Translational profiling of bladder sensory nerves and their cell type identities using dissociation free single nucleus sequencing

使用无解离单核测序对膀胱感觉神经及其细胞类型进行翻译分析

基本信息

批准号：
10309020
负责人：
Sanjay Jain
金额：
$ 23.63万
依托单位：
WASHINGTON UNIVERSITY
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：
2021-08-01 至 2023-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10309020
关键词：
Acute Address Adoption Adult Affect Afferent Neurons Affinity Chromatography Aging Anatomy Area Axon Basic Science Benchmarking Biological Bladder Bladder Diseases Bladder Dysfunction Blood Vessels Brain Cell Nucleus Cells Communication Communities Complex Cues Data Development Diabetes Mellitus Disease Dissociation Distal Drug Targeting Efferent Neurons Enterobacteria phage P1 Cre recombinase Etiology Female Frequencies Functional disorder Gene Expression Genes Genetic Genetic Transcription Genetic Translation HIV Health Homeostasis Immune Impairment Incontinence Infection Inflammation Informatics Injury Interphase Interstitial Cystitis Intractable Pain Knowledge Label Lead Longevity Lower urinary tract Maintenance Maps Messenger RNA Methods Molecular Morphologic artifacts Motor Mus Muscle Natural regeneration Nerve Nerve Fibers Nervous system structure Neuroanatomy Neurodegenerative Disorders Neurogenic Bladder Neurologic Neurons Neuropathy Nociceptors Organ Overactive Bladder Pathogenesis Pathologic Peripheral Peripheral Nerves Peripheral Nervous System Physiological Physiology Play Presynaptic Terminals Procedures Property Proteins Protocols documentation RNA Recurrence Regulation Relaxation Research Ribosomes Role Sensory Signal Transduction Site Smooth Muscle Myocytes Source Spinal Dysraphism Spinal Ganglia Stress Structure Symptoms Synapses System Technology Time Tissues Transcript Translating Translational Research Translations Traumatic CNS injury Urethral sphincter Urinary system Urinary tract Urinary tract infection Urination Urologic Diseases Urothelium Validation Viral Withdrawal afferent nerve axonal degeneration cell type clinical Diagnosis comparative design implementation framework innovation insight interest lower urinary tract symptoms male malformation nerve supply nervous system disorder neuronal cell body neurophysiology normal aging novel preservation repaired response satellite cell sensory neuropathy sex somatosensory targeted treatment tool transcriptome sequencing translational scientist translatome

项目摘要

Abstract Peripheral axons need sustained maintenance and repair to maintain connectivity with their targets. Local axonal translation plays a role in regulation of protein and energy homeostasis across lifespan. The repertoire of active mRNA translation during development and adulthood, and the ability to repair, regenerate and translate specific subsets of mRNAs in relation to environmental cues is stored in distal axons. In many instances these activities take place rapidly for immediate response at target sites before new transcripts localize from the cell bodies. In disease states, altered axonal translation may affect elongation, withdrawal, pruning, branching or survival. In the bladder innervation by afferent (sensory) and efferent (parasympathetic and sympathetic) nerves is critical for its function. These nerves establish intricate connections with different regions of the bladder and cell types including the urothelium, resident immune cells, smooth muscle cells and blood vessels and likely employ local translation to maintain homeostasis and respond to injury or physiologic changes. The identity of genes that are actively translated in bladder nerve terminal and subsequently their role in bladder has not been studied. Defining actively translated mRNA in healthy and disease states in bladder nerves and the identity of the soma source will provide insights into the mechanisms of neuroanatomical and physiological interactions between the peripheral nervous and urinary systems, their roles in a number of bladder diseases with neuronal basis and aid in rationally designing targeted therapies aimed at preserving neuro-uro connections. This has potential to impact understanding of declining bladder function in ageing, debilitating bladder symptoms and dysfunction (such as intractable pain, incontinence, frequency and urgency to urinate) that affect millions of people in the US, neurodegenerative diseases and neuropathies (diabetes, HIV) affecting bladder function. We will establish methods and proof of concept studies to delineate the translatome in bladder sensory, autonomic axons in healthy male and female mice and in female mice with UTI (Aim 1) and identify their cognate cell bodies by developing artifact –free single nucleus RNA expression maps of DRG sensory neurons projecting to the bladder (Aim 2). The methods developed will overcome challenges unique to composition, function and anatomy of bladder, its nerves and corresponding soma that are different from the CNS or somatosensory system and serve as benchmark for application to other lower urinary tract organs. The methods developed, genetic tools used and novel data generated will be publicly shared for rapid adoption and accelerating studies to better understand regulation of nerve terminals with their target sites by diverse set of users including basic and translational researchers, computational biologists and clinicians interested in urinary tract disorders and normal physiology.

抽象的外周轴突需要持续的维护和修复，以维持与其局部目标的连接。轴突翻译在整个生命周期的蛋白质和能量稳态调节中发挥着作用。发育和成年期间的活性 mRNA 翻译，以及修复、再生和再生的能力翻译与环境线索相关的特定 mRNA 子集存储在许多远端轴突中。在这种情况下，这些活动会迅速发生，以便在新转录本之前在目标站点立即做出反应在疾病状态下，轴突翻译的改变可能会影响伸长、收缩、在膀胱神经支配中，传入神经（感觉神经）和传出神经（副交感神经）。和交感神经）对其功能至关重要，这些神经与不同的神经建立了复杂的联系。膀胱区域和细胞类型，包括尿路上皮、常驻免疫细胞、平滑肌细胞和血管并可能利用局部翻译来维持体内平衡并对损伤或生理做出反应膀胱神经末梢中活跃翻译的基因的特性及其随后的变化。尚未研究在健康和疾病状态下积极翻译的 mRNA 在膀胱中的作用。膀胱神经和体细胞来源的身份将提供对膀胱神经的机制的见解周围神经和泌尿系统之间的神经解剖学和生理学相互作用，在许多具有神经学基础的膀胱疾病中的作用，并有助于合理设计靶向治疗旨在保护神经-尿路连接，这有可能影响对膀胱衰退的理解。缓解衰老、膀胱衰弱症状和功能障碍（如顽固性疼痛、失禁、尿频和尿急）影响着美国数百万人、神经退行性疾病和我们将建立影响膀胱功能的神经病（糖尿病、艾滋病毒）的方法和概念验证。研究描绘健康雄性和雌性小鼠膀胱感觉、自主神经轴突中的翻译组在患有 UTI 的雌性小鼠中（目标 1），并通过开发无伪影的单一细胞体来识别其同源细胞体投射到膀胱的 DRG 感觉神经元的核 RNA 表达图（目标 2）。所开发的技术将克服膀胱及其神经和膀胱的组成、功能和解剖结构所特有的挑战与中枢神经系统或体感系统不同的相应体体，并作为基准开发的方法、使用的遗传工具和新数据的应用。生成的结果将公开共享，以供快速采用和加速研究，以更好地了解监管包括基础研究人员和转化研究人员在内的不同用户组的神经末梢及其目标位点，对尿路疾病和正常生理学感兴趣的计算生物学家和粉丝。