NADPH oxidase-associated transition from Barrett's esophagus to adenocarcinoma

描述(由申请人提供)：胃食道反流病(GERD)影响超过十分之一的40岁以上的成年人和四分之一的60岁以上的成年人。大约10%的GERD患者患有巴雷特食道(BE)，这与发生食管腺癌(EA)的风险增加近30-125倍有关。从BE到异型增生和EA的进展机制尚不清楚。因此，我们将研究这一进程中可能涉及的一些途径。BE和EA患者体内的ROS水平升高，可能在BE向EA的转化过程中起关键作用。我们发现一种新的NADPH氧化酶亚型NOX5-S在高度不典型增生的EA细胞系、EA组织和BE粘膜中高表达。NOX5-S基因最先在人胎肾中发现，在其他组织中未见报道。在Seg1EA细胞中，脉冲酸暴露引起NOX5-S表达上调，并增加过氧化氢的产生。NOX5-S来源的ROS有助于促进SEG1细胞的增殖和减少细胞凋亡。然而，酸诱导NOX5-S上调和依赖NOX5-S促进细胞增殖和减少凋亡的机制尚不完全清楚。因此，我们将测试这一假设，即酸暴露通过激活Barrett化生细胞中的Rho激酶和MAP激酶上调NADPH氧化酶，导致自由基产生过剩，进而可能通过激活核因子kappaB(NF-kB)进一步增强NADPH氧化酶的表达。此外，ROS还可上调细胞周期蛋白D1和死亡结构域沉默蛋白(SODD)的表达。细胞周期蛋白D1和SODD的表达上调可促进细胞增殖，减少细胞凋亡。因此，BE患者持续的酸反流可能导致高水平的ROS，细胞增殖增加，细胞凋亡减少，从而导致DNA损伤和突变增加，从而促进BE向异型增生和EA的进展。为了验证这一假设，我们将：1)在BE细胞系、Barrett‘s粘膜活检组织和EA细胞(Seg1、Flo和OE33)中检测酸诱导NADPH氧化酶表达的信号转导途径。我们将重点介绍Rho激酶、MAP激酶和核因子-kB的作用。2)研究NADPH氧化酶依赖的细胞增殖增加和细胞凋亡减少的机制，重点研究细胞周期蛋白D1和SODD的作用，以及NADPH氧化酶衍生的ROS是否导致DNA损伤。3)研究NOX5-S发挥功能所需的调控蛋白，重点是Rac1和p22Phox。更好地了解酸诱导NADPH氧化酶上调，导致细胞增殖增加和细胞凋亡减少的信号转导途径，可能为预防EA的发展提供合理的途径。此外，NOX5-S有可能成为早期发现食管异型增生的潜在生物标志物。我们的模型也可能适用于其他与炎症相关的癌症。 公共卫生相关性：这项建议研究了一种特殊的酶NOX5-S在巴雷特食道(一种食道细胞被酸反流改变的情况)发展到食道癌过程中的作用。该项目旨在寻找潜在的治疗靶点来防止这种进展。

DESCRIPTION (provided by applicant): Gastroesophageal reflux disease (GERD) affects more than one in ten adults over 40 years of age and one in four adults over 60. Approximately 10% of GERD patients develop Barrett's esophagus (BE), which is associated with nearly a 30-125-fold increased risk for the development of esophageal adenocarcinoma (EA). The mechanism of progression from BE to dysplasia and to EA is unknown. We will therefore examine some of the pathways that may be involved in this progression. Reactive oxygen species (ROS) are elevated in BE and EA, and may play a key role in the progression from BE to EA. We found that a novel isoform of NADPH oxidase NOX5-S is overexpressed in EA cell lines, EA tissues, and BE mucosa with high grade dysplasia. NOX5-S was first identified in human fetal kidney and has not been described in other tissues. In SEG1 EA cells pulsed acid exposure causes upregulation of NOX5-S and increases H2O2 production. NOX5-S-derived ROS contribute to increased cell proliferation and decreased apoptosis in SEG1 cells. However, the mechanisms of acid-induced NOX5-S upregulation and NOX5-S-dependent increase in cell proliferation and decrease in apoptosis are not fully understood. We will therefore test the hypothesis that acid exposure upregulates NADPH oxidases via activation of Rho kinases and MAP kinases in Barrett's metaplastic cells, causing overproduction of free radicals, which in turn may further enhance the expression of NADPH oxidases through activation of the nuclear factor kappaB (NF-kB). In addition, ROS may upregulate cyclin D1 and the silencer of the death domain (SODD). Upregulation of cyclin D1 and SODD will increase cell proliferation and decrease apoptosis in these cells. Thus the persistent acid reflux present in BE patients may cause high levels of ROS, increased cell proliferation and decreased apoptosis, which may lead to DNA damage and increased mutations, contributing to the progression from BE to dysplasia and to EA. To test this hypothesis we will: 1) Examine the signal transduction pathway of acid-induced expression of NADPH oxidases in a BE cell line, in Barrett's mucosal biopsies, and in EA cells (SEG1, FLO, and OE33). We will focus on the role of Rho kinase, MAP kinases and NF-kB. 2) Examine the mechanisms of NADPH oxidase-dependent increase in cell proliferation and decrease in apoptosis, concentrating on the role of cyclin D1 and SODD, and examine whether NADPH oxidase-derived ROS cause DNA damage. 3) Examine the regulatory proteins required for NOX5-S to function, focusing on Rac1 and p22phox. A better understanding of the signal transduction pathway of acid induced upregulation of NADPH oxidases, leading to increased cell proliferation and decreased apoptosis, may provide a rational approach to the prevention of development of EA. In addition, NOX5-S may be a potential biomarker for early detection of esophageal dysplasia. Our model may also be applicable to other inflammation-associated cancers. PUBLIC HEALTH RELEVANCE: This proposal studies the role of a particular enzyme NOX5-S in the progression from Barrett's esophagus, a condition where cells in the esophagus have been altered by acid reflux, to esophageal cancer. This project is designed to find potential therapeutic targets to prevent this progression.