Linkage of Lung Inflammation to 8-oxoguanine and OGG1

描述（由申请人提供）：8-氧-7,8-二氢鸟嘌呤（8-oxoG）通常被用作氧化应激的标志物，是由环境和内源性活性氧（ROS）在核酸中产生的。它是DNA中的一种突变前病变，因为它在复制过程中可能与腺嘌呤错配。碱基8-oxoG在DNA碱基切除修复（BER）途径中被8-oxoG DNA糖基酶1 （OGG1）从DNA中去除。8-oxoG修复的减少和由此产生的积累与各种人类疾病和衰老有关，尽管其病因学作用尚不清楚。炎症是大多数疾病的根源，包括呼吸系统疾病、心血管疾病、中枢神经系统疾病和致癌疾病。豚草花粉提取物（RWPE：具有促氧化和抗原性成分）增加了基因组中8-oxoG的水平和小鼠气道中OGG1的活性。在RWPE暴露前，致敏小鼠肺中OGG1（但不包括其他氧化碱基特异性DNA糖基酶）的下调显著降低了过敏性气道炎症。重要的是，EG8-oxoG（基因组外8-oxoG）单独诱导小鼠肺部趋化因子的表达，以及中性粒细胞的积累。我们的数据还表明，EG8-oxoG增加了1)活化的小gtpase的水平；2) Ras与Raf-1结合；3) MEK1,2的磷酸化；4) ERK1 2;5) RelA-Ser276。EG8-oxoG诱导由CXCL-8启动子驱动的荧光素酶表达。值得注意的是，其他氧化嘌呤碱没有这种影响。这些意想不到的观察结果使我们假设，OGG1从DNA中释放的8-oxoG作为一种信号分子发挥作用，因为它能够增加激活的小gtpase的水平，从而启动细胞激活事件的级联反应，导致促炎介质表达增加和炎症加剧。我们将通过追求三个具体目标来检验这一假设。我们将研究是否：Aim 1)缺乏8-oxoG修复使小鼠对炎症难以抵抗；目的2)OGG1的糖基酶活性在翻译后被调节，从而从DNA中释放EG8-oxoG；3) EG8-oxoG通过NF-？B，由Ras-Raf-MEK/ERK- MSK1通路激活。肺部炎症的小鼠疾病模型将用于建立我们在培养细胞中产生的结果的病因学相关性。这些目标的完成将提供第一个证据，证明EG8-oxoG是氧化应激介导的DNA损伤/修复和细胞反应之间的联系，作为一个信号分子诱导促炎趋化因子的表达。我们的机制研究也应该为新的治疗方法奠定基础。例如，捕获、清除或将EG8-oxoG转化为非信号形式的药物应该不仅有利于预防气道的炎症过程，而且有益于心血管、中枢神经系统或肥胖相关的炎症性疾病等。

DESCRIPTION (provided by applicant): 8-oxo-7,8-dihydroguanine (8-oxoG), often used as a marker of oxidative stress, is generated in nucleic acids by environmental and endogenous reactive oxygen species (ROS). It is a premutagenic lesion in DNA because of its mispairing potential with adenine during replication. The base 8-oxoG is removed from the DNA by 8-oxoG DNA glycosylase 1 (OGG1) in the DNA base excision repair (BER) pathway. Decreased repair and resulting accumulation of 8-oxoG have been related to various human diseases and aging, although its etiological role is poorly understood. Inflammation is the root of most diseases including those of the respiratory, cardiovascular, central nervous systems and of carcinogenesis. Ragweed pollen extract (RWPE: has pro-oxidant and antigenic components) increases the 8-oxoG level in the genome and OGG1 activity in the mouse airways. Downregulation of OGG1 (but not of other oxidized-base specific DNA glycosylases) in the lungs of sensitized mice before RWPE exposure significantly decreased allergic airway inflammation. Importantly, EG8-oxoG (extragenomic 8-oxoG) alone induced chemokine expression in mouse lungs, along with neutrophil accumulation. Our data also show that EG8-oxoG increased the levels of 1) activated small GTPases; 2) Ras to Raf-1 binding; and phosphorylation of 3) MEK1,2; 4) ERK1,2; and 5) RelA-Ser276. EG8-oxoG induced luciferase expression driven from the CXCL-8 promoter. Notably, other oxidized purine bases had no such effects. These unexpected observations led us to hypothesize that 8-oxoG liberated from DNA by OGG1 functions as a signaling molecule by virtue of its ability to increase levels of activated small GTPases, thereby initiating cascades of cellular activation events leading to increased pro-inflammatory mediator expression and exacerbation of inflammation. We will test this hypothesis by pursuing three Specific Aims. We will investigate whether: Aim 1) deficiency in 8-oxoG repair renders mice refractory to inflammation; Aim 2) OGG1's glycosylase activity is post-translationally modulated for release of EG8-oxoG from DNA; and Aim 3) EG8-oxoG enhances expression of pro-inflammatory mediators via NF-?B, activated by the Ras-Raf-MEK/ERK- MSK1 pathway. A mouse disease model for lung inflammation will be used to establish the etiological relevance of our results generated in cultured cells. Completion of these aims will provide the first evidence that EG8-oxoG is the link between oxidative stress- mediated DNA damage/repair and cellular responses, by acting as a signaling molecule inducing pro- inflammatory chemokine expression. Our mechanistic studies should also lay the foundation for novel therapeutic approaches. For example, drugs that trap, scavenge, or convert EG8-oxoG into a non-signaling form should be beneficial for the prevention of inflammatory processes not only in airways, but also in cardiovascular, and central nervous systems or in obesity-associated inflammatory diseases, among others. PUBLIC HEALTH RELEVANCE: Respiratory diseases affect >eight hundred million people worldwide, and in the US there are approximately twenty million outpatient visits, two million emergency room visits, and half million hospitalization per year related to these diseases. Our novel observations linking oxidative genome damage repair to inflammation and subsequently identifying its molecular mechanism provide an opportunity to explore unconventional preventive therapeutic approaches to inhibit inflammation and thereby subsequent pathogenesis.