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: 10200663
• 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/10200663
• 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; efficacy evaluation; energy balance; human model; impaired capacity; insight; lower urinary tract symptoms; male; mouse model; novel; novel diagnostics; novel therapeutic intervention; 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.

这项更新研究计划的总体目标是确定缺血在发展中的作用 另一个目标是检查靶向细胞的功效。 预防或逆转膀胱功能障碍的应激反应途径是。 一系列令人烦恼的排尿症状，对生活质量有重大影响。 退伍军人中膀胱功能障碍和 LUTS 的发生率几乎是普通人的五倍 研究表明，患有 LUTS 的退伍军人的生活质量得分低于退伍军人。 在大多数情况下，尤其是老年退伍军人，LUTS 会导致失眠、焦虑、疲劳和疲劳。 大多数男性 LUTS 病例归因于良性膀胱出口梗阻 (BOO)。 然而，研究表明，大约三分之一到一半以上都有前列腺肥大。 在大多数情况下，LUTS 与 BPE 或 BOO 无关。这些观察结果表明，除了 BOO 之外， 越来越多的基础和临床证据表明，与年龄相关的膀胱局部变化会导致 LUTS。 研究表明，与衰老相关的膀胱缺血在人体膀胱血液损伤中发挥着关键作用。 流量和膀胱缺血的发展已在老年 LUTS 患者中得到证实。 导致膀胱缺血时逼尿肌过度活动的潜在机制仍然很大程度上未知。 初步数据表明，缺血会引发细胞应激并损害细胞防御能力 通过损害细胞能量传感器腺苷单磷酸激活蛋白激酶 α-2 (AMPK- α2)。细胞应激和缺陷的 AMPK-α2 产生具有两个互补的独特应激反应 RNA。 链，即双链 RNA (dsRNA)，导致 AMPK-α2/dsRNA 应激反应的激活 AMPK-α2/dsRNA 途径似乎会损害毒蕈碱 M2 和 M3 受体，引发 收缩蛋白的翻译后修饰，增加平滑肌收缩并产生 我们敢说“慢性缺血是逼尿肌过度活动的一个独立因素”。 无梗阻膀胱逼尿肌过度活动的机制涉及细胞应激的激活。 通过 AMPK-α2/dsRNA 途径做出反应，通过修饰平滑肌细胞来触发膀胱过度收缩 肌肉收缩元件”使用完善的膀胱缺血模型以及基因敲除小鼠和 在细胞培养转染和基因删除技术中，我们提出了三个具体目标。 我们将确定膀胱缺血中 AMPK-α2/dsRNA 应激反应途径的分子调节。 AMPK-α2 损伤的机制，量化和克隆膀胱缺血中的 dsRNA 并定义串扰 AMPK-α2 和 dsRNA 之间的机制在目标 II 中，我们将通过以下方式定义过度活跃收缩的调节。 AMPK-α2/dsRNA 通路在膀胱缺血中的作用 我们将确定 AMPK-α2/dsRNA 通路如何引发。 毒蕈碱 M2 和 M3 的转录和翻译后修饰导致膀胱过度收缩 我们将研究防止 M2 和 M3 的应激调节修饰的治疗策略 在目标 III 中，我们将定义 AMPK-α2/dsRNA 下游的信号传导机制。 我们将确定 AMPK-α2/dsRNA- 的作用。 肌动蛋白-α1 和肌球蛋白重链 (MHC) 修饰和调节中氧化还原和 PKR 信号传导的调节 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 修饰以及逆转膀胱缺血中的逼尿肌过度活动。 通过这些研究，我们将：（1）为逼尿肌过度活动的病理生理学提供新的见解 膀胱无梗阻；(2)阐明了AMPK-α2/dsRNA通路与过度活跃之间的分子联系 膀胱收缩和3）明确的膀胱过度收缩的压力敏化机制。 拟议的研究可能会导致针对逼尿肌过度活动/LUTS 的新诊断和治疗策略。