Roles of Nitric Oxide and Superoxide in Cystitis

描述（由申请方提供）：盆腔区域的辐照可导致膀胱炎症和功能障碍。这种膀胱炎或其可能性也增加了膀胱癌的发病率，禁止对膀胱肿瘤进行放射治疗，并限制了治疗其他盆腔恶性肿瘤的允许放射剂量。放射性膀胱炎的发病机制尚不清楚。它可能涉及伞细胞特有的线粒体一氧化氮（NO）合酶（mtNOS）的激活，渗透性屏障的破坏和尿液渗入固有层。这反过来又会导致炎症和固有层中胶原III沉积增加。膀胱顺应性降低和功能障碍的结果。我们已经开发了放射性膀胱炎的啮齿动物模型，其中照射导致在12小时内跨上皮阻力降低和尿素和水渗透性增加。在6个月时，膀胱测压显示膀胱顺应性和收缩间期减少，而残余容量和基线压力增加，这些特征表明纤维化。用辐射防护剂锰超氧化物歧化酶（MnSOD）进行的先前转染仅部分有效，最可能是由于过氧化物酶活性降低和过量的过氧化氢形成。然而，在照射期间使用NOS抑制剂的新型膀胱内治疗，或照射缺乏mtNOS的膀胱，提供了几乎完全的保护。抑制NO可以阻止其与超氧化物（O2）反应形成过氧亚硝酸盐（ONO2-），过氧亚硝酸盐（ONO2-）可以破坏呼吸链的复合物I和III并导致凋亡/坏死细胞死亡。具体目标1将检验电离辐射激活mtNOS，导致活性氮和氧物质（RNS和ROS）破坏尿路上皮通透性屏障的假设。我们已经开发了NO和ONO2微传感器，使我们能够同时测量实时，这些代谢物在完整的小鼠膀胱和培养的尿路上皮细胞的水平变化。这些测量将与线粒体酶功能的测定变化相关。具体目标2将检验膀胱内施用NOS拮抗剂或MnSOD或具有过氧化物酶活性的SOD模拟物保护膀胱免受放射性膀胱炎的假设。将在1至6个月时通过采用渗透性测量、膀胱测压和胶原沉积的组织化学分析来评估这些疗法在辐照小鼠膀胱中的有效性。

DESCRIPTION (provided by applicant): Irradiation of the pelvic region can result in bladder inflammation and dysfunction. This cystitis or its likelihood also increases the incidence of bladder cancer, prohibits radiation treatment for bladder tumors, and limits the allowable radiation dose for treating other pelvic malignancies. The mechanism of radiation cystitis is unclear. It may involve activation of a mitochondria nitric oxide (NO) synthase (mtNOS) unique to the umbrella cells, disruption of the permeability barrier and infiltration of urine into the lamina propria. This in turn can lead to inflammation and increased collagen III deposition in the lamina propria. Decreased bladder compliance and dysfunction result. We have developed rodent models of radiation cystitis where irradiation results in decreased transepithelial resistance and increased urea and water permeabilities within 12 hours. At six months, cystometrograms show that bladder compliances and intercontractile intervals are decreased while residual volumes and baseline pressures are increased-features indicative of fibrosis. Prior transfection with the radioprotectant manganese superoxide dismutase (MnSOD) is only partially effective, most likely due to decreased peroxidase activity and excess hydrogen peroxide formation. However, novel intravesical therapy with a NOS inhibitor during irradiation, or irradiation of bladders devoid of mtNOS, offers almost complete protection. Inhibition of NO can prevent its reaction with superoxide (O2) to form peroxynitrite (ONO2-), which can damage complexes I and III of the respiratory chain and lead to apoptotic/necrotic cell death. Specific Aim 1 will test the hypothesis that ionizing radiation activates mtNOS, resulting in reactive nitrogen and oxygen species (RNS and ROS) which disrupt the urothelial permeability barrier. We have developed NO and ONO2- microsensors which allow us to simultaneously measure in real-time, the changing levels of these metabolites in intact mouse bladders and cultured urothelial cells. These measurements will be correlated with assayed changes in mitochondrial enzyme functions. Specific Aim 2 will test the hypothesis that the intravesical administration of NOS antagonists or MnSOD or SOD mimetics with peroxidase activity protect the bladder against radiation cystitis. The effectiveness of these therapies in irradiated mouse bladders will be assessed at 1 to 6 months by employing permeability measurements, cystometry and histochemical analyses for collagen deposition.