丹参凝胶治疗对特应性皮炎小鼠模型皮肤屏障功能、表皮增生以及免疫功能的影响

摘要 目的：研究丹参凝胶治疗对特应性皮炎小鼠模型皮肤屏障功能、表皮增生以及免疫功能的影响。方法：30只C57BL/6小鼠被随机分为Control组、AD组和SG组,每组10只。AD组和SG组背部涂抹对二硝基氟苯建立特应性皮炎小鼠模型,SG小鼠在模型建立成功后涂抹丹参凝胶治疗3周,Control组和AD组涂抹凡士林作为对照。3周后,测量所有小鼠经皮水分丢失量( TEWL) 、皮肤厚度、脾脏指数、胸腺指数,血清IgE、IFN-γ和IL-4,脾脏树突状细胞、Th1和Th2细胞比例。结果：丹参凝胶治疗3周后,AD组和SG组小鼠TEWL、皮肤厚度、脾脏指数、胸腺指数,血清IgE、IFN-γ和IL-4含量,以及脾脏Th2细胞比例均显著高于对照组正常小鼠（P<0.05）,而脾脏树突状细胞、Th1细胞和Th1/Th2细胞比例均显著低于对照组正常小鼠（P<0.05）;与AD组小鼠相比,SG组小鼠TEWL、皮肤厚度、脾脏指数、胸腺指数,血清IgE、IFN-γ和IL-4含量,以及脾脏Th2细胞比例均显著降低（P<0.05）,而脾脏树突状细胞、Th1细胞和Th1/Th2细胞比例均显著升高（P<0.05）。结论：丹参凝胶具有保护特应性皮炎样小鼠皮肤屏障功能和抑制表皮增生的功能,并且可以影响特应性皮炎样小鼠脾脏树突状细胞和辅助性T细胞比例。     

The Skin Commensal Yeast Malassezia Triggers a Type 17 Response that Coordinates Anti-fungal Immunity and Exacerbates Skin Inflammation

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Suitability of Nicotinic Acetylcholine Receptor α7 and Muscarinic Acetylcholine Receptor 3 Antibodies for Immune Detection: Evaluation in Murine Skin

Recent evidence reveals a crucial role for acetylcholine and its receptors in the regulation of inflammation, particularly of nicotinic acetylcholine receptor α7 (Chrna7) and muscarinic acetylcholine receptor 3 (Chrm3). Immunohistochemistry is a key tool for their cellular localization in functional tissues. We evaluated nine different commercially available antibodies on back skin tissue from wild-type (Wt) and gene-deficient (KO) mice. In the immunohistochemical analysis, we focused on key AChR-ligand sensitive skin cells (mast cells, nerve fibers and keratinocytes). All five antibodies tested for Chrm3 and the first three Chrna7 antibodies stained positive in both Wt and respective KO skin. With the 4th antibody (ab23832) nerve fibers were unlabeled in the KO mice. By western blot analysis, this antibody detected bands in both Wt and Chrna7 KO skin and brain. qRT-PCR revealed mRNA amplification with a primer set for the undeleted region in both Wt and KO mice, but none with a primer set for the deleted region in KO mice. By 2D electrophoresis, we found β-actin and β-enolase cross reactivity, which was confirmed by double immunolabeling. In view of the present results, the tested antibodies are not suitable for immunolocalization in skin and suggest thorough control of antibody specificity is required if histomorphometry is intended.     

Prevalence of positive atopy patch test in an unselected pediatric population

Background

In the latest decades, epidemiological studies on allergic disorders in children, including atopic dermatitis, rhinitis and asthma, demonstrated a continuous increase in prevalence. However, such studies are usually performed by questionnaires and, sometimes, by skin prick test or in vitro IgE tests, while the portion of allergy sustained by the cell-mediated mechanism is neglected, because the essential test, i.e. the atopy patch test is not performed.

Methods

This cross-sectional survey studied by a specific questionnaire, skin prick test and atopy patch test, an unselected population, represented by the entire scholastic population attending a Primary school and a Junior Secondary school in the rural town of San Marco in Lamis, 12.000 inhabitants (Puglia, Italy).

Results

Among the 456 subjects included, 78 (17.1 %) had a positive skin prick test and 57 (12.5 %) had a positive atopy patch test. In particular, 13.4 % of subjects were positive only to skin prick test and 8.8 % were positive only to atopy patch test. The allergen most frequently positive was the house dust mite, with 41 positive results to skin prick test and 55 to atopy patch test, while for pollen positive results concerned almost exclusively the skin prick test.

Conclusions

This survey on an unselected population of children detected a prevalence of positive results to atopy patch test not so distant from the positive results to skin prick test, and in 8.8 % of subjects the atopy patch test was the only positive test. This would suggest to add the atopy patch test in future epidemiological studies on allergy, in order to avoid to overlook the not negligible portion of patients with T-cell-mediated allergy.     

Oral Administration of a Hop Water Extract Ameliorates the Development of Dermatitis Induced by the Periodical Topical Application of a Mite Antigen in Atopic Dermatitis Model NC/Nga Mice

We investigated the inhibitory effect of an oral administration of a hop water extract (HWE) on the development of dermatitis by using NC/Nga atopic dermatitis model mice. The induction of allergic dermatitis was conducted by tape-stripping and topical application of a mite antigen (Dermatophagoides farinae) on to the ear once a week for 10 weeks. HWE was orally administered at a dose of 100 or 500 mg/kg. The total immunoglobulin E (IgE) concentration in serum and the ear thickness were periodically examined. Finally, the antigen-specific IgE level in the serum and the production of interleukin (IL)-4, IL-12 and interferon (IFN)-γ from splenocytes and cervical lymph node cells were measured. The oral administration of HWE significantly inhibited the increase of total IgE production and ear swelling throughout the experimental period. The production of IL-12 was significantly lower in the HWE administered group than in the control group. The results suggest that the intake of HWE may be effective in preventing and alleviating the development of atopic dermatitis-like skin disease.     

Inhibitory Effects of Heartwood Extracts of Broussonetia kazinoki Sieb on the Development of Atopic Dermatitis in NC/Nga Mice

We investigated the effects of a topically applied extract of the heartwood of Broussonetia kazinoki Sieb (B. kazinoki) on atopic dermatitis (AD)-like skin lesions induced by an extract of the house-dust mite Dermatophagoides farina in NC/Nga mice. We found that topically applied B. kazinoki extract suppressed the histological manifestations of AD-like skin lesions, and decreased the levels of plasma immunoglobulin E (IgE) and interleukin-4 (IL-4) in the mice. Moreover, B. kazinoki inhibited the induction of thymus-and-activation-regulated chemokine (TARC/CCL17), macrophage-derived chemokine (MDC/CCL22), and regulated-on-activation-normal T cell-expressed-and-secreted chemokine (RANTES/CCL5) in HaCaT cells activated by tumor necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma). In conclusion, our results suggest that B. kazinoki extract has therapeutic advantages in the treatment of AD.     

Proliferative responses towards native,heat-denatured and pepsin-treated ovalbumin by peripheral blood mononuclear cells from patients with hen's egg-sensitive atopic dermatitis

In order to clarify the mechanism of food-antigen recognition, the proliferative responses of peripheral blood mononuclear cells (PBMCs) to native, heat-denatured or pepsin-treated ovalbumin (OA) were investigated in 16 hen's egg-sensitive patients with atopic dermatitis (AD). Seven of them had hypersensitivity to boiled hen's egg and others had not. The responses of PBMCs to heat-denatured OA were lower than those to native OA in the patients without hypersensitivity to boiled hen's egg. However, there were no differences of the responses of PBMCs between heat-denatured OA and native OA in the patients with hypersensitivity to boiled hen's egg. Moreover, the reduction of the responses of PBMCs to pepsin-treated OA was recognized in six out of seven patients. The primary structure of OA did not change by heating or pepsin treatment according to sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). These results suggested that the secondary structure of OA changed in connection with the reduction of the responses of PBMCs to denatured OA. In addition, we demonstrated the suppressive effect of anti-HLA-DR and anti-HLA-DQ monoclonal antibodies on the proliferative response of PBMCs to OA. The results suggested that the proliferative responses of PBMCs to OA were restricted by HLA-DR or HLA-DQ in hen's egg-sensitive patients with AD.Abbreviations AD atopic dermatitis - BSA bovine serum albumin - OA ovalbumin - PBMC peripheral blood mononuclear cell - SDS-PAGE sodium dodecyl sulfate polyacrylamide gel electrophoresis     

Novel flexible vesicles based topical formulation of levocetirizine: in vivo evaluation using oxazolone-induced atopic dermatitis in murine model

Background: Levocetirizine, an active enantiomer of cetirizine is third generation antihistaminic agent used for treating various allergies like atopic dermatitis, chronic idiopathic urticaria and allergic rhinitis.

Objective: Development of novel topical formulation of levocetirizine based on flexible vesicles (FVs) with an aim to have targeted peripheral antihistaminic effect.

Materials and methods: The FVs were prepared by thin film hydration method and characterized for drug content, entrapment efficiency, pH, vesicular size, spreadability, morphological characteristics and drug leakage studies. Franz diffusion cell assembly was used to carry out the ex vivo permeation studies through mice skin and the permeation profile of the developed FV formulation was compared with conventional formulations of levocetirizine.

Results and discussion: The ex vivo permeation studies revealed 1.78-fold increase in percent permeation of levocetirizine from FV formulation as compared to conventional formulations of levocetirizine in 8?h. Further, oxazolone induced atopic dermatitis murine model was selected to study the in vivo pharmacodynamic activity. The developed formulation was evaluated for scratching score, erythema score and histological evaluation. There was marked reduction in scratching score from 15.25 scratches/20?min with conventional levocetirizine cream to 6.75 scratches/20?min with application of levocetirizine FV formulation. Also, there was significant reduction in erythema score as well as dermal eosinophil count. Results of skin sensitivity and toxicity studies suggest that the developed formulation was dermally safe and nontoxic.

Conclusion: A novel FVs based topical formulation of levocetirizine was successfully developed for treatment of atopic dermatitis.