Functional integration of neurons derived from reprogrammed pericytes isolated from the adult human brain

从成人大脑中分离出的重编程周细胞衍生的神经元的功能整合

• 批准号: 252361766
• 负责人: Professor Dr. Benedikt Berninger
• 金额: --
• 依托单位: Institut für Physiologische Chemie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2013
• 资助国家: 德国
• 起止时间: 2012-12-31 至 2014-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/252361766?language=en
• 关键词: Functional; integration; neurons; derived; reprogrammed

Cellular reprogramming of somatic cells into induced neurons could pave the way for new therapies of currently uncurable neurodegenerative diseases. We could previously show that pericytes isolated from the adult human brain can be reprogrammed into induced neurons by forced expression of the two transcription factors Sox2 and Ascl1. This proposal aims at investigating whether such reprogramming can also take place in vivo and whether thereby newly generated neurons can functionally integrate into a neuronal network. We will examine this by using a transplantation model established by Prof. Marcos R Costa at the Federal University of Rio Grande do Norte (Brazil). Hereby in vitro retrovirus-transduced human pericytes are transplanted into the lateral ventricle of mouse embryos. The retroviruses encode Sox2 and/or Ascl1 as well as a reporter gene (GFP or RFP). After different survival times, the transplanted cell will be located in the host tissue based on their reporter expression and examined for their marker expression (neuronal versus pericyte markers). If we can show that reprogramming can also occur in the in vivo context, we will next examine by means of a rabies virus-based tracing technique whether and which type of presynaptic inputs impinge on neurons derived from pericytes. This study will be complemented by classical patch-clamp electrophysiology for synaptic activity. The experiments described here will provide evidence whether neurons derived from reprogrammed pericytes of human origin can be recognized by authentic neurons as target cells and thus proof of principle evidence for or against their synaptic integration. This represents a major advancement in our understanding of the potential of this cell-based approach. This project will be conducted in close collaboration with Prof. Marcos R. Costa, who has applied for a grant at the CNPq to study the integration of neurons of reprogrammed mouse astrocytes. Both project will be highly complementary and require the expertise of both laboratories.

将体细胞重新编程为诱导神经元，可能为目前无法治愈的神经退行性疾病的新疗法铺平道路。我们以前可以证明，从成人大脑中分离的周细胞可以通过强制表达两个转录因子Sox2和Ascl1而重新编程为诱导神经元。这项提议旨在研究这种重新编程是否也可以在体内发生，以及由此产生的新产生的神经元是否可以在功能上整合到神经元网络中。我们将使用北里奥格兰德联邦大学(巴西)的Marcos R Costa教授建立的移植模型来研究这一点。因此，体外逆转录病毒转导的人周细胞被移植到小鼠胚胎的侧脑室。逆转录病毒编码Sox2和/或Ascl1以及一个报告基因(GFP或RFP)。在不同的存活时间后，移植的细胞将根据它们的报告表达定位在宿主组织中，并检查它们的标记表达(神经细胞标记与周细胞标记)。如果我们能证明在活体环境中也可以发生重编程，我们接下来将通过基于狂犬病病毒的跟踪技术来检查是否以及哪种类型的突触前输入会影响周细胞来源的神经元。这项研究将与经典的膜片钳突触活动电生理学相补充。这里描述的实验将提供证据，证明来自人类来源的重新编程的周细胞的神经元是否可以被真正的神经元识别为靶细胞，从而证明支持或反对突触整合的原则证据。这代表着我们在理解这种基于细胞的方法的潜力方面取得了重大进展。该项目将与马科斯·R·科斯塔教授密切合作，科斯塔教授已在CNPQ申请拨款，研究重新编程的小鼠星形胶质细胞神经元的整合。这两个项目将具有很强的互补性，需要两个实验室的专业知识。