【作者】 秦大强；

【导师】 罗巍；

【作者基本信息】 浙江大学 ， 神经病学， 2010， 硕士

【摘要】 研究背景：腓骨肌萎缩症(Charcot-Marie-Tooth Disease, CMT)是一类最常见的遗传性周围神经病,慢性进行性病程,致残率高。临床上依据其病理和电生理特点可分为两型：脱髓鞘型(CMT1型)和轴突型(CMT2型)。CMT2L型被发现是由小分子热休克蛋白22基因(HSP22)的点突变(423G→T)所致。HSP22蛋白变异如何致病目前机制尚不明确,有两种不同的理论推测：1、突变后功能缺失；2、获得的毒性作用。因HSP22蛋白变异所造成的神经肌肉疾病发病年龄均较晚,提示组织功能丧失是一个损伤效应逐渐累积的过程,疾病的发生更可能与突变造成的保护作用下降有关,而非突变直接造成组织损伤。研究目的：从腓肠神经筛选出CMT2L患者和正常人的的差异表达蛋白质,探讨CMT2L的可能发病机制,为探索CMT的治疗和预防打下基础。研究方法：1、采用2DE-MS技术体系,对人类正常腓肠神经组织和CMT2L患者腓肠神经组织提取蛋白后行双向电泳,PDQUEST7.0软件分析选择差异表达蛋白质,并通过基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS)进行蛋白质鉴定结果,筛选差异表达蛋白质。2、通过免疫荧光染色分析差异表达蛋白质和HSP22是否存在共定位。结果：通过2-DE方法及PDQUEST筛选出6个表达明显增高的蛋白,并通过MS成功鉴定出6个差异蛋白：Tubulin alpha-1B.Actin cytoplasmic 2、Tubulin alpha-1A、POTE ankyrin domain family member F、Glutathione S-transferase omega-1及EH domain-containing protein 2。其中Glutathione S-transferase omega-1及EH domain-containing protein 2可能与轴突的退行性病变有一定相关性。这两种蛋白均与HSP22在细胞胞浆内存在共定位。结论：Glutathione S-transferase omega-1及EH domain-containing protein 2可能参与了CMT2L的发生和发展,其相关机制尚待进一步阐明。双向电泳一质谱分析技术为CMT发生发展过程中差异表达蛋白的研究提供了有效的技术手段。更多还原

【Abstract】 Background:Charcot-Marie-Tooth disease (CMT) is the most common inherited peripheral neuropathy presenting with the phenotype of a chronic progressive neuropathy affecting both the motor and sensory nerves and high morbidity. Electrophysiological studies and pathological basis of clinical electrophysiological or anatomical pathology findings distinguish two major types-the demyelinating form and the axonal form. CMT2L was found to be caused by a novel 423G->T (Lys141Asn) missense mutation of HSP22. The mechanism of the HSP22 variation is still unknown, there are two different theoretical speculation:1) the loss of function mutation,2) acquired the toxic effects. The relatively late onset of the disease, which suggests an accumulative tissue damage pattern, indicates a more indirect, perhaps protective role of the wild-type sHSPs rather than an immediate role in proper tissue function.Objective:To screen differentially expressed proteins in human sural nerve between CMT2L patients and normal controls for understanding the pathogenesis of CMT. Results:Six differential protein spots were identified by PDQUEST analysis. Six proteins were successfully identified by MS, which are Tubulin alpha-1B. Actin cytoplasmic 2、Tubulin alpha-1A、POTE ankyrin domain family member F、Glutathione S-transferase omega-1 and EH domain-containing protein 2. The last two proteins may be of significant implications in axon degeneration, and may have some correction with axon degeneration. They both co-localized with HSP22 in the cell cytoplasm.Conclusions:Glutathione S-transferase omega-1 and EH domain-containing protein 2 may be involved in the occurrence and development of CMT2L and the related mechanisms remain to be elucidated. Two-dimensional electrophoresis and mass spectrometry technique provide an effective technique for screening the differentially expressed proteins in the development and progression of CMT.更多还原