title: PPARγ as a sensor of lipase activity and a target for the lipase inhibitor orlistat.
abstract: A PPARγ fluorescence polarization (FP) assay was used to measure the release of fatty acid products from triglyceride emulsions during digestion with pancreatic and yeast lipases in a real-time, homogenous assay. Using the same FP assay we show the anti-obesity drug Orlistat is a PPARγ ligand with an IC50 of 2.84 ± 0.16 μM. Analytical Mass Spectrometry confirms that Orlistat does not bind covalently to PPARγ. The PPARγ FP assay is shown to be a simple method for measuring real-time lipase activity using a number of triglyceride substrates including olive oil and grape seed oil emulsions. Incubation of Orlistat with the human intestinal epithelial cell line Caco-2, at concentrations of 1 - 100 μM, leads to induction of genes regulated by PPARγ. At 100 μM Orlistat, transcription of β-defensin 1 (hDB1) & Adipose Differentiation Related Protein (ADRP) increase by up to 2.6 fold and 6.8 fold, respectively. Although at 1 μM and 100 μM Orlistat did not significantly increase defensin protein synthesis, at 10 μM Orlistat induced a 1.5 fold increase in hDB1 protein secretion in the human colonic adenocarcinoma cell line HT-29. Thus Orlistat is similar to the anti-diabetic drug Rosiglitazone in its ability to induce defensin gene expression. The antimicrobial peptide β-defensin 1 protects against pathogenic micro-organisms in the gut and PPARγ suppresses inflammatory gene expression. These may be beneficial side effects of Orlistat consumption on gut epithelial cells.
authors.forename: Harry
authors.initials: H
authors.lastname: Martin
authors.forename: Tony K
authors.initials: TK
authors.lastname: McGhie
authors.forename: Kerry
authors.initials: K
authors.lastname: Bentley-Hewitt
authors.forename: John
authors.initials: J
authors.lastname: Christeller
mesh_terms: D019440:Anti-Obesity Agents / Q000494:pharmacology; D018938:Caco-2 Cells / Q000187:drug effects / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005454:Fluorescence Polarization / Q000379:methods; D005786:Gene Expression Regulation / Q000187:drug effects; D019073:HT29 Cells / Q000187:drug effects; D006801:Humans; D007783:Lactones / Q000494:pharmacology*; D008049:Lipase / Q000032:analysis* / Q000037:antagonists & inhibitors / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics; D000077403:Orlistat; D047495:PPAR gamma / Q000235:genetics / Q000378:metabolism*; D000070780:Perilipin-2; D023083:beta-Defensins / Q000235:genetics
chemical_list: D019440:Anti-Obesity Agents; C406407:DEFB1 protein, human; D004791:Enzyme Inhibitors; D007783:Lactones; D008565:Membrane Proteins; C000602982:PLIN2 protein, human; D047495:PPAR gamma; D000070780:Perilipin-2; D023083:beta-Defensins; D000077403:Orlistat; D008049:Lipase