Nitroxides (lead compound, Tempol), which are potent antioxidants, are proving to have broad utility in a number of disease processes and/or conditions that represent excessive oxidative stress. The fact that nitroxides exert activity over such a range of disease conditions speaks to the importance of free radical reactions in tissue. Likewise, it is becoming apparent that free radicals are important in normal molecular signaling pathways and related gene expression. We have further demonstrated that Tempol application can provide protection of normal tissues exposed to radiation. Using a miniature pig (minipig) model for irradiation-induced oral mucositis, animals were exposed to daily fractionated radiation (5 x 6 Gy), where the nitroxide Tempol was administered i.p. 10 min before each radiation fraction. Tempol provided protection against radiation-induced mucositis and ulceration. These findings are consistent with mouse studies conducted recently showing comparable radioprotective effects. We have also finished studies evaluating Tempol to reduce the severity and progression of multiple sclerosis (MS). Several reactive oxygen (ROS) and reactive nitrogen species (RNS) are implicated in inflammatory-mediated damage to the central nervous system (CNS) in MS and its animal model, experimental autoimmune encephalomyelitis (EAE). The goal of these studies was to investigate the immunomodulatory effects and therapeutic potential of orally-delivered Tempol in the mouse EAE model. Mice receiving TEMPOL chow ad libitum for 2 weeks prior to induction of active EAE showed delayed onset and reduced incidence of disease compared to control-fed animals. Reduced disease severity was associated with limited microglial activation and fewer inflammatory infiltrates. Tempol's effects were immunomodulatory, not immunosuppressive: T cells produced less interferon-gamma and tumor necrosis factor-alpha. TEMPOL administration was associated with an enrichment of CD8+ T cell populations and CD4+FoxP3+ regulatory populations. Tempol treatment also reduced the severity of clinical disease when administered after the induction of disease, and also after the onset of clinical symptoms. The ability of oral TEMPOL to reduce inflammation and axonal damage and loss demonstrate both anti-inflammatory and protective properties, with significant promise for the treatment of MS and related neurological disorders. We have recently initiated a repeat study where ear-tagged mice which received either 0 Gy (n = 25) or 5.4 Gy (n = 75) TBI. Multiple urine samples from each mouse were collected and stored over a three-month period post-IR. Animals were followed to the end of their lifespan and pathology assessment will be performed on each mouse to determine cause of death. Urine samples for control and irradiated mice have been processed by UHPLC-ESI-QTOFMS. Preliminary assessment of metabolites of the repeat study have been compared with results from the initial study. Between the two experiments 304 metabolites were identified as being important in separating control and IR samples. As the number of relevant metabolites become identified, chemical synthesis will be required carried out as required. The overall goal is to chemically identify a group of metabolites which may be predictive of people exposed to non-lethal IR that would predict for carcinogenesis within this group.
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