Regulation of Detrusor Overactivity by Cellular Stress in Bladder Ischemia

膀胱缺血时细胞应激对逼尿肌过度活动的调节

• 批准号: 9976982
• 负责人: KAZEM M AZADZOI
• 金额: --
• 依托单位: VA BOSTON HEALTH CARE SYSTEM
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-07-01 至 2023-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9976982
• 关键词: Actins; Adenosine Monophosphate; Aging; Agonist; Animal Model; Anxiety; Basic Science; Benign; Bladder; Bladder Dysfunction; Bladder Tissue; Blood flow; Calcium; Cell Culture Techniques; Cells; Cellular Stress; Cellular Stress Response; Chronic; Clinical Research; Contractile Proteins; Cultured Cells; Data; Development; Double-Stranded RNA; Elderly; Elements; Fatigue; Female; Functional disorder; Gene Deletion; Gene Silencing; General Population; Genetic Transcription; Goals; Human; Impairment; Incidence; Increased frequency of micturition; Ischemia; Knock-out; Knockout Mice; Lead; Link; Mediating; Mental Depression; Modeling; Modification; Molecular; Motor; Mus; Muscarinic M2 Receptor; Muscarinic M3 Receptor; Muscle Contraction; Myosin Heavy Chains; Nocturia; Obstruction; Overactive Bladder; Oxidation-Reduction; Pathway interactions; Pelvic Pain; Phosphorylation; Play; Population; Post-Translational Protein Processing; Prevalence; Prostatic; Protein Kinase; Quality of life; RNA; Regulation; Reporting; Research; Research Proposals; Resources; Role; Signal Transduction; Sleeplessness; Smooth Muscle; Smooth Muscle Myocytes; Stress; Symptoms; Technology; Testing; Therapeutic; Transfection; Urge Incontinence; Veterans; associated symptom; base; biological adaptation to stress; cellular targeting; energy balance; human model; impaired capacity; insight; lower urinary tract symptoms; male; mouse model; novel; novel diagnostics; novel therapeutics; older patient; prevent; prophylactic; protein kinase R; sensor; uptake

The overall goal of this renewal research proposal is to determine the role of ischemia in the development of detrusor overactivity in the non-obstructed bladder. Another goal is to examine the efficacy of targeting cellular stress response pathways to prevent or reverse bladder dysfunction. Lower urinary tract symptoms (LUTS) are bothersome constellation of voiding symptoms with significant impact on quality of life. The prevalence of bladder dysfunction and LUTS among Veterans is almost five folds higher in comparison with its incidence in the general population. It was shown that Veterans with LUTS had a worse quality of life score than Veterans without LUTS. In most cases, particularly in elderly Veterans, LUTS resulted in insomnia, anxiety, fatigue, and depression. Most cases of LUTS in male are attributed to bladder outlet obstruction (BOO) due to benign prostatic enlargement (BPE). However, it has been shown that in approximately one third to more than one half of cases, LUTS are not associated with BPE or BOO. These observations suggest that, in addition to BOO, aging-related local changes in the bladder contribute to LUTS. Growing evidence from basic and clinical research suggests that aging-associated bladder ischemia play a key role. Impairment of human bladder blood flow and the development of bladder ischemia have been verified in elderly patients with LUTS. However, the underlying mechanisms contributing to detrusor overactivity in bladder ischemia remain largely unknown. Our preliminary data suggest that ischemia provokes cellular stress and compromises cellular defensive capacity by impairing the cellular energy sensor adenosine monophosphate-activated protein kinase alpha-2 (AMPK- α2). Cell stress and defective AMPK-α2 give rise to a unique stress response RNA with two complementary strands namely double-stranded RNA (dsRNA) leading to activation of the AMPK-α2/dsRNA stress response pathway. The AMPK-α2/dsRNA pathway seems to compromise muscarinic M2 and M3 receptors, provoke post-translational modifications of contractile proteins, increase smooth muscle contractions and engender detrusor overactivity. We hypothesize that “chronic ischemia is an independent factor in the development of detrusor overactivity in the non-obstructed bladder. The mechanism involves activation of cellular stress response via AMPK-α2/dsRNA pathway that triggers overactive bladder contractions by modification of smooth muscle contractile elements”. Using a well-established bladder ischemia model along with knockout mice and cell culture transfection and gene deletion technologies, we propose three specific aims. In aim I, we will define molecular regulation of the AMPK-α2/dsRNA stress response pathway in bladder ischemia. We will determine the mechanism of AMPK-α2 impairment, quantify and clone dsRNA in bladder ischemia and define crosstalk mechanisms between AMPK-α2 and dsRNA. In aim II, we will define regulation of overactive contractions by AMPK-α2/dsRNA pathway in bladder ischemia. We will determine how AMPK-α2/dsRNA pathway provokes overactive bladder contractions by transcriptional and post-translational modifications of muscarinic M2 and M3 receptors. We will examine therapeutic strategies to prevent stress-regulated modifications of M2 and M3 and reverse detrusor overactivity. In aim III, we will define signaling mechanisms downstream of AMPK-α2/dsRNA pathway that modify contractile proteins in bladder ischemia. We will determine the role of AMPK-α2/dsRNA- regulated redox and PKR signaling in modifications of actin-α1 and myosin heavy chain (MHC) and regulation of smooth muscle calcium uptake. We will examine therapeutic strategies to prevent redox- and PKR-mediated actin-α1 and MHC modifications and reverse detrusor overactivity in bladder ischemia. At the conclusion of these studies, we will have: (1) provided new insights into the pathophysiology of detrusor overactivity in the non-obstructed bladder; (2) elucidated the molecular link between AMPK-α2/dsRNA pathway and overactive bladder contractions and 3) defined stress sensitization mechanisms in overactive bladder contractions. Our proposed research may lead to novel diagnostic and therapeutic strategies against detrusor overactivity/LUTS.

该更新研究建议的总体目标是确定缺血在发展中的作用 非刺激性膀胱中的逼尿肌过度活动性。另一个目标是检查靶向细胞的效率 预防或逆转膀胱功能障碍的压力响应途径。较低的尿道符号（LUTS）为 两种缺失症状的星座都对生活质量产生了重大影响。流行率 与之相比，退伍军人中的膀胱功能障碍和LUTs几乎高出五倍 普通人群。结果表明，LUTS的退伍军人的生活质量比退伍军人更差 没有路。在大多数情况下，特别是在老兵中，LUTS导致失眠，动画，疲劳和 沮丧。大多数男性中的LUT案件归因于良性造成的膀胱出口异常（BOO） 前列腺增强（BPE）。但是，已经表明，大约三分之一到一半以上 在情况下，LUT与BPE或BOO无关。这些观察结果表明，除了BOO外， 膀胱与衰老相关的局部变化有助于LUTS。越来越多的证据来自基本和临床 研究表明，与衰老相关的膀胱缺血起着关键作用。人类膀胱损害 在古老的LUT患者中，已经验证了流量和膀胱缺血的发育。但是， 导致膀胱局部缺血过度活动的基本机制仍然很大未知。我们的 初步数据表明，缺血会引起细胞应力并损害细胞防御能力 通过损害细胞能传感器腺苷一磷酸激活的蛋白激酶α-2（AMPK-- α2）。细胞应力和有缺陷的AMPK-α2产生了独特的应力反应RNA，具有两个完整性 链，即双链RNA（dsRNA），导致AMPK-α2/dsRNA应力响应的激活 路径。 AMPK-α2/DSRNA途径似乎损害了毒蕈碱M2和M3受体 收缩蛋白的翻译后修饰，增加平滑肌收缩和产生 迫害者过度活动。我们假设“慢性缺血是发展的独立因素 非刺激性膀胱中的逼尿肌过度活动性。该机制涉及细胞应激的激活 通过AMPK-α2/dsRNA途径的响应，通过修饰光滑，触发过度活动的膀胱收缩 肌肉收缩元素”。使用良好的膀胱缺血模型以及淘汰小鼠和 细胞培养转化和基因缺失技术，我们提出了三个具体目标。在目标一世中，我们将定义 AMPK-α2/dsRNA应激反应途径的分子调节膀胱缺血。我们将确定 AMPK-α2损伤，量化和克隆DSRNA的机制在膀胱缺血中并定义串扰 AMPK-α2和DSRNA之间的机制。在AIM II中，我们将通过 膀胱缺血中的AMPK-α2/DSRNA途径。我们将确定AMPK-α2/DSRNA途径如何挑衅 毒蕈碱M2和M3的转录和翻译后修饰过度活跃的膀胱收缩 受体。我们将研究理论策略，以防止M2和M3的应力调节修改和 反向逆转录者过度活动。在AIM III中，我们将定义AMPK-α2/DSRNA下游的信号传导机制 修改膀胱缺血中收缩蛋白的途径。我们将确定AMPK-α2/dsRNA-的作用 在肌动蛋白-α1和肌球蛋白重链（MHC）的修饰中，调节的氧化还原和PKR信号传导以及调节 平滑肌钙吸收。我们将研究预防氧化还原和PKR介导的治疗策略 肌动蛋白-α1和MHC的修饰和反向膀胱缺血中的过度运动。结束 这些研究，我们将有：（1）对迫害者过度活动的病理生理提供了新的见解。 非遮挡的膀胱； （2）阐明了AMPK-α2/DSRNA途径与过度活跃之间的分子链接 膀胱收缩和3）在过度活跃的膀胱收缩中定义了压力敏感性机制。我们的 拟议的研究可能导致针对迫害者过度活动/LUT的新型诊断和治疗策略。