非标记定量技术检测特应性皮炎患儿尿液中差异蛋白的应用

目的应用非标记定量技术研究特应性皮炎患儿尿液中的差异表达蛋白,筛选特应性皮炎患儿尿液中的特异性蛋白质标记物。方法收集特应性皮炎患儿和健康对照者的尿液各6份,处理和提取尿液蛋白质,经一维SDS-PAGE和蛋白胶内酶解获得肽段,应用高效液相色谱串联质谱鉴定,使用Mascot软件查询并用Scaffold软件作相对定量分析。结果特应性皮炎患儿组和健康对照组的尿液共检测出103种表达差异倍数大于1.5倍的蛋白质,患儿组表达升高的有48种蛋白质,表达降低的有55种蛋白质,其中10种蛋白质仅在患儿组检测到,17种蛋白质仅在对照组检测到。结论特应性皮炎患儿与正常儿童尿液间存在蛋白质表达差异。     

菊芋乙醇提取物对粉尘螨致小鼠特应性皮炎模型的作用

目的:探究菊芋乙醇提取物(ethanol extract of Jerusalem artichoke, JAE)对粉尘螨致小鼠特应性皮炎模型作用。方法:选取30只健康的小鼠随机分成对照组、模型组和JAE组。对照组用生理盐水处理;模型组予粉尘螨变应原浸液处理制作特应性皮炎模型;JAE组予JAE干预特应性皮炎皮损。给小鼠外用粉尘螨变应原浸液致过敏性皮炎4周,通过测量小鼠背部和耳郭皮肤反应对小鼠特应性皮炎严重程度进行评估;通过测定血浆免疫球蛋白E(IgE)和组胺水平研究JAE的保护作用;通过反转录(RT)-PCR和蛋白质印迹测定角蛋白(CK)1、CK10 mRNA的表达和蛋白的水平。结果:特应性皮炎小鼠经JAE处理后,其皮炎的严重程度包括红斑和(或)出血、水肿、结痂,血浆IgE及组胺水平均降低。与对照组相比,模型组中CK1和CK10的mRNA表达上调(P0.05),与模型组相比,经过JAE干预后CK1和CK10 mRNA和蛋白表达均下降(P0.05),表明JAE可下调CK1和CK10蛋白表达。结论:JAE可降低特应性皮炎小鼠皮肤中CK1和CK10蛋白表达。     

窄谱中波紫外线对特应性皮炎的疗效和免疫功能的影响

目的: 评价窄谱中波紫外线对成人特应性皮炎(AD)临床和免疫功能的影响.方法: 55例特应性皮炎患者分为NB-UVB治疗组(30例)和常规治疗组(25例),治疗前后分别测定血清IL-4和IFN-γ的含量,并对其临床疗效进行判断.同时选10名健康查体者作为正常对照组.结果: 特应性皮炎患者血清IL-4水平高于正常对照组,IFN-γ低于正常对照组(P<0.05);NB-UVB治疗后血清IL-4水平降低,IFN-γ水平升高,与治疗前相比均具有显著性差异(P<0.05);常规治疗组治疗前后血清IL-4、IFN-γ含量变化无显著性差异(P>0.05);NB-UVB治疗后临床疗效显著,与常规治疗组相比亦有显著性差异(P<0.05).结论: NB-UVB能影响特应性皮炎患者的免疫功能,具有显著的临床疗效.     

延边朝鲜族AD患者FLG基因SNPs与血清生化指标相关性分析

目的研究延边地区朝鲜族特应性皮炎患者丝聚蛋白基因单核苷酸多态性与血清IgE及IL-13相关性分析。方法选择70例特应性皮炎患者和90例正常对照人群作为研究对象,使用PCR法,研究丝聚蛋白基因2个SNPs位点(rs11584340及rs3126085)的基因型分布,并进行生化指标在各基因型间比较。结果延边朝鲜族特应性皮炎组血清IgE及IL-13水平显著高于正常组(P<0.01);rs11584340多态在特应性皮炎组内基因型AG有增高血清IgE趋势(P=0.05),但与IL-13不相关(P>0.05)。结论血清IgE及IL-13与延边朝鲜族人群特应性皮炎有明显相关性,是特应性皮炎的危险因素;延边朝鲜族特应性皮炎患者中rs11584340多态基因型AG有增加血清IgE趋势。     

咪唑斯汀治疗湿疹和特应性皮炎临床疗效及免疫机制的初步研究

目的:了解咪唑斯汀和西替利嗪治疗湿疹和特应性皮炎的有效性和安全性;并观察治疗前、后湿疹和特应性皮炎患者血清中白介素(IL)-4、IL-5、干扰素(IFN)-γ和肿瘤坏死因子(TNF)-α的水平变化.方法:采用随机对照方法,将患者随机分为咪唑斯汀组、西替利嗪组和对照组,外搽1%达克罗宁霜,治疗3周后观察、比较各组的疗效和安全性,并应用ELISA法检测治疗前、后湿疹和特应性皮炎患者的IL-4、IL-5、IFN-γ和TNF-α水平 .结果:入组前各组的症状、体征无明显差异.治疗后咪唑斯汀组疗效优于对照组和西替利嗪组.治疗前湿疹和特应性皮炎患者血清IL-4、IL-5和IFN-γ水平高于健康对照组,差异有统计学意义(P<0.05);而TNF-α水平与健康人差异无统计学意义(P>0.05);经咪唑斯汀和西替利嗪治疗后血清IL-4、IL-5和IFN-γ水平下降.结论:咪唑斯汀治疗特应性皮炎和湿疹安全、有效;咪唑斯汀对IL-4、IL-5的影响作用强于西替利嗪.     

利用血清蛋白质组学技术筛选银屑病相关蛋白

目的 研究常见亚型银屑病患者血清中差异表达蛋白,筛选银屑病特异性蛋白质标记物。 方法 收集首次发病进展期寻常性银屑病6例、红皮病性银屑病5例及健康人6例的血清,分别将各组标本混合,并去除高丰度白蛋白和免疫球蛋白IgG后,应用双向电泳技术分离血清,比较差异蛋白点。利用基质辅助激光解析电离飞行时间质谱,对候选差异表达蛋白点进行肽质量指纹图谱分析,通过NCBI数据库搜索鉴定蛋白。 结果 获得了3组较好的血清双向电泳图谱。应用胶图分析软件分析,发现寻常性银屑病组、红皮病性银屑病组与健康对照组间差异蛋白点分别为33个和17个,寻常性银屑病组和红皮病性银屑病组之间差异蛋白点26个。共鉴定得到14种蛋白。在银屑病组中表达升高的有补体成分3、白介素16、维生素D结合蛋白、α1抗胰蛋白酶等;红皮病组中补体成分3、补体因子H、α1抗胰蛋白酶、血液结合素、触珠蛋白的表达量高于健康人对照,α1抗胰蛋白酶、补体因子H、补体成分4和触珠蛋白的表达量高于寻常性银屑病组。在红皮病性患者血清中表达下降的有血清淀粉样蛋白P。 结论 寻常性银屑病、红皮病性银屑病与健康人对照间血清蛋白表达谱存在差异。     

湘籍汉族特应性皮炎患者FcεRIβ基因启动子-109位基因多态性分析

目的 探讨湘籍汉族特应性皮炎患者IgE高亲和力受体β链(FcεRIβ)基因启动子-109位基因多态性及与血清总IgE的关系.方法 采用PCR-单链构型多态性(SSCP),DNA测序分析FcεRIβ基因多态性,ELISA检测血清总IgE.结果 20例特应性皮炎患者中有4例FcεRIβ基因启动子-109位泳动异常,其中有2例特应性皮炎患者第138位碱基发生C→A替换,159位碱基发生T→C替换;2例患者165位碱基发生A→G替换.突变组特应性皮炎患者血清总的IgE为(1.88±1.30)mg/L,无突变组特应性皮炎患者血清总的IgE为(0.32±0.12)mg/L,正常人对照组血清总的IgE为(0.20±0.05)mg/L;突变组较无突变组及正常人对照组差异有统计学意义(P＜0.05).结论 特应性皮炎患者IgE高亲和力受体β链启动区-109位多态性伴有IgE高表达.     

湘籍汉族特应性皮炎患者FcεRIβ因启动子-109位基因多态性分析

目的探讨湘籍汉族特应性皮炎患者IgE高亲和力受体β链(FcεRIβ)基因启动子-109位基因多态性及与血清总IgE的关系。方法采用PCR-单链构型多态性(SSCP),DNA测序分析FcεRIβ基因多态性,ELISA检测血清总IgE。结果20例特应性皮炎患者中有4例FcεRIβ基因启动子-109位泳动异常,其中有2例特应性皮炎患者第138位碱基发生G→A替换,159位碱基发生T→C替换；2例患者165位碱基发生A→G替换。突变组特应性皮炎患者血清总的IgE为(1．88±1．30)mg/L,无突变组特应性皮炎患者血清总的IgE为(0．32±0．12)mg/L,正常人对照组血清总的IgE为(0．20±0．05)mg/L；突变组较无突变组及正常人对照组差异有统计学意义(P＜0．05)。结论特应性皮炎患者IgE高亲和力受体β链启动区-109位多态性伴有IgE高表达。     

儿童特应性皮炎患者外周血单一核细胞IL-31与疾病严重程度相关性分析

目的 探讨IL-31在儿童特应性皮炎发病机制中的作用以及与特应性皮炎瘙痒的相关性。方法 22例特应性皮炎患儿与22例健康儿童外周血单一核细胞在葡萄球菌肠毒素B（SEB）刺激或非刺激状态下，应用实时PCR方法分析IL-31表达情况；酶联免疫吸附法测定其血清IgE水平；对患儿病情进行病情严重程度评分，分析IL-31 mRNA与IgE水平、疾病严重程度及瘙痒的相关性。结果 特应性皮炎患儿外周血单一核细胞IL-31表达显著增加，是对照组的23.2倍（P < 0.01）。特应性皮炎组和对照组外周血单一核细胞受SEB刺激后IL-31表达均有不同程度升高，以特应性皮炎组IL-31表达增加更显著，是对照组的20.44倍。患儿血清总IgE水平中位数为260.05 IU/mL（范围5.9 ～ 1131.01 IU/mL），对照组为17.7 IU/mL（范围5 ～ 140.7 IU/mL），两组比较，P < 0.01。IL-31与患儿病情严重程度以及血清总IgE水平无显著相关性（r = 0.07，P > 0.05；r = 0.22，P > 0.05）。结论 IL-31可能参与儿童特应性皮炎发病，其作用机制可能不依赖血清IgE；SEB能诱导正常人外周血单一核细胞快速表达IL-31，是IL-31产生的重要调节因素。     

儿童特应性皮炎严重程度与血清25-羟维生素D和总IgE水平的相关性

目的：明确儿童特应性皮炎患者特应性皮炎严重程度与血清25-羟维生素D和IgE水平的相关性。方法：参考SCORAD评分法评估60例特应性皮炎患者疾病严重程度，并检测特应性皮炎患者及55例对照组血清25-羟基维生素D3水平以及特应性皮炎患者总IgE水平。结果：特应性皮炎组患者血清25-羟维生素D水平(16.13±6.68)ng/mL低于对照组(19.81±8.24)ng/mL，差异有统计学意义（P<0.05）。根据SCORAD评分，特应性皮炎患者中33例为轻度、20例为中度，7例为重度。轻度组血清25-羟基维生素D3水平为(18.69±7.01)ng/mL，高于中度(12.81±4.35)ng/mL，差异有统计学意义（P<0.05）。血清25-羟维生素D与SCORAD评分之间有显著负相关（P<0.05）。IgE水平与SCORAD评分之间无相关性（P>0.05）。结论：AD患者血清25-羟基维生素D与AD严重程度呈负相关。     

Detection of circulating immune complexes in patients with atopic dermatitis and psoriasis

Sera from 32 patients with atopic dermatitis and 22 patients with psoriasis were examined for the presence of circulating immune complexes (CIC) in comparison to 51 healthy controls using a PEG-precipitation laser nephelometer technique. Different patterns of the precipitated proteins were found in both diseases. In atopic dermatitis C3 and IgG were significantly elevated in CIC. Furthermore, significantly increased amounts of IgE were found in the precipitates. Groups with high and low serum IgE levels showed no significant differences in the quantity of precipitated proteins. Skin involvement did not correlate with CIC. In psoriasis patients, a different pattern with significantly increased IgA, IgG, IgM and C3 was found in the precipitates. IgE was also significantly increased in comparison to the controls. No difference was found between patients with psoriasis vulgaris and psoriasis guttata. CIC in psoriasis and in atopic dermatitis thus showed a characteristic composition. However, a detection of CIC was not directly related to the cutaneous manifestation of the disease.     

IgE-mediated hypersensitivity against human sweat antigen in patients with atopic dermatitis

Sweating aggravates itch in atopic dermatitis, but the mechanism is unclear. In this study, we examined the involvement of type I hypersensitivity in the aggravation of atopic dermatitis by sweating. Skin tests with autologous sweat were positive in 56 of 66 patients (84.4%) with atopic dermatitis, but only in 3 of 27 healthy volunteers (11.1%). Sweat samples from both patients and healthy volunteers induced varying degrees of histamine release from basophils of patients with atopic dermatitis. However, the histamine release was impaired by removal of IgE on the basophils. Incubation of basophils with myeloma IgE before sensitization with serum of patients blocked the ability to release histamine-induced sweat. IgE antibody against antigen(s) in sweat may be present in serum of patients with atopic dermatitis. Key words:     

Immunofluorescence of the skin in allergic diseases: an investigation of patients with contact dermatitis, allergic vasculitis and atopic dermatitis.

Skin biopsies for immunofluorescent studies were taken from patients with contact dermatitis (positive patch tests), atopic dermatitis and allergic vasculitis for comparison with normal-appearing skin from the same patients, and from healthy controls. A variety of deposits of immunoglobulins, complement components and fibrinogen were demonstrated in 6 out of 20 patients with contact dermatitis, 7 out of 10 with atopic dermatitis, 8 out of 10 with allergic vasculitis, and in 4 out of 20 control individuals. No diagnostic pattern of deposits was found. Elevated serum IgE and eosinophilic counts were found in patients with atopic dermatitis, and high serum IgA and fibrinogen levels were found in the allergic vasculitis group.     

Production of interleukin-2 by mononuclear cells in vitro in patients with atopic dermatitis and psoriasis. Comparison with serum interleukin-2 receptor levels.

Atopic dermatitis is associated with profound immunological alterations, in particular decreased lymphoproliferative responses upon stimulation with T-cell mitogens. T-cell blastogenesis involves the production of the soluble cytokine interleukin-2 (IL-2), which in turn upregulates the expression of its own receptor. To investigate the potential role of this cytokine for the pathomechanisms present in atopic dermatitis, 24-h supernatants of PHA-stimulated peripheral blood mononuclear cells from patients with atopic dermatitis (n = 30) of a moderate to severe disease activity were tested for IL-2 activity. In addition, serum concentrations of soluble interleukin-2 receptor (IL-2R) were measured. Non-atopic healthy controls (n = 19) and patients with psoriasis (n = 20), an inflammatory skin disorder with distinct pathogenesis, served as controls. In comparison with psoriasis patients and normal controls, PHA-stimulated mononuclear cells of atopic dermatitis patients released significantly less IL-2 into supernatants. Moreover, there was an inverse correlation between IL-2 concentrations and body surface involvement or serum IgE levels. In contrast, serum IL-2R levels were significantly elevated in both atopic dermatitis and psoriasis, as compared with healthy controls. Furthermore, IL-2R levels in atopic dermatitis patients showed a significant correlation with IgE levels and body surface involvement. The data indicate that T cell activation may occur in both skin diseases. Atopic dermatitis, however, is further characterized by the decreased capacity of mononuclear cells to release IL-2 upon stimulation in vitro.     

Impaired responses of peripheral blood mononuclear cells to staphylococcal superantigen in patients with severe atopic dermatitis: a role of T cell apoptosis

Staphylococcus aureus colonization is an almost universal feature of atopic dermatitis. In order to investigate the role of staphylococcal enterotoxin B in the pathogenesis of atopic dermatitis, we assessed the correlation between clinical disease severity and proliferative response of peripheral blood mononuclear cells to staphylococcal enterotoxin B in patients with atopic dermatitis. Peripheral blood mononuclear cells from patients with mild atopic dermatitis showed significantly increased proliferative responses to staphylococcal enterotoxin B compared to controls. In contrast, peripheral blood mononuclear cells from patients with severe atopic dermatitis showed markedly suppressed proliferative responses. Additionally, longitudinal evaluation of peripheral blood mononuclear cell samples from the same patient demonstrated that proliferative responses were suppressed only at times of severe disease exacerbation. Mixing experiments, using autologous T cells and antigen presenting cells that were isolated at different time points from the same patient, demonstrated that T cells of severe atopic dermatitis patients were dysfunctional, but their antigen presenting cell function remained intact. We found no significant differences of interleukin-2 levels in the culture supernatants between healthy controls and atopic dermatitis groups. Fluorescence-activated cell sorter analysis for APO2.7 antigen, an early apoptosis cell marker, demonstrated that approximately 60% of staphylococcal-enterotoxin-B-stimulated T cells expressed APO2.7 antigen in severe atopic dermatitis cases. By contrast, 5%-20% of T cells expressed APO2.7 after staphylococcal enterotoxin B stimulation in cases of mild atopic dermatitis and in healthy controls. Nuclear staining with Hoechst 33258 also showed approximately 40% apoptotic cells in the CD19-CD16-PBMC of severe atopic dermatitis patients, compared with only 5%-10% in the mild atopic dermatitis group and in healthy controls. Blocking monoclonal antibody to Fas ligand partially prevented the staphylococcal-enterotoxin-B-induced apoptosis detected by APO2.7 expression and Hoechst 33258 staining. Suppressed proliferation of peripheral blood mononuclear cells in severe atopic dermatitis patients may be secondary to T cell death by apoptosis. These results suggest that an infection of S. aureus producing staphylococcal enterotoxin B may play a role in aggravation of atopic dermatitis by inducing apoptosis in T cells.     

The prevalence of Malassezia yeasts in patients with atopic dermatitis, seborrhoeic dermatitis and healthy controls

Cultures for Malassezia yeasts were taken from both normal-looking skin and lesional skin in 124 patients with atopic dermatitis, 16 patients with seborrhoeic dermatitis and from normal skin of 31 healthy controls. Positive Malassezia growth was found in fewer patients with atopic dermatitis (56%) than in patients with seborrhoeic dermatitis (88%) or in healthy controls (84%, p<0.01). In the patients with atopic dermatitis, fewer positive cultures were found in lesional (28%) than in non-lesional skin (44%, p<0.05), while positive cultures were found in 75% of both lesional and non-lesional skin of patients with seborrhoeic dermatitis (not significant). M. sympodialis dominated in patients with atopic dermatitis (46%) and in healthy controls (69%). In patients with seborrhoeic dermatitis both M. sympodialis and M. obtusa were cultured in 43%. A Malassezia species extract mixture would increase the possibility of detecting IgE sensitization to Malassezia in patients with atopic dermatitis.     

The changing face of dermatology out-patient referrals in the south-east of Scotland

The distribution and localization of several neuropeptides were investigated in the lichenified lesions of 11 patients with atopic dermatitis using indirect immunofluorescence. Substance P-positive nerve fibres were observed in most of the cases of atopic dermatitis, but not in normal controls. Somatostatin immunoreactive nerves were not found in the skin of atopic dermatitis, whereas a normal pattern of immunoreactivity could be detected in most of the healthy subjects. Neuropeptide Y-positive dendritic epidermal cells were observed in lesional skin from patients with atopic dermatitis, but not in controls. Calcitonin gene-related peptide and vasoactive intestinal polypeptide immunoreactivity in patients with atopic dermatitis did not differ from that in healthy subjects. With galanin antiserum a diffuse intracellular staining was observed in the epidermis of both atopic patients and controls, while no positive staining was found with either neurotensin or neurokinin A antibodies in either group. These findings suggest a possible involvement of some neuropeptides in the pathomechanisms of atopic dermatitis.