Induction of oligodendrocyte lineage cells by direct cellular reprogramming.

直接重编程是再生医学领域的一项最新和令人兴奋的技术，它用于将丰富且容易获得的细胞(例如成纤维细胞)直接转化为不同类型的感兴趣细胞。类似于诱导多能干细胞的产生，强制表达一组确定的转录因子被用来诱导所需的细胞身份转换。第一批将成纤维细胞直接重编程为不同神经细胞类型以及神经干细胞的报道现已发表。到目前为止，还没有关于成纤维细胞直接重编程为神经胶质细胞的报道。然而，少突胶质细胞是外胚层起源的主要神经胶质细胞类型之一，对神经元功能至关重要。它们与神经细胞轴突紧密结合，并赋予这些轴突以髓鞘，这是神经系统快速信号传播的先决条件。在这项研究方案中，我假设一组不同的转录因子的过度表达足以诱导成纤维细胞(和多能干细胞)直接转化为少突胶质细胞系的细胞。我自己的初步数据有力地支持了这种方法的可行性。报告鼠成纤维细胞将作为起始细胞群，以确定最有效的转录因子组合来诱导重编程过程。诱导细胞在体外和体内都有广泛的特点。此外，在阶段特异性标记的控制下表达的荧光报告将允许定义转录因子组合，这些转录因子组合介导少突胶质细胞谱系的不同阶段的转换(即少突胶质前体细胞与分化的少突胶质细胞)。随后，结果将被转化到人体系统中，在人体系统中，多能干细胞将作为初始种群。基因打靶方法将被应用于获得人类多能报告细胞系，这将有助于诱导人类少突胶质细胞系的研究。一旦建立了强大的人少突胶质细胞培养系统，将会有一些后续项目。直接重新编程的少突胶质细胞需要与通过传统的胚胎干细胞定向分化获得的相应细胞进行比较。比较来自野生型和脑白质营养不良的重编程细胞-患者将能够在体外建立脱髓鞘疾病的模型，并为药物和毒理学测试以及移植研究铺平道路。

Direct reprogramming is a recent and exciting technology in the field of regenerative medicine that is used to convert abundant and easily accessible cells (e.g. fibroblasts) directly into different cell-types of interest. Similar to the generation of induced pluripotent stem cells, forced expression of a defined set of transcription factors is used to induce the desired switch of cellular-identity. The first reports of direct reprogramming of fibroblasts into different neuronal cell-types as well as into neural stem cells have now been published. To date, there are no reports about direct reprogramming of fibroblasts into glial cells. However, oligodendrocytes, one of the principal glial cell-types of ectodermal origin, are critical for neuronal function. They engage tightly with nerve cell axons and endow these with myelin-sheaths, a prerequisite for rapid signal-propagation in the nervous system.In this research proposal I hypothesise that overexpression of a distinct set of transcription factors is sufficient to induce the direct conversion of fibroblasts (and pluripotent stem cells) into cells of the oligodendrocyte-lineage. My own preliminary data strongly support the feasibility of this approach. Reporter mouse fibroblasts will be used as starting cell population to define the most efficient combination of transcription factors to induce the reprogramming process. Induced cells will be characterised extensively both in vitro and in vivo. Furthermore, fluorescent reporters that are expressed under the control of stage-specific markers will allow the definition of transcription factor-combinations that mediate a conversion into distinct stages of the oligodendrocyte-lineage (i.e. oligodendrocyte precursor cells versus differentiated oligodendrocytes). Subsequently, the results will be translated into the human system, where pluripotent stem cells will serve as initial starting population. Gene-targeting methods will be applied to derive human pluripotent reporter cell-lines that will facilitate the study of induced human oligodendrocyte-lineage cells.Once a robust human oligodendrocyte-culture system has been established, a number of follow-up projects will follow. Directly reprogrammed oligodendrocytes need to be compared with their counterparts derived by conventional directed differentiation from embryonic stem cells. Comparison of reprogrammed cells derived from wild-type versus leukodystrophy-patients will allow in-vitro modelling of demyelinating disease and pave the way for drug- and toxicology testing as well as transplantation studies.