Increasing dopamine neuron survival during grafting

对啮齿动物和非人类灵长类动物的研究表明，高达95%的多巴胺(DA)神经元移植到纹状体后第一周内死亡，这可能是迄今为止研究中临床结果不佳的原因之一。此外，当细胞被移植到老年宿主体内时，移植细胞的糟糕存活率似乎被夸大了。与移植过程和移植后早期间隔相关的多种损伤可能会使移植细胞容易死亡，包括机械创伤、缺氧、氧化应激和神经营养因子停用。尽管存在这些问题，识别DA移植物中的细胞死亡机制及其可能的触发因素，为有效的干预提供了途径 限制移植神经元的死亡。本项目旨在研究四种可能限制移植神经元死亡的干预措施，优化DA替代和功能恢复的可能性：1)通过细胞黏附因子L1抗体和Tenascin的处理来减少由解剖和植入组织准备所引发的凋亡，以及，2)使用甲磺酸氮卓酮和褪黑素来减少氧化应激，3)通过血管内皮生长因子(VEGF)治疗促进移植物的新生血管，从而减少缺氧/缺血，4)通过暴露于周期AMP刺激移植细胞的DA表型。在啮齿动物实验中，所有建议的干预措施都产生了显著的移植物增强作用。在本申请中，每一种干预措施都将在经MPTP治疗的幼年圣基茨绿猴身上进行功能疗效测试。年龄适中的胎儿中脑组织，经过不同的处理，将被植入纹状体，以及 定量的行为测量将评估功能结果，并与更广泛移植物的组织学和生化证据相关联。最后一个实验将结合单独证明可以增强年轻成年猴子移植物功能的治疗，并比较这种联合治疗在年轻成年猴子和老年MPTP治疗猴子中的功能结果。成功的提高移植细胞存活和生长的方法可能会改善移植结果，并适用于干细胞或其他以细胞为基础的帕金森病治疗。

Studies in rodents and nonhuman primates suggest that up to 95% of grafted dopamine (DA) neurons die within the first week after implantation into the striatum, a possible contributor to poor clinical outcome in studies to date. In addition, poor survival of grafted cells appears to be exaggerated when cells are implanted into an elderly host. Multiple insults associated with the transplant procedure and early post-graft interval could render grafted cells susceptible to death, including mechanical trauma, hypoxia, oxidative stress, and neurotrophic factor withdrawal. Despite these problems, the identification of cell death mechanisms operating in DA grafts, and their probable triggers, provides access to cogent interventions to limit death of grafted neurons. This project aims to investigate four interventions that may limit death of grafted neurons, optimizing the potential for DA replacement and recovery of function: 1) reducing the apoptosis triggered by dissection and preparation of the tissue for implantation, termed "anoikis", via treatment with the cell adhesion factors L1 antibody and tenascin, and, 2) utilizing treatment with the lazaroid tirilazad mesylate and melatonin to reduce oxidant stress, 3) reducing hypoxia/ischemia by accelerating neovascularization of grafts using treatment with vascular endothelial growth factor (VEGF), 4) stimulating the DA phenotype of grafted cells by exposure to cyclic AMP. All interventions proposed have yielded significant graft augmentation in rodent experiments. In the present application, each intervention will be tested for functional efficacy in young adult MPTP-treated St. Kitts green monkeys. Optimally aged fetal mesencephalic tissue, with varying treatments, will be implanted into the striatum, and quantitative behavioral measurements will assess functional outcome, correlated with histological and biochemical evidence of more extensive grafts. A final experiment will combine treatments proven individually to augment graft function in young adult monkeys, and compare the functional outcome of this combination therapy in young adult and aged MPTP-treated monkeys. Successful methods for augmentation of grafted cell survival and growth may improve transplantation results and be applicable to stem cells or other cell-based therapies for Parkinson's disease.