Role of Nitric Oxide in Interstitial Cystitis

描述(申请人提供)：痛性膀胱综合症/间质性膀胱炎(PBS/IC)是一种慢性疼痛的膀胱疾病，没有确凿的病因，也没有有效的治疗方法能够改善症状，包括尿频、尿急、夜尿和疼痛。在家猫中也有一种类似的疾病，称为猫间质性膀胱炎(FIC)，它几乎表现出人类PBS/IC的所有特征和症状。与啮齿动物模型中的急性损伤/炎症不同，患有这种自然发生疾病的猫可能会允许对人类PBS/IC的病理生理学进行更相关的研究。我们在FIC膀胱的尿路上皮细胞中发现了一些异常，包括屏障功能、增殖和生长的改变，以及对化学和物理刺激的高度敏感性/反应。这可能是细胞内钙释放/封存和/或细胞内信号通路改变的结果。FIC膀胱传入神经也有异常，包括形态/电学性质改变、神经放电和神经纤维分布。此外，我们还有证据表明，在盆腔传入区域，FIC脊髓胶质细胞的形态和功能长期发生变化，这表明这些细胞在异常感觉机制的产生中发挥了作用。在长期培养过程中，这些FIC细胞类型发生了持续的变化，这给了我们一个不同的视角，表明沿着感觉通路发生多重变化的可能性。最近的研究结果支持了脊髓神经胶质细胞的激活在持续性疼痛的起始和扩大中的重要作用，因为已知这些细胞可以增强和延长二级脊髓感觉神经元对外周刺激的反应。因此，感觉通路中的多个部位可能具有类似类型的“原发”缺陷，或者尿路上皮中的一个原发缺陷可能会在更近端的部位依次引发继发缺陷。我们在这一更新应用中的目标是进一步了解在尿路上皮(和神经胶质细胞)功能观察到的持续变化背后的信号通路，以及负责各种细胞-细胞相互作用的信号机制，以及这些机制在FIC中可能如何改变。这些目标将使用包括成像技术、分子生物学和发射器释放测量在内的多学科方法来实现。目的1评价PBS/FIC中调节尿路上皮小泡循环和递质释放的细胞内机制。我们推测，在诊断为FIC的猫中，尿路上皮细胞对机械或化学刺激的反应释放了介质/信号因子，从而导致了感觉症状。这一目标将利用膜封闭染料的成像，以探索化学和机械诱导的介质从尿路上皮细胞释放的机制。目的#2将评估PBS/FIC中伞状细胞形态、信号和通讯变化的潜在机制。我们将使用完整的膀胱片和极化的尿路上皮细胞培养来测量尿路上皮层之间顶端细胞特性和细胞信号的变化。目的3评价PBS/FIC中脊髓胶质细胞的形态和功能特性。我们将对培养的脊髓星形胶质细胞的化学、形态和功能以及正常和FIC中神经胶质细胞与神经元的相互作用进行研究。了解这些类型的改变所涉及的机制可能会为PBS/IC临床治疗的新靶点的开发提供重要的见解。 公共卫生意义：痛性膀胱综合征/间质性膀胱炎(PBS/IC)是一种慢性、特发性的膀胱疼痛状况，通常被描述为尿路上皮(膀胱腔内的上皮层)疾病。猫的一种类似疾病，称为猫间质性膀胱炎(FIC)，表现出与人类PBS/IC几乎所有的特征和症状。这是一项授权的更新申请，我们确定FIC-尿路上皮对化学和物理刺激表现出高度的敏感性，随后尿路上皮来源的递质释放增加。这些研究将检验这种类型的非神经性痛觉异常/痛敏的潜在机制。了解导致和维持这些类型变化的机制可能为确定未来PBS/IC等膀胱疾病临床治疗的新靶点提供重要的洞察力。

DESCRIPTION (provided by applicant): Painful bladder syndrome/Interstitial cystitis (PBS/IC) is a chronic painful condition of the urinary bladder in which there are no proven etiologies and no effective treatments that are able to ameliorate the symptoms, which include urinary frequency, urgency, nocturia and pain. There is a comparable disease in domestic cats termed feline interstitial cystitis (FIC), which demonstrates nearly all of the characteristics and symptoms of human PBS/IC. Cats with this naturally occurring disease, as opposed to acute injury/inflammation in rodent models, may permit more relevant studies of the pathophysiology of PBS/IC in humans. We have identified a number of abnormalities in the urothelium of FIC bladders including alterations in barrier function, proliferation and growth, and a heightened sensitivity/response to both chemical and physical stimuli. These may be a result of changes in intracellular Ca2+ release/sequestration and/or intracellular signaling pathways. There are also abnormalities in FIC bladder afferents including altered morphological/electrical properties, nerve firing and distribution of nerve fibers. Moreover, we also have evidence of long-term alterations in FIC spinal cord glial cell morphology and function in regions of pelvic afferent input suggesting that these cells play a role in the generation of abnormal sensory mechanisms. The persistent change that takes place in these FIC cell types during long-term culture has given us a different perspective suggesting the probability of multiple changes along the sensory pathway. Recent findings support an important role for activation of spinal cord glial cells in both initiation and amplification of persistent pain, as it is known that these cells can enhance and prolong the response of second order spinal sensory neurons to peripheral stimulation. Thus, multiple sites in the sensory pathway may have a similar type of 'primary' defect, or a primary defect in the urothelium may sequentially trigger a secondary defect at more proximal sites. Our goals in this renewal application are to further understand the signaling pathways underlying the persistent changes observed in urothelial (and glial cell) function, as well as signaling mechanisms responsible for various cell-cell interactions and how these mechanisms may be altered in FIC. These goals will be accomplished using a multidisciplinary approach including imaging techniques, molecular biology and measurement of transmitter release. Aim #1 will evaluate intracellular mechanisms regulating urothelial vesicle recycling and transmitter release in PBS/FIC. We hypothesize that the release of mediators/signaling factors from the urothelium in response to mechanical or chemical stimuli contributes to the sensory symptoms in cats diagnosed with FIC. This aim will utilize imaging with membrane-impermeant dyes in order to explore the mechanisms responsible for chemical and mechanical evoked release of mediators from urothelial cells. Aim #2 will evaluate mechanisms underlying changes in umbrella cell morphology, signaling and communication in PBS/FIC. We will use intact bladder sheets and polarized urothelial cultures to measure changes in apical cell properties and cell signaling between urothelial layers. Aim #3 will evaluate morphological/functional properties of spinal cord glial cells in PBS/FIC. We will characterize the chemistry/morphology and function of cultured spinal cord astrocytes as well as glial-neuronal interactions in normal and FIC. Understanding the mechanisms involved in these types of changes may provide important insights for the development of novel targets for the clinical management of PBS/IC. PUBLIC HEALTH RELEVANCE: Painful Bladder Syndrome/Interstitial Cystitis (PBS/IC) is a chronic, idiopathic painful condition of the urinary bladder which is often described as a disease of the urothelium (the epithelial layer lining the bladder lumen). A comparable disease in cats, termed feline interstitial cystitis (FIC), exhibits nearly all of the characteristics and symptoms to that of human PBS/IC. This is a renewal application of a grant where we determined that FIC-urothelium exhibits a heightened sensitivity to chemical and physical stimuli with subsequent augmentation of urothelial-derived transmitter release. These studies will examine mechanisms underlying this type of nonneuronal allodynia/hyperalgesia. Understanding the mechanisms contributing to and maintaining these types of changes may provide important insight for the identification of novel targets for the future clinical management of bladder diseases such as PBS/IC.