副肿瘤性天疱疮患者的脱发机制

目的探讨副肿瘤性天疱疮(PNP)患者血清中抗桥粒芯蛋白(DSG)抗体与头皮脱发之间的关系.方法收集我科收治的2例PNP,2例大疱性类天疱疮患者和5例正常人血清,及2例正常人头皮组织.采用酶联免疫吸附(ELISA)方法检测PNP患者血清中DSG 1和3抗体水平,间接免疫荧光法检测PNP患者血清与正常人头皮毛囊结合情况,同时采用正常人血清和大疱性类天疱疮患者血清作对照.结果2例PNP患者中,有1例抗DSG 3抗体强阳性,抗DSG1抗体弱阳性;另1例患者两种抗体均为阴性.2例患者血清均可与正常人头皮毛囊结合,表皮细胞间和毛囊的外毛跟鞘均可见荧光信号.大疱性类天疱疮患者血清只能与表皮基底膜带结合,正常人血清在基底膜带、表皮细胞间和毛囊均无任何荧光信号.结论PNP患者出现脱发可能与患者血清中抗DSG3抗体水平有关.     

副肿瘤性天疱疮的基础与临床研究

副肿瘤性天疱疮(PNP)是一种自身免疫性大疱性皮肤病,常伴发肿瘤.临床表现为严重的黏膜糜烂和多形性皮肤损害.PNP的主要病理学特点为表皮基底细胞层上棘细胞松解性水疱及个别坏死的角质形成细胞.患者血清中可存在致病性的自身抗体,其产生与伴发的肿瘤有关.早期诊断及手术及时切除肿瘤是成功治疗的关键.     

Purification and identification of natural antikeratin autoantibody from normal human sera and its effects on the cultured keratinocytes

Objective: To purify the natural antikemtin autoantibody (AK auto-Ab) and observe its effects on the proliferation of the cultured keratinocytes. Methods: Natural AK auto-Ab was purified by using keratin affinity column, and then the titre and specificity of the Abs were studied by ELISA, immunoperoxidase staining and immuno-electronicmi-croscope. The effect of the purified Abs on the cultured keratino-cytes was assayed by ^3H-TdR incorporation. Results:Natural AK auto-Ab was obtained. The binding activity oflgG AK auto-Ab in purified Ab remained similar to that in pooled human sera, and the specificity of the obtained antibody is strong. The purified antibody could decrease the ^3H-TdR incorporation of the cultured keratinocytes in a dose-dependent manner. Conclusion: The method of purifying AK auto-Ab is simple, practicable and reliable. Natural AK auto-Ab, existing in normal human individuals, has inhibitory effect on the proliferation of the cultured keratinocytes.     

应用酶联免疫吸附试验检测血清桥粒芯蛋白抗体水平监测寻常型天疱疮患者病情变化

目的:研究较大样本寻常型天疱疮(pemphigus vulgaris,PV)患者,血清抗桥粒芯蛋白(desmoglein,Dsg)1和抗Dsg3特异性抗体酶联免疫吸附试验(enzyme-linked immunosorbent assay,ELISA)指数长期的变化情况,观察其与病情变化的相关性,探讨其用于病情监测、预测疾病复发和指导治疗的可行性。方法:对20例确诊PV的患者进行长期随访,收集患者在各随访时间点的血清,记录每次随访时的病情并进行评分(autoimmune bullous skin disorderintensity score,ABSIS);检测患者血清的间接免疫荧光(indirect immunofluorescence,IIF)滴度、Dsg3 ELISA指数和Dsg1 ELISA指数;利用统计学方法和做图方法分析病情评分与Dsg3 ELISA指数、Dsg1 ELISA指数和IIF滴度之间的关系。结果:PV患者皮肤和口腔的病情评分,分别与Dsg3 ELISA指数、Dsg1 ELISA指数和IIF滴度均具有显著性关联(P<0.01),患者疾病活动期和临床缓解期两组间Dsg3 ELISA指数、Dsg1 ELISA指数和IIF滴度差异均有统计学意义(P<0.01)。各组患者相关性分析发现,Dsg3 ELISA指数、Dsg1 ELISA指数和IIF滴度几乎均与病情平行波动变化,ELISA指数优于IIF的平行性,并且ELISA指数可以预测病情是否会反复,从而指导治疗。结论:PV患者血清抗Dsg3抗体ELISA指数与病情变化平行波动,可以反映疾病的活动程度,可用于病情监测,并可为临床上预测疾病复发,指导临床调整治疗提供有利的实验室证据。     

纤溶酶原激活剂在天疱疮发病中的作用

目的探讨尿激酶型纤溶酶原激活剂（ｕ－ＰＡ）在天疱疮发病过程中的变化以及尿激酶对培养的表皮片的影响。方法应用免疫组织化学法检测天疱疮器官模型中尿激酶抗原的表达情况；发色底物法测定棘层松解模型中纤溶酶原激活剂（ＰＡ）的活性；在培养基中加入尿激酶，观察其对表皮的影响。结果棘层松解模型中，ＰＡ的活性在２４小时后即升高，在４８小时至７２小时内成直线上升；天疱疮器官模型中尿激酶抗原表达增强；纯化的尿激酶可致棘层松解样变化。结论天疱疮抗体激活角朊细胞过多地表达ｕ－ＰＡ，且活性增高，在角朊细胞周围产生一个局限性的蛋白水解过程，引起细胞粘着性的解离，产生棘层松解。     

SSA/Ro antigen expressed on membrane of UVB-induced apoptotic keratinocytes is pathogenic but not detectable in supernatant of cell culture

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伴发副肿瘤性天疱疮的Castleman瘤B细胞免疫球蛋白基因重排与超变分析

目的:我科近来研究了副肿瘤性天疱疮(PNP)患者肿瘤在发病中的作用,证明1例Castleman瘤产生针对表皮连接蛋白的抗体.本文进一步探讨肿瘤和抗体在疾病发生过程中的作用和PNP的发病机制.方法:对1例副肿瘤性天疱疮伴发已经证明可分泌自身反应性抗体的Castleman瘤患者肿瘤B细胞进行培养,免疫组化分析其表面标志;扩增患者肿瘤组织的RNA,将PCR产物克隆、测序.对B细胞克隆的免疫球蛋白可变区基因重排和超变区突变特点进行分析.结果:培养肿瘤细胞多数为CD20、HLA-DR、smIgM、smIgG阳性.克隆得到IgVH与IGHV3-9*01胚系基因片段接近,Ig VL与IGKV4-1*01胚系基因片段同源.VH和VL基因CDRs区的核苷酸改变均明显多于FRs区,并且在VH和VL基因观察到的CDRs区替换突变数目远大于可能发生随机突变的值.结论:本例伴发Castleman瘤的副肿瘤性天疱疮患者的肿瘤中存在特殊B细胞克隆,其已重排的免疫球蛋白可变区基因CDRs超变区发生明显体细胞突变,可能是抗原选择的结果.该克隆经过免疫球蛋白类别转换,可能直接产生与表皮自身抗原特异性结合的自身反应性IgG.     

自身免疫性副肿瘤性天疱疮发病机制的研究进展

副肿瘤性天疱疮(paraneoplastic pemphigus,PNP)是由Anhalt等[1,2]于1990年命名的一种自身免疫性皮肤黏膜疾病,该病特点为伴发肿瘤及血清中存在能识别表皮蛋白成分的独特的IgG自身抗体.我科自1999年诊断了我国首例PNP患者后[3],至今已收治了12例.本文就我科所见PNP结合文献对临床特点作一介绍,并重点探讨其发病机制.     

The mechanism of signal transduction during vascular smooth muscle cell proliferation induced by autoantibodies against angiotensin AT1 receptor from hypertension

Background Autoantibodies against angiotensin AT1 receptor have been discovered in patients with preeclampsia or malignant hypertension. Some studies have demonstrated that the autoantibodies are involved in the immunopathogenesis of hypertension and have an agonist effect similar to angiotensin II. Methods Autoantibodies against AT1 receptor were purified from sera of patients with primary hypertension by affinity chromatography. Proliferation of cultured rat vascular smooth muscle cells was detected by bromodeoxyuridine incorporation and activation of signalling molecules detected by Western blotting and electrophoretic mobility shift assay. Results The AT1-RAb caused a significant proliferation similar to the Ang II during first 24 hours. The levels of nuclear factor-KB （NF-KB）, phosphorylated JAK2., phosphorylated STAT1 （pSTAT1） and phosphorylated STAT3 （pSTAT3） molecules were increased in response to the autoantibodies. In contrast, the activations of NF-KB and JAK-STAT were blocked by Iosartan, pyrrolidinedithiocarbamate （a specific inhibitor of NF-KB） and AG490 （a specific inhibitor of the JAK2. tyrosine kinase）. The expressions of NF-KB, pSTAT1 and pSTAT3 reached peak levels at different times. Moreover, the relative densities of electrophoretic bands showed that activation of pSTAT3 was more significant than STAT1 induced by AT1 -RAb. Conclusions These results suggest that the autoantibodies against AT1 receptor have an agonist effect similar to Ang II in proliferation of VSMCs and the NF-KB and JAK-STAT proteins play essential roles. The effect is different from Angll in that STAT3 is the main downstream activating molecule in JAK-STAT signalling pathway.     

Autoantibodies to survivin in the sera of patients with infiltrating ductal carcinoma of the breast

Autoantibodies to survivin have been reported in lung cancers and in gastrointestinal cancers. A few reports have also described a low prevalence of autoantibodies to survivin and at low titres in the sera of breast cancer patients with no implications for their clinical usefulness. This study was designed to re-examine the prevalence and the clinical correlations of autoantibodies to the tumour protein survivin, in the sera of patients with infiltrating ductal carcinoma of the breast using an ELISA assay. In spite of the low prevalence of autoantibodies to survivin (7%, n = 57), their presence was associated with grade III tumours, with tumour sizes exceeding 10cm, with axillary lymph nodal involvement and with metastases. Moreover, all the autoantibody-positive cases were estrogen and progesterone receptors negative. Furthermore, all the autoantibody-positive cases expressed survivin with high scores.     

Pemphigoid: clinical, histologic, immunopathologic, and therapeutic considerations

Autoimmune blistering diseases are generally distinct entities characterized by relatively consistent clinical, histologic, and immunopathologic findings. These disorders may cause impaired adhesion of epidermis to epidermal basement membrane (eg, the pemphigoid group of disorders [bullous, gestational, and mucous membrane]) or impaired adhesion of epidermal cells to each other (eg, the pemphigus group of disorders). Recent studies have shown that these disorders are characterized by autoantibodies that often display pathogenic (ie, blister-forming) activity in passive transfer models. Interestingly, the autoantigens targeted by these patients' autoantibodies represent important structural proteins that promote cell matrix (eg, pemphigoid) or cell-to-cell (eg, pemphigus) adhesion in skin. Autoimmune blistering diseases are characterized by substantial morbidity (pruritus, pain, disfigurement), and in some instances, mortality (secondary to loss of epidermal barrier function). Treatment with systemic immunosuppressives has reduced morbidity and mortality in patients with these diseases. JAMA. 2000;284:350-356     

原发性高血压与心脏β2、α1及AT1受体自身抗体的关系

目的探讨抗G-蛋白偶联型β2、α1及AT1受体的自身抗体在原发性高血压病程中的分布情况。方法应用酶联免疫吸附测定(ELISA)技术,以细胞外第二环表位肽段的合成肽作为抗原,检测50例高血压心脏病、40例单纯高血压和40例正常人血清中抗G-蛋白偶联家族中β2、α1和AT1受体的自身抗体。结果高血压心脏病患者血清中抗G-蛋白偶联型β2、α1和AT1受体自身抗体的阳性率明显高于单纯高血压和正常组(P<0.05);高血压心脏病自身抗体阳性的平均几何滴度与单纯性高血压比较无明显差异(P>0.05),但两组阳性抗体的平均几何滴度均明显高于正常组。高血压心脏病抗β2受体自身抗体阳性患者中,81.0%同时具有α1受体的自身抗体,76.2%同时具有AT1受体的自身抗体,52.4%存在上述3种受体的自身抗体。结论抗G-蛋白偶联型β2、α1及AT1受体的自身抗体参与原发性高血压的病理生理过程,可能与心肌和血管重构有关。     

从牛舌中提取寻常型天疱疮抗原的方法及其应用

目的：探讨从牛舌黏膜上皮中提取寻常型天疱疮抗原的可行性及其在天疱疮研究中的意义。方法：应用8mol/L尿素浸泡法制备牛舌黏膜上皮提取物,免疫印迹分析寻常型天疱疮患者血清中天疱疮抗体结合的靶抗原。结果：13例寻常型天疱疮全部检出IgG型和IgA型天疱疮抗体,它们分别识别牛舌黏膜上皮提取物中260ku,230ku,210ku,190ku,165ku,160ku,135ku,115ku,97ku,85ku,65ku和62ku多肽中一种或几种抗原。结论：从牛舌中提取寻常型天疱疮抗原的方法可行,适合于天疱疮抗体的免疫印迹检测。     

心脏β1和M2受体自身抗体与心力衰竭相关性的研究

目的:本研究拟对不同心脏病所致的心力衰竭(CHF)患者,检测心脏β1和M2受体的自身抗体,探讨心功能发生病理变化时,这两种自身抗体的产生与疾病发生、发展的相关性。方法:以细胞外第2环表位肽段的合成肽作为抗原,应用酶联免疫吸附测定(ELISA)技术,随机检测265例受试者血清中心脏β1和M2受体的自身抗体。结果:CHF组β1受体自身抗体的阳性率为45.7%(86/188),明显高于对照组的10.4%(8/77)(P<0.01);CHF组M2受体自身抗体的阳性率为49.5%(93/188),明显高于对照组的11.7%(9/77)(P<0.01);心功能Ⅱ-Ⅲ级(NYHA心功能分级)的患者自身抗体的阳性率及抗体滴度明显高于Ⅳ级;CHF组β1受体自身抗体阳性血清中高达56.1%的患者同时具有M2受体的自身抗体。结论:心脏β1和M2受体自身抗体存在于多种心脏病所致心力衰竭患者的血清中,可能与心力衰竭时心肌结构变化和功能下降有关;β1和M2受体的双抗体阳性可能是自身免疫反应的多重性表现,提示免疫学机制参与心力衰竭和/或心肌重构的病理生理过程,参与的程度在疾病的早、中期大于晚期。     

利用重组抗原检测抗SSB自身抗体及其临床意义

目的：利用重组抗原检测自身免疫疾患者血清中抗ＳＳＢ自身抗体并探讨其临床意义。方法：应用ＤＮＡ重组技术构建ＳＳＢ基因工程菌，并表达ＳＳＢ融合蛋白。经ＧＳＴ柱层析法化后，分别经蛋白质印迹（ＷＢＴ）法和酶联免疫吸附（ＥＬＩＳＡ）法检测２０坐干燥综合征患者、６０份系统性红斑狼疮患者和３０份正常人血清。结果：经常规试剂盒检测为ＳＳＢ（＋）的２４份患者血清，利用重组抗原检测均阳性，经常剂剂盒检测为ＳＳＢ（－）     

天然抗角蛋白自身抗体对体外培养角质形成细胞细胞周期的影响

目的：研究天然抗角蛋白自身抗体对体外培养角质形成细胞细胞周期的影响。方法亲和层析法从混和人血清中提取纯化天然抗角蛋白自身抗体,然后通过SDS聚丙烯酰胺凝胶电泳对纯化得到的抗体进行纯度分析,利用MTT比色法与流式细胞仪分析,观察纯化抗体对培养人角质形成细胞细细胞周期的影响。结果：纯化的抗角蛋白自身抗体纯度已达到的电泳级,MTT比色法的结果表明天然抗角蛋白身身抗体对培养的人角质形成细胞具有显著的剂量依赖性的抑制作用。相关系数为－0．797,流式细胞仪分析显示,浓度为0．01125g.L^-1的抗角蛋白自身抗体处理72h后,处于G2／M＋S期的角度质形成细胞细比例从56．2％下降至46．7％,而处于G1／G0期的细胞比例相应从43．8％升至53．3％,结论从正常人血清中提取的天然抗角蛋白身身抗体对体外培养的人角质形成细胞细的增殖具有抑制作用,使多数角质形成细胞细滞留于G1／G0期。     

儿童天疱疮抗体IgG亚型及与天疱疮抗原的反应性

目的观察寻常型天疱疮（Pemphigus vulgaris,PV）患儿和一级亲属血清中PV自身抗体（PV-IgG）亚型及其与桥粒芯糖蛋白（Desmoglein,Dsg）反应性,探讨PV-IgG检测的临床意义和PV的发病机制。方法用间接免疫荧光法（Indirect immunofluorescenceI,IF）检测5例PV患儿、12例一级亲属和15例正常对照者血清PV-IgG,并用免疫印迹法（Western blot,WB）检测PV-IgG亚型及其与Dsg1、Dsg3反应性。结果 PV患儿的PV-IgG阳性率和平均抗体滴度明显高于其一级亲属者（P〈0.01）,活动期患儿的PV-IgG阳性率和平均抗体滴度也明显高于缓解期（P〈0.01）;PV患儿中PV-IgG阳性率（80.0%）明显高于一级亲属（25.0%）和正常对照组（0.0%）（P〈0.01）,PV患儿中Dsg3反应性IgG4水平明显高于一级亲属（P〈0.01）,且其在活动期病例中水平也明显高于缓解期（P〈0.05）。结论寻常型天疱疮的发生与PV-IgG水平有关;Dsg3是PV的主要自身抗原I;gG4是PV的主要致病抗体。     

Vaccination with a Recombinant Chicken FGFR-1 Bypasses Immunological Tolerance against Self-FGFR-1 in Mice

It has been generally believed that that the growth and metastasis of solid tumors depend on tumor angiogenesis, the generation of new blood vessels. At present, basic fibroblast growth factor (bFGF) has been shown to be one of the most important angiogenic growth factors for tumor angiogenesis. bFGF serves its biological functions through interaction with its high-affinity receptor, fibro- blast growth factor receptor-1(FGFR-1)[1–3]. FGFR-1 is a member of the receptor tyrosine kinase co…     

鼻咽癌患者血清中自身抗体的初步检测

目的:探讨鼻咽癌患者血清中是否存在抗鼻咽癌自身抗原的抗体,为寻找鼻咽癌新的肿瘤标志物提供依据.方法:制备Epstein-Barr病毒阴性的鼻咽癌细胞株CNE1细胞板,酶联免疫吸附实验(ELISA)分析其与32例鼻咽癌患者血清和54例正常血清反应的差异性;提取CNEl总蛋白,Western印迹分析其与鼻咽癌患者血清反应是否存在特异性的蛋白质.结果:ELISA分析显示鼻咽癌患者血清平均滴度(0.904±0.032)明显高于正常人血清平均滴度(0.736±0.028)(P＜0.01);Western印迹分析发现鼻咽癌患者血清与正常人血清相比,部分阳性条带一致,但条带强度增加,还发现出现了部分新的阳性条带,这些条带可能属于鼻咽癌相关抗原或鼻咽癌特异性抗原.结论:鼻咽癌患者血清中存在抗鼻咽癌自身抗原的抗体,这些抗体并非针对Epstein-Barr病毒,为寻找鼻咽癌血清肿瘤标志物提供了依据.