天疱疮抗体滴度与疾病活动性及CD19、CD20的相关性研究
发布时间：2018-08-03 11:15
【摘要】： 天疱疮是一组以天疱疮抗体介导的器官特异性自身免疫性疾病,发病率虽然很低,但呈上升趋势。好发于30-50岁中年人群,无明显性别差异。临床上主要分为四型：寻常型、增殖型、落叶型、红斑型,但四型之间可相互转化。此外,还有疱疹样天疱疮、IgA天疱疮、药物诱发的天疱疮、副肿瘤性天疱疮、流行性性天疱疮(又名巴西天疱疮)等亚型。该组疾病的共同特征有：临床表现为疱壁薄、松弛易破的水疱或大疱形成；组织病理为棘层松解所致表皮内水疱形成；直接免疫荧光检查(indirect immunofluorescence,IIF)可见棘层细胞间IgG和C3沉积,取患者血清做间接免疫荧光检查,可显示血清中有抗表皮棘细胞间物质抗体。 天疱疮的发病机理尚未完全清楚。越来越多研究认为在天疱疮发病过程中,T细胞和B细胞相互作用、相互影响,产生针对自身细胞间物质的抗体,从而导致疾病的发生。天疱疮的抗原主要集中在桥粒,桥粒是角质形成细胞间连接的主要结构,由相邻细胞的细胞膜发生卵圆形致密增厚而共同构成。由两类蛋白质构成：一类是跨膜蛋白,位于桥粒芯(desmosomal core),主要由桥粒芯蛋白(desmoglein,Dsg)和桥粒芯胶蛋白(desmocollin,Dsc)构成,它们形成桥粒的电子透明细胞间隙和细胞间接触层；另一类为胞质内的桥粒斑(desmosomal plaque)蛋白,是盘状附着板的组成部分,主要成分为桥粒斑蛋白(desmoplakin,DP)和桥粒斑珠蛋白(plakogloubin,PG)。天疱疮抗体(Pemphigus antibody)与抗原结合后,通过细胞信号传导途径激活一系列蛋白水解酶,导致细胞间链接结构水解,从而引起表皮棘层细胞相互分离、棘层松解及表皮内水疱形成。钙粘素是Ca2+依赖的细胞间连接分子,它对表皮细胞间粘附连接起重要作用。天疱疮抗原的c DNA序列与钙粘素有明显的同源性,因此天疱疮抗体也损害了表皮细胞间的粘连功能,导致棘层松解。棘层松解在PF患者较表浅,损害仅达表皮颗粒层。而在PV患者则深达表皮生发层。几乎所有天疱疮患者免疫病理均示角质形成细胞间IgG和C3呈网状沉积。 天疱疮的诊断主要结合四个方面因素考虑：1、典型的临床皮损表现；2、皮肤组织病理所示的棘层松解；3、免疫病理示角质形成细胞间IgG和C3呈网状沉积；4、组织上发现患者血清中或原位皮肤存在自身抗桥粒芯糖蛋白抗体。目前天疱疮治疗方法很多,但皮质类固醇激素仍是目前的首选,开始剂量需较大,以期在1-2周内抑制新的水疱形成,必要时可应用皮质类固醇激素冲击疗法或加用免疫抑制剂治疗。根据患者皮损面积分为轻、中、重度并给与适合剂量的激素,以不出现新的水疱为宜。皮损面积体表面积10%为轻度,激素量为30mg/d；体表面积30%皮损面积体表面积50%为中度,激素量为60-80 mg/d；皮损面积体表面积50%为重度,激素量80 mg/d。此外,部分皮损面积很局限的天疱疮患者在口服少量激素的同时加用激素软膏或他克莫司软膏外用可促进皮损的愈合。 但目前天疱疮诊治中最大的问题是明确诊断和判断疾病的活动程度及预后。尼氏征对于天疱疮的诊断具有较高的特异性,但其敏感性较差。直接免疫荧光(direct immunofluorescence,DIF)检查是诊断天疱疮的金标准,但反复的皮肤活检术给病人造成较大的痛苦,难以为广大的患者所接受,也难以作为随访的标准。根据已知的天疱疮发病机制,本实验尝试检测血循环中的天疱疮抗体对天疱疮患者进行诊断,并探讨其与疾病的活动性的关系,以寻找出适合临床诊断、判断疗效和随访的指标。 目的： 1、通过酶联免疫吸附(Enzyme-linked immunosorbent assay,ELISA)检测天疱疮患者血清中的抗体IgG滴度,了解其敏感性和特异性。 2、对天疱疮患者的病情活动度进行评分,探讨其与天疱疮抗体滴度的相关性。 3、通过流式细胞仪(Flow cytometry,FCM)检测天疱疮患者血清中CD19、CD20的表达水平,探讨天疱疮抗体滴度与CD19、CD20表达水平的相关性。 方法： 1、收集2009-1至2010-2之间门诊及住院并确诊的38例天疱疮患者及非天疱疮患者22人的血清,离心后用天疱疮抗体检测试剂盒(MBL公司),通过ELISA法检测60份血清中天疱疮抗体IgG的滴度。了解该方法与临床的相符性,并探讨其敏感性和特异性。 2、对确诊的天疱疮患者进行病情活动度评分,分析疾病病情活动评分与天疱疮抗体滴度进行相关性。 3、将38例天疱疮患者中的28位患者的血清经流式细胞技术检测其中CD19、CD20的表达水平,并与之前检测的相应的天疱疮抗体滴度进行多元相关性分析。 结果： 1、诊断的符合度、敏感性和特异性：38例天疱疮患者中有37例阳性,1例阴性。22例非天疱疮患者则均为阴性。该方法的诊断符合率为98.3%,敏感性为97.4%,特异性为100%。 2、病情活动度评分与天疱疮抗体滴度的Pearson双变量相关性分析：病情活动度评分与天疱疮抗体滴度之间的相关性具有统计学意义(P0.001),相关系数r=0.671,直线回归方程为：病情活动评分=1.079+0.671×抗体滴度。 3、天疱疮抗体滴度与血清CD19、CD20表达水平的多元回归分析：天疱疮抗体滴度与血清CD19、CD20表达水平的的相关性亦具有统计学意义(P0.05),将数据进行对数转换后,其复相关系数R=0.55,R2=0.302,lgCD19(x1)的偏回归系数为0.518,lgCD20(x2)的偏回归系数为0.066,截距为1.674,得出回归方程为：Lg抗体滴度=1.674+0.518×LgCD19+0.066×LgCD20。 结论： 1、该实验进一步证实了天疱疮抗体用于天疱疮的诊断具有较高的敏感性及特异性,与临床符合率较高,对疾病的诊断具有重要的意义。 2、天疱疮抗体抗体滴度越高,疾病的活动性越强,与疾病活动度相平行,具有较高的相关性。可用于疾病活动度的判断、治疗效果的评价随访的指标。 3、天疱疮抗体滴度与患者血清中的CD19、CD20的表达水平亦具有相关性,CD19、CD20的表达水平越高,血清中的抗体的滴度也越高。
[Abstract]:Pemphigus is an organ specific autoimmune disease mediated by pemphigus antibody. Although the incidence of pemphigus is very low, the incidence of pemphigus is on the rise. There are no significant gender differences in the 30-50 year old middle-aged people. It is mainly divided into four types: vulgaris, proliferating, deciduous, erythema, but the type four can be transformed from each other. In addition, there are herpes samples. Pemphigus, pemphigus IgA, pemphigus induced by drugs, paraneoplastic pemphigus, and pemphigus epidemic (also known as pemphigus Brazil). The common characteristics of the disease are: the clinical manifestation is the thin wall of the blister, the loosening blister or bulla; the histopathology is the formation of the blister caused by the spinous release of the epidermis; direct immunofluorescence. Indirect immunofluorescence (IIF) shows the deposition of IgG and C3 between the spinous cells, and the indirect immunofluorescence examination of the patient's serum shows that there are anti epidermal acanthosis antibodies in the serum.
The pathogenesis of pemphigus is not completely clear. More and more studies have shown that in the process of pemphigus, T and B cells interact and interact with each other to produce antibodies against their own intercellular substances, which lead to the occurrence of disease. The antigen of pemphigus is mainly concentrated in the pemphigus, and the pemphigus is the main junction of the keratinocytes. The structure, composed of two kinds of proteins, consists of two kinds of proteins: a kind of transmembrane protein, located in the grain core (desmosomal core), mainly composed of desmoglein, Dsg and desmocollin, Dsc, which form the transparent space and intercellular space of the bridging particles. The contact layer; the other kind is the desmosomal plaque protein in the cytoplasm, which is a component of the disc attachment plate. The main components are desmoplakin, DP and plakogloubin, PG. Pemphigus antibody (Pemphigus antibody) activates a series of protein water through the cell signal transduction pathway after the combination of the antibody (Pemphigus antibody) with the anti original. The hydrolytic enzyme leads to the hydrolysis of intercellular links, resulting in the isolation of the epidermal acanthosis cells, the acanthosis and the formation of the blisters in the epidermis. The cadherin is a Ca2+ dependent intercellular junction molecule, which plays an important role in the adhesion and connection between the epidermis cells. The C DNA sequence of pemphigus antigen has obvious homology with the cadherin, so pemphigus is resistant to pemphigus. The body also damaged the adhesion function between the epidermal cells and resulted in the acanthosis. The acanthosis was superficial in the PF patients and the damage was only to the epidermis. In the patients with PV, the epidermis was deep. The immunhistopathology of almost all pemphigus showed that the IgG and C3 were meshed between the keratinocytes.
The diagnosis of pemphigus was mainly combined with four factors: 1, typical clinical skin lesions, 2, acanthosis of the skin histopathology; 3, IgG and C3 in the keratinocytes of the immunhistopathology showed reticular deposition; 4, the tissue was found in the sera or in situ in its own anti pemphigoprotein antibody. There are many treatments, but corticosteroids are still the first choice. The initial dose should be large in order to inhibit the formation of new blisters within 1-2 weeks. It is necessary to use corticosteroid shock therapy or immunosuppressive therapy when necessary. According to the area of skin lesions, the patients are divided into light, medium, severe and given the suitable dose of hormone, in order not to appear. The surface area of the skin area was mild, the surface area of the skin lesion was 10%, the volume of the body surface area was 30mg/d, the body surface area 30% skin area 50% was moderate, the hormone amount was 60-80 mg/d, the skin area surface area 50% was 50%, the hormone was 80 mg/d., and the patients with the limited area of skin lesions were combined with a small amount of hormone at the same time. External application of plain ointment or Tacrolimus Ointment can promote the healing of skin lesions.
But at present, the biggest problem in the diagnosis and treatment of pemphigus is to diagnose and judge the degree of activity and prognosis of the disease. Nil's sign has a high specificity for the diagnosis of pemphigus, but its sensitivity is poor. Direct immunofluorescence (direct immunofluorescence, DIF) is the gold standard for the diagnosis of pemphigus, but repeated skin biopsy is given to the patients. According to the known pathogenesis of pemphigus, this experiment is trying to detect pemphigus antibodies in the blood circulation to diagnose pemphigus, and to explore the relationship between pemphigus and the activity of the disease in order to find a suitable clinical diagnosis, to judge the curative effect and follow it. The target of the visit.
Objective:
1. Enzyme-linked immunosorbent assay (ELISA) was used to detect the antibody IgG titer in the serum of patients with pemphigus.
2, we assessed the degree of disease activity in pemphigus patients and explored the correlation between pemphigus antibody titer and pemphigus antibody titer.
3, the expression level of CD19 and CD20 in the serum of pemphigus patients was detected by Flow cytometry (FCM), and the correlation between the tipper titer antibody titer and the expression of CD19 and CD20 was investigated.
Method:
1, the serum of 38 pemphigus patients and 22 pemphigus patients were collected from 2009-1 to 2010-2 in outpatients and hospitalized. After centrifugation, the titer antibody test kit (MBL company) was used to detect the titer of the pemphigus antibody IgG in 60 sera and to understand the coincidence of this method with the clinical and its sensitivity and specificity.
2. The patients with pemphigus were assessed by disease activity score, and the correlation between disease activity score and pemphigus antibody titer was analyzed.
3, the serum level of CD19 and CD20 was detected by serum flow cytometry in 28 patients with pemphigus, and the correlation was analyzed with the correlation of the antibody titer of pemphigus which was detected before.
Result:
1, diagnostic accuracy, sensitivity and specificity: 37 cases were positive in 38 pemphigus patients and 1 negative.22 cases were negative. The diagnostic coincidence rate of this method was 98.3%, sensitivity was 97.4%, and the specificity was 100%..
2, the correlation of Pearson bivariate analysis of disease activity score and pemphigus antibody titer: the correlation between disease activity score and pemphigus antibody titer was statistically significant (P0.001), the correlation coefficient was r=0.671, and the linear regression equation was =1.079 +0.671 x antibody titer.
3, multiple regression analysis of the titer's titer antibody titer and the level of serum CD19, CD20 expression: the correlation between the tipper titer antibody titer and the serum CD19, CD20 expression level also has statistical significance (P0.05). After the logarithmic conversion of the data, the partial regression coefficient of the correlation coefficient R=0.55, R2=0.302, lgCD19 (x1) is 0.518, lgCD20 (x2) partial regression The coefficient is 0.066 and the intercept is 1.674. The regression equation is: Lg antibody titer =1.674+0.518 * LgCD19+0.066 * LgCD20.
Conclusion:
1, the experiment further confirmed that pemphigus antibody has high sensitivity and specificity in the diagnosis of pemphigus, and it has a higher coincidence with the clinic. It is of great significance for the diagnosis of the disease.
2, the higher the titer antibody titer, the stronger the activity of the disease, and the higher correlation with the degree of disease activity. It can be used to judge the degree of disease activity and evaluate the outcome of the treatment.
3, the titer of pemphigus was also associated with the expression level of CD19 and CD20 in the patient's serum. The higher the expression level of CD19, the higher the level of CD20, the higher the titer of the antibody in the serum.
【学位授予单位】：南方医科大学
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R758.66
本文编号：2161573
[Abstract]:Pemphigus is an organ specific autoimmune disease mediated by pemphigus antibody. Although the incidence of pemphigus is very low, the incidence of pemphigus is on the rise. There are no significant gender differences in the 30-50 year old middle-aged people. It is mainly divided into four types: vulgaris, proliferating, deciduous, erythema, but the type four can be transformed from each other. In addition, there are herpes samples. Pemphigus, pemphigus IgA, pemphigus induced by drugs, paraneoplastic pemphigus, and pemphigus epidemic (also known as pemphigus Brazil). The common characteristics of the disease are: the clinical manifestation is the thin wall of the blister, the loosening blister or bulla; the histopathology is the formation of the blister caused by the spinous release of the epidermis; direct immunofluorescence. Indirect immunofluorescence (IIF) shows the deposition of IgG and C3 between the spinous cells, and the indirect immunofluorescence examination of the patient's serum shows that there are anti epidermal acanthosis antibodies in the serum.
The pathogenesis of pemphigus is not completely clear. More and more studies have shown that in the process of pemphigus, T and B cells interact and interact with each other to produce antibodies against their own intercellular substances, which lead to the occurrence of disease. The antigen of pemphigus is mainly concentrated in the pemphigus, and the pemphigus is the main junction of the keratinocytes. The structure, composed of two kinds of proteins, consists of two kinds of proteins: a kind of transmembrane protein, located in the grain core (desmosomal core), mainly composed of desmoglein, Dsg and desmocollin, Dsc, which form the transparent space and intercellular space of the bridging particles. The contact layer; the other kind is the desmosomal plaque protein in the cytoplasm, which is a component of the disc attachment plate. The main components are desmoplakin, DP and plakogloubin, PG. Pemphigus antibody (Pemphigus antibody) activates a series of protein water through the cell signal transduction pathway after the combination of the antibody (Pemphigus antibody) with the anti original. The hydrolytic enzyme leads to the hydrolysis of intercellular links, resulting in the isolation of the epidermal acanthosis cells, the acanthosis and the formation of the blisters in the epidermis. The cadherin is a Ca2+ dependent intercellular junction molecule, which plays an important role in the adhesion and connection between the epidermis cells. The C DNA sequence of pemphigus antigen has obvious homology with the cadherin, so pemphigus is resistant to pemphigus. The body also damaged the adhesion function between the epidermal cells and resulted in the acanthosis. The acanthosis was superficial in the PF patients and the damage was only to the epidermis. In the patients with PV, the epidermis was deep. The immunhistopathology of almost all pemphigus showed that the IgG and C3 were meshed between the keratinocytes.
The diagnosis of pemphigus was mainly combined with four factors: 1, typical clinical skin lesions, 2, acanthosis of the skin histopathology; 3, IgG and C3 in the keratinocytes of the immunhistopathology showed reticular deposition; 4, the tissue was found in the sera or in situ in its own anti pemphigoprotein antibody. There are many treatments, but corticosteroids are still the first choice. The initial dose should be large in order to inhibit the formation of new blisters within 1-2 weeks. It is necessary to use corticosteroid shock therapy or immunosuppressive therapy when necessary. According to the area of skin lesions, the patients are divided into light, medium, severe and given the suitable dose of hormone, in order not to appear. The surface area of the skin area was mild, the surface area of the skin lesion was 10%, the volume of the body surface area was 30mg/d, the body surface area 30% skin area 50% was moderate, the hormone amount was 60-80 mg/d, the skin area surface area 50% was 50%, the hormone was 80 mg/d., and the patients with the limited area of skin lesions were combined with a small amount of hormone at the same time. External application of plain ointment or Tacrolimus Ointment can promote the healing of skin lesions.
But at present, the biggest problem in the diagnosis and treatment of pemphigus is to diagnose and judge the degree of activity and prognosis of the disease. Nil's sign has a high specificity for the diagnosis of pemphigus, but its sensitivity is poor. Direct immunofluorescence (direct immunofluorescence, DIF) is the gold standard for the diagnosis of pemphigus, but repeated skin biopsy is given to the patients. According to the known pathogenesis of pemphigus, this experiment is trying to detect pemphigus antibodies in the blood circulation to diagnose pemphigus, and to explore the relationship between pemphigus and the activity of the disease in order to find a suitable clinical diagnosis, to judge the curative effect and follow it. The target of the visit.
Objective:
1. Enzyme-linked immunosorbent assay (ELISA) was used to detect the antibody IgG titer in the serum of patients with pemphigus.
2, we assessed the degree of disease activity in pemphigus patients and explored the correlation between pemphigus antibody titer and pemphigus antibody titer.
3, the expression level of CD19 and CD20 in the serum of pemphigus patients was detected by Flow cytometry (FCM), and the correlation between the tipper titer antibody titer and the expression of CD19 and CD20 was investigated.
Method:
1, the serum of 38 pemphigus patients and 22 pemphigus patients were collected from 2009-1 to 2010-2 in outpatients and hospitalized. After centrifugation, the titer antibody test kit (MBL company) was used to detect the titer of the pemphigus antibody IgG in 60 sera and to understand the coincidence of this method with the clinical and its sensitivity and specificity.
2. The patients with pemphigus were assessed by disease activity score, and the correlation between disease activity score and pemphigus antibody titer was analyzed.
3, the serum level of CD19 and CD20 was detected by serum flow cytometry in 28 patients with pemphigus, and the correlation was analyzed with the correlation of the antibody titer of pemphigus which was detected before.
Result:
1, diagnostic accuracy, sensitivity and specificity: 37 cases were positive in 38 pemphigus patients and 1 negative.22 cases were negative. The diagnostic coincidence rate of this method was 98.3%, sensitivity was 97.4%, and the specificity was 100%..
2, the correlation of Pearson bivariate analysis of disease activity score and pemphigus antibody titer: the correlation between disease activity score and pemphigus antibody titer was statistically significant (P0.001), the correlation coefficient was r=0.671, and the linear regression equation was =1.079 +0.671 x antibody titer.
3, multiple regression analysis of the titer's titer antibody titer and the level of serum CD19, CD20 expression: the correlation between the tipper titer antibody titer and the serum CD19, CD20 expression level also has statistical significance (P0.05). After the logarithmic conversion of the data, the partial regression coefficient of the correlation coefficient R=0.55, R2=0.302, lgCD19 (x1) is 0.518, lgCD20 (x2) partial regression The coefficient is 0.066 and the intercept is 1.674. The regression equation is: Lg antibody titer =1.674+0.518 * LgCD19+0.066 * LgCD20.
Conclusion:
1, the experiment further confirmed that pemphigus antibody has high sensitivity and specificity in the diagnosis of pemphigus, and it has a higher coincidence with the clinic. It is of great significance for the diagnosis of the disease.
2, the higher the titer antibody titer, the stronger the activity of the disease, and the higher correlation with the degree of disease activity. It can be used to judge the degree of disease activity and evaluate the outcome of the treatment.
3, the titer of pemphigus was also associated with the expression level of CD19 and CD20 in the patient's serum. The higher the expression level of CD19, the higher the level of CD20, the higher the titer of the antibody in the serum.
【学位授予单位】：南方医科大学
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R758.66
【参考文献】
相关期刊论文 前3条
1 杨卫兵,刁庆春,叶庆佾;间接ELISA和ABCELISA法对天疱疮抗体检测的研究[J];中华皮肤科杂志;1998年04期
2 李渝,刁庆春,王鲁,钟白玉,唐书谦,叶庆佾,刘荣卿;天疱疮抗体亚类在天疱疮发病中的作用[J];中华皮肤科杂志;2000年04期
3 聂祝湘,刘荣卿,叶庆佾,唐书谦,钟白玉;天疱疮抗体对桥粒影响的动态观察[J];中华医学杂志;1996年09期
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