摘要
自诱导多能干细胞的方法建立以来,干细胞重编程为再生医学带来了希望。在神经科学领域,已有研究证明神经干细胞的适应性强、成瘤风险低,可以定向地治疗神经系统相关的疾病。因此,通过体外诱导得到有效的应用于临床治疗的神经干细胞具有重要的研究意义。越来越多的证据表明,与遗传学方法相比,小分子化合物可以提高体细胞重编程效率,而且其应用、优化、制造及开发成药物相对容易。本研究尝试使用一组特定的9种小分子化合物组合(M9),在无须低氧的条件下,对小鼠(Mus musculus)14 d的胚胎成纤维细胞(mouse embryonic fibroblasts,MEFs)持续诱导12 d,检测诱导前后的神经干细胞有关表面标志物并提取RNA做转录组谱系分析。结果显示,诱导12 d后的细胞形态呈现与原代神经干细胞相似的球状,SOX2等神经干细胞表面标志物在免疫荧光以及RT-qPCR检测中均上调。转录组分析结果表明,M9诱导产生的细胞与原代神经干细胞具有非常相似的基因表达谱和自我更新能力。综上所述,M9可以诱导小鼠MEFs重编程成神经干细胞(neural stem cells,NSCs)。本研究意义在于使用小分子化合物替代转录因子,在体外简便的条件下实现了神经干细胞的直接重编程,避免了引入病毒转导载体带来的致瘤风险,促进了神经干细胞临床治疗的研究。
Stem cell reprogramming has held promise for regenerative medicine since the creation of induced pluripotent stem cell methods.In the field of neuroscience,it has been proved that neural stem cells have strong adaptability,low tumorigenesis risk,and can be used to treat diseases related to the nervous system in a targeted manner.Therefore,it is of great significance to induce neural stem cells for clinical treatment in vitro.Compared with genetic methods,there is growing evidence that small molecule compounds can improve the efficiency of somatic cell reprogramming and are relatively easier to apply,optimize,manufacture,and develop into drugs.In this study,mouse embryonic fibroblasts(MEFs)of 14 days were induced for 12 days by a specific combination of 9 small molecular compounds(M9)without hypoxic conditions.Surface markers of neural stem cells were detected before and after induction and RNA was extracted for transcriptome analysis.The results showed that after induction for 12 days,the cell morphology was spherical similar to that of the primary neural stem cells.The levels of SOX2 and other surface markers of the neural stem cells were up-regulated in immunofluorescence and RT-qPCR.Transcriptome analysis showed that the M9-induced cells had a very similar gene expression profile and self-renewal ability to the primary neural stem cells.In conclusion,M9 can induce MEFs to reprogram into neural stem cells.The significance of this study lies in the use of small molecular compounds to replace transcription factors,and the direct reprogramming of neural stem cells is realized in vitro under simple conditions,avoiding the tumorigenesis risk caused by the introduction of viral transduction vectors,and promoting the research on clinical treatment of neural stem cells.
作者
付金金
王光兴
张慧娜
柴子佳
杨哲
邵瑾良
孙毅
Fu Jinjin;Wang Guangxing;Zhang Huina;Chai Zijia;Yang Zhe;Shao Jinliang;Sun Yi(Beijing Institute of Brain Disorders,Laboratory of Brain Disorders,Ministry of Science and Technology,Collaborative Innovation Center for Brain Disorders,Capital Medical University,Beijing,100069;School of Medicine,Tongji University,Shanghai,200092)
出处
《基因组学与应用生物学》
CAS
CSCD
北大核心
2022年第3期671-681,共11页
Genomics and Applied Biology
基金
国家重点研发计划课题(2016YFA0100801)
国家自然科学基金重点项目(82030035)共同资助
