title: Establishment of a cell-based drug screening model for identifying agonists of human peroxisome proliferator-activated receptor gamma (PPARγ).
abstract: OBJECTIVE
  Peroxisome proliferator-activated receptor gamma (PPARγ) plays a critical role in regulation of diverse biological processes, including lipid metabolism and adipogenesis, cell division and apoptosis, and is involved in variety of disease conditions, such as obesity, atherosclerosis, inflammation and tumour. Developing a cell-based reporter gene model targeting PPARγ would be useful to screen human PPARγ agonists that could be beneficial to patients with these diseases.
  
  
  METHODS
  We stably co-transfected human embryonic kidney (HEK) cell line 293T cells with phPPARγ-IRES2-EGFP vector to express human PPARγ (hPPARγ), a reporter vector pPPRE×3-TK-LUC, and control vector pRL-CMV. The efficiency of the co-transfection was evaluated with flow cytometry of hPPARγ expressing cells. Specificity of hPPARγ activity was determined by dual luciferase reporter assay of co-transfected cells exposed to PPARγ agonist rosiglitazone, PPARα agonist WY14643 and retinoic acid receptor alpha (RARα) agonist all-trans-retinoic acid (ATRA).
  
  
  RESULTS
  The phPPARγ-IRES2-EGFP co-transfected HEK293T cells showed concentration- and time-dependent luciferase induction upon exposure to the rosiglitazone, while WY14643 and ATRA were unable to activate the co-transfected HEK293T cells.
  
  
  CONCLUSIONS
  These data indicated that the HEK293T cells could be stably transfected with hPPARγ. This cell-based drug screening platform could be used targeting specific nuclear receptor of hPPARγ with effectiveness and specificity for hPPARγ agonists discovery.
authors.affiliation: Key Laboratory of Cell Engineering of Guizhou Province, the Affiliated Hospital of Zunyi Medical College, China.
authors.forename: Jing-Jing
authors.initials: JJ
authors.lastname: Ma
authors.forename: Tao
authors.initials: T
authors.lastname: Zhang
authors.forename: Ning
authors.initials: N
authors.lastname: Fang
authors.forename: Yan
authors.initials: Y
authors.lastname: Zou
authors.forename: Qi-Hai
authors.initials: QH
authors.lastname: Gong
authors.forename: Li-Mei
authors.initials: LM
authors.lastname: Yu
authors.forename: Dai-Xiong
authors.initials: DX
authors.lastname: Chen
mesh_terms: D004353:Drug Evaluation, Preclinical / Q000379:methods*; D005434:Flow Cytometry; D017930:Genes, Reporter; D005822:Genetic Vectors; D057809:HEK293 Cells; D006801:Humans; D008156:Luciferases / Q000378:metabolism; D008954:Models, Biological; D047495:PPAR gamma / Q000819:agonists* / Q000235:genetics; D018168:Receptors, Retinoic Acid / Q000819:agonists; D000072482:Retinoic Acid Receptor alpha; D000077154:Rosiglitazone; D045162:Thiazolidinediones / Q000494:pharmacology; D014162:Transfection / Q000379:methods*; D014212:Tretinoin / Q000494:pharmacology
chemical_list: D047495:PPAR gamma; C000607071:RARA protein, human; D018168:Receptors, Retinoic Acid; D000072482:Retinoic Acid Receptor alpha; D045162:Thiazolidinediones; D000077154:Rosiglitazone; D014212:Tretinoin; D008156:Luciferases