Research paper Chronic gastric ulceration causes matrix metalloproteinases-9 and -3 augmentation: Alleviation by melatonin Krishnendu Ganguly 1 , Snehasikta Swarnakar * Department of Physiology, Drug Development Diagnostics and Biotechnology Division, CSIR-Indian Institute of Chemical Biology, 4, Raja S.C. Mullick Road, Jadavpur, Kolkata 700032, India article info Article history: Received 16 March 2012 Accepted 7 August 2012 Available online 13 August 2012 Keywords: Indomethacin Chronic ulcer Cytokine Matrix metalloproteinase Melatonin Reactive oxygen species Signaling NF-kB AP-1 abstract Matrix metalloproteinases (MMPs) are a family of zinc-dependent enzymes capable of degradation of extracellular matrix (ECM) and key player in various inflammatory diseases. We investigated the regu- lation of MMPs in chronic gastric ulceration in mice. We generated chronic gastric ulcers in mice by indomethacin and examined the activity and expression of MMP-9 and -3 in stomach. Melatonin (N- acetyl-5-methoxytryptamine) treatment has also been applied to mice to characterize the changes in expression and activities of MMPs in gastric tissues. We observed significant upregulation of MMP-9 and -3 expressions and activities in stomach with increasing doses and duration of indomethacin that corroborated with increased activity of activator protein (AP)-1. Substantial damage in gastric epithelial layer was found during chronic ulceration. Melatonin suppressed MMP-9 and -3 expressions and activities during prevention and healing of chronic gastric ulcers. It also suppressed protein oxidation, lipid peroxidation and antioxidant enzymes. Additionally, expression of tumor necrosis factor (TNF)-a, interleukin (IL)-1b and IL-8 was significantly high in ulcerated stomachs while melatonin treatment blocked them to control level. We found elevated phosphorylation of extracellular-regulated kinase (ERK)1/2 and c-Jun N-terminal kinase (JNK) during chronic gastric ulceration, which were significantly reversed by melatonin. Moreover, expression of NF-kB, c-fos and c-jun were inhibited by melatonin resulting down regulation of MMP-9 and -3 expressions. In summary, oxidative stress is preceded by chronic inflammation that enhances the expression of MMP-9 and -3, while melatonin arrests both of them via reduction of AP-1 activity during protection of ulcer. Ó 2012 Elsevier Masson SAS. All rights reserved. 1. Introduction Gastric inflammation is more prevalent than gastric ulcer which predisposes to both ulceration and in more advanced stages to adenocarcinoma [1]. Apart from the infection with Helicobacter pylori, indiscriminate use of nonsteroidal anti-inflammatory drugs (NSAIDs) are major risk factors for gastritis and gastric ulcers because both types of damages are accompanied by tissue inflammation and secretion of cytokines, including interleukin (IL)- 1b and tumor necrosis factor (TNF)-a [2e4]. Indomethacin causes injury in the gastroduodenal tract through expression of interleu- kins (IL)-1, -6, and -8 and infiltration of polymorphonuclear leukocyte and in turn apoptosis in lesion area of gastric mucosa [2,5,6]. Indomethacin also activates the infiltrated inflammatory cells in lesion areas that generates reactive oxygen species (ROS) i.e., superoxide (O 2  ), which is rapidly converted to hydrogen peroxide (H 2 O 2 ) by superoxide dismutase (SOD) or hydroxyl radical (  OH) by Fenton reaction and the superoxide driven HabereWeiss reaction [7e9]. This ROS can induce lipid peroxidation, protein oxidation and glutathione depletion and, thereby cause tissue injury in gastric mucosa [10,11]. Melatonin, (N-acetyl-5 methoxy- tryptamine), a hormone produced from the pineal gland, retina and enterochromaffin cells of gastrointestinal tract that maintains circadian rhythmicity, seasonal reproduction, sleep and gastroin- testinal homeostasis [15,16]. It also acts as an antioxidant and thus protects against various types of gastric ulceration through Abbreviations: AP, activator protein; b.w., body weight; ECM, extracellular matrix; ERK, extracellular-regulated kinase; i.p., intraperitoneal; IL, interleukin; JNK, c-Jun N-terminal kinase; MMPs, matrix metalloproteinases; NSAIDs, non- steroidal anti-inflammatory drugs; NF-kB, nuclear factor kappa beta; O 2  , super- oxide radical;  OH, hydroxyl radical; H 2 O 2 , hydrogen peroxide; ROS, reactive oxygen species; RT-PCR, reverse transcriptase-PCR; TIMP, tissue inhibitor of metalloproteinase. * Corresponding author. Tel.: þ91 33 2473 3491x159; fax: þ91 33 2473 5197. E-mail address: snehasiktas@hotmail.com (S. Swarnakar). 1 Present address: Laboratory of Molecular Neurobiology, Nencki Institute, Pas- teura 3, 02-093 Warsaw, Poland. Contents lists available at SciVerse ScienceDirect Biochimie journal homepage: www.elsevier.com/locate/biochi 0300-9084/$ e see front matter Ó 2012 Elsevier Masson SAS. All rights reserved. http://dx.doi.org/10.1016/j.biochi.2012.08.004 Biochimie 94 (2012) 2687e2698