Dystrophin、Utrophin和nNOS在不同类型肌营养不良症中的表达研究

发布时间：2018-02-28 22:04

本文关键词： 抗肌萎缩蛋白抗肌萎缩蛋白相关蛋白神经元型一氧化氮合酶肌营养不良症免疫荧光　出处：《郑州大学》2016年硕士论文　论文类型：学位论文

【摘要】：研究背景在欧盟,肌营养不良症的发病率为1/2000,病情进展缓慢、致残率高、预后差,目前尚无有效的治疗手段,有关疾病诊断和治疗方面的研究显得尤为重要。抗肌萎缩蛋白(dystrophin)对维持肌纤维的结构完整性具有重要作用,dystrophin基因突变引起Duchenne型肌营养不良症(DMD)和Becker型肌营养不良症(BMD)。抗肌萎缩蛋白相关蛋白(utrophin)是dystrophin的同源蛋白,两种蛋白结构高度相似,utrophin主要在再生肌纤维中表达并维持肌纤维结构的完整性,在成熟肌纤维中几乎全部被dystrophin替代,近年来通过动物实验发现utrophin在成熟肌纤维中的表达上调可能减少DMD患者肌纤维的损伤。神经元型一氧化氮合酶(n NOS)能产生一氧化氮(NO)增加骨骼肌运动中的血供,且在结构上与dystrophin关系密切相关,n NOS的异常表达可能与患者的运动不耐受症状有关。目前国内外关于dystrophin、utrophin和n NOS在不同类型肌营养不良症中的表达研究较少,该研究为明确上述3种蛋白在不同类型肌营养不良症中的表达情况为肌营养不良症的诊断、分型提供依据,并进一步了解3种蛋白表达与DMD患者病情严重程度的关系。探索dystrophin、utrophin和n NOS在不同类型肌营养不良症中的表达,进一步研究3种蛋白的表达水平与Duchenne型肌营养不良症(DMD)患者病情严重程度的关系,及DMD患者中dystrophin、n NOS和utrophin之间的表达关系。目的方法1.收集从2013年3月到2015年5月来我院就诊并诊断明确的DMD 26例、BMD7例、肢带型肌营养不良症(LGMD)10例、面肩肱型肌营养不良症(FSHD)5例、远端型肌营养不良症5例和强直性肌营养不良症(DM)11例。收集同时间段行肌肉活检病理结果正常者或急诊外伤患者作为对照组21例。利用免疫荧光方法检测3种蛋白在肌营养不良症患者和对照组中的表达水平并进行荧光强度分级,比较组间蛋白的表达水平。2.收集DMD患者病情严重程度指标:Hammersmith功能运动评分(HFMS),患者出现下述症状时的年龄:首发运动症状、上下楼梯困难、不能独立行走。根据dystrophin、n NOS的免疫荧光染色将DMD患者分为2组:完全缺失组和部分缺失组,根据utrophin的免疫荧光染色分为3组:正常阴性组、轻度增加组和明显增加组,比较组间患者病情严重程度指标的差异。3.用spearman秩相关分析DMD患者的dystrophin、utrophin和n NOS表达的相关性。结果1 3种蛋白在不同类型肌营养不良症中的表达与对照组相比,3种蛋白在LGMD、FSHD、远端型肌营养不良症和DM中的表达差异均无统计学意义;与对照组和其他类型肌营养不良症相比,DMD和BMD中的dystrophin和n NOS的表达均减少(P0.002),utrophin的表达均增多(P0.002),且DMD中dystrophin减少较BMD严重(P0.002)。2 DMD患者中3种蛋白不同表达水平组间患者病情严重程度的分析Dystrophin和n NOS的2组间患者病情严重程度指标的差异均无统计学意义。utrophin的3组间比较:utrophin明显增加组患者出现上下楼梯困难和不能独立时的年龄较正常阴性组和轻度增加组晚(P0.05),其他组间病情严重程度指标比较差异无统计学意义。3 DMD中3种蛋白的相关性相关分析结果:utrophin与dystrophin的表达呈负相关(r=-0.598,P0.05),而n NOS与utrophin、dystrophin均无相关性。结论1.Dystrophin、utrophin和n NOS在DMD和BMD中表达异常,有助于为DMD和BMD的诊断和分型提供依据。2.DMD中utrophin的表达增加能延缓病情进展,而dystrophin和n NOS的表达对病情无明确影响。3.DMD患者中utrophin和dystrophin的表达呈负相关,提示dystrophin的表达减少可能反馈性引起utrophin的表达上调。
[Abstract]:Research background in the EU, the incidence of muscular dystrophy rate is 1/2000, slow progression, high disability rate, poor prognosis, there is no effective treatment at present, research on disease diagnosis and treatment is very important. Dystrophin (dystrophin) plays an important role in maintaining the structural integrity of muscle fibers. Mutations in the dystrophin gene of Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD). The dystrophin associated protein (utrophin) is dystrophin homologous protein, two protein structures are highly similar, utrophin mainly in regenerating muscle fibers express and maintain the integrity of the muscle fiber structure, almost all replaced by dystrophin in mature muscle fibers, in recent years through animal experiments found that utrophin expression was up-regulated in mature muscle fibers in DMD patients may reduce muscle damage of neuronal oxygen. Nitrogen synthase (n NOS) can produce nitric oxide (NO) increase in skeletal muscle blood supply, and the structure and relationship of dystrophin is closely related to the abnormal expression of n NOS may not exercise tolerance and symptoms. At home and abroad about the expression of dystrophin, utrophin and N less NOS in different type muscular dystrophy in the study, to clarify the 3 proteins in different types of muscular dystrophy in the expression for the diagnosis of muscular dystrophy, provide the basis for the classification, and further understand the 3 protein expression in patients with DMD disease severity. The relationship between the expression of utrophin and N on dystrophin, NOS in different types of muscular dystrophy in the further study of 3 protein expression with Duchenne muscular dystrophy (DMD) relationship between the severity of the disease, and in patients with DMD dystrophin, n NOS and utrophin. The relationship between the expression of objective methods. 1. collection from March 2013 to May 2015 in our hospital and diagnosed 26 DMD cases, BMD7 cases, limb girdle muscular dystrophy (LGMD) in 10 cases, facioscapulohumeral muscular dystrophy (FSHD) in 5 cases, distal muscular dystrophy and 5 cases of ankylosing muscular dystrophy (DM) in 11 cases. Collect the same time muscle biopsy results were normal or emergency trauma patients as the control group. 21 cases were detected by fluorescence intensity grading immunofluorescence method of 3 proteins in muscular dystrophy patients and the control group in the expression level and compared between groups, protein the expression level of.2. in patients with DMD severity index: Hammersmith motor function score (HFMS), patients have the following symptoms: the age of starting motor symptoms, stairs difficult, unable to walk independently. According to dystrophin, N and NOS immunofluorescence staining in DMD patients 2 group: complete deletion group and partial deletion group according to immunofluorescence staining for utrophin were divided into 3 groups: normal group and negative group, a slight increase in.3. increased group, differences were compared between patients with the severity of the disease index by Spearman rank correlation analysis with DMD dystrophin, n NOS and the correlation between the expression of utrophin. Results 13 kinds of protein in different types of muscular dystrophy in expression compared with the control group, 3 proteins in LGMD, FSHD, differential expression of distal muscular dystrophy and DM were not statistically significant; compared with the control group and other types of muscular dystrophy, the expression of DMD and BMD in dystrophin n and NOS were decreased (P0.002), the expression of utrophin was increased (P0.002), and DMD dystrophin BMD (P0.002) is a serious reduction in expression level between the groups with the severity of Dystrophin and N NOS analysis of 3 different kinds of protein.2 in patients with DMD 2 The difference between the 3 groups was the severity of the disease index were not statistically significant.Utrophin utrophin significantly increased in patients receiving the stairs difficult and can not be independent of age than normal group and negative group increased slightly later (P0.05), the other group of 3 protein disease severity index had no difference the significance of.3 DMD in the correlation analysis results: the expression of utrophin was negatively correlated with dystrophin (r=-0.598, P0.05), n NOS and utrophin dystrophin were not correlated. Conclusion 1.Dystrophin, abnormal expression of utrophin and N NOS in DMD and BMD, helps to provide the basis for the expression of.2.DMD in the increase of utrophin can delay the disease progress for the diagnosis of DMD and BMD and the type, and the expression of dystrophin and N NOS on the condition of no clear effect on the expression of utrophin was negatively related to dystrophin and.3.DMD patients, suggesting that the expression of dystrophin decreased Less likely feedback caused the up-regulated expression of utrophin.

【学位授予单位】：郑州大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R746.2

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