title: Hepatic gene expression response to acute indomethacin exposure.
abstract: BACKGROUND
  Rising morbidity and mortality related to the use of NSAIDs has led to the withdrawal of some of these agents and reconsideration of the adverse effects and usage paradigms of commonly available NSAIDs. Our objective in this study was to assay molecular indicators of acute hepatic injury associated with the administration of indomethacin, a prototypical NSAID, metabolized by the liver that undergoes enterohepatic circulation with associated gastrointestinal adverse effects.
  
  
  METHODS
  Analysis of gene expression, using high-throughput, ADME (absorption, distribution, metabolism, excretion)-specific microarrays, was performed on RNA extracted from the livers of control or indomethacin treated rats, in parallel with serum enzyme tests and histological analysis of paraffin-embedded liver specimens. Male Sprague-Dawley rats (n = 45) were administered intraperitoneal injections of indomethacin for 3 days at the recommended normal dose (6.7 mg/kg), indomethacin at a high dose (20 mg/kg) or vehicle alone (controls).
  
  
  RESULTS
  Upon termination of the study on day 4, serum gamma-glutamyl transferase activity and alkaline phosphatase/alanine aminotransferase ratios were significantly elevated in both high- and normal-dose cohorts compared with vehicle-treated animals. Diffuse microvascular steatosis was present in hepatic serial sections obtained from all animals subjected to the high-dosage regimen. High-resolution microarray analysis (six replicates/gene/animal) identified 256 genes, after outlier removal, in 17 functional classifications that were significantly altered by the high, but not by the normal dosage. These included depression of 10 of 11 cytochrome P450 genes (2B3, 2C70, 1A2-P2, 4F1, 2E1, 3A1, 2F1, 3AP7, 2C11, phenobarb-inducible P6) and 7 of 9 genes involved in the response to reactive oxygen species (e.g. glutathione reductase, glutathione transferase, and superoxide dismutase). Of 16 genes associated with toxin removal, nine exhibited significantly decreased transcripts. There was a marked shift away from lipid metabolism (decreased expression of eight genes) towards glucose utilization associated with steatosis. Despite the compromise of detoxification programs and a shift in metabolic substrate utilization, a compensatory remodeling response was activated, including genes for metalloproteases (ADAM10, MMP10, MMP11), integrins (integrin alpha-1 and alpha-E1), and extracellular matrix molecules (platelet/endothelial cell adhesion molecule-1 and heparan sulfate proteoglycan, perlecan), as well as transcripts associated with cell proliferation. The expression levels of only five genes were significantly altered among animals receiving the normal indomethacin dosage.
  
  
  CONCLUSIONS
  These data confirmed that even brief exposure to indomethacin altered serum enzymatic activities and that high levels significantly altered gene expression in the liver and hepatic histology (by interfering with the clearance of toxins and xenobiotic substrates) and the regulation of basal metabolism.
authors.affiliation: Department of Pathology, University of Pittsburgh School of Medicine, Shadyside Hospital, Pittsburgh, Pennsylvania 15232, USA. laframboisewa@wpmc.edu
authors.forename: William A
authors.initials: WA
authors.lastname: LaFramboise
authors.forename: Kelly L
authors.initials: KL
authors.lastname: Bombach
authors.forename: Andrew R
authors.initials: AR
authors.lastname: Pogozelski
authors.forename: Ryan F
authors.initials: RF
authors.lastname: Cullen
authors.forename: Noah
authors.initials: N
authors.lastname: Muha
authors.forename: James
authors.initials: J
authors.lastname: Lyons-Weiler
authors.forename: Scott J
authors.initials: SJ
authors.lastname: Spear
authors.forename: Rajiv J
authors.initials: RJ
authors.lastname: Dhir
authors.forename: Robert D
authors.initials: RD
authors.lastname: Guthrie
authors.forename: James A
authors.initials: JA
authors.lastname: Magovern
mesh_terms: D000818:Animals; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000008:administration & dosage / Q000633:toxicity*; D056486:Chemical and Drug Induced Liver Injury / Q000209:etiology / Q000378:metabolism; D003577:Cytochrome P-450 Enzyme System / Q000097:blood / Q000378:metabolism; D020869:Gene Expression Profiling; D007213:Indomethacin / Q000008:administration & dosage / Q000633:toxicity*; D008099:Liver / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D008297:Male; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D000894:Anti-Inflammatory Agents, Non-Steroidal; D012333:RNA, Messenger; D003577:Cytochrome P-450 Enzyme System; D007213:Indomethacin