SOD1-V14M基因突变的肌萎缩侧索硬化患者特异的诱导性多能干细胞系的建立

摘要

Objective To generate familial ALS patient specific induced pluripotent stem (iPS) cell lines and motor neurons and provide a cell-based disease model for amyotrophic lateral sclerosis (ALS) in Han Chinese.Methods iPS cells were derived from familial ALS patient by introducing 4 transcription factors OCT3/4,SOX2,KLF4 and c-MYC into fibroblast cells by retroviruses.Karyotypic analysis,immunofluorescence staining and quantitative reverse transcription-polymerase chain reaction (RT-PCR) were used to identify the pluripotency of these iPS cell lines.In addition,motor neurons were derived from these iPS cells by inhibiting SMAD pathway.Results IPS cell lines were established from ALS patient carrying SOD1-V14M mutation.They had pluripotency and were similar to human ES cells.Furthermore,motor neurons were successfully induced from these iPS cells.Conclusion SOD1-V14M mutation does not affect the reprogramming of fibroblast cells and pluripotency of iPS cells,nor does it prevent differentiation of motor neurons.Furthermore,the above cell-based disease model can recapitulate key aspects of ALS pathogenesis so that it provides an indispensible resource for further elucidating ALS disease pathogenesis and screening appropriate drug candidates in Han Chinese.%目的 建立家族性肌萎缩侧索硬化(ALS)患者特异性的诱导性多能干细胞(iPS细胞)系,并诱导其分化为运动神经元,为研究中国人群ALS发病机制及药物筛选提供疾病模型.方法 通过逆转录病毒导入外源性的yamanaka转录因子,将SOD1-V14M基因突变的ALS患者成纤维细胞诱导为iPS细胞,并采用核型分析、免疫荧光染色、定量反转录PCR、畸胎瘤形成等实验对iPS细胞进行全能性鉴定,通过抑制SMAD信号通路诱导iPS细胞分化为运动神经元.结果 建立了SOD1-V14M基因突变的ALS病人特异性的iPS细胞系,验证具有全能性,并成功分化为运动神经元.结论 SOD1基因突变不影响成纤维细胞的重编程及iPS细胞的多能性,也不阻碍iPS细胞定向分化为运动神经元.本研究成功建立了SOD1-V14M基因突变的ALS患者iPS细胞系,为中国汉族人群家族性ALS疾病的研究及药物筛选提供了不可替代的资源.