Roles of Nitric Oxide and Superoxide in Cystitis

描述(申请人提供)：照射骨盆区域会导致膀胱炎和功能障碍。这种膀胱炎或其可能性还会增加膀胱癌的发病率，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。它可能涉及伞状细胞特有的线粒体一氧化氮合酶(MtNOS)的激活，通透性屏障的破坏和尿液向固有层的渗透。这反过来会导致炎症和III型胶原在固有层的沉积增加。导致膀胱顺应性下降和功能障碍。我们已经建立了放射性膀胱炎的啮齿动物模型，在12小时内，辐射导致跨上皮阻力降低，尿素和水的通透性增加。在6个月时，膀胱测量显示，膀胱顺应性和收缩间期减少，而残余容量和基线压力增加-这是纤维化的特征。预先用放射防护剂锰超氧化物歧化酶(MnSOD)转染只有部分效果，很可能是由于过氧化物酶活性降低和过氧化氢生成过量所致。然而，在照射期间使用一氧化氮合酶抑制剂的新型膀胱内治疗，或照射没有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)反应生成过氧亚硝酸盐(ONO2-)，过氧亚硝酸盐可破坏呼吸链的复合体I和III，导致细胞凋亡/坏死。具体目标1将测试电离辐射激活线粒体一氧化氮合酶，导致活性氮和氧物种(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微型传感器，使我们能够同时实时测量这些代谢物在完整的小鼠膀胱和培养的尿路上皮细胞中的变化水平。这些测量将与线粒体酶功能的检测变化相关联。具体目标2将验证这样一种假设，即膀胱内注射一氧化氮合酶拮抗剂或具有过氧化物酶活性的MnSOD或SOD模拟物可以保护膀胱免受放射性膀胱炎的伤害。这些疗法在1到6个月后将通过渗透性测量、膀胱测定法和胶原沉积的组织化学分析来评估这些疗法在受照射的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.