QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) Data Supplement All Versions of this Article: jpet.107.127720v1 325/2/389    most recent Submit a response Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Raveendran, N. N. Articles by Lillich, J. D. Search for Related Content PubMed PubMed Citation Articles by Raveendran, N. N. Articles by Lillich, J. D.

GASTROINTESTINAL, HEPATIC, PULMONARY, AND RENAL

Drug-Induced Alterations to Gene and Protein Expression in Intestinal Epithelial Cell 6 Cells Suggest a Role for Calpains in the Gastrointestinal Toxicity of Nonsteroidal Anti-Inflammatory Agents

Departments of Anatomy and Physiology (N.N.R., L.C.F., S.G.) and Clinical Sciences (K.S., D.N., K.W., J.D.L.), College of Veterinary Medicine, Kansas State University, Manhattan, Kansas

Nonsteroidal anti-inflammatory drugs (NSAIDs) are used extensively as therapeutic agents, despite their well documented gastrointestinal (GI) toxicity. At this time, the mechanisms responsible for NSAID-associated GI damage are incompletely understood. In this study, we used microarray analysis to generate a novel hypothesis about cellular mechanisms that underlie the GI toxicity of NSAIDs. Monolayers of intestinal epithelial cells (IEC-6) were treated with NSAIDs that either exhibit (indomethacin, NS-398 [N-[2-(cyclohexyloxy)-4-nitrophenyl]-methanesulfonamide]) or lack (SC-560 [5-(4-chlorphenyl)-1-(4-methoxyphenyl)-3-(trifluoromethyl)-1H-pyrazole]) inhibitory effects on IEC-6 migration. Bioinformatic analysis of array data identified the calpain cysteine proteases and their endogenous inhibitor calpastatin as potential targets of NSAIDs shown previously to retard IEC-6 migration. Accordingly, quantitative real-time reverse transcription polymerase chain reaction and immunoblotting were performed to assess the effects of NSAIDs on the expression of mRNA and protein for calpain 8, calpain 2, calpain 1, and calpastatin. In treated IEC-6 monolayers, NS-398 decreased the expression of mRNA for calpain 2 and calpain 8. Both NS-398 and indomethacin decreased the protein expression of calpains 8, 2, and 1. None of the NSAIDs affected expression of calpastatin mRNA or protein. The calpain inhibitors, N-acetyl-Leu-Leu-methioninal and N-acetyl-Leu-Leu-Nle-CHO, retarded IEC-6 cell migration in a concentration-dependant fashion, and these inhibitory effects were additive with those of indomethacin and NS-398. Our experimental results suggest that the altered expression of calpain proteins may contribute to the adverse effects of NSAIDs on intestinal epithelial restitution.

Address correspondence to: Dr. James D. Lillich, Department of Clinical Sciences, VCS-Q-203 Moiser Hall, College of Veterinary Medicine, Kansas State University, Manhattan, KS 66506. E-mail: lillich{at}vet.ksu.edu