Nitroxides as Protectors Against Oxidative Stress

摘要事实证明，氮氧化物在代表过度氧化应激的许多疾病过程和/或条件下具有广泛的用途。氮氧化物在这样一系列条件下发挥活性的事实说明了组织中自由基反应的重要性。同样，很明显，自由基在正常的分子信号通路和相关基因表达中也是重要的。我们继续探索(S)长期服用坦波尔(在食物或饮用水中)如何导致小鼠体重显著减轻和自发肿瘤发病率下降的机制(S)。对选定的组织进行基因表达谱研究，以获得在体重变化之前(14天)和在动物体重减轻后(60天)，Tempoll对小鼠的早期基因表达变化。由坦波尔饮食引起的大多数基因变化(上调和下调)发生在肝脏。坦波尔饮食上调了与线粒体功能以及脂肪酸和/或脂肪合成相关的基因。谷胱甘肽代谢基因和Nrf2基因(提供对自由基的保护)上调。我们目前正在评估Tempoll对脂肪组织基因表达谱的影响，特别是如果可以观察到上调或下调的基因对蛋白质的调节。无论是在饮食中还是通过灌胃给药，小鼠体内都会产生数百种尿液代谢产物，其中包括大量的坦波尔代谢物。特别令人感兴趣的是代谢物，如2，8-二羟基喹啉及其葡萄糖苷，与对照组相比升高，而代谢物，如泛甲酸和异丁基肉碱，与对照组相比显著减弱。目前的研究重点是2，8-二羟基喹啉的存在是否与肠道微生物区系有关，以及坦普尔可能如何影响肠道微生物区系。2，8-二羟基喹啉的进一步升高可能是坦普尔治疗诱导脂肪酸β氧化的信号。最后，在一个与氧化应激相关的项目中，我们评估了小鼠体内肿瘤的存在是否会改变尿液代谢物的特征。在荷瘤小鼠的尿液和血液中发现了许多不同的代谢物，包括己酰甘氨酸、烟酰胺1-氧化物、二甲基精氨酸(氧化应激生物标记物)和C反应蛋白(炎症生物标记物)。此外，肝脏的mRNA分析表明，炎性细胞因子、肿瘤坏死因子和转化生长因子-β的诱导以及肝脏细胞色素P450的减少。升高的血清甘油三酯升高，提示肝脏功能障碍。总而言之，这些发现表明，肿瘤生长诱导了炎症反应，导致肝功能障碍。在小鼠体内给药是否可以抑制这种反应并影响肿瘤的生长目前正在研究中。

Summary Nitroxides 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 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 continue to search for the mechanism(s) of how long-term administration of Tempol (in the food or drinking water) results in dramatic weight reduction and a decrease in spontaneous tumor incidence in mice. Gene expression profile studies of selected tissues were conducted to obtain early gene expression changes by Tempol in mice prior to changes in weight (14 days) and later when animal weights were reduced by Tempol (60 days). The majority of gene changes (both up- and down-regulated) resulting from the Tempol diet occurred in the liver. Tempol diet up-regulated genes related to mitochondrial function and fatty acid and/or lipid synthesis. Glutathione metabolism genes and Nrf2 genes (which provide protection against free radicals) were up-regulated. We are currently evaluating the effect of Tempol on gene expression profiles in adipose tissue, in particular if proteins modulation can be observed from genes either up- or down-regulated. Tempol administration to mice either in the diet or by gavage resulted in hundreds of urine metabolic products including numerous Tempol metabolites. Of particular interest were metabolites such as 2,8-dihydroxylquinoline and its glucuonide, which were elevated and metabolites such as panthothenic acid and isobutrylcarnitine which were significantly attenuated compared to control. Current studies are focusing on whether the presence of 2,8-dihydroxylquinoline is related to the gut microflora and how Tempol might be impacting the gut microflora. Further elevation of 2,8-dihydroxylquinoline may signal that Tempol treatment induces fatty acid beta-oxidation. Lastly, in a related project dealing with oxidative stress, we evaluated whether the presence of a tumor in a mouse alters urine metabolite profiles. A number of different urine and blood metabolites were found to be elevated in tumor-bearing mice including hexanoylglycine, nicotinamide 1-oxide, dimethylargine (biomarker for oxidative stress) and c-reactive protein (inflammatory biomarker). Further, mRNA analysis of liver indicated the induction of inflammatory cytokines, TNF, and TGF-beta and decrease in liver cytochromes P450. Elevated serum triglycerides were elevated suggesting liver dysfunction. Collectively, these findings suggest that tumor growth induces an inflammatory response resulting in liver dysfunction. Whether Tempol administration in mice could dampen this response and impact tumor growth is being studied currently.