Nonsteroidal anti-inflammatory drugs (NSAIDs) are among the most widely used drugs worldwide for the relief of inflammation and pain. However, NSAIDs cause adverse gastrointestinal (GI) side-effects in a large proportion of people who use them, many of whom require hospitalization to recover. Traditionally, NSAID- induced GI toxicity was associated with the inhibition of cyclooxygenase (COX) enzymes, particularly COX-1. Recent evidence however, indicates that COX inhibition does not account for the full range of toxic effects observed in the GI tract. Additionally, the discovery of the adverse cardiovascular effects of COX-2 specific NSAIDs has demonstrated the importance of identifying mechanisms through which NSAIDs cause GI toxicity. Accordingly, we propose to investigate the molecular mechanisms underlying inhibition of restitution by NSAIDs, one of the negative effects that NSAIDs have on GI epithelia. GI epithelia form a barrier that separates the internal milieu from the harsh conditions of the lumen of the GI. Restitution is the cell proliferation-free process by which wounds or gaps in the epithelial barrier are repaired by the spreading and migration of neighboring cells into the wound. NSAIDs with ulcerogenic potential inhibit cell migration in intestinal epithelial cells. Our research group has shown that this inhibition is associated with a decrease in the expression and activity of calpain proteases and depolarization of membrane potential (Em). On the basis of these data, we hypothesize that NSAIDs impair intestinal epithelial cell migration and inhibit calpain activity through a signaling pathway that is highly sensitive to drug-induced changes in Em. Therefore, in Specific Aim 1 we will determine if NSAIDs impair de-adhesion of cell contacts by measuring the number and distribution of cellular adhesions as well as the expression and rate of turnover of specific adhesion components.
In Specific Aim 2, we will determine if NSAIDs or Em depolarization modulate calpain activity through disruption of EGF-stimulated phosphorylation of calpains or disruption of PIP balance by measuring expression and activation of the EGF pathway, PIP levels, and calpain activity and localization to the membrane.
In Specific Aim 3, we will extend our in vitro experiments by determining the effects of NSAIDs on cell migration, calpain and Kv channel expression, and calpain participation in membrane localized complexes in rats treated with NSAIDs. These innovative experiments have the potential to identify molecular mechanisms through which NSAIDs decrease calpain activity, in addition to determining the effects of reductions in calpain activity on cellular de-adhesion during migration both in vitro and in vivo. This project will provide basic information on wound healing processes in intestinal cells that are applicable to other tissues, document the effects of NSAIDs on these processes, and give insight into how NSAIDs may affect the healing process in other tissues. The research also has great potential for aiding in the development of new drugs in this class capable of ameliorating pain without the adverse side effects of current drugs.

Public Health Relevance

The widespread use of nonsteroidal anti-inflammatory drugs (NSAIDs) for the alleviation of inflammation and pain poses a major medical risk to patients due to the common toxic effects these drugs have on the gastrointestinal tract. Our research is designed to elucidate novel mechanisms through which these toxic effects occur in order to eliminate the side effects associated with NSAID use and enhance the safety of future drugs of this class. This research may help reduce or eliminate the loss of billions of dollars per year due to medical complications resulting from NSAID-induced GI toxicity.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Academic Research Enhancement Awards (AREA) (R15)
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Gastrointestinal Mucosal Pathobiology Study Section (GMPB)
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Hamilton, Frank A
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Kansas State University
Other Clinical Sciences
Schools of Veterinary Medicine
United States
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Silver, Kristopher; Littlejohn, A; Thomas, Laurel et al. (2017) Suppression of calpain expression by NSAIDs is associated with inhibition of cell migration in rat duodenum. Toxicology 383:1-12
Silver, Kristopher; Littlejohn, Alaina; Thomas, Laurel et al. (2015) Inhibition of Kv channel expression by NSAIDs depolarizes membrane potential and inhibits cell migration by disrupting calpain signaling. Biochem Pharmacol 98:614-28