Original Contribution OXYGEN TENSION REGULATES REACTIVE OXYGEN GENERATION AND MUTATION OF Helicobacter pylori AH-MEE P ARK, *QUAN LI,* KUMIKO NAGATA, y TOSHIHIDE TAMURA, y KUNIO SHIMONO, z EISUKE F. SATO, * and MASAYASU INOUE* ,b *Department of Biochemistry and Molecular Pathology, Osaka City University Medical School, 1-4-3 Asahimachi, Abeno, Osaka 545-8585, Japan; y Department of Bacteriology, Hyogo College of Medicine, 1-1 Mukogawa, Nishinomiya, Hyogo 663-8501, Japan; and z Hinode-En Elder Health Center, Ohishi-Minamimachi, Nada, Kobe, Hyogo 657-0852, Japan (Received 3 October 2003; Revised 21 January 2004; Accepted 2 February 2004) Abstract—Although both bacillary and coccoid forms of Helicobacter pylori reside in human stomach, the pathophysiological significance of the two forms remains obscure. The present work describes the effect of oxygen tension on the transformation and reactive oxygen species (ROS) metabolism of this pathogen. Most H. pylori cultured under an optimum O 2 concentration (7%) were the bacillary form, whereas about 80% of cells cultured under aerobic or anaerobic conditions were the coccoid form. The colony-forming unit of H. pylori decreased significantly under both aerobic and anaerobic culture conditions. The bacillary form of H. pylori generated predominantly superoxide radical, whereas the coccoid form generated preferentially hydroxyl radical. Specific activities of cellular respiration, urease, and superoxide dismatase decreased markedly after transformation of the bacillary form to the coccoid form, with concomitant generation of protein carbonyls and 8-hydroxyguanine. The frequency of mutation of cells increased significantly during culture under nonoptimum O 2 conditions. These results indicate that ROS generated by H. pylori catalyze the oxidative modification of cellular DNA, thereby enhancing the transformation from the bacillary to the coccoid form. The enhanced generation of mutagenic hydroxyl radicals in the coccoid form might accelerate mutation and increase the genetic diversity of H. pylori. D 2004 Elsevier Inc. All rights reserved. Keywords—Helicobacter pylori, Oxidative stress, Superoxide, Mutation, Gastric cancer, Free radicals INTRODUCTION Helicobacter pylori is a gram-negative and microaerobic bacterium that plays important roles in the pathogenesis of gastritis, peptic ulcer, and gastric cancer [1,2]. Two types of H. pylori, bacillary and coccoid forms, are seen in human gastric mucosa; the bacillary form is the predominant form found in human stomach, and the coccoid form is the major form found in extragastric environments [3]. Both oral-to-oral and fecal-to-oral routes have been postulated to be involved in the mechanism of infection of human subjects [4–6]. We previously reported that the oxygen tension in gastric juice changes significantly depending on the injected solutions [7]. Gastric mucosal oxygen tension has been reported to decrease significantly in animals exposed to restraint stress [8]. When the bacillary form of H. pylori is exposed to various stresses, such as unfavorable O 2 tension and antibiotics, it undergoes transformation to the coccoid form [9]. The coccoid form of H. pylori has been postulated to be viable but difficult to grow under culture conditions [10]. The mechanism and pathophysiological significance of the transformation of H. pylori remain unknown. It has been postulated that the transformation of H. pylori from the bacillary to the coccoid form might occur irreversibly, and, hence, the latter form might represent a dormant stage in the life cycle [11,12]. However, the coccoid form of H. pylori has been known Address correspondence to: Dr. Masayasu Inoue, Department of Biochemistry and Molecular Pathology, Osaka City University Medical School, 1-4-3 Asahimachi, Abeno, Osaka 545-8585, Japan; Fax: +81-6- 6645-3721; E-mail: inoue@med.osaka-cu.ac.jp. Free Radical Biology & Medicine, Vol. 36, No. 9, pp. 1126 –1133, 2004 Copyright D 2004 Elsevier Inc. Printed in the USA. All rights reserved 0891-5849/$-see front matter doi:10.1016/j.freeradbiomed.2004.02.001 1126