Role of astrocyte-secreted pleiotrophin in dendritic spine phenotypes in Down Syndrome

星形胶质细胞分泌的多效蛋白在唐氏综合症树突棘表型中的作用

基本信息

批准号：
10478860
负责人：
Ashley N Brandebura
金额：
$ 3.49万
依托单位：
SALK INSTITUTE FOR BIOLOGICAL STUDIES
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-06-01 至 2022-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10478860
关键词：
Age Architecture Astrocytes Autopsy Biological Caliber Cells Clinical Coculture Techniques Complement Data Dendritic Spines Development Down Syndrome Down-Regulation Environment Exhibits Failure Filopodia Functional disorder Future Genetic Goals Histologic Human Impairment In Vitro Injections Institutes Intellectual functioning disability Investigation Knockout Mice Knowledge Light Link Maintenance Mass Spectrum Analysis Mediating Messenger RNA Morphology Mus Neck Neurodevelopmental Disorder Neurons Pathology Pattern Phenotype Physiological Process Protein Secretion Proteins Pyramidal Cells Quality of life Rattus Regulation Research Role Seminal Signal Transduction Small Interfering RNA Synapses Testing Thinness Thrombospondin 1 Tissues Transcript Transfection Up-Regulation Vertebral column Viral Work clinically relevant density dentate gyrus equipment training experimental study in vivo in vivo two-photon imaging insight mRNA Expression mRNA sequencing molecular sequence database molecular targeted therapies mouse Ts65Dn mouse model normal aging novel overexpression pleiotrophin ranpirnase receptor synaptogenesis syndecan 3 therapeutic target

项目摘要

Mouse models of Down Syndrome (DS) and DS human postmortem tissue display an overall decrease in dendritic spine density with an increased number of spines with immature morphology. Although the spine deficits observed in DS were traditionally viewed as a failure of neuronal cell intrinsic signaling, more recent studies suggest that astrocyte-neuron signaling contributes to the spine phenotypes. One seminal study utilizing co-cultures of human astrocytes and rat primary neurons demonstrated that wildtype (WT) neurons cultured in the presence of DS astrocytes possessed a significantly decreased total spine density with a shift towards a higher ratio of thin filopodia-like spines (immature morphology) when compared to WT neurons cultured with WT astrocytes. These results echo the findings of a multitude of studies demonstrating that proteins secreted from astrocytes can modulate neuronal synaptogenesis, maturation and maintenance. In order to identify the full complement of changes in the astrocyte secretome in DS, the lab conducted an unbiased mass spectrometry screen on the conditioned media of cultured astrocytes isolated from trisomic Ts65Dn mice (a well-characterized mouse model of DS) compared to their euploid controls on the same background. Pleiotrophin (Ptn) was identified among the top 10 proteins decreased in DS (over 4-fold decrease). Ptn is an attractive protein candidate to investigate for a role in spine phenotypes in DS due to (i) its highly enriched mRNA expression pattern in astrocytes during cortical synaptogenesis and (ii) a recently demonstrated role for Ptn in neuronal spinogenesis in dentate gyrus. There are many intriguing questions that remain regarding the involvement of Ptn in various aspects of cortical synapse and spine formation, maturation and maintenance, the role of Ptn in DS pathology, as well as which receptor(s) may be transducing effects on neuronal synaptic architecture. The goals of this proposal are to investigate the role of Ptn in regulating synapse and spine density, spine morphology and spine stability in normal cortical development, to determine if restoring physiological levels of Ptn secretion in DS can rescue the spine phenotypes observed, and to define the receptor(s) that mediate the effects of Ptn on dendritic spines. The main hypothesis is that Ptn regulates spine density, morphology and stability in cortical development and that down-regulation of secreted Ptn is a key mechanism leading to spine phenotypes in DS. Aim 1 will investigate if Ptn regulates cortical synapse and spine density, spine morphology and spine stability using Ptn knockout mice. Aim 2 asks if up-regulation of Ptn can rescue spine phenotypes in DS utilizing in vitro and in vivo approaches. Aim 3 investigates which neuronal receptor(s) mediate effects of Ptn utilizing an in vitro siRNA screen. These experiments will expand the current knowledge of the role of Ptn in cortical development and have significant clinical relevance to elucidate a mechanism by which dysregulation of Ptn levels leads to spine dysgenesis in DS. The proposed work will take place at the Salk Institute for Biological Studies, a high caliber, collaborative research environment which provides access to all necessary equipment and training.

唐氏综合征（DS）和DS人类后组织组织的小鼠模型显示，树突状脊柱密度的总体降低，脊柱数量增加，具有未成熟的形态。尽管传统上将DS中观察到的脊柱缺陷视为神经元细胞固有信号的失败，但最近的研究表明，星形胶质细胞 - 神经元信号传导有助于脊柱表型。一项使用人类星形胶质细胞和大鼠原发性神经元共培养的开创性研究表明，在存在DS星形胶质细胞的存在下培养的野生型（WT）神经元具有显着降低的总脊柱密度，而与WT神经培养的薄丝状脊柱（无效的形态学）相比，较高的薄丝状脊柱（无效的形式）的转变与较高的比例。这些结果与大量研究的发现相呼应，表明从星形胶质细胞分泌的蛋白质可以调节神经元突触发生，成熟和维持。为了确定DS中星形胶质细胞分泌组的完整补充，该实验室在与同一背景上的真皮对照中相比，该实验室在从TRISOMIC TS65DN小鼠的条件培养基上进行了无偏的质谱屏幕。在DS的前10种蛋白质中鉴定出多叶酸（PTN）（PTN）（降低4倍）。 PTN是一种有吸引力的蛋白质候选者，可以研究（i）在皮质突触发生过程中其在星形胶质细胞中高度富集的MRNA表达模式，并且（ii）最近证明了PTN在齿状回和齿状回合中神经元旋转中的作用。关于PTN参与皮层突触和脊柱形成，成熟和维持，PTN在DS病理学中的作用以及哪些受体对神经元突触结构的影响。该提案的目标是研究PTN在调节正常皮质发育中调节突触和脊柱密度，脊柱形态和脊柱稳定性中的作用，以确定DS中PTN分泌的生理水平是否可以挽救观察到的脊柱表型的生理水平，并定义了介导Ptn on denderritic Spine denderritic Spine的受体。主要假设是PTN调节皮质发育中的脊柱密度，形态和稳定性，而分泌PTN的下调是导致DS中脊柱表型的关键机制。 AIM 1将研究PTN是否使用PTN基因敲除小鼠调节皮质突触和脊柱密度，脊柱形态和脊柱稳定性。 AIM 2询问PTN的上调是否可以在使用体外和体内方法的DS中挽救脊柱表型。 AIM 3研究了哪种神经元受体利用体外siRNA筛选介导PTN的作用。这些实验将扩大当前对PTN在皮质发育中作用的知识，并具有显着的临床相关性，以阐明PTN水平失调导致DS中脊柱发病障碍的机制。拟议的工作将在Salk生物学研究所进行，这是一个高素质的协作研究环境，可访问所有必要的设备和培训。