Clinical and Experimental Pharmacology and Physiology (2007) 34, 787–791 doi: 10.1111/j.1440-1681.2007.04645.x Blackwell Publishing Asia Original Article HBO-induced oxidative stress H Ay et al. TIME-DEPENDENT COURSE OF HYPERBARIC OXYGEN-INDUCED OXIDATIVE EFFECTS IN RAT LUNG AND ERYTHROCYTES ‡ Hakan Ay,* Turgut Topal, † Bülent Uysal, † Mehmet Özler, † Sükrü Öter, † Ahmet Korkmaz † and Kadir Dündar* *Department of Undersea and Hyperbaric Medicine, † Department of Physiology, Gülhane Military Medical Academy, Ankara, Turkey SUMMARY 1. The oxygen toxicity of hyperbaric oxygen (HBO) treatment has long been of interest. There is an extensive amount of informa- tion regarding the role oxidative stress plays after HBO exposure in different tissues, but the question of the persistence of this oxidative effect has not been thoroughly elucidated. 2. The present study was performed to elucidate the persistence of the oxidative effects of HBO on rat lungs and erythrocytes after they had been subjected to 100% oxygen exposure. 3. Rats were divided into five groups. All animals, except those in the control group, were subjected to 100% oxygen for 2 h at 3 ATA (∫ 300 kPa). Rats were killed at 30, 60, 90 or 120 min after exposure and thiobarbituric acid-reactive substances (TBARS) levels and the activity of superoxide dismutase (SOD) and glutathione peroxidase (GPx) were determined. 4. Thiobarbituric acid-reactive substances levels and SOD and GPx levels were found to be significantly increased in lung tissue up to 60 min after exposure. Superoxide dismutase activity persisted at significantly high values for 90 min after exposure in erythrocytes and the lung. The TBARS levels in erythrocytes were also significantly higher for 60 min, whereas increased GPx activity was observed to persist for only 30 min. 5. The oxidative effect of HBO exposure declines to physiological levels within 90 min at most for erythrocytes and in lung tissue in rats. Further studies should focus on the molecular mechanisms that can be activated during this time interval. Key words: anti-oxidant enzymes, erythrocyte, hyperbaric oxygen, lung, oxidative stress. INTRODUCTION Hyperbaric oxygen (HBO) therapy has been recommended for and has been used in the treatment of a wide variety of medical conditions over the past 40 years. The Undersea and Hyperbaric Medical Society (UHMS) approves the use of HBO for a few conditions, based on reasonable scientific evidence or well-validated clinical experience, namely air or gas embolism, carbon monoxide poisoning, clostridial myositis and myonecrosis (gas gangrene), crush injury, compartment syndrome and other acute traumatic ischaemias, decompression sickness, enhancement of healing in selected problem wounds, excep- tional blood loss (anaemia), necrotizing soft tissue infections, refractory osteomyelitis, delayed radiation injury (osteoradionecrosis), com- promised skin grafts and flaps, thermal burns and intracranial abscess. 1 The most important reason for this limitation is the potential risk of oxygen toxicity from the 100% oxygen breathed during the session. 2 Experimental studies focusing on HBO-induced toxicity have reported toxic effects with relatively higher atmospheric pressures, such as 4–5 ATA (atmospheres absolute; 1 ATA = 100 kPa) and/or longer exposure durations of over 2 h. 3–7 However, it is impossible to extrapolate these results to the approved therapeutic limits of a maximum 3 ATA pressure for 2 h. 1 Therefore, we have tested the ability of HBO to induce oxidative stress within these maximal limits determined in previous studies. 8–10 First, we observed that exposure to HBO at 3 ATA for 2 h resulted in significant oxidative stress in the rat lung and that melatonin, a strong anti-oxidant molecule, is capable of blocking this effect. 9 We then determined that increasing exposure pressure also increased the oxidative effect of HBO: signi- ficant oxidative effects were initiated with normobaric 100% oxygen, but with incremental pressure the oxidative levels increased significantly. 10 Although it is well known that HBO exposure causes oxida- tive stress, the molecular mechanisms responsible remain unclear. In addition, it has also been suggested that some of the beneficial effects of HBO may occur via the action of the stimulated reactive molecules, especially superoxide ( ) and hydrogen peroxide (H 2 O 2 ). 11 In a previous study, published in 1983, Noda et al. demonstrated that increased tissue levels of HBO-induced lipid peroxides were not detected 3 h after exposure. 12 Since that report until the present time, there seems to be no additional detailed data in the literature regarding the persistence of the oxidative effects of HBO. Therefore, we undertook the present study to elucidate the time-course of the effects of HBO and to provide further information to help determine the molecular actions of HBO therapy. Taking into consideration our previous studies, the rat seemed to be a useful species in which to undertake these investigations. 8–10 The lung was given preference because it is the primary target of high oxygen levels, as are the oxygen-carrying cells, the erythrocytes. Oxidative status was determined by measurement of thiobarbituric acid-reactive substances (TBARS), as an index of lipid peroxidation, and the activity of the anti-oxidant enzymes superoxide dismutase Correspondence: Dr Sükrü Öter, Gülhane Askeri Tip Akademisi, Fizyoloji Anabilim Dali, 06018, Etlik, Ankara, Türkiye. Email: fizyoter@gmail.com ‡ Preliminary outcomes of this study were presented at the UHMS 2006 Annual Meeting (Ay H, Topal T, Uysal B et al. Oxidative effect of hyperbaric oxygen in rat lung and erythrocytes declines in a time-dependent manner after exposure. In: Proceedings of the Undersea and Hyperbaric Society Scientific Meeting, 22–24 June 2006, Orlando, FL. 2006; 113 (Abstract K6).) Received 25 September 2006; revision 14 January 2007; accepted 29 January 2007. © 2007 The Authors Journal compilation © 2007 Blackwell Publishing Asia Pty Ltd O 2 -