to investigate the effect of berberine on allergic contact dermatitis (ACD) in rats and explore its underlying mechanisms. Firstly, ACD model was established by sensitizing and challenging with 2,4-dinitrofluorobenzene (DNFB) topically, and the rats were treated with berberine. Ear swelling was assessed, and cytokine, IgE, and histamine productions were measured. The ear biopsies were obtained for histology analysis. Additionally, rat peritoneal mast cells (RPMCs) were isolated for detection of microRNA-21 (miR-21) expression, mitogen-activated protein kinase (MAPK) signaling, and MC degranulation. Lastly, RPMCs were transfected with miR-21 mimic or miR-21 inhibitor to investigate the relationship between miR-21 and p38 pathway in MC. Our results showed that berberine significantly attenuated ear swelling in DNFB-induced ACD (ACD vs high dose of berberine 0.48 ± 0.03 vs. 0.33 ± 0.03 mm, P < 0.01), inhibited inflammatory cell infiltration (86 ± 5.16 vs. 58 ± 4.32 cells/mm2, P < 0.01), reduced MC recruitment (61 ± 4.07 vs. 39 ± 3.42 mast cells/mm2, P < 0.01), as well as decreased inflammatory cytokine, IgE, and histamine productions (all P < 0.05). Berberine treatment inhibited miR-21 expression, suppressed β-hexosaminidase and histamine release, and prevented p38 phosphorylation (all P < 0.05), which was abrogated by pretreatment with miR-21 overexpression. These findings indicate that miR-21-mediated inhibition of MC degranulation is involved in the anti-ACD effect of berberine via inhibiting p38 pathway, which provide a new insight into the immunopharmacological role of berberine and suggest its potential application for the treatment of allergic inflammation, such as ACD. [ABSTRACT FROM AUTHOR]

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The skin is an important site for local or systemic application of drugs. However, most of the drugs have poor permeability through the skin’s outermost layer, stratum corneum. The aim of this study was to develop a method to enable transdermal delivery of morin (3, 5, 7, 2, 4-pentahydroxyflavone), which is a poorly water-soluble drug with anti-inflammatory properties obtained from natural products. Morin phospholipid complex (MPC) was prepared and then loaded in Carbopol 940 hydrogel (MPC-gel), which can significantly increase the transdermal flux of morin based on the in vitro skin penetration data presented in this paper. To further enhance permeation, different compositions of penetration enhancers were dispersed in the gel and screened. After applied onto the mouse skin, MPC-gel showed apparent reduction of ear swelling in 2, 4-dinitrofluorobenzene (DNFB)-induced allergic contact dermatitis (ACD). Further determination of cytokines levels, histopathological analysis and T lymphocytes proliferation indicates that the MPC-gel is potent enough to reduce the inflammatory response mediated by the DNFB in ACD mice model. Collectively, we anticipate that such an approach may provide a new treatment for topical ACD. [ABSTRACT FROM AUTHOR]

Acupuncture has shown beneficial effect in the treatment of multiple dermatologic conditions including dermatitis, pruritus, urticaria, and hyperhidrosis; however, the detailed mechanisms are still kept unclear. This study aimed to investigate if electro-acupuncture (EA) treatment prevents 2,4-dinitrofluorobenzene (DNFB)-induced allergic contact dermatitis (ACD) in rats and explore its underlying mechanisms. ACD was induced by sensitizing and challenging with DNFB topically. Rats were treated daily following bilateral subcutaneous stimulation of EA at Zusanli acupoint (ST36) for 1 week. Ear swelling and serum IgE levels were measured. The ear biopsies were obtained for histology. Inflammatory cytokines on the dermatological ear and local acupoint tissue were assayed. Spleen lymphocytes and the homogenized supernatant of local acupuncture area were used to co-culture for flow cytology and immune analysis, respectively. EA treatment at ST36 notably inhibited ear swelling and inflammatory cell infiltration on DNFB-induced ACD. EA also decreased serum IgE concentrations and alleviated the production of inflammatory cytokines in dermatological ear. Additionally, EA treatment attenuated the percentage of CD4IFN-γ and CD4IL-4 T cells associated with ACD. Interestingly, secretion of interleukin (IL)-10 in the local acupoint tissue following EA stimulation was increased and showed suppressive function when co-cultured with the spleen lymphocytes from DNFB group. Lastly, EA treatment demonstrably suppressed p38 MAPK activation in DNFB-treated rats. Our findings suggest that EA treatment at ST36 may ameliorate inflammation associated with DNFB-induced ACD via triggering local IL-10 production and inhibiting p38 MAPK activation, which provide an alternative and promising therapy for ACD. [ABSTRACT FROM AUTHOR]

Background Angiotensin II (Ang II), the main peptide of the renin-angiotensin system ( RAS), has been suggested to be involved in inflammatory bowel diseases. Since RAS has emerged as gut motility regulator, and dysmotility is associated with intestinal inflammation, our objective was to investigate in rat 2,4-dinitrobenzenesulfonic acid ( DNBS)-induced colitis the functionality of RAS and its contribution to colonic motor alterations. Methods The effects of Ang II on the longitudinal colonic muscular contractility of control and DNBS-treated rats were characterized in vitro. Transcripts encoding for Ang II receptors were investigated by RT- PCR. Key Results Inflamed preparations showed a longitudinal muscle marked hypocontractility. Angiotensin II caused contractile effects in both preparations, but the responses in DNBS preparations were reduced compared to controls. In both preparations, Losartan, AT1 receptor antagonist, reduced Ang II effects. PD123319, AT2 receptor antagonist, enhanced Ang II responses only in DNBS rats, as well as Nω-Nitro-L-arginine (L- NNA), nitric oxide ( NO) synthase inhibitor, or tetrodotoxin ( TTX), neural toxin. The co-administration of PD123319 and TTX or L- NNA produced no additive effects. PD123319 per se improved colonic contractility in inflamed tissues. The effect was reduced in the presence of L- NNA or TTX. All Ang II receptor subtypes were expressed in both preparations. Conclusions & Inferences AT1 receptors mediate Ang II contractile responses in rat colon. During inflammation a recruitment of Ang II AT2 receptors would counteract AT1-contractile activity. A tonic activation of AT2 receptors would contribute to the general reduction in muscle contractility during experimental inflammation. A role for enteric neurons and NO is also suggested. [ABSTRACT FROM AUTHOR]

In mice, 2,4-dinitrofluorobenzene (DNFB) induces contact allergic dermatitis (CAD), which, in a late phase, is characterized by mast cell (MC) infiltration and angiogenesis. Palmitoylethanolamide (PEA), an endogenous anti-inflammatory molecule, acts by down-modulating MCs following activation of the cannabinoid CB 2 receptor and peroxisome proliferator-activated receptor-α (PPAR-α). We have previously reported the anti-inflammatory effect of PEA in the early stage of CAD. Here, we examined whether PEA reduces the features of the late stage of CAD including MC activation, angiogenesis and itching. After sensitization to DNFB, female C57BL/6J mice underwent to three DNFB challenges at days 5, 12 and 19 and treatments were given at each challenge and for two more days. CAD was expressed as Δ increase in ear thickness between challenged and un-challenged mice. PEA (5 mg/kg/i.p.) reduced: i) the DNFB-induced Δ increase; ii) the number of MCs per tissue area; iii) the expression of VEGF and its receptor Flk-1. These effects were reversed by co-administration of AM630 (1 mg/kg/i.p.), a CB 2 antagonist, but not GW6471 (1 mg/kg/i.p.), a PPAR-α antagonist. Finally, PEA reduced the number of ear scratchings 48 h after DNFB challenge and this effect was reversed by both CB2 and PPAR-α antagonists, suggesting the involvement of both receptors. PEA, by reducing the features of late stage CAD in mice, may be beneficial in this pathological condition. [ABSTRACT FROM AUTHOR]

The objective of the study was to investigate the possibility of modulation of skin inflammation by topical treatment with a novel compound: an opioid-neurotensin hybrid peptide PK20 encompassing endomorphin-2 analog and modified fragment of neurotensin (8–13). Contact sensitivity response was induced in mice by skin sensitization with dinitrofluorobenzene (DNFB) followed by topical hapten application on ears. Mice were treated locally with PK20 or pure cream 2 h after the challenge with DNFB. 2 and 24 h after hapten exposure, ear thickness was determined. Ears were collected for histology and homogenization. Supernatants were used for measurement of contents of cytokines and lipid peroxidation products. Treatment with PK20 reduced significantly the late phase of contact sensitivity response, which was revealed by ear thickness diminution and reduction of inflammatory cell infiltration. The average concentrations of IL-1α, MCP-1, TNF-α and thiobarbituric acid-reactive substances were significantly decreased in the ears treated with the chimera in comparison to the control cream treated ears in DNFB sensitized/DNFB challenged group. We found that PK20 topical treatment alleviates hypersensitivity responses triggered by DNFB challenge and usage of the hybrid peptide may be a novel therapeutic strategy in the treatment of chronic inflammatory diseases. However, the mechanism remains unclear and needs further investigation. [ABSTRACT FROM AUTHOR]

Cannabinoids have anti-inflammatory effects and can produce bronchodilation in the airways. We have investigated the effects of cannabinoids on tracheal hyperreactivity and airway inflammation in dinitrofluorobenzene (DNFB)-induced experimental non-atopic asthma in mice. 5-hydroxytryptamine (5-HT)-induced contraction response was enhanced while carbachol- and electrical field stimulation-induced contractions, and isoprenaline-induced relaxation responses were remained unchanged in DNFB group. The increased 5-HT-induced contractions were inhibited by incubation with either atropine or tetrodotoxin. DNFB application resulted in increased macrophage number in the bronchoalveolar lavage fluid (BALF). In vivo ACEA (CB 1 agonist) treatment prevented the increase in 5-HT contractions, while JWH133 (CB 2 agonist) had no effect. However, neither ACEA nor JWH133 prevented the increase in macrophage number in BALF. In vitro ACEA incubation also inhibited the increase in 5-HT contraction in DNFB group. These results show that cannabinoid CB 1 receptor agonist can prevent tracheal hyperreactivity to 5-HT in DNFB-induced non-atopic asthma in mice. [ABSTRACT FROM AUTHOR]

We examined the therapeutic effect of the extract fromSasa veitchiileaves (Sasa extract: SE) on the allergic skin inflammation model. SE derived from 0.141 g (low dose) or 0.562 g (high dose) of the leaves was administered orally for 15 days. High dose of SE suppressed ear swelling response induced by repeated 2,4-dinitrofluorobenzene (DNFB) paintings, and both low and high doses of SE reduced infiltration of immune cells into the ear dermis. Although the number of T cells, Treg, CD4+T cells and CD8+T cells in draining lymph nodes (dLNs) was not changed by SE, IFN-γ secretion from dLN induced by concanavalin A or 2,4-dinitrobenzenesulfonic acid was suppressed by a high dose of SE while IL-10 levels remained unchanged. These results suggest that SE suppresses allergic skin inflammation via reducing IFN-γ secreted from immune cells, especially T cells. [ABSTRACT FROM AUTHOR]

Four simple and sensitive spectrophotometric methods were developed for the determination of the sulfhydryl containing drug mesna (MSN). Methods I and II rely on nucleophilic aromatic substitution reactions using two UV tagging reagents namely: 4-chloro-7-nitrobenzo-2-oxa-1,3-diazole (NBD-Cl) for method I and 2,4-dinitrofluorobenzene (DNFB) for method II. Both reactions took place in alkaline buffered medium and the obtained yellowish products were measured at 414 and 332 nm for methods I and II, respectively. Methods III and IV are indirect spectrophotometric methods based on the suppressive effect of MSN on the absorption of two ternary complex systems which are composed of 1,10-phenanthroline, silver and eosin for method III and 1,10-phenanthroline, silver and bromopyrogallol red for method IV. The decrease in absorbance of the ternary complexes was measured at 547 and 635 nm for methods III and IV, respectively. All the experimental parameters affecting these reactions were carefully studied and optimized. The methods were validated as per the ICH guidelines. The methods were applicable in the linearity ranges 4–18 μg/mL for method I, 4–16 μg/mL for method II, 0.25–2.25 μg/mL for method III and 0.25–1.75 μg/mL for method IV. The proposed methods were successfully applied for the analysis of MSN in its commercial ampoules and no interference was encountered from the present excipients as indicated by the satisfactory percentage recoveries. The results obtained were in a good agreement with those obtained from a previously published method of the investigated drug. [ABSTRACT FROM AUTHOR]

The mature fruit of Kochia scoparia (L.) Schrad. is widely administered in China and Korea as a medicinal herb for treatment of skin diseases, diabetes mellitus and rheumatoid arthritis. The present study investigated the effects of methanol extracts of K. scoparia dried fruit (MEKS) on ear swelling, histopathological changes (such as epidermal acanthosis, spongiosis and immune cell infiltration) and cytokine production in 1-fluoro-2,4-dinitrofluorobenzene (DNFB)-induced contact dermatitis mice. Topical application of MEKS inhibited DNFB-induced ear thickness and weight increases, as well as DNFB-induced epidermal acanthosis, spongiosis and immune cell infiltration. In addition, treatment with MEKS significantly decreased the levels of tumor necrosis factor-a, interferon-γ and monocyte chemotactic protein-1 in inflamed tissues. These data indicate that the mature fruit of K. scoparia has the potential to be administered for the treatment of inflammatory skin diseases and that the anti-inflammatory action of K. scoparia is involved in the inhibition of type 1 T helper cell skewing reactions. [ABSTRACT FROM AUTHOR]

Atopic dermatitis (AD) is a common multifactorial chronic skin disease that has a multiple and complex pathogenesis. AD is gradually increasing in prevalence globally. In NC/Nga mice, repetitive applications of 2, 4-dinitrofluorobenzene (DNFB) evoke AD-like clinical symptoms similar to human AD. Aspartame (N-L-α-aspartyl-L-phenylalanine 1-methyl ester) is a methyl ester of a dipeptide, which is used as an artificial non-nutritive sweetener. Aspartame has analgesic and anti-inflammatory functions that are similar to the function of nonsteroidal anti-inflammatory drugs such as aspirin. We investigated whether aspartame can relieve AD-like clinical symptoms induced by DNFB treatment in NC/Nga mice. Sucrose did not relieve AD-like symptoms, whereas aspartame at doses of 0.5 μg kg−1 and 0.5 mg kg−1 inhibited ear swelling and relieved AD-like clinical symptoms. Aspartame inhibited infiltration of inflammatory cells including eosinophils, mast cells, and CD4+ T cells, and suppressed the expression of cytokines including IL-4 and IFN-γ, and total serum IgE levels. Aspartame may have therapeutic value in the treatment of AD. [ABSTRACT FROM AUTHOR]

Astaxanthin (AST) is known to exhibit antioxidative and antitumor properties, therefore, the present study investigated its other potential medical applications. AST was observed to exhibit anti-allergic and anti-inflammatory effects in a dinitrofluorobenzene (DNFB)-induced contact dermatitis (CD) mouse model and RBL-2H3 cell lines. The topical application of AST effectively inhibited the enlargement of ear thickness and increase in weight, which occurred following repeated application of DNFB. Furthermore, topical application of different concentrations of AST inhibited inflammatory hyperplasia, edema, spongiosis, and the infiltration of mononuclear cells and mast cells in the ear tissue. In addition, the levels of TNF-α and IFN-γ produced were decreased by application of AST in vivo, and treatment of RBL-2H3 cells with AST inhibited the release of histamine and β-hexosaminidase in vitro. Taken together, these data suggested that AST may be used to treat patients with allergic skin diseases through a mechanism, which may be associated with that involved in anti-inflammatory or anti-allergic activities. [ABSTRACT FROM AUTHOR]

UV radiation (UVR) induces DNA damage, leading to the accumulation of mutations in epidermal keratinocytes and immunosuppression, which contribute to the development of nonmelanoma skin cancer. We reported previously that the TLR4-MyD88 signaling axis is necessary for UV-induced apoptosis. In the dinitrofluorobenzene contact hypersensitivity model, UV-irradiated MyD88-deficient (MyD88-/-) C57BL/6 mice had intact ear swelling, exaggerated inflammation, and higher levels of dinitrofluorobenzene-specific IgG2a compared with wild-type (WT) mice. Even with normal UV-induced, dendritic cell migration, DNA damage in the local lymph nodes was less pronounced in MyD88-/- mice compared with WT mice. Cultured, UV-irradiated WT APCs showed cleavage (inactivation) of the DNA damage-recognition molecule PARP, whereas PARP persisted in MyD88-/- and TLR4-/- APCs. Epidermal DNA from in vivo UV-irradiated MyD88-/- mice had an increased resolution rate of cyclobutane pyrimidine dimers. Both in vitro treatment of MyD88-/- APCs with and intradermal in vivo injections of PARP inhibitor, PJ-34, caused WT-level cyclobutane pyrimidine dimer repair. Lymphoblasts deficient in DNA repair (derived from a xeroderma pigmentosum group A patient) failed to augment DNA repair after MyD88 knockdown after UVR, in contrast to lymphoblasts from a healthy control. These data suggest that interference with the TLR4/MyD88 pathway may be a useful tool in promoting DNA repair and maintaining immune responses following UVR-induced damage. [ABSTRACT FROM AUTHOR]

Of the biogenic polyamines, spermidine is a natural constituent of living cells and organisms. Spermidine is associated with regulation of cell growth, proliferation and differentiation, and with the suppression of oxidation and inflammation. Atopic dermatitis ( AD) is a chronic inflammatory skin disease that has a complex and multiple pathogenesis, which includes genetic abnormality, modified or abnormal immune response and the production of nitric oxide and reactive oxygen species. We investigated whether spermidine can relieve AD-like clinical manifestation induced by the continual application of 2,4-dinitrofluorobenzene ( DNFB) in NC⁄Nga mice. Spermidine at concentrations of 1 or 10 mg/kg reduced increasing ear swelling and attenuated oedema, haemorrhage and hyperkeratosis in AD-like skin lesions. Repetitive application of DNFB induced inflammatory cell infiltration to skin lesions, whereas intraperitoneal injection of spermidine inhibited DNFB-evoked infiltration of eosinophils, mast cells and T lymphocytes. Furthermore, spermidine suppressed mast cell degranulation and production of interferon-gamma by activated CD4+ T cells in AD-like skin lesions. Spermidine may be a potential therapeutic agent for treatment of AD. [ABSTRACT FROM AUTHOR]

The present study was conducted with the aim to investigate the immuno-modulatory and histological stabilization effects of nanocarrier–based transcutaneous co-delivery of hydrocortisone (HC) and hydroxytyrosol (HT). In this investigation, the clinical and pharmacological efficacies of nanoparticle (NP)-based formulation to alleviate 2,4-dinitrofluorobenzene (DNFB)-induced atopic dermatitis (AD) was explored by using an NC/Nga mouse model. Ex vivo visual examination of AD induction in experimental mice indicated remarkable control of NP-based formulations in reducing pathological severity of AD-like skin lesions. Therapeutic effectiveness of NP-based formulations was also evaluated by comparing skin thickness of AD-induced NP-treated mice (456±27 µm) with that of atopic mice (916±37 µm). Analysis of the immuno-spectrum of AD also revealed the dominance of NP-based formulations in restraining immunoglobulin-E (IgE), histamine, prostaglandin-E2 (PGE2), vascular endothelial growth factor-α (VEGF-α), and T-helper cells (TH1/TH2) producing cytokines in serum and skin biopsies of tested mice. These anti-AD data were further supported by histological findings that revealed alleviated pathological features, including collagen fiber deposition, fibroblasts infiltration, and fragmentation of elastic fibers in experimental mice. Thus, NP-mediated transcutaneous co-delivery of HC and HT can be considered as a promising therapy for managing immunological and histological spectra associated with AD. [ABSTRACT FROM AUTHOR]

Background: Morinda citrifolia (Rubiaceae), commonly known as noni is distributed throughout tropical and sub-tropical regions of the world. Anti-allergic effects of noni have not been reported despite the clinical usage as an anti-allergic agent. Materials and Methods: To investigate the anti-allergic effects of the 50% ethanolic extract of M. citrifolia fruits and leaves (MCF-ext and MCL-ext), dinitrofluorobenzene (DNFB)-induced triphasic cutaneous reaction and picryl chloride-induced contact dermatitis (PC-CD) tests were performed. Results: In DNFB-induced triphasic cutaneous reaction, oral administration of MCF-ext and MCL-ext exhibited dose-dependent inhibition of cutaneous reaction at 1 h (immediate phase response) after the DNFB challenge. MCF-ext also inhibited ear swelling at 24 h (late phase response) and 8 days (very late phase response) after the DNFB challenge. The effect of MCL-ext on the immediate phase response was attributed to the anti-degranulation from RBL-2H3 cells, while MCF-ext had no significant effect on degranulation. The active components of anti-degranulation activity in MCL-ext were determined to be ursolic acid, rutin and kaempferol-3-O-α-L-rhamnopyranosyl-(1→6)-β-D-glucopyranoside. In the PC-CD test, both MCF-ext and MCL-ext showed an anti-swelling effect but the potency of MCF-ext was stronger than MCL-ext. Conclusion: These data suggest that noni fruits and leaves can be a daily consumable material for the prevention of allergic symptoms. [ABSTRACT FROM AUTHOR]

Memantine is chemically a tricyclic amine and is used for Parkinson's disease and movement disorders. Although several HPLC methods with different derivatization reagents have been developed for the determination of memantine in biological fluids, there are some complications which limit the use of these methods in routine analysis of memantine in in vitro tests. We established a simple, sensitive, precise, and accurate HPLC method for the quantification of memantine in dosage forms. Pre-column derivatization of memantine was performed with 1-fluoro-2,4-dinitrobenzene and the reaction product was separated on a Nova-Pak C18 column. A mixture of acetonitrile and sodium dihydrogenphosphate (pH 2.5; 0.05 M) (70: 30, v/v) was used as the mobile phase. UV detection was performed at 360 nm. Forced degradation studies were performed on a powdered tablet sample of memantine hydro-chloride using acidic (0.1 M hydrochloric acid), basic (0.1 M sodium hydroxide), oxidative (10% hydrogen peroxide), thermal (105°C), photolytic, and humidity conditions. Good linearity (r2=0.999) was obtained over the range of 1-12 μg mL-1 of memantine hydrochloride with acceptable within-day and between-day precision values in the range of 0.05-0.95%. The proposed method was used for the assay determination and dissolution rate study of memantine dosage forms with excellent specificity. [ABSTRACT FROM AUTHOR]

Highlights: [•] Fisetin has various physiological effects such as anti-allergic and anti-inflammation. [•] Fisetin relieves DNFB-induced AD-like clinical symptoms in murine model. [•] Fisetin inhibited infiltration of inflammatory cells and suppressed the expressions of various inflammatory mediators. [•] Fisetin inhibits phosphorylation of NF-κB p65 which associated with inflammation. [•] These results implicate fisetin as a potential therapeutic for AD. [ABSTRACT FROM AUTHOR]

Background: The slough shed of Cryptotympana atrata Fabricius is widely used to treat skin diseases in China, Japan, and Korea. Objective: To investigate the anti-inflammatory effects of C. atrata on contact dermatitis. Materials and Methods: We investigated the effects of C. atrata methanol extract (MECA) on ear swelling, histophathological changes and cytokine production in 1-fluoro-2,4-dinitrofluorobenzene (DNFB)-induced contact dermatitis (CD) mice. Results: Topical application of MECA effectively inhibited enlargement of ear swelling (30 and 100 μ/ear, P < 0.05; 300 μg/ear, P < 0.01). MECA treatment also inhibited hyperplasia, spongiosis (100 and 300 μg/ear, P < 0.001), and immune cell infiltration (30 μg/ear, P < 0.05; 100 and 300 μg/ear, P < 0.001) induced by DNFB. In addition, treatment with MECA suppressed the increase in the levels of TNF-α (P < 0.05), IFN-γ (3, 100 μg/ear, P < 0.05; 300 μg/ear, P < 0.01), and IL-6 (100 μg/ear, P < 0.05; 300 μg/ear, P < 0.01) production. Conclusion: These data suggest that MECA has the potential for use in the treatment of inflammatory skin diseases, including CD. Moreover, the results presented herein indicate that anti-inflammatory actions of MECA are mediated by decreasing production of TNF-α, IFN-γ, and IL-6 in inflamed tissues. [ABSTRACT FROM AUTHOR]

We developed a new local lymph node assay (LLNA) that includes the elicitation phase termed LLNA:DAE for discrimination of borderline-positive chemicals as classified by the LLNA modified by Daicel based on ATP content (LLNA:DA) and for cross-sensitization testing. Although the LLNA:DA method could help identify skin sensitizers, some skin irritants classified as non-sensitizers by the LLNA were classified as borderline positive. In addition, the evaluation for the cross-sensitization potential between chemicals was impossible. In the LLNA:DAE procedure, test group of mice received four applications of chemicals on the dorsum of the right ear for induction and one application on the dorsum of the left ear for elicitation. Control group of mice received one chemical application on the dorsum of the left ear. We evaluated the sensitizing potential by comparing the weights of the lymph nodes from the left ears between the two groups. The results of using the LLNA:DAE method to examine 24 chemicals, which contained borderline-positive chemicals, were consistent with those from the LLNA method, except for nickel chloride (NiCl2). Two chemical pairs, 2,4-dinitrochlorobenzene (DNCB) with 2,4-dinitrofluorobenzene (DNFB) and hydroquinone (HQ) with p-benzoquinone (p-BQ), showed clear cross-sensitization with each other, while another chemical pair, DNFB with hexylcinnamic aldehyde (HCA) did not. Taken together, our results suggest that the LLNA:DAE method is useful for discriminating borderline-positive chemicals and for determining chemical cross-sensitization. [ABSTRACT FROM AUTHOR]

In this study, a sensitive, simple, and reliable HPLC method for quantification of pregabalin in human plasma was developed and validated. 1-Fluoro-2,4-dinitrobenzene was used as precolumn derivatization agent. For chromatography, an analytical reversed phase (C18) column and a mixture of Na2HPO410 mM (pH 8.0)--methanol (35: 65 v/v) were used as stationary and mobile phase, respectively. Detection was performed using a UV detector tuned at 360 nm. The linearity of the method was tested over the concentration range 1-4500 ng/mL in 500 μL of human plasma and satisfactory results were obtained (r² > 0.999). The method showed good precision and accuracy in terms of within--between days relative standard deviations and percent deviation from nominated values (in the range of 4.3-12.7% and 2.6-8.0%, resp.). The limit of quantification of the method was found to be 1 ng/mL which is better than previously reported method and indicates its potential application for sensitive bioanalysis. [ABSTRACT FROM AUTHOR]

Background: Previous studies have indicated that colitis increases intestinal permeability to food antigens. This condition also generates an immunoreactive milieu in the gut, which may exacerbate or counteract allergy reactions. This, along with the fact that both colitis and allergy are being codiagnosed more frequently, means the scientific interest on the immune relation between these pathologies is increasing. We evaluated the immune response to an internalized food antigen that was initiated during a concomitant active intestinal inflammatory response. Methods: An ovalbumin (OVA)-induced immune response was analyzed in healthy mice and in mice suffering from colitis induced by the administration of dinitrofluorobenzene/dinitrosulfonic acid (DNFB/DNS) at the moment of OVA challenge. The OVA-induced clinical score and allergy response both in plasma and in splenocyte cultures from these animals were compared. Results: Although no differences were observed in the allergy clinical score, the concomitant active colitis led to an increase in the immune response to OVA antigen, as shown by increased spleen size and OVA-induced splenocyte proliferation, exacerbated expression of total and OVA-specific IgG1 levels, increased colonic IL-4 expression and OVA-induced IL-4 and IL-5 cytokine expression in spleen cells. Conclusions: Our results indicate that animals with active colitis undergo an exacerbated immune response to an internalized antigen. This finding could be relevant for the allergy management of patients presenting simultaneously with chronic colitis. © 2013 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Allergic diseases are chronic inflammatory conditions with specific immune and inflammatory mechanisms. Scientific interest in understanding the mechanisms and discovering novel agents for the prevention and treatment of allergic disease is increasing. Streptochlorin, a small compound derived from marine actinomycete possesses anti-angiogenic and anti-tumor activities. However, the anti-allergic effects and underlying mechanisms remain to be elucidated. In the present study, we investigated the effect of streptochlorin on allergic responses in vitro and in vivo. Streptochlorin inhibited degranulation and production of tumor necrosis factor-α and IL-4 by antigen-stimulated mast cells. Streptochlorin also inhibited the phosphorylation of Akt and the mitogen-activated protein kinases (MAPKs), including p38, ERK, and JNK. Further, streptochlorin reduced the phosphorylation of Syk in RBL-2H3 cells and inhibited the activity of Lyn and Fyn. Furthermore, administration of streptochlorin suppressed the allergic reactions in both passive cutaneous anaphylaxis reaction and 2, 4-dinitrofluorobenzene (DNFB)-induced allergic dermatitis in mice model. Considering the data obtained, we report for the first time that streptochlorin possess anti-allergic properties. The underlying mechanism of streptochlorin in exhibiting potent anti-allergic activity might be through the inhibition of the Lyn/Fyn and Syk signaling pathways. [ABSTRACT FROM AUTHOR]

Background ∆9-Tetrahydrocannabinol ( THC), the active constituent of Cannabis sativa, exerts its biological effects in part through the G-protein-coupled CB1 and CB2 receptors, which were initially discovered in brain and spleen tissue, respectively. However, THC also has CB1/2 receptor-independent effects. Because of its immune-inhibitory potential, THC and related cannabinoids are being considered for the treatment of inflammatory skin diseases. Here we investigated the mechanism of the anti-inflammatory activity of THC and the role of CB1 and CB2 receptors. Methods We evaluated the impact of topically applied THC on DNFB-mediated allergic contact dermatitis in wild-type and CB1/2 receptor-deficient mice. We performed immunohistochemical analyses for infiltrating immune cells and studied the influence of THC on the interaction between T cells, keratinocytes and myeloid immune cells in vitro. Results Topical THC application effectively decreased contact allergic ear swelling and myeloid immune cell infiltration not only in wild-type but also in CB1/2 receptor-deficient mice. We found that THC (1) inhibited the production of IFNγ by T cells, (2) decreased the production of CCL2 and of IFNγ-induced CCL8 and CXL10 by epidermal keratinocytes and (3) thereby limited the recruitment of myeloid immune cells in vitro in a CB1/2 receptor-independent manner. Conclusions Topically applied THC can effectively attenuate contact allergic inflammation by decreasing keratinocyte-derived pro-inflammatory mediators that orchestrate myeloid immune cell infiltration independent of CB1/2 receptors. This has important implications for the future development of strategies to harness cannabinoids for the treatment of inflammatory skin diseases. [ABSTRACT FROM AUTHOR]

Contact hypersensitivity (CHS) mouse model induced by 2, 4-dinitrofluorobenzene (DNFB) is thought to be a T helper 1 (Th1)-dominant response and used for investigating anti-inflammatory and immunosuppressive agents. However, it is hardly used for screening large-scale drugs because of the large number of animals and complex mechanisms involved in-vivo. In this study, we focused on whether T lymphocytes from CHS mouse model could maintain the state of immune response in-vitro and explored a suitable time for drugs screening. The results showed that CD4+ T cells of CHS mice were higher compared with those in normal group. The expression of T-bet and GATA3 showed a Th1 shift and the levels of interleukin (IL)-2 and IL-4 also showed similar trend. Furthermore, IL-6 produced by T lymphocytes from CHS mice had a high level too. Then, we detected the effects of dexamethasone (DEX), cyclosporine A (CsA) and mycophenolate mofetil (MMF) on T lymphocytes in-vitro, and the data displayed that these immunosuppressive drugs could all inhibit the proliferation of T lymphocytes significantly. These findings suggested that T lymphocytes from CHS mice could mimic a similar immune response in-vitro, and it's also a suitable method for screening antiinflammatory and immunosuppressive agents. [ABSTRACT FROM AUTHOR]

Sensitization and challenge using dinitrofluorobenzene (DNFB) induce contact hypersensitivity (CHS) with Th1 cell infiltration, whereas those using FITC generate CHS with Th2 cell infiltration. In this study, we attempted to determine the role of CXCR3, a chemokine receptor, in Th1- and Th2-type CHS induced by DNFB or FITC using CXCR3-deficient (CXCR3-/-) mice. Ear swelling was prolonged after DNFB challenge in CXCR3-/- mice, which was accompanied by increased Th1 cytokines and decreased TGF-β and IL-10 expression at a late time point of CHS, whereas there was no significant difference between wild-type and CXCR3-/- mice in FITC-induced CHS. In Th1-type CHS, the number of regulatory T cells (Tregs) was decreased in the challenged ear of CXCR3-/- mice compared with that of wild-type mice, suggesting that CXCR3 would be important in migration of Tregs into the site of inflammation. Moreover, we examined the characteristics of CXCR3+ Tregs both in vitro and in vivo, revealing that CXCR3+ Tregs expressed high levels of TGF-β and IL-10 as well as IFN-? compared with CXCR3- Tregs. When CXCR3-/- mice were injected with CXCR3+ Tregs, the prolonged ear swelling induced by DNFB was normalized. Taken together, our results suggest that CXCR3+ Tregs play a key role for quenching Th1-type CHS. [ABSTRACT FROM AUTHOR]

Epidermal keratinocytes (KCs) and cannabinoid (CB) receptors both participate in the regulation of inflammatory responses in a mouse model for allergic contact dermatitis, the contact hypersensitivity (CHS) response to the obligate sensitizer 2,4-dinitrofluorobenzene. In this study, we investigated the cellular and molecular mechanisms how CB1 receptors attenuate CHS responses to 2,4-dinitrofluorobenzene. We used a conditional gene-targeting approach to identify the relative contribution of CB1 receptors on epidermal KCs for the control of CHS responses. To determine the underlying cellular and molecular mechanisms that regulate inflammatory responses in the effector phase of CHS, we performed further investigations on inflamed ear tissue and primary KC cultures using morphologic, molecular, and immunologic methods. Mice with a KC-specific deletion of CB1 receptors developed increased and prolonged CHS responses. These were associated with enhanced reactive epidermal acanthosis and inflammatory KC hyperproliferation in the effector phase of CHS. In vitro, primary cultures of CB1 receptor-deficient KC released increased amounts of CXCL10 and CCL8 after stimulation with IFN-γ compared with controls. In vivo, contact allergic ear tissue of CB1 receptor-deficient KCs showed enhanced expression of CXCL10 and CCL8 compared with controls. Further investigations established CCL8 as a proinflammatory chemokine regulated by CB1 receptors that promotes immune cell recruitment to allergen-challenged skin. Taken together, these results demonstrate that CB1 receptors are functionally expressed by KCs in vivo and help to limit the secretion of proinflammatory chemokines that regulate T cell-dependent inflammation in the effector phase of CHS. [ABSTRACT FROM AUTHOR]

Abstract: Silymarin inhibits UVB-induced immunosuppression in mouse skin. To identify the molecular mechanisms underlying this effect, we used an adoptive transfer approach in which dendritic cells (DCs) from the draining lymph nodes of donor mice that had been UVB-exposed and sensitized to 2,4,-dinitrofluorobenzene (DNFB) were transferred into naïve recipient mice. The contact hypersensitivity (CHS) response of the recipient mice to DNFB was then measured. When DCs were obtained from UVB-exposed donor mice that were not treated with silymarin, the CHS response was suppressed confirming the role of DCs in the UVB-induced immunosuppression. Silymarin treatment of UVB-exposed donor mice relieved this suppression of the CHS response in the recipients. Silymarin treatment was associated with rapid repair of UVB-induced cyclobutane pyrimidine dimers (CPDs) in DCs and silymarin treatment did not prevent UV-induced immunosuppression in XPA-deficient mice which are unable to repair UV-induced DNA damage. The CHS response in mice receiving DCs from silymarin-treated UV-exposed donor mice also was associated with enhanced secretion of Th1-type cytokines and stimulation of T cells. Adoptive transfer of T cells revealed that transfer of either CD8+ or CD4+ cells from silymarin-treated, UVB-exposed donors resulted in enhancement of the CHS response. Cell culture study showed enhanced secretion of IL-2 and IFNγ by CD8+ T cells, and reduced secretion of Th2 cytokines by CD4+ T cells, obtained from silymarin-treated UVB-exposed mice. These data suggest that DNA repair-dependent functional activation of DCs, a reduction in CD4+ regulatory T-cell activity, and stimulation of CD8+ effector T cells contribute to silymarin-mediated inhibition of UVB-induced immunosuppression. [Copyright &y& Elsevier]

Cytotoxic T lymphocyte-associated antigen-4 ( CTLA-4)-immunoglobulin ( Ig) has immunosuppressive properties both in vivo and in vitro, but much is still unknown about the mechanisms by which CTLA-4- Ig exerts its immunosuppressive activities in vivo. The aim of this study was to investigate the effect of CTLA-4- Ig in a mouse model of contact hypersensitivity ( CHS). The inflammatory response in the presence or absence of CTLA-4- Ig was evaluated by measuring the increase in ear thickness in sensitized animals after challenge. We observed a dose-dependent suppression of the ear swelling in both dinitrofluorobenzene ( DNFB)- and oxazolone-induced CHS. The suppressive effect was still present 3 weeks after administration, even in the absence of circulating levels of CTLA-4- Ig. It was further shown that CTLA-4- Ig inhibits activation of T cells in the draining lymph node after sensitization and affects the maturation level of both dendritic cells and B cells. Furthermore, CTLA-4- Ig reduces infiltration of activated CD8+ T cells into the inflamed ear tissue and suppresses both local and systemic inflammation, as illustrated by reduced expression of cytokines and chemokines in the inflamed ear and a reduced level of acute-phase proteins in circulation. Finally, our results suggest that CTLA-4- Ig has a mainly immunosuppressive effect during the sensitization phase. We conclude that CTLA-4- Ig induces long-term immunosuppression of both DNFB- and oxazolone-induced inflammation and our data are the first to compare the effect of this compound in both DNFB- and oxazolone-induced CHS and to show that CTLA-4- Ig exerts an immunosuppressive effect on both local and systemic inflammatory mediators which is mediated principally during the sensitization phase. [ABSTRACT FROM AUTHOR]

Abstract: In our previous study, it was found that linoleoyl ethanolamide (LE) is present in sake lees, which are produced as a byproduct during the making of Japanese sake. LE is a fatty acid ethanolamide, which have been demonstrated to exert a variety of biological functions, and in this study, the anti-inflammatory effects of LE were examined using in vitro cell culture and in vivo animal experiments. In mouse RAW264.7 macrophages, LE suppressed the lipopolysaccharide (LPS)-induced expression of pro-inflammatory cytokines, such as tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, and IL-6. In addition, LE inhibited LPS-induced increases in the levels of cyclooxygenase enzyme-2 and prostaglandin E2, which are indicators of inflammation. The inhibitory effect of LE on the release of TNF-α was stronger than that of dipotassium glycyrrhizinate, which is widely used in external human skin care treatments. LE also suppressed the LPS-induced activation of Toll-like receptor 4 signaling and nuclear translocation of nuclear factor-κB (NF-κB) p65. In a contact dermatitis animal model, applying LE to affected ear skin ameliorated 2,4-dinitrofluorobenzene-induced contact dermatitis and pro-inflammatory cytokine expression at inflamed sites. These results indicate that LE exerts anti-inflammatory effects by inhibiting NF-κB signaling, and LE is proposed to be a useful therapeutic agent against contact dermatitis. [Copyright &y& Elsevier]

With 2,4-dinitrofluorobenzene–acetonitrile pre-column derivatization, taurine and its intermediate (2-aminoethyl hydrogen sulfate) in beverages and milk samples were determined by high-performance liquid chromatography coupled with diode array detection. Chromatographic separation was performed by isocratic reverse phase model with acetonitrile-phosphate solution (pH 7.0) (19: 81, v/v) as the mobile phase on a C18column. By precipitation protein and filter, the sample could be directly used for precolumn derivatization and detection. Good linearity was observed in a range of 0.4–500 mg/L for all analytes with the correlation coefficients (r2) ≥ 0.9991. Overall recoveries of the two compounds ranged from 82% to 103% with repeatability (RSD) less than 4.6% at three different spiked levels (5 to 200 mg/L, n = 5). The proposed method offered a good alternative for routine analysis due to its simplicity and at the same time reliability. [ABSTRACT FROM AUTHOR]

Abstract: Atopic dermatitis (AD) is a common inflammatory skin disease for which few effective treatments are available. Resolvin E1 (RvE1; 5S,12R,18R-trihydroxy-6Z,8E,10E,14Z,16E-eicosapentaenoic acid) is an endogenous lipid mediator derived from omega-3 fatty eicosapentaenoic acid, which is a potent inhibitor of inflammation. AD-like skin lesion was induced by repetitive skin contact with DNFB in NC/Nga mice and the effects of RvE1 were evaluated on the basis of histopathological findings of skin, ear swelling and cytokine production of CD4+ T cells. Intraperitoneal injection of RvE1 for one week after DNFB challenge significantly lowered ear swelling and improved back skin lesions. In addition, RvE1 significantly suppressed production of interferon-gamma (IFN-γ) and interleukin-4 (IL-4) by activated CD4+ T cells and serum IgE level. Furthermore, RvE1 reduced DNFB-induced infiltration of eosinophils, mast cells, CD4+ T cells, and CD8+ T cells in skin lesions. Therefore, RvE1 may suppress the development of AD-like skin lesions in DNFB-treated NC/Nga mice by reducing IL-4 and IFN-γ of activated CD4+ T cells and serum IgE levels and infiltration of immune cells to skin lesion. [Copyright &y& Elsevier]

Abstract: Ethnopharmacological relevance: The dried root of Sophora flavescens Aiton (Sophorae radix, SR) has long been used in traditional medicine for the treatment of fever and swelling in eastern countries. Materials and methods: The present study investigated the anti-allergic and anti-inflammatory effects of SR using 1-fluoro-2,4-dinitrofluorobenzene (DNFB)-induced contact dermatitis mouse model and in vitro using RBL-2H3 cells. Results: In mice, the topical application of 10mg/mL of SR effectively inhibited enlargement of ear thickness and weight induced by repeated painting with DNFB. Topical application of SR also inhibited hyperplasia, edema, spongiosis and infiltration of mononuclear cells in ear tissue. In addition, production levels of interferon-gamma and tumor necrosis factor-alpha were decreased by SR in vivo. Finally, the release of histamine and β-hexosaminidase, and migration were inhibited by treatment with SR. Conclusions: These data indicate the potential of SR in treating patients with allergic skin diseases and also suggest that related mechanisms are involved in anti-inflammatory action on the Th 1 skewing reaction and inhibition against recruitment and degranulation of mast cells. [Copyright &y& Elsevier]

Abstract: Contact hypersensitivity (CHS) is a delayed-type hypersensitivity reaction which is mediated by hapten-specific T cells. Strong haptens, such as 2, 4-dinitrofluorobenzene (DNFB) can induce it. Grape seed proanthocyanidins extract (GSPs), which is an antioxidant derived from grape seeds, has been reported to possess a variety of potent properties. However, few reports demonstrated the effects of GSPs on contact hypersensitivity. Therefore, the present study was devised to describe the role of GSPs on a mouse model of experimental CHS induced by DNFB and try to explore the possible underlying mechanisms. We observed that, GSPs when orally administrated into the CHS mice, inhibited the aggravation of inflammation. After administration of GSPs, there was obvious fewer inflammatory cell infiltration in the inflamed ears. Ear swelling after challenge was significantly reduced. In addition, we investigated the effects of GSPs on T cells in vitro, which play critical role during the progress of CHS. It was found that GSPs inhibited proliferative activity of T cells by blocking the activation of mitogen-activated protein kinases (MAPK) and NF-кB signaling pathways. Collectively, these results showed that GSPs has protective effect on CHS induced by DNFB and it also could inhibit the proliferation ability of T cells in vitro, suggesting the potential of GSPs as new and effective compound for the treatment of T-cell mediated inflammatory diseases. [Copyright &y& Elsevier]

Abstract: Extrusion of artificially contaminated food is reported to degrade aflatoxins to varying degrees depending on the extrusion conditions. This work sought to determine the (1) efficacy of extrusion cooking in destroying naturally contaminated peanuts and (2) nutritional quality of decontaminated peanut meal. Naturally contaminated peanut meal was extruded by varying the moisture (20, 28, 35g/100g), pH (7.5, 9.5) and extruder die diameter (2.5, 3, 3.5, 4.0mm). Aflatoxins levels in the extrudates were determined using HPLC procedures, and the nutritional quality was assessed using in-vitro methods. The highest aflatoxin reduction in naturally contaminated peanut meal was 59% at feed moisture content of 35g/100g. Higher (91%) reduction was achieved in the artificially contaminated peanut meal at moisture of 20g/100g. In-vitro protein digestibility and Fluorodinitrobenzene (FDNB)-available lysine of the extrudates were not significantly different from non-extruded peanut meal. Extrusion conditions for aflatoxin reduction did not adversely affect protein nutritional quality. [Copyright &y& Elsevier]

The article offers information on a study conducted on spectrophotometric determination of some central nervous system (CNS) stimulants in spike human urine and dosage form, through derivatization with 2,4-Dinitrodluorobenzene. It states that shimadzu UV-Visible 1601 PC spectrophotometer was used for spectrophotometric measurements. It mentions several reagents used, including Phenylpropanolamine Hydrogen Chloride(HCl), borate buffer solution, and methanol, hydrochloric acid, and boric acid. It presents that the method showed simple and suitable technique to quantify the drugs, that may get employed for quality control analysis.

A validated, selective, and sensitive chromatographic method for determination of γ-aminobutyric acid (GABA) has been developed by precolumn derivative of the sample with 2,4-dinitrofluorodinitrobenzene (FDNB). The derivatives were separated with RP-HPLC on a C18 column and UV detection (360 nm). The calibration curve was linear over a range of 0.2-5.0 mg/mL GABA with a correlation coefficient of 0.9954. The precision of this method is high (RSD = 0.061%). This method also possessed high recovery rate (>99%) and good stability over a period of four weeks. The method was successfully used to determine the glutamate decarboxylase (GAD) activity in several plant tissues. [ABSTRACT FROM AUTHOR]

Abstract: Taking into account the structural similarities of amino alcohols with amino acids and in order to reduce time for derivatization unconventional approaches viz. microwave irradiation, ultrasonication and ultra centrifugation were applied for synthesis of dinitrophenyl derivatives of nine amino alcohols and to work out a method of choice for sample preparation for direct enantioseparation. The enantiomeric dinitrophenyl derivatives so synthesized were separated on α1-acid glycoprotein and β-cyclodextrin columns with detection at 230nm using photodiode array detection system. Derivatization methods and chromatographic parameters were optimized. β-Cyclodextrin column was found better compared to AGP column for enantioseparation. Limit of detection, quantification, accuracy and precision were also determined. The developed method was successfully applied to determine enantiomeric purity of 1-amino-2-propanol obtained from vitamin B12 hydrolysate. [Copyright &y& Elsevier]

The effect of moist and dry heat processing on lysine content of selected foods was investigated. Two moist heat (open and closed vessel) and dry heat (microwave and hot air oven) methods were chosen for processing samples from cereal, oilseed and pulse groups with whey protein concentrate as reference sample. Unheated samples served as controls. The samples were analysed for protein content, available lysine by fluorodinitrobenzene method and for in vitro protein digestibility (IVPD). Results indicated that in moist heat-treated samples lysine retention was between 69 – 83%. Dry heat treatment resulted in a greater loss of lysine, which was proportional to the degree of heating. The retention of lysine in dry heat-treated samples ranged from 31 to 65 %. All heated samples had higher digestibility than their raw controls. In between methods of heating, the IVPD of dry heat-treated samples were slightly lower than moist heat-treated samples. It can be concluded that heat treatments increase the in vitro digestibility of proteins but lysine content was affected adversely. [ABSTRACT FROM AUTHOR]

Summary: Bacterial virus entry and cell wall depolymerization require the breakdown of peptidoglycan (PG), the peptide-cross-linked polysaccharide matrix that surrounds bacterial cells. Structural studies of lysostaphin, a PG lytic enzyme (autolysin), have suggested that residues in the active site facilitate hydrolysis, but a clear mechanism for this reaction has remained unsolved. The active-site residues and a structural pattern of β-sheets are conserved among lysostaphin homologs (such as LytM of Staphylococcus aureus) and the C-terminal domain of gene product 13 (gp13), a protein at the tail tip of the Bacillus subtilis bacteriophage ϕ29. gp13 activity on PG and muropeptides was assayed using high-performance liquid chromatography, and gp13 was found to be a d,d-endopeptidase that cleaved the peptide cross-link. Computational modeling of the B. subtilis cross-linked peptide into the gp13 active site suggested that Asp195 may facilitate scissile-bond activation and that His247 is oriented to mediate nucleophile generation. To our knowledge, this is the first model of a Zn2 + metallopeptidase and its substrate. Residue Asp195 of gp13 was found to be critical for Zn2 + binding and catalysis by substitution mutagenesis with Ala or Cys. Circular dichroism and particle-induced X-ray emission spectroscopy showed that the general protein folding and Zn2 + binding were maintained in the Cys mutant but reduced in the Ala mutant. These findings together support a model in which the Asp195 and His247 in gp13 and homologous residues in the LytM and lysostaphin active sites facilitate hydrolysis of the peptide substrate that cross-links PG. Thus, these autolysins and phage-entry enzymes have a shared chemical mechanism of action. [Copyright &y& Elsevier]

Heme oxygenase (HO)-1 may have an important role in the resolution of T cell-mediated inflammation. The authors elucidated the role of the anti-inflammatory HO-1 in the pathogenesis of skin inflammation, using a mouse contact hypersensitivity (CHS) induced by 2,4-dinitrofluorobenzene (DNFB). Ear swelling was induced by challenge with DNFB, accompanied by infiltration of inflammatory cells in the challenged ear skin. DNFB challenge induced low levels of HO-1 mRNA and protein expression. Ear swelling induced by DNFB challenge was significantly reduced by topical treatment with cobalt protoporphyrin IX (CoPP), a HO-1 inducer, but exaggerated by blockage of HO-1 activity with tin protoporphyrin IX (SnPP), a HO-1 inhibitor. Similarly, the number of infiltrated cells in DNFB-challenged ear skin were reduced by CoPP but increased by SnPP. Our findings suggest that HO-1 plays an important role in CHS and is an important pharmacological target for the treatment of CHS. [ABSTRACT FROM AUTHOR]

Abstract: Six new functionalized oxacalix[4]arene porphyrins have been synthesized via a high-yielding ‘3+1’ condensation between meso-(3,5-dihydroxyphenyl)triphenylporphyrin and readily available new fluorodinitrobenzene-containing trimers. The X-ray structure of one linear trimer is presented. The synthesis of a porphyrin containing two oxacalix[4]arene moieties is also reported using a similar strategy. 1H NMR data and computer calculations using the AM1 semiempirical method incorporated into the Spartan program indicate that the oxacalix[4]arene porphyrins adopt 1,3-alternating conformations. The photophysical properties of the oxacalix[4]arene porphyrins were investigated. [Copyright &y& Elsevier]

In the present study, the effect of iron-deficiency anemia on delayed cutaneous hypersensitivity was measured using weanling C57BL/6 female mice which were fed either an ad libitum control diet supplemented with 25 to 30 mg Fe/kg diet (FePO4), an iron-deficient test diet (5 to 6 mg Fe/kg diet), or a pairfed control diet (25 to 30 mg Fe/kg diet). When skin sensitizing agent (dinitrofluorobenzene) was applied to these animals and skin responses were measured 3 to 5 days later, anemic mice showed a significantly decreased inflammatory skin respone than either control or pairfed mice. Five days after sensitization, the animals were challenged with dinitrofluorobenzene painted on the right ear and an equal dose of only the solvent on the left ear followed by 125I-deoxyuridine injected intraperitoneally. The ratio of either total or DNA associated radioactivity incorporated into the right over the left ears was significantly lower in anemic mice than either control or pairfed mice. A single dose of Imferon injected 24 h before the recall dose of dinitrofluorobenzene restored the ratio of 125I-dUR incorporated in anemic mice without having any significant effect on either the control or pairfed groups. The results suggest that iron is not required for sensitization but is required for an effective inflammatory response. [ABSTRACT FROM AUTHOR]

Seventeen test foods were each analyzed by four methods. Total lysine was measured by conventional amino acid analysis. Reactive lysine was measured with either fluorodinitrobenzene, o-phthalaldehyde or a differential dye-binding procedure. The results were then compared with another group's results from rat growth assays of the same samples for availably lysine. A sample of deliberately heat-damaged milk powder gave a rat assay value corresponding to 64% of its total lysine content; other values were all higher and on average 99% for 7 animal products, and 87% for 9 vegetable products. The correlation coefficient between the two sets of values was 0.95. The 'reactive lysine' procedures failed to give a better prediction of the rat values. [ABSTRACT FROM AUTHOR]