技术与方法

脊髓性肌萎缩症患者尿液细胞模型的建立

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  • 福建医科大学附属第一医院神经内科,福州 350005
陈万金,副主任医师,研究方向:神经遗传病。

收稿日期: 2014-07-01

  网络出版日期: 2014-10-28

基金资助

国家自然科学基金项目(编号:81322017,81371261),福建省自然科学基金杰出青年基金项目(编号:2012J06016)和国家临床重点专科建设项目资助

The construction of urine-derived cell lines from patients with spinal muscular atrophy

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  • Department of Neurology, First Affiliated Hospital, Fujian Medical University, Fuzhou 350005, China

Received date: 2014-07-01

  Online published: 2014-10-28

摘要

脊髓性肌萎缩症(Spinal muscular atrophy, SMA)大多数在儿童或婴幼儿期发病,表现为进行性、对称性的肢体无力和肌肉萎缩,迄今尚无有效的治疗方法,是婴幼儿最常见的致死性遗传病之一。患者来源的细胞系是该病研究的重要工具,但依赖于肌肉或皮肤活检等创伤性手术的成纤维细胞培养较难被患者及家属接受。文章收集SMA患者及健康对照的新鲜尿液,进行离心、尿液沉渣培养,观察尿液细胞的生长状况,用酶联免疫吸附实验(Enzyme-linked immunosorbent assay,ELISA)分析患者尿液细胞中SMN(Survival of motor neuron)蛋白的表达量,应用免疫荧光染色观察SMN蛋白在细胞内的定位。共建立了11例SMA患者和14例健康对照的尿液细胞系,尿液细胞体外增殖旺盛,细胞形态及生长速度较稳定。患者来源的尿液细胞SMN1(Survival of motor neuron 1) 基因缺失突变、SMN蛋白表达量降低,荧光染色提示SMN蛋白在胞浆和胞核中均有定位。尿液细胞培养步骤简单、无创伤性、患儿及其家属的依从性好,是获取和保存病人来源标本的有效方法,在脊髓性肌萎缩症发病机制研究和临床应用方面具有较好的应用价值。

本文引用格式

陈万金, 张奇杰, 何瑾, 林翔, 王柠 . 脊髓性肌萎缩症患者尿液细胞模型的建立[J]. 遗传, 2014 , 36(11) : 1168 -1172 . DOI: 10.3724/SP.J.1005.2014.1168

Abstract

Spinal muscular atrophy (SMA) is a common neurodegenerative disease in childhood and infancy, clinically characterized by progressive and symmetric muscular weakness and atrophy. Few effective therapies are available now, and SMA is one of the most common genetic causes of infantile mortality. SMA patient-derived cells are beneficial in basic research on this disease, but the most common model cell, fibroblasts can only be obtained through invasive procedures such as muscle or skin biopsy, which are unwelcome to patients and their families. In this study, fresh urine from SMA patients and healthy controls was collected and centrifuged, and the urine sediment was cultured in vitro. The growth characteristics of urine-derived cells were observed, and the survival of motor neuron (SMN) gene, and the amount and localization of SMN protein in different urine cell lines were investigated. In total, 25 urine cell lines from 11 SMA patients and 14 healthy controls were established. These urine-derived cells expand robustly in vitro with stable cell morphological characteristics. The urine cell lines derived from patients carry the SMN1 gene defect and express a low level of SMN protein, while the intracellular localization of SMN protein is normal. Urine-derived cell culture technology is simple, non-invasive and highly reproducible, a way of obtaining and storing rare cell samples from SMA patients with which to study the pathogenesis of SMA.

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