神经精神狼疮患者脑脊液及血清蛋白质组学研究

发布时间：2018-03-22 21:23

本文选题：神经精神狼疮　切入点：MALDI-TOF-MS　出处：《北京协和医学院》2009年博士论文　论文类型：学位论文

【摘要】：第一部分：神经精神狼疮脑脊液蛋白质指纹图谱及分析目的筛选神经精神狼疮(NPSLE)脑脊液生物标志物,并建立相应NPSLE疾病的树形分类模型。材料和方法用MALDI-TOF-MS联合WCX磁珠生成脑脊液蛋白质谱,比较NPSLE治疗前组(27例)与脊柱侧凸组(17例)、无神经精神症状的系统性红斑狼疮组(non-NPSLE,10例)之间的差异蛋白峰；建立NPSLE的树形分类模型,并用其他中枢神经系统受累疾病(腰椎间盘突出12例,其他结缔组织病相关脑病4例)进行盲法检验。比较17例NPSLE患者治疗前后脑脊液蛋白质谱的差异蛋白峰。结果NPSLE治疗前组与非精神神经狼疮组(脊柱侧凸和non-NPSLE)脑脊液蛋白质谱共有25个差异峰,用其中质荷比为8595、7170、7661、7740和5806差异峰建立NPSLE树形分类模型,其对源数据的敏感性92.6%,特异性92.6%；其他中枢神经系统受累疾病(16例)盲法检验显示该模型分类NPSLE的特异性为81.3%。17例NPSLE患者治疗前后脑脊液质谱有4个差异峰,其中质荷比为4963和8595蛋白峰显著降低,质荷比为6637和6896蛋白峰显著升高。结论MALDI-TOF-MS联合WCX磁珠可用于脑脊液蛋白质组研究,发现了多个潜在脑脊液生物标志物可能用于NPSLE检测、诊断和病情评估,建立了敏感性及特异性均好的NPSLE树形分类模型。第二部分：神经精神狼疮血清蛋白质指纹图谱及分析目的筛选神经精神狼疮(NPSLE)血清生物标志物。材料和方法用MALDI-TOF-MS联合WCX磁珠生成血清蛋白质谱,比较NPSLE治疗前组(24例)与健康对照组(21例)、无神经精神症状的狼疮组(non-NPSLE,8例)之间的差异蛋白峰；比较13例NPSLE治疗前后血清蛋白质谱的差异蛋白峰；比较同一NPSLE患者同期血清与脑脊液质谱相同质荷比差异蛋白峰的相互关系。结果NPSLE治疗前组与健康对照组血清蛋白质谱有35个差异峰,与non-NPSLE组有13个差异峰；non-NPSLE组与健康对照组有15个差异峰,其中质荷比为6438、8122、8690、9280、28081蛋白峰在上述多组之间均存在统计学及质谱图差异。13例NPSLE患者治疗前后血清质谱中质荷比为4959、6189蛋白峰治疗后升高显著。同一NPSLE患者同期血清与脑脊液质谱相同质荷比差异蛋白峰共4对：4959(血清)4963(脑脊液),NPSLE治疗后4959显著升高,而4963显著降低；5905(血清)5866(脑脊液),NPSLE治疗前组均高于正常对照组；7754(血清)7740(脑脊液)和8581(血清)8595(脑脊液),均为NPSLE治疗前组血清水平降低,而脑脊液水平升高。结论发现多个潜在血清生物标志物可能用于神经精神狼疮检测、诊断和病情评估,其中同—NPSLE患者同期血清与脑脊液质谱相同质荷比的4对差异蛋白峰可能更有研究和临床应用价值。第三部分：神经精神狼疮脑脊液蛋白质指纹图谱差异蛋白峰鉴定研究背景及目的血清淀粉样蛋白A(SAA)是一种急性期蛋白,参与激活疾病的多种前炎症反应。多项研究鉴定血清SELDI-TOF-MS质谱的质荷比为11.7KD蛋白峰对应SAA蛋白。本试验拟鉴定脑脊液质谱质荷比11.7KD的差异蛋白峰为SAA蛋白。材料和方法SAA抗体进行脑脊液免疫沉淀后,分别运用MALDI-TOF-MS联合WCX磁珠法和免疫印迹法检测免疫沉淀反应后上清及沉淀蛋白。结果SAA抗体免疫沉淀后上清蛋白质谱中质荷比11.7KD的蛋白峰强度显著下降；SAA抗体免疫沉淀后琼脂糖珠捕捉的蛋白在免疫印迹中出现分子量约为12KD的目的条带。结论脑脊液蛋白质谱的质荷比11.7KD蛋白峰对应SAA蛋白,神经精神狼疮(NPSLE)患者脑脊液SAA水平升高,提示SAA可能成为NPSLE的生物标志物。
[Abstract]:Part one: protein fingerprint and analysis of cerebrospinal fluid in neuromental lupus erythematosus
Objective to screen the neuropsychiatric systemic lupus erythematosus (NPSLE) in cerebrospinal fluid biomarkers, tree classification model and established the corresponding NPSLE disease. Materials and methods of using MALDI-TOF-MS combined with WCX beads cerebrospinal fluid protein mass spectrometry generates NPSLE, before the treatment group (27 cases) and scoliosis group (17 cases), systemic lupus erythematosus group without neuropsychiatric symptoms (10 non-NPSLE cases) between different protein peaks; tree classification model based on NPSLE, and other central nervous system involvement disease (12 cases of lumbar disc herniation and other connective tissue disease related encephalopathy in 4 cases) were blind test. 17 cases of NPSLE patients before and after treatment in different protein peaks. Cerebrospinal fluid protein mass spectrometry results before NPSLE treatment group and non neuropsychiatric lupus group (scoliosis and non-NPSLE) cerebrospinal fluid protein consists of 25 distinct peaks, with the establishment of the NPSLE / Z 8595717076617740 and 5806 differential peaks The tree classification model, the sensitivity of the 92.6% sources of data, the specificity was 92.6%; the other central nervous system involvement disease (16 cases) blind test show the specificity of the NPSLE classification model for 81.3%.17 NPSLE patients before and after treatment of cerebrospinal fluid mass spectrometry 4 differential peaks, the mass ratio of 4963 and 8595 protein peaks were significantly reduced M / Z 6637, and 6896 protein peaks were significantly increased. Conclusion MALDI-TOF-MS combined with WCX can be used for beads of cerebrospinal fluid proteomic research, found a number of potential CSF biomarkers may be used for NPSLE detection, diagnosis and evaluation, established the NPSLE tree model sensitivity and specificity are good.
The second part: the fingerprint and analysis of serum protein of neuromental lupus erythematosus
Objective to screen the neuropsychiatric systemic lupus erythematosus (NPSLE) serum biomarkers. Materials and methods of using MALDI-TOF-MS combined with WCX beads generated serum protein mass spectrometry, comparison of NPSLE before the treatment group (24 cases) and control group (21 cases), lupus group without neuropsychiatric symptoms (8 cases non-NPSLE) between different protein peaks; comparison 13 cases of NPSLE between before and after treatment of serum protein protein interaction peak; compared with the same period of NPSLE patients serum and cerebrospinal fluid mass the same mass of protein peaks. The former group and the healthy control group of serum protein spectrum has 35 distinct peaks results in the treatment of NPSLE, there are 13 differences between the peak and the non-NPSLE group; non-NPSLE group and health the control group had 15 distinct peaks, including 643881228690928028081 / Z protein peaks in the group are in NPSLE patients before and after treatment of.13 and statistical difference spectra of serum in mass spectrometry 49596189 protein peaks after treatment increased significantly. The same year Xue Qing NPSLE patients with cerebrospinal fluid mass mass the same protein peaks were 4 to 4963: 4959 (Xue Qing), NPSLE (CSF) after treatment 4959 significantly increased, and 4963 decreased significantly; 5905 (Xue Qing 5866), NPSLE (CSF) before treatment group were higher than that of the normal control group; 7754 (Xue Qing) 7740 (cerebrospinal fluid) and 8581 (Xue Qing, 8595) (cerebrospinal fluid) were NPSLE group before treatment reduced the level of Xue Qing, and cerebrospinal fluid levels. Conclusion found a number of potential biomarkers for Xue Qing may God by the spirit of lupus detection, diagnosis and evaluation, of which 4 of the difference NPSLE with the same period of Xue Qing and the same mass mass cerebrospinal fluid protein peaks may be more research and clinical application value.
The third part: identification of differential protein peaks of protein fingerprints in cerebrospinal fluid of neuromental lupus erythematosus
Background and objective: serum amyloid A (SAA) is an acute phase protein, is involved in many inflammatory diseases. Many studies identified the activation of serum SELDI-TOF-MS MS / Z 11.7KD protein peaks corresponding to SAA protein. The aim of this study is to identify the different mass mass 11.7KD cerebrospinal fluid protein peak is SAA protein. SAA antibody material and the methods of cerebrospinal fluid after immunoprecipitation, respectively using MALDI-TOF-MS WCX combined detection of immune magnetic beads and Western blotting precipitation supernatant and protein precipitation after the reaction. Results the protein peaks of the supernatant protein mass spectrometry SAA antibodies after immunoprecipitation in mass of 11.7KD decreased significantly; SAA antibodies after immunoprecipitation agarose beads to capture protein in immunoblots in the molecular weight of about 12KD to Z band. 11.7KD protein peaks of SAA protein in cerebrospinal fluid protein. Conclusion neuropsychiatric systemic lupus erythematosus (NPSL E) the level of SAA in the cerebrospinal fluid (CSF) increases, suggesting that SAA may be a biomarker for NPSLE.

【学位授予单位】：北京协和医学院
【学位级别】：博士
【学位授予年份】：2009
【分类号】：R593.241;R3411

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