天疱疮患者病情与抗桥粒芯蛋白抗体水平的相关性研究

目的:研究天疱疮患者血清中抗桥粒芯蛋白(desmoglein,Dsg)1和抗Dsg3抗体水平与其皮肤、口腔黏膜损害严重程度的相关性,同时对间接免疫荧光(IIF)检测的天疱疮抗体滴度与治疗中使用皮质类固醇控制剂量的相关性进行分析。方法:采用酶联免疫吸附试验(ELISA)试剂盒测定55例天疱疮患者血清中抗Dsg1和抗Dsg3抗体水平。结果:抗Dsg1抗体水平与患者皮肤损害严重程度有显著相关性(P<0.01),抗Dsg3抗体水平与口腔黏膜损害严重程度有显著相关性(P<0.01)。天疱疮患者血清IIF滴度与抗Dsg1抗体水平相关(P<0.01),寻常型天疱疮患者IIF滴度与抗Dsg1和抗Dsg3抗体水平均有相关性(P分别<0.01和<0.05)。寻常型天疱疮患者皮质类固醇控制剂量与抗Dsg1抗体水平和IIF滴度显著相关(P<0.05)。结论:ELISA方法检测天疱疮患者抗Dsg1和抗Dsg3抗体对天疱疮的临床诊断、分型、衡量口腔黏膜和皮肤损害严重程度具有一定意义。     

天疱疮患者皮损局部B淋巴细胞及其产生特异性抗体的功能研究

目的 检测寻常型天疱疮患者皮损局部B淋巴细胞及其产生特异性抗体的功能。方法 寻常型天疱疮患者35例,健康对照22例。取健康对照皮肤和寻常型天疱疮患者初发水疱或糜烂皮损组织,分离获得单个核细胞。流式细胞仪检测患者皮损局部淋巴细胞、CD19+ B细胞比例,以及特异性识别桥粒芯糖蛋白1（Dsg1）和Dsg3的CD19+ B淋巴细胞比例。体外培养寻常型天疱疮患者皮损局部淋巴细胞,ELISA法检测培养上清液中抗Dsg1和Dsg3抗体滴度,采用受试者工作特征曲线（ROC）分析阳性率。结果 寻常型天疱疮患者皮损局部淋巴细胞、CD19+ B细胞比例分别为17.95% ± 3.85%、4.27% ± 1.13%,高于健康对照组（7.83% ± 1.29%、0.61% ± 0.31%）,差异有统计学意义（t = 2.49,U = 13.00,均P < 0.05）。天疱疮患者皮损局部CD19+ B淋巴细胞中,表达IgG的细胞比例为（38.33 ± 5.56）%,表面识别Dsg1与Dsg3的细胞比例分别为12.87% ± 1.267%、10.42% ± 1.243%。局部淋巴细胞体外培养6 d后,培养上清液中抗Dsg1与Dsg3抗体滴度分别为（4.89 ± 1.56） U/ml、（35.45 ± 13.03） U/ml,阳性率分别为85%（17/20）和95%（19/20）。 结论 寻常型天疱疮患者皮损局部有可与Dsg1及Dsg3特异性结合的B淋巴细胞聚集,体外培养后可产生特异性抗Dsg1与抗Dsg3自身抗体。     

桥粒芯糖蛋白抗体与天疱疮临床表型和疾病活动度的关系及其变化规律

【摘要】 目的 评估酶联免疫吸附试验（ELISA）检测桥粒芯糖蛋白1（Dsg1）、Dsg3抗体与天疱疮患者临床表型、疾病活动度的关系及变化规律。方法 收集2015年1月至2018年1月在中国医学科学院皮肤病医院就诊的天疱疮患者111例,按临床分型,采用ELISA测定患者初发时、控制阶段、维持阶段及复发时血清Dsg1、Dsg3抗体水平变化,并分析其变化规律。采用SPSS22软件,多组间比较采用单因素方差分析,组间两两比较采用LSD?t检验。结果 天疱疮初发时、控制阶段、维持阶段及复发时分别有92例、53例、33例、9例患者完成检测。92例初发患者中,36例落叶型天疱疮患者Dsg1和Dsg3抗体水平阳性率分别为100%、2.77%,10例黏膜型寻常型天疱疮分别为20%、80%,46例黏膜皮肤型寻常型天疱疮分别为97.82%、95.65%。初发时、控制阶段、维持阶段和复发时落叶型天疱疮患者Dsg1抗体水平分别为（137.43 ± 77.74）、（13.94 ± 14.81）、（21.50 ± 58.33）、（121.13 ± 86.89） U/ml;黏膜型寻常型天疱疮患者Dsg3抗体水平分别为（125.61 ± 94.81）、（34.5 ± 16.26）、0.6、258 U/ml;黏膜皮肤型寻常型天疱疮患者Dsg1抗体水平分别为（115.39 ± 70.62）、（15.74 ± 25.10）、（3.62 ± 12.09）、（78.60 ± 92.25） U/ml;Dsg3抗体水平分别为（137.98 ± 81.25）、（58.14 ± 63.46）、（29.26 ± 64.70）、（136.9 ± 101.47） U/ml。落叶型天疱疮Dsg1抗体水平和黏膜型寻常型天疱疮、黏膜皮肤型寻常型天疱疮Dsg3抗体水平在控制阶段、维持阶段均低于初发时、复发时（P < 0.05）。治疗过程中2例患者出现表位扩展现象,4例患者病情稳定期时出现Dsg抗体高滴度现象。结论 Dsg抗体谱与天疱疮临床表型相关,其ELISA值可用于监测疾病活动,并可对治疗的有效性作出评价。     

抗构象表位桥粒芯蛋白抗体和乙酰胆碱受体抗体滴度与寻常型天疱疮病情活动度的相关性研究

【摘要】 目的 研究寻常型天疱疮患者血清中相关抗体滴度与病情严重程度和病情活动度的相关性。方法 收集2012—2015年于中国医学科学院皮肤病医院首次就诊的24例活动期寻常型天疱疮（PV）患者,评估患者活动期和稳定期天疱疮疾病面积指数（PDAI）,并采集血清标本。采用酶联免疫吸附实验（ELISA）检测血清标本中具有致病作用的抗构象表位桥粒芯蛋白（Dsg）抗体滴度、总Dsg抗体滴度和乙酰胆碱受体（AChR）抗体滴度。计量资料比较采用t检验,计数资料比较采用Fisher 精确检验法,相关性比较采用Pearson分析。结果 活动期患者的Dsg1抗体滴度（611.4 ± 136.8）与抗构象表位Dsg1抗体滴度（585.5 ± 134.7）差异无统计学意义（t = 0.13,P = 0.89）,Dsg3抗体滴度（708.6 ± 130.7）高于抗构象表位Dsg3抗体滴度（297.2 ± 54.4,t = 2.90,P < 0.01）。活动期患者的Dsg1抗体滴度及抗构象表位Dsg1抗体滴度与PDAI评分均呈正相关（r = 0.54、0.54,均P < 0.01）;Dsg3抗体滴度与PDAI评分无相关性（r = 0.11,P = 0.62）,抗构象表位Dsg3抗体滴度与PDAI评分呈正相关（r = 0.53,P < 0.01）。20例稳定期患者血清中Dsg1抗体与抗构象表位Dsg1抗体滴度与首次就诊时比较均明显下降。Dsg3抗体滴度仅7例明显下降;13例仍存在较高滴度的Dsg3抗体,其中6例抗构象表位Dsg3抗体滴度明显下降,5例AChR抗体滴度由阳性转为阴性。结论 Dsg1抗体及抗构象表位Dsg1抗体滴度都可以反映病情活动度。部分患者病情活动度与Dsg3抗体滴度不一致,抗构象表位Dsg3抗体或者AChR抗体可能有助于反映病情活动度。     

天疱疮发病机制的研究进展

天疱疮是一种自身抗体引起的自身免疫性疾病,以往观点认为,抗桥粒芯蛋白抗体在天疱疮发病中处于核心地位,研究发现,疾病中的一些现象不能完全用桥粒芯抗体的理论解释,对这一观点提出补充,并发现了一系列新的天疱疮抗体,丰富了新的天疱疮发病理论.其中皮肤乙酰胆碱系统在介导角质形成细胞间黏附和天疱疮发病中具有重要作用,基底细胞收缩和松解凋亡理论也使得天疱疮的发病机制进一步完善.基于这些新的发病机制,天疱疮的非糖皮质激素治疗取得了一些进展,有望改善天疱疮患者预后.     

棘层松解性棘细胞瘤

组织病理特征 本文2张组织病理图片的主要特征是：表皮增厚,呈乳头瘤样增生,皮突延长;表皮棘层松解,主要位于表皮的下部：棘层松解不伴有角化不良：未见明显的角化过度和角化不全。[第一段]     

棘层松解性棘细胞瘤

组织病理特征本文2张组织病理图片的主要特征是:表皮增厚,呈乳头瘤样增生,皮突延长;表皮棘层松解,主要位于表皮的下部;棘层松解不伴有角化不良;未见明显的角化过度和角化不全.     

ELISA技术检测寻常型天疱疮桥粒芯蛋白3抗体水平研究

目的:评价酶联免疫吸附试验(ELISA)检测抗桥粒芯蛋白(desmoglein,Dsg)3抗体在寻常型天疱疮(PV)诊断中的意义.方法:对来自不同中心的106例PV患者和106例对照人群血清标本编盲后,进行ELISA检测抗Dsg3自身抗体和间接免疫荧光(IIF)法检测血清天疱疮自身抗体.结果:ELISA法检测Dsg3抗体敏感度为77.4%,特异性为94.3%;IIF法检测抗体敏感度为79.2%,特异性为94.3%.两组间的差异无统计学意义.结论:ELISA方法检测Dsg3抗体对于寻常型天疱疮的诊断是一种较好的辅助方法.     

生殖器丘疹样棘层松解性皮病一例

报道1例生殖器丘疹样棘层松解性皮病.患者,男,30岁.2年前肛周出现灰白色丘疹,部分融合,伴瘙痒.实验室检查:HPV6阳性,醋酸白试验、梅毒螺旋体明胶凝集试验(TPPA)、HIV均阴性.皮损组织病理示:基底层上方棘层下方多处裂隙,可见棘层松解细胞,中等量淋巴细胞浸润.直接免疫荧光:IgG、IgM、IgA、C3阴性.予以...     

桥粒芯蛋白1酶联免疫吸附试验检测落叶型天疱疮血清抗体的评价

目的 探讨桥粒芯蛋白1（Dsg1）酶联免疫吸附试验（ELISA）检测落叶型天疱疮（PF）血清学抗体的意义。方法 将80例PF患者和132例对照人群的血清标本进行编盲,用ELISA法检测所有标本中抗Dsg1自身抗体,同时应用间接免疫荧光（IIF）法检测所有标本的抗体滴度,操作过程为随机检测,最后将两方法的结果进行比较。结果 75例PF患者和5例对照人群（包括1例不确定值,大疱性类天疱疮、SLE、皮肌炎、湿疹及健康者各1例）Dsg1 ELISA为阳性,71例PF患者和0例对照人群IIF为阳性。Dsg1 ELISA的敏感性为93.8%（95%可信区间0.85 ~ 0.98）,特异性为96.2%（95%可信区间0.91 ~ 0.99）。IIF的敏感性为88.8%（95%可信区间0.82 ~ 0.96）,特异性为100%（95%可信区间0.96 ~ 1.00）。两者相比,敏感性（P = 0.289）和特异性（P = 1.000）差异均无统计学意义。结论 Dsg1 ELISA是一种简便、敏感、特异的血清学检测方法,可作为诊断PF的一种辅助手段。     

天疱疮抗体致棘刺松解量化研究

观察1例寻常型天疱疮不同浓度天疱疮抗体（Pab)致表皮棘刺松解的作用,及对组织尿激酶型纤溶酶原（uPA)表达的影响。结果提示间接免疫荧光（IIF）检出Pab量接近Pab致棘刺松解量,IIF阴性可作为病情缓解的可靠指标,而直接免疫荧光（DIF）更适于作为活动性天疱疮的诊断指标。     

Autoantibodies against desmoglein 2 are not pathogenic in pemphigus

BackgroundAnti-desmoglein 1 and 3 autoantibodies justify acantholysis in pemphigus; however, the pathogenesis of anti-desmoglein 2 is hypothetical.ObjectiveTo compare the participation of desmogleins 1, 2 and 3 through the production of serum autoantibodies, and protein and gene expression in the skin/mucosa of patients with pemphigus foliaceus and pemphigus vulgaris.MethodsThe autoantibodies were titrated by ELISA in 202 samples of pemphigus foliaceus, 131 pemphigus vulgaris, 50 and 57 relatives of patients with pemphigus foliaceus and pemphigus vulgaris, respectively, and 114 controls. Protein and gene expressions were determined by immunohistochemistry and qPCR in the skin/mucosa of 3 patients with pemphigus foliaceus and 3 patients with pemphigus vulgaris.ResultsHigher titers of anti-desmoglein 2 (optical density) resulted in pemphigus foliaceus and pemphigus vulgaris, when compared to controls (0.166; 0.180; 0.102; respectively; p < 0.0001). There was a correlation between anti-desmoglein 2 and anti-desmoglein 1 titers in pemphigus foliaceus (r = 0.1680; p = 0.0206). There was no cross-reaction of anti-desmoglein 2 with desmoglein 1 and 3. Protein overexpression of desmoglein 2 was observed in intact and lesional skin of patients with pemphigus compared to the skin of controls. Internalization granules of desmoglein 1 and 3, but not of desmoglein 2, were observed in lesions of pemphigus foliaceus and pemphigus vulgaris, respectively. Gene overexpression of desmoglein 2 was observed in the mucosa.Study limitationsSmall sample size for the statistical analysis of protein and gene expression.ConclusionAutoantibodies against desmoglein 2 are not pathogenic in pemphigus; protein and gene overexpression of desmoglein 2 in the skin and mucosa may be involved in acantholysis repair.     

寻常型天疱疮患者血清骨桥蛋白的检测北大核心CSCD

目的 探讨寻常型天疱疮（PV）患者血清中骨桥蛋白在PV发病中的作用.方法 用酶联免疫吸附试验对31例寻常型天疱疮患者和35例健康对照组血清骨桥蛋白的浓度进行测定.用SPSS17.0及Excell统计软件对数据进行处理.结果 PV患者血清骨桥蛋白的表达水平显著高于健康对照组（P＜0.01）.皮肤黏膜同时受累患者血清骨桥蛋白水平高于仅皮肤受累组（P＜0.05）.合并感染组血清骨桥蛋白水平高于未合并感染组（P＜ 0.01）; PV患者的血清骨桥蛋白水平与抗Dsg3抗体呈正相关（rs=0.489,P＜ 0.01）.结论 骨桥蛋白在PV患者血清中高表达,特别是皮肤黏膜同时受累的患者,且与天疱疮抗体滴度呈正相关.     

Use of recombinant pemphigus vulgaris antigen in development of ELISA and IB assays to detect pemphigus vulgaris autoantibodies

Aim/Objective The objectives of this study are: (1) to measure the titers of pemphigus vulgaris (PV) autoantibody in the sera of patients with active disease, using three different assays: (a) Indirect immunofluorescence (IIF) using monkey esophagus as a substrate, (b) immunoblot (IB) and, (c) enzyme-linked immunosorbent assay (ELISA) using recombinant PV antigen (rPVA). (2) To compare the sensitivity of these three assays. Background The titer of PV autoantibodies and disease severity and extent do not always correlate. This could be due to the lack of consistency and specificity of the substrate. Different results are obtained using different substrates. A standard substrate with uniformly controlled source of antigen would be more useful and clinically beneficial. Methods In this study we studied 25 PV patients, six each with bullous pemphigoid (BP), ocular cicatricial pemphigoid (OCP), mucous membrane pemphigoid (MMP), and herpes gestationis (HG), and sera from 16 normal subjects. IIF was used to determine the PV autoantibody using monkey esophagus. IB assay was used according to standard protocol using normal human epidermis and rPVA as substrates. ELISA was performed using rPVA as antigens expressed in E. coli. Results Sera of all 25 PV patients showed binding to the rPVA, normal human sera and the sera from the six BP, six OCP, six MMP, and six HG patients did not show any binding. In addition, we used antisera from rabbits immunized with PVA peptides (Bos-1, Bos-6) which also showed binding to rPVA, whereas normal rabbit sera did not show any reactivity. ELISA and IB titers in all the patients were 2.5 to 160 times higher than with the conventionally used IIF assay. The titers of the PV specific autoantibody measured using the rPVA did not show statistically significant differences between the ELISA and IB assays. Conclusions IB and ELISA are superior to IIF in evaluating the antibody levels in PV patients. ELISA is more practical and is preferable to IB and is recommended for clinical use.     

凋亡在寻常型天疱疮发病中的作用

以往认为寻常型天疱疮仅由致病性抗体作用导致的一类自身免疫性皮肤黏膜疾病.但近年来其发病机制不断被揭示,凋亡在疾病中的作用亦越来越明确,如角质形成细胞经抗体直接作用可产生凋亡现象;棘层松解后形成的细胞间连接丧失可触发失巢凋亡.松解凋亡的提出进一步阐明棘层松解和细胞凋亡之间的关联性,也证明寻常型天疱疮的凋亡与传统的凋亡现象不同,具有其特殊性,对寻找疾病治疗的新靶点具有重要意义.     

Activation of keratinocyte muscarinic acetylcholine receptors reverses pemphigus acantholysis

We studied neuroendocrine mediator regulation of adhesion and motility of human epidermal keratinocytes (EK) and found that cholinergic compounds control EK cell-matrix and cell-cell attachment. In this study, we tested the anti-acantholytic activity of muscarinic agonists in pemphigus and non-pemphigus acantholysis, and investigated the effects of pemphigus antibody (Pab) on the the keratinocyte muscarinic acetylcholine receptors (mAChR). Acantholysis produced by Pab from two patients with pemphigus vulgaris was compared with acantholysis induced by the serine protease trypsin. the calcium chelator EOT A or the muscarinic antagonist atropine. Trypsinized EK first lost contact with microplate surface and then retracted their intercellular filaments. EDTA-treated cells first detached from each other and then from the dish. EK cultures treated with Pab or atropine rounded up and retracted their intercellular filaments simultaneously, although it took hours to obtain acantholysis with Pab treatment compared to several seconds with atropine treatment. Addition of acetylcholine or other muscarinic agonists (bethanechol. carbachol or methacholine) to acantholytic cultures reversed both pemphigus and non-pemphigus acantholysis. Acantholysis induced by atropine reversed spontaneously. Short-term preexposure of EK to Pab significantly increased, and long-term preexposure significantly decreased, the [³H]atropine binding to keratinocyte mAChR. We conclude that muscarinic agonists reverse various types of acantholysis. including acantholysis induced by Pab, and that binding of Pab to EK may affect the ability of keratinocyte mAChR to bind its ligands.     

The value of trichoscopy in the differential diagnosis of scalp lesions in pemphigus vulgaris and pemphigus foliaceus

BACKGROUND

Trichoscopy is becoming increasingly popular in diagnosing hair and scalp diseases. Scalp involvement in pemphigus is common. The scalp may be the first or only site of clinical manifestation of the disease.

OBJECTIVE

The aim of this study was to analyze whether trichoscopy may be useful in aiding differential diagnosis of scalp lesions in patients with pemphigus vulgaris and pemphigus foliaceus.

METHODS

Trichoscopy was performed in 19 patients with scalp lesions in the course of pemphigus (9 patients with pemphigus vulgaris and 10 with pemphigus foliaceus). In all patients, the diagnosis of scalp pemphigus was confirmed by histopathology. The working magnification was 20-fold and 70-fold.

RESULTS

The most frequently observed trichoscopy features of pemphigus lesions were: extravasations (18/19; 94.7%) and yellow hemorrhagic crusts (11/19; 57.9%). Yellow dots with whitish halo were observed in 6/19 (31.6%) patients with pemphigus. White polygonal structures were observed in pemphigus foliaceus (6/10; 60%), but not in pemphigus vulgaris. Vascular abnormalities were more frequent in pemphigus vulgaris, when compared to pemphigus foliaceus, and were associated with a severe course of disease. Linear serpentine vessels were the most frequent vascular abnormality in patients with pemphigus vulgaris and pemphigus foliaceus (77.8% and 30%, respectively).

CONCLUSION

Trichoscopy may serve as a useful supplementary method in the differential diagnosis of pemphigus, especially in cases of desquamative or exudative lesions limited to the scalp. Extravasations, yellow hemorrhagic crusts, yellow dots with whitish halo, white polygonal structures and linear serpentine vessels are trichoscopy features which may suggest the diagnosis of pemphigus.     

Intraperitoneal injection of pemphigus vulgaris-IgG into mouse depletes epidermal keratinocytes of desmoglein 3 associated with generation of acantholysis

Pemphigus vulgaris is an autoimmune blistering disease caused by antibodies against desmoglein (Dsg) 3. We previously reported that pemphigus vulgaris (PV)-IgG caused the formation of Dsg3-depleted desmosomes in normal human cultured keratinocytes and DJM-1, a human squamous cell carcinoma cell line. In the present study, we injected PV-IgG and normal human IgG into neonatal mice and examined the quantities of Dsg3 in the mouse skin. We showed that injection of PV-IgG into neonatal mice caused suprabasal blister formation and approximately 30% reduction of Dsg3 in mouse epidermal keratinocytes, compared to mice injected with normal human IgG. In addition, we showed that the quantity of Dsg3 in the skin of patients with PV did decrease, as compared to that in healthy volunteers. Our data suggests the reduction of Dsg3 might be relevant to blister formation. These results also suggest that even a partial depletion of Dsg3 may contribute to blistering in PV patients.