Reactive Nitrogen Species Mediate DNA Damage in Helicobacter pylori-Infected Gastric Mucosa Masaki Katsurahara,* Yoshinao Kobayashi,* Motoh Iwasa,* Ning Ma, † Hiroyuki Inoue,* Naoki Fujita,* Kyosuke Tanaka,* Noriyuki Horiki,* Esteban C. Gabazza* and Yoshiyuki Takei* *Department of Gastroenterology and Hepatology, Division of Clinical Medicine and Biochemical Sciences, Mie University Graduate School of Medicine, Mie, Japan, † Faculty of Health Sciences, Suzuka University of Medical Science, Mie, Japan Introduction Helicobacter pylori infection of the stomach occurs in roughly half of the human population worldwide. Understanding of the etiology of gastritis has been pro- foundly altered by the discovery of H. pylori, which is now recognized as the major cause of chronic gastritis. Chronic nonatrophic gastritis, chronic atrophic gastritis, intestinal metaplasia (IM), and dysplasia represent an inflammatory sequence towards the intestinal type of gastric carcinoma [1]. Helicobacter pylori induces the recruitment and activa- tion of monocytes and neutrophils. These cells produce reactive oxygen species (ROS), such as superoxide anion radical, hydrogen peroxide, and subsequently hydroxyl radical [2,3]. Increased levels of ROS are evident in H. pylori-infected gastric mucosa [4,5]. In parallel with ROS production, inducible nitric oxide synthase (iNOS) is expressed in gastric mucosa with H. pylori infection and the increase in iNOS expression is closely correlated with inflammatory cell infiltration in the gastric mucosa [6]. iNOS catalyzes the production of NO • , which Keywords Helicobacter pylori, oxidative stress, nitrative stress, reactive nitrogen species, 8-nitroguanine, chronic gastritis, eradication. Reprint requests to: Yoshinao Kobayashi, Department of Gastroenterology and Hepatology, Division of Clinical Medicine and Biochemical Sciences, Mie University Graduate school of Medicine, 2-174 Edobashi, Tsu-city, Mie 514-8507, Japan. E-mail: yoshinao@clin.medic.mie-u.ac.jp Abstract Background: Reactive oxygen species (ROS) and reactive nitrogen species (RNS) can play an important role in cellular injury and carcinogenesis of gastric epithelial cells infected with Helicobacter pylori. 8-OH-deoxy guanosine (8-OHdG) and 8-nitroguanine (8-NG) are markers for ROS- and RNS- mediated DNA oxidation, respectively. In this study, RNS-mediated DNA damage in gastric mucosa was observed directly using a newly developed antibody to 8-NG to clarify how H. pylori infection causes nitrative DNA damage to gastric epithelial cells. Methods: Immunohistochemistry with anti-8-OHdG and anti-8-NG antibod- ies was performed on gastric tissue samples from 45 patients (25 men and 20 women) with H. pylori-positive gastritis and 19 patients (11 men and 8 women) exhibiting successful H. pylori eradication. Histologic factors for gastric mucosal inflammation were graded according to the guidelines of the Updated Sydney system. Results: In corpus mucosa, 8-OHdG and 8-NG production were significantly associated with the degree of glandular atrophy, infiltration of chronic inflammatory cells and intestinal metaplasia in the glandular epithelial cells. Successful H. pylori eradication resulted in a significant reduction of chronic inflammatory cell infiltration and neutrophilic activity. Mean 8-OHdG production was lower after H. pylori eradication in both corpus and antral mucosa (p = .022 and .049, respectively). However, the reduction in 8-NG exhibited was more pronounced than the reduction of 8-OhdG (p = .004 and .007, respectively). Conclusions: Helicobacter pylori infection can induce inflammatory cells infiltration, which evokes DNA damage of gastric epithelial cells through ROS and RNS production. 8-NG might be a more sensitive biomarker than 8-OHdG for H. pylori-induced DNA damage in gastric mucosa. Helicobacter ISSN 1523-5378 552 ª 2009 Blackwell Publishing Ltd, Helicobacter 14: 552–558