RAD-Induced Nitric Oxide & Cellular Radiosensitivity

描述(由申请人提供)：最近的几项研究表明，电离辐射会激活代谢过程，导致活性氧(ROS)的瞬时产生。细胞质电离事件的这种放大和信号机制的潜在机制似乎涉及到细胞内一个线粒体向另一个线粒体传播的钙依赖信号。在时间上符合的是一种结构性的、依赖钙离子的一氧化氮合酶(NOS*)的激活。目前认为，辐射诱导的一氧化氮合酶的激活是一种机制：(1)限制辐射后形成的细胞毒性、致突变的ROS(02-/H202)的数量；(2)激活参与细胞生长调节的信号转导通路。由于其独特的物理化学性质，增强的NO。辐射后的产生还提供了一种机制，用于在单个细胞中发生的电离事件所产生的信号在组织内的细胞内和细胞间传播。为了测试这一提议，有两个具体目标： 确定通过分子和药物手段调节一氧化氮合酶活性是否增强辐射诱导的细胞杀伤和/或诱变，而辐射诱导的细胞杀伤和/或诱变又可以通过过度表达超氧化物歧化酶(SOD)或用超氧化物歧化酶模拟物处理细胞来减少。确定一氧化氮合酶活性的操纵是否调节辐射诱导的ROS产生，以02为单位 研究目的：(A)NO-/sGC/PKG信号转导通路的调控是否改变细胞的放射敏感性；(B)辐射刺激的sGC/PKG与下游结果之间的关系，包括c-fos和p21Waf1/Cip1的转录激活以及MAPK家族成员的激活；(C)辐射刺激的瞬时S亚硝化以及胞浆和核蛋白酪氨酸磷酸酶的抑制是否参与细胞对辐射的反应(SHP-2、SHP-1和CDC25)。

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).