BARRETT'S ESOPHAGUS AND OXIDATIVE STRESS: ROLE OF GASTRIC ACID AND BILE ACID

我们的主要转化目标是开发一种化学预防策略，用于抑制Barrett's的进展。 食管腺癌。我们假设疏水性胆汁的有害作用 酸和低pH可被细胞保护性胆汁酸熊去氧胆酸（UDCA）抑制， 保护细胞免受氧化损伤。 在该提议中待检验的中心假设是胆汁酸与低pH组合诱导 氧化应激导致DNA损伤、基因组不稳定性和Barrett's细胞凋亡抗性 食道因此，熊去氧胆酸治疗引起的胆汁酸组成的改变可能是 有益于Barrett食管患者预防肿瘤进展。 提出了三个具体目标来检验这一假设。首先，我们将评估胆汁酸 鸡尾酒和/或低pH对活性氧（ROS）的产生、氧化、DNA损伤， DNA修复蛋白、抗氧化和抗凋亡蛋白在食管癌细胞系中的体外表达 和离体BE活组织检查。我们将确定急性、反复和慢性暴露于胆汁的影响 酸和/或低pH（1）对超氧化物的诱导，（2）形成8-OH-dG，氧化DNA的标记物 损伤，（3）DNA修复蛋白（即Mlh 1、Pms 2和PARP）、抗氧化剂（即超氧化物）的表达 歧化酶和过氧化氢酶）和抗凋亡蛋白（即Mcl-1、存活素、Bcl-xL）。此外，我们计划 使用基于微阵列的比较基因组杂交（CGH）鉴定DNA改变模式， 针对胆汁酸和/或低pH的抗性开发的新型食管细胞系。在具体目标#2中，我们计划 评价UDCA及其结合形式[（牛磺熊去氧胆酸（TUDCA），甘氨- 熊去氧胆酸（GUDCA）]对ROS产生、氧化性DNA损伤、基因组不稳定性和 低pH和细胞毒性诱导的DNA修复蛋白、抗氧化和抗凋亡蛋白的表达 体外胆汁酸。由于UDCA是一种优秀的ROS清除剂，我们预计UDCA将减少 最后，在具体目标#3中，我们建议进行试点临床试验， 评价UDCA作为一种潜在预防剂的试验，该药物可改变 回流，使得毒性较小的胆汁酸回流到食道。因此，胆汁酸组成在 将确定基线和UDCA治疗后。此外，氧化应激标志物（8-OH-dG） 将在基线、UDCA治疗6个月和UDCA治疗6个月时在BE活检中评估增殖（Ki-67） UDCA停药后，使用免疫组织化学结合图像分析。 我们的长期目标是评估导致BE发展和/或 发展为食管腺癌并抑制这些过程。

Our major translational goal is to develop a chemopreventive strategy for inhibiting progression of Barrett's esophagus to esophageal adenocarcinoma. We hypothesize that the deleterious effects of hydrophobic bile acids and low pH may be inhibited by the cytoprotective bile acid, ursodeoxycholic acid (UDCA), that was shown to protect cells against oxidative injury. The central hypothesis to be tested in this proposal is that bile acids in combination with low pH induce oxidative stress that leads to DMA damage, genomic instability and apoptosis resistance in Barrett's esophagus. Therefore, an alteration of bile acid composition by ursodeoxycholic acid treatment may be beneficial to Barrett's esophagus patients in the prevention of neoplastic progression. Three specific aims are proposed to test this hypothesis. First, we will evaluate the effect of a bile acid cocktail and/or low pH on the generation of reactive oxygen species (ROS), oxidative, DNA damage, expression of DNA repair proteins, anti-oxidant and anti-apoptotic proteins in vitro in esophageal cell lines and ex vivo, in BE biopsies. We will determine the effect of acute, repeated and chronic exposure to bile acids and/or low pH (1) on the induction of superoxide, (2) formation of 8-OH-dG, a marker of oxidative DNA damage, (3) expression of DNA repair proteins (i.e. Mlh1, Pms2 and PARP), anti-oxidant (i.e. superoxide dismutases and catalase) and anti-apoptotic proteins (i.e. Mcl-1, survivin, Bcl-xL). Furthermore, we plan to identify patterns of DNA alterations using microarray-based comparative genomic hybridization (CGH) in novel esophageal cell lines developed for resistance to bile acids and/or low pH. In specific aim #2 we plan to evaluate the effects of UDCA and its conjugated forms [(tauro-ursodeoxycholic acid (TUDCA), glyco- ursodeoxycholic acid (GUDCA)] on ROS production, oxidative DNA damage, genomic instability and the expression of DNA repair proteins, anti-oxidant and anti-apoptotic proteins induced by low pH and cytotoxic bile acids in vitro. Since UDCA is an excellent scavenger of ROS we anticipate that UDCA will reduce damage caused by bile acids and low pH. Finally, in specific aim #3 we propose to conduct a pilot clinical trial to evaluate UDCA as a potential preventive agent that changes the composition of bile acids in the refluxate so that less toxic bile acids are refluxed to the esophagus. Thus, bile acid composition at the baseline and post UDCA treatment will be determined. Furthermore, markers of oxidative stress (8-OH-dG) and proliferation (Ki-67) will be assessed in BE biopsies at the baseline, 6 months on UDCA and 6 months after discontinuation of UDCA using immunohistochemistry in conjunction with image analysis. Our long-term goal is to evaluate factors and molecular mechanisms leading to BE development and/or progression to esophageal adenocarcinoma and to inhibit these processes.