Department of Chemistry, National Taiwan University, Taipei, 106, Taiwan.
Department of Medical Research, Taipei Veterans General Hospital, Taipei, 112, Taiwan.
Sci Rep. 2018 Jan 8;8(1):11. doi: 10.1038/s41598-017-18324-8.
Direct-lineage conversion of the somatic cell by reprogramming, in which mature cells were fully converted into a variety of other cell types bypassing an intermediate pluripotent state, is a promising regenerative medicine approach. Due to the risk of tumorigenesis by viral methods, a non-viral carrier for the delivery of reprogramming factors is very desirable. This study utilized the mesoporous silica nanoparticles (MSNs) as a non-viral delivery system for transduction of the three key factors to achieve conversion of mouse fibroblasts (MFs) into functional dopaminergic neuron-like cells (denoted as fDA-neurons). At the same time, a neurogenesis inducer, ISX-9, was co-delivered with the MSNs to promote the direct conversion of neuron-like cells. Good transfection efficiency of plasmid@MSN allowed repeated dosing to maintain high exogenous gene expression analyzed by qPCR and the changes in neural function markers were monitored. To further validate the dopaminergic function and the electrophysiological properties of fDA-neurons, the results of ELISA assay showed the high levels of secreted-dopamine in the conditional medium and rich Na/K-channels were observed in the fDA-neurons on Day 22. The results demonstrated that MSN nanocarrier is effective in delivering the reprogramming factors for the conversion of functional dopaminergic neurons from adult somatic cells.
体细胞通过重编程进行直接谱系转化，其中成熟细胞完全转化为多种其他细胞类型，而绕过中间多能状态，这是一种很有前途的再生医学方法。由于病毒方法存在致癌风险，因此非常需要非病毒载体来递送重编程因子。本研究利用介孔硅纳米粒子（MSNs）作为非病毒递送系统来转导三个关键因子，以实现将小鼠成纤维细胞（MFs）转化为功能性多巴胺能神经元样细胞（记为 fDA-神经元）。同时，与 MSNs 共递送神经发生诱导剂 ISX-9 以促进神经元样细胞的直接转化。质粒@MSN 的良好转染效率允许重复给药以维持 qPCR 分析的高外源基因表达，并监测神经功能标记物的变化。为了进一步验证 fDA-神经元的多巴胺能功能和电生理特性，ELISA 测定结果表明，在第 22 天的条件培养基中观察到高分泌多巴胺水平，并且在 fDA-神经元中观察到丰富的 Na/K 通道。结果表明，MSN 纳米载体可有效递送电子重编程因子，从而将功能性多巴胺能神经元从成年体细胞中转化而来。
