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Magnetic nanoparticle mediated delivery of neurotherapeutic genes to multipotent neural stem cell transplant populations

磁性纳米颗粒介导将神经治疗基因递送至多能神经干细胞移植群体

基本信息

批准号：
BB/J017590/1
负责人：
Divya Chari
金额：
$ 38.79万
依托单位：
Keele University
依托单位国家：
英国
项目类别：
Research Grant
财政年份：
2013
资助国家：
英国
起止时间：
2013 至无数据
项目状态：
已结题

来源：
https://gtr.ukri.org/projects?ref=BB%2FJ017590%2F1
关键词：
Magnetic nanoparticle mediated delivery neurotherapeutic

项目摘要

The adult brain and spinal cord (together called the central nervous system or CNS) repair poorly after injury and disease. This leads to limited recovery for patients with severe brain/spinal cord injury and disease with profound consequences for their quality of life. Stem cells found in the nervous system called neural stem cells or NSCs have enormous potential for increasing repair at injury/disease sites in the CNS as they can give rise to new stem cells, replace lost cell types of the CNS and can also suppress injury responses. These cells can migrate long distances in the CNS and are particularly attracted into areas of damage in the nervous system. Therefore, an important use for these engineered stem cells is to function as as transport 'vehicles' to deliver therapeutic molecules to injury/disease areas in the CNS. In experimental neurology research, the most common way of delivering genes to stem cells is to use modified viruses as vehicles. Infection of cells with these viruses enables the transfer of genes of interest into the infected cells. However, experimental methods using viruses can be be technically difficult, time consuming and expensive and can alter the basic cell properties of neural stem cells such as their division capability and their genesis of new cell types. This method also has other drawbacks including major safety issues (such as reactions from the immune system and cancer causing effects) and difficulties in achieving the large scale production that is required for human and veterinary clinical therapies. State-of-the-art delivery systems using small particles with an iron core called magnetic nanoparticles (MNPs) have many benefits in this regard and can be used effectively for delivery of genes. Preliminary clinical trials have shown that injecting MNPs into the body appears to be safe. MNPs can be linked with genes and taken up into cells to mediate gene delivery. Although MNP based systems are thought to be less effective than viruses for delivering genes, we have shown that applying static or oscillating magnetic fields (a method called 'magnetofection') can dramatically increase gene delivery by MNPs. DMC's group proved recently that MNPs can be used to safely deliver genes to several major cell types (of both rodent and canine origin) that are used in neural cell transplantation therapies. The methods used were not associated with safety issues and did not alter basic stem cell properties like their division or migration capability or the types/numbers of daughter cells they give rise to. The methods we have developed for MNP based gene transfer are very safe, technically simple, quick and relatively inexpensive, therefore their use will provide an effective and convenient method for delivering genes and can result in significant cost savings to laboratories and funding agencies.This study will build on recent findings to develop methods to use MNPs to deliver genes coding for therapeutic molecules to stem cells (that will then be used for transplantation). We will assess different strategies to optimise uptake of MNPs coated with genes into stem cells and establish if this procedure has adverse effects on the survival and development of the cells. A further goal is to develop a new, technically easy and cheap method to examine the interactions of stem cells and MNPs at high magnification using a special 'electron' microscope. We will also investigate the underlying mechanisms by which oscillating magnetic fields increase gene delivery by MNPs. The repair enhancing potential of the engineered stem cells will be tested in a 'living' 3D slice model of the spinal cord, that can function well as injured 'host' tissue for transplant cells. This method provides a robust alternative to the use of surgical transplantation to study stem cell transplantation therapies in living animals, thereby significantly reducing animal usage and suffering in experimental research.

成年人的大脑和脊髓（统称为中枢神经系统或CNS）在受伤和疾病后修复不良。这导致严重脑/脊髓损伤和疾病患者的恢复有限，对其生活质量产生深远影响。在神经系统中发现的干细胞称为神经干细胞或NSC，具有增加CNS损伤/疾病部位修复的巨大潜力，因为它们可以产生新的干细胞，取代CNS丢失的细胞类型，还可以抑制损伤反应。这些细胞可以在CNS中长距离迁移，特别是被吸引到神经系统的损伤区域。因此，这些工程化干细胞的一个重要用途是作为运输“载体"将治疗分子递送到CNS中的损伤/疾病区域。在实验神经学研究中，将基因传递到干细胞的最常见方法是使用修饰病毒作为载体。用这些病毒感染细胞能够将感兴趣的基因转移到感染的细胞中。然而，使用病毒的实验方法在技术上可能是困难的，耗时和昂贵的，并且可以改变神经干细胞的基本细胞特性，例如它们的分裂能力和它们的新细胞类型的发生。该方法还具有其他缺点，包括主要的安全问题（例如来自免疫系统的反应和致癌作用）和难以实现人类和兽医临床治疗所需的大规模生产。使用具有铁核的小颗粒的最先进的递送系统称为磁性纳米颗粒（MNP）在这方面具有许多益处，并且可以有效地用于递送基因。初步临床试验表明，将MNP注射到体内似乎是安全的。MNP可以与基因连接并进入细胞以介导基因递送。虽然基于MNP的系统被认为在传递基因方面不如病毒有效，但我们已经证明，施加静态或振荡磁场（一种称为“磁转染”的方法）可以显著增加MNP的基因传递。DMC的研究小组最近证明，MNP可用于安全地将基因传递到几种主要的细胞类型（啮齿动物和犬源），这些细胞类型用于神经细胞移植治疗。使用的方法与安全性问题无关，并且不会改变基本的干细胞特性，如其分裂或迁移能力或其产生的子细胞的类型/数量。我们开发的基于MNP的基因转移方法非常安全，技术简单，快速且相对便宜，因此，它们的使用将提供一种有效和方便的方法来递送基因，并可以为实验室和资助机构节省大量成本。这项研究将建立在最近的发现基础上，以开发使用MNP将编码治疗分子的基因递送到干细胞的方法（然后用于移植）。我们将评估不同的策略来优化干细胞对基因包被的MNP的吸收，并确定这一过程是否对细胞的生存和发育产生不利影响。另一个目标是开发一种新的，技术上容易和廉价的方法，使用特殊的“电子”显微镜在高放大倍数下检查干细胞和MNP的相互作用。我们还将研究振荡磁场增加MNP基因递送的潜在机制。工程干细胞的修复增强潜力将在脊髓的“活”3D切片模型中进行测试，该模型可以作为移植细胞的受损“宿主”组织发挥良好作用。该方法为使用手术移植来研究活体动物的干细胞移植疗法提供了一种稳健的替代方案，从而显著减少了实验研究中的动物使用和痛苦。