H(2)S-Releasing Aspirin Protects against Aspirin-Induced Gastric Injury via Reducing Oxidative Stress Lei Liu1, Jie Cui1, Cheng-Jie Song1, Jin-Song Bian2, Anna Sparatore3, Piero Del Soldato4, Xin-Yu Wang5*, Chang-Dong Yan1* 1 Department of Physiology, Xuzhou Medical College, Xuzhou, Jiangsu, People’s Republic of China, 2 Department of Pharmacology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore, Singapore, 3 Dipartimento di Scienze Farmaceutiche ‘‘Pietro Pratesi,’’ Universita` degli Studi di Milano, Milan, Italy, 4 CTG Pharma, Milan, Italy, 5 Department of Endocrinology, Shenzhen Second People’s Hospital, Guangdong Province, People’s Republic of China Abstract The aim of this study was to examine the effect of ACS14, a hydrogen sulfide (H2S)-releasing derivative of aspirin (Asp), on Asp-induced gastric injury. Gastric hemorrhagic lesions were induced by intragastric administration of Asp (200 mg/kg, suspended in 0.5% carboxymethyl cellulose solutions) in a volume of 1 ml/100 g body weight. ACS14 (1, 5 or 10 mg/kg) was given 30 min before the Asp administration. The total area of gastric erosions, H2S concentration and oxidative stress in gastric tissues were measured three hours after administration of Asp. Treatment with Asp (200 mg/kg), but not ACS14 (430 mg/kg, at equimolar doses to 200 mg/kg Asp), for 3 h significantly increased gastric mucosal injury. The damage caused by Asp was reversed by ACS14 at 1–10 mg/kg in a concentration-dependent manner. ACS14 abrogated Asp-induced upregulation of COX-2 expression, but had no effect on the reduced PGE2 level. ACS14 reversed the decreased H2S concentrations and blood flow in the gastric tissue in Asp-treated rats. Moreover, ACS14 attenuated Asp-suppressed superoxide dismutase-1 (SOD-1) expression and GSH activity, suggesting that ACS14 may stimulate antioxidants in the gastric tissue. ACS14 also obviously inhibited Asp-induced upregulation of protein expression of oxidases including XOD, p47phox and p67phox. In conclusion, ACS14 protects Asp induced gastric mucosal injury by inhibiting oxidative stress in the gastric tissue. Citation: Liu L, Cui J, Song C-J, Bian J-S, Sparatore A, et al. (2012) H(2)S-Releasing Aspirin Protects against Aspirin-Induced Gastric Injury via Reducing Oxidative Stress. PLoS ONE 7(9): e46301. doi:10.1371/journal.pone.0046301 Editor: Peiwen Fei, University of Hawaii Cancer Center, United States of America Received June 6, 2012; Accepted August 29, 2012; Published September 28, 2012 Copyright: 2012 Liu et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This study was supported by research grants from China National Science Fund in Jiangsu Province (BK2009088) and National Natural Science Foundation of China (30873055). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: Dr. Piero Del Soldato is a shareholder and employee of CTG Pharma, Milan, Italy. This company has patent on ACS14 used in this study. There are no further patents, products in development or marketed products to declare. This does not alter the authors’ adherence to all the PLOS ONE policies on sharing data and materials, as detailed online in the guide for authors. * E-mail: yancd55@163.com (CDY); wangxy926@yahoo.com (XYW) Introduction Non-steroidal anti-inflammatory drugs (NSAIDs) are among the most commonly prescribed drugs due to their high efficacy in reducing of pain, fever, inflammation and protection against stroke and myocardial infarction [1]. However, their clinical use is commonly associated with the occurrence of adverse effects at the level of digestive tract, ranging from dyspeptic symptoms, gastrointestinal erosions and peptic ulcers to more serious complications, such as overt bleeding or perforation [2]. Redox imbalances appear to play a major pathogenic role in aspirin (Asp) toxicity and gastropathy [3]. To overcome the adverse effects related to NSAID-induced gastrointestinal toxicity, different therapeutic strategies have been evaluated [4–6]. This may include reducing the risk of gastrointestinal damage induced by NSAIDs and enhancing the protective function of the gastric mucosa. Hydrogen sulfide (H2S) is now well recognized to be an endogenous gaseous mediator. Like nitric oxide, another ‘‘gaso- transmitter’’, H2S also regulates various physiological functions [7,8]. It was recently found that H2S produces strong anti- oxidative [9], anti-apoptotic [10] and anti-inflammatory effects [11] in different tissue injuries. H2S and H2S-releasing molecules are able to enhance intracellular antioxidant activities by means of several mechanisms, including stimulation of glutathione and induction of the antioxidant and tissue protective protein heme oxygenase-1 [12–14]. ACS14 is a H2S releasing compound, 2-acetyloxybenzoic acid 4-(3-thioxo-3H-1, 2-dithiol-5-yl) phenyl ester (ACS 14, S-aspirin) [15,16]. The pharmacological profile of ACS14 was described recently [15]. It contains a dithiolethione moiety which gradually releases H2S for a sustained period [17]. In the present study, we therefore investigated the effect of ACS14 on Asp-induced gastric mucosal injury by examining whether ACS14 can prevent Asp- induced redox imbalances in rats. Methods Animals Male Sprague-Dawley rats, 200–240 g, were obtained from the Animal Center of Xuzhou Medicine College (Xuzhou, China) and were housed at 22uC in a controlled environment with 12 h of artificial light per day. They were fasted for 20–24 h before the experiments but had free access to drinking water. All animal experiments were conducted in accordance with international ethical guidelines and the experimental protocols for using rats PLOS ONE | www.plosone.org 1 September 2012 | Volume 7 | Issue 9 | e46301