Role of Nitric Oxide in Interstitial Cystitis

描述（由申请人提供）：膀胱疼痛综合征/间质性膀胱炎（PBS/IC）是一种慢性膀胱疼痛状况，没有明确的病因，也没有有效的治疗方法可以改善症状，包括尿频、尿急、夜尿和疼痛。在家猫中有一种类似的疾病，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。与啮齿类动物模型中的急性损伤/炎症相反，患有这种自然发生疾病的猫可能允许对人类PBS/IC的病理生理学进行更相关的研究。我们已经确定了FIC膀胱尿路上皮的一些异常，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。这些可能是细胞内Ca2+释放/封存和/或细胞内信号通路变化的结果。FIC膀胱传入神经也有异常，包括形态/电特性、神经放电和神经纤维分布的改变。此外，我们也有证据表明，骨盆传入输入区域FIC脊髓胶质细胞形态和功能的长期改变表明这些细胞在异常感觉机制的产生中起作用。在长期培养过程中，这些FIC细胞类型发生的持续变化为我们提供了一个不同的视角，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脊髓胶质细胞的激活在持续疼痛的起始和扩增中发挥重要作用，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相似类型的“原发性”缺陷，或者尿路上皮的原发性缺陷可能依次引发更近端部位的继发性缺陷。我们的目标是进一步了解尿路上皮（和神经胶质细胞）功能持续变化背后的信号通路，以及负责各种细胞间相互作用的信号机制，以及这些机制如何在FIC中改变。这些目标将通过多学科的方法来实现，包括成像技术、分子生物学和测量递质释放。目的1将评估PBS/FIC中调节尿路上皮囊泡循环和递质释放的细胞内机制。我们假设，在机械或化学刺激下，尿路上皮释放的介质/信号因子有助于诊断为FIC的猫的感觉症状。该目的将利用膜外染料成像来探索尿路上皮细胞化学和机械诱发释放介质的机制。目的2将评估PBS/FIC中伞细胞形态、信号和通信变化的潜在机制。我们将使用完整的膀胱片和极化尿路上皮培养来测量顶端细胞特性和尿路上皮层之间细胞信号传导的变化。目的3将评估PBS/FIC中脊髓胶质细胞的形态学/功能特性。我们将描述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中胶质-神经元的相互作用。了解这些类型变化的机制可能为PBS/IC临床管理的新靶点的开发提供重要的见解。

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.