Phenotypic & Functional Analysis of Human Schwann Cells for Potency Assay Dvlpmt

描述（由申请人提供）：雪旺细胞（SC）是细胞移植治疗的首选细胞类型，旨在恢复受损CNS和PNS的功能。在动物模型中的工作一直表明，SC减少脊髓损伤后形成的空腔的大小;促进损伤部位的轴突再生和髓鞘再生轴突，从而改善功能恢复。2012年7月，FDA批准了一项I期临床试验，以测试脊髓损伤患者自体成人神经源性人类SC移植的安全性。 在细胞治疗中使用原代SC的可行性依赖于细胞培养方法的可用性，从而从外周神经活检中获得高数量的移植质量纯化的SC。准确评估人SC产品的身份和功能对于确定细胞的潜在治疗价值至关重要。 事实上，在移植前评估细胞功能是临床试验后期产品放行的监管要求。尽管有大量的文献报道 关于啮齿动物SC的研究，关于人类SC的研究很少。这至少部分是由于培养的人SC在最初为啮齿动物细胞设计的标准功能测定中的表现不佳。该项目旨在克服体外试验开发中的技术障碍，以评估人类干细胞的生物活性。由于SC的身份和功能是不可分割地交织在一起，我们建议开始解决这一挑战，通过开发简单的分析方法来评估分化潜力的SC成为髓鞘细胞的分化标志物之前和之后的管理cAMP，分化的指示信号（目的1）。 因为轴突的鞘化和髓鞘形成可能是体外和体内SC功能的两个最关键的决定因素，我们还提出通过对影响共培养系统的SC和神经元组分的当前方案进行合理修改来开发人类SC和神经元的髓鞘形成共培养物（目的2）。鉴定体外扩增的SC中与其分化为髓鞘形成细胞的潜能相关的最重要的细胞特征将使我们能够 开发和实施可靠的检测方法，评估细胞的治疗价值。如果成功，允许在体外定量髓鞘形成的共培养系统可以作为SC效力的直接测量。

DESCRIPTION (provided by applicant): The Schwann cell (SC) is a cell type of choice for cell transplantation therapies that aim to restore function in the injured CNS and PNS. Work in animal models has consistently shown that SCs reduce the size of the cavities that form after spinal cord injury; promote axon regeneration across the lesion site and myelinate regenerating axons, thereby improving functional recovery. In July 2012, the FDA approved a Phase I clinical trial to test the safety of autologous adult nerve-derived human SC transplantation in patients suffering from spinal cord injury. The feasibility to use primary SCs in cell therapy relies on th availability of cell culture methods rendering high numbers of transplantation quality purified SCs from peripheral nerve biopsies. An accurate assessment of the identity and function of the human SC product is essential to determine the potential therapeutic value of the cells. Indeed, the assessment of cell function prior to transplantation is a regulatory requirement for product release in later phases of clinical testing. Even though an extensive literature exists that report studies on rodent SCs, studies on human SCs are scarce. This is at least in part due to the poor performance of cultured human SCs in standard functional assays that were originally designed for rodent cells. This project seeks to overcome technical barriers in the development of in vitro assays to evaluate the biological activity of human SCs. Because SC identity and function are inextricably intertwined, we propose to begin addressing this challenge by developing simple analytical methods to evaluate the differentiating potential of SCs to become myelinating cells by the identification of differentiation markers before and after administration of cAMP, an instructing signal for differentiation (Aim 1). Because ensheathment and myelination of axons are perhaps the two most critical determinants for SC function in vitro and in vivo, we also propose to develop myelinating co-cultures of human SCs and neurons by implementing rational modifications to current protocols affecting both the SC and the neuronal components of the co-culture system (Aim 2). Identifying the most important cellular characteristics of in vitro expanded SCs relevant to their differentiating potential into myelin-forming cells will allow us to develop and implement reliable assays that assess the therapeutic value of the cells. If successful, co-cultures systems that allow for a quantification of myelin formation in vitro can serve as a direct measure of SC potency.