RAD-Induced Nitric Oxide & Cellular Radiosensitivity

描述（由申请人提供）：最近的几项研究表明，电离辐射激活代谢过程，导致活性氧（ROS）的短暂产生。这种放大的潜在机制和细胞质电离事件的信号传导机制似乎涉及细胞内Ca2+依赖信号从一个线粒体传播到另一个线粒体。时间上一致的是一个组成的，Ca2+依赖性一氧化氮合酶（NOS*）的激活。提出辐射诱导的NOS激活机制是：(1)限制辐射后形成的细胞毒性、致突变性ROS （02-/H202）的数量；(2)激活参与细胞生长调节的信号转导通路。由于其独特的理化性质，增强了NO。辐照后的产生也为单个细胞中发生的电离事件产生的信号在组织内的细胞内和细胞间传播提供了一种机制。为了测试这一提议，有两个具体目标：

DESCRIPTION (provided by applicant): Several recent studies have demonstrated that ionizing radiation activates a metabolic process resulting in the transient generation of reactive oxygen (ROS). The underlying mechanism for this amplification and signaling mechanism of cytoplasmic ionization events appears to involve a Ca2+ dependent signal propagated from one mitochondria to another within a cell. Temporally coincident is the activation of a constitutive, Ca2+ dependent nitric oxide synthase (NOS*). It is proposed that the radiation-induced activation of NOS is a mechanism that (1) limits the amount of cytotoxic, mutagenic ROS (02-/H202) formed after radiation; and (2) activates signal transduction pathways involved in cell growth regulation. Because of its unique physiochemical properties, the enhanced NO. generation after irradiation also provides a mechanism for the intracellular and intercellular propagation within a tissue of a signal generated by an ionization event that has occurred in a single cell. To test this proposal, there are two specific aims: Determine whether modulation of NOS activity by molecular and pharmacological means enhances radiation-induced cell killing and/or mutagenesis that in turn can be reduced by over expression of superoxide dismutase (SOD) or treating cells with SOD mimetics. Determine whether manipulation of NOS activity modulates radiation-induced ROS generation measured as 02 Test (a) whether manipulation of the NO-/sGC/PKG signal transduction pathway alters cellular radiosensitivity; (b) the relationships between radiation-stimulated sGC/PKG and downstream consequences including transcriptional activation of c-fos and p21 Waf1/Cip1 and activation of members of the MAPK family and (c) whether radiation-stimulated transient S-nitrosylation and inhibition of cytosolic and nuclear protein tyrosine phosphatases (PIP) are involved in cellular responses to radiation (SHP-2, SHP-1 and cdc25).