Peroxisome Proliferator-activated Receptors α and γ Are Activated by Indomethacin and Other Non-steroidal Anti-inflammatory Drugs

To gain insight into the function of peroxisome proliferator-activated receptor (PPAR) isoforms in rodents, we disrupted the ligand-binding domain of the alpha isoform of mouse PPAR (mPPAR alpha) by homologous recombination. Mice homozygous for the mutation lack expression of mPPAR alpha protein and yet are viable and fertile and exhibit no detectable gross phenotypic defects. Remarkably, these animals do not display the peroxisome proliferator pleiotropic response when challenged with the classical peroxisome proliferators, clofibrate and Wy-14,643. Following exposure to these chemicals, hepatomegaly, peroxisome proliferation, and transcriptional-activation of target genes were not observed. These results clearly demonstrate that mPPAR alpha is the major isoform required for mediating the pleiotropic response resulting from the actions of peroxisome proliferators. mPPAR alpha-deficient animals should prove useful to further investigate the role of this receptor in hepatocarcinogenesis, fatty acid metabolism, and cell cycle regulation.

To gain insight into the function of peroxisome proliferator-activated receptor (PPAR) isoforms in mammals, we have cloned and characterized two PPAR alpha-related cDNAs (designated PPAR gamma and -delta, respectively) from mouse. The three PPAR isoforms display widely divergent patterns of expression during embryogenesis and in the adult. Surprisingly, PPAR gamma and -delta are not activated by pirinixic acid (Wy 14,643), a potent peroxisome proliferator and activator of PPAR alpha. However, PPAR gamma and -delta are activated by the structurally distinct peroxisome proliferator LY-171883 and linoleic acid, respectively, indicating that each of the isoforms can act as a regulated activator of transcription. These data suggest that tissue-specific responsiveness to peroxisome proliferators, including certain fatty acids, is in part a consequence of differential expression of multiple, pharmacologically distinct PPAR isoforms.

Constitutive cyclooxygenase (COX-1; prostaglandin-endoperoxide synthase, EC 1.14.99.1) is present in cells under physiological conditions, whereas COX-2 is induced by some cytokines, mitogens, and endotoxin presumably in pathological conditions, such as inflammation. Therefore, we have assessed the relative inhibitory effects of some nonsteroidal antiinflammatory drugs on the activities of COX-1 (in bovine aortic endothelial cells) and COX-2 (in endotoxin-activated J774.2 macrophages) in intact cells, broken cells, and purified enzyme preparations (COX-1 in sheep seminal vesicles; COX-2 in sheep placenta). Similar potencies of aspirin, indomethacin, and ibuprofen against the broken cell and purified enzyme preparations indicated no influence of species. Aspirin, indomethacin, and ibuprofen were more potent inhibitors of COX-1 than COX-2 in all models used. The relative potencies of aspirin and indomethacin varied only slightly between models, although the IC50 values were different. Ibuprofen was more potent as an inhibitor of COX-2 in intact cells than in either broken cells or purified enzymes. Sodium salicylate was a weak inhibitor of both COX isoforms in intact cells and was inactive against COX in either broken cells or purified enzyme preparations. Diclofenac, BW 755C, acetaminophen, and naproxen were approximately equipotent inhibitors of COX-1 and COX-2 in intact cells. BF 389, an experimental drug currently being tested in humans, was the most potent and most selective inhibitor of COX-2 in intact cells. Thus, there are clear pharmacological differences between the two enzymes. The use of such models of COX-1 and COX-2 activity will lead to the identification of selective inhibitors of COX-2 with presumably less side effects than present therapies. Some inhibitors had higher activity in intact cells than against purified enzymes, suggesting that pure enzyme preparations may not be predictive of therapeutic action.