PT  - JOURNAL ARTICLE
AU  - Martin Schubert
AU  - Stefanie Becher
AU  - Maria Wallert
AU  - Marten B. Maeβ
AU  - Masoumeh Abhari
AU  - Knut Rennert
AU  - Alexander S. Mosig
AU  - Silke Groβe
AU  - Regine Heller
AU  - Michael Grün
AU  - Stefan Lorkowski
TI  - The PPARγ antagonist 2-chloro-5-nitro-N-phenylbenzamide (GW9662) triggers PLIN2 expression via PPARδ and induces lipogenesis and triglyceride accumulation in human THP-1 macrophages
AID  - 10.1124/mol.119.117887
DP  - 2019 Jan 01
TA  - Molecular Pharmacology
PG  - mol.119.117887
4099  - http://molpharm.aspetjournals.org/content/early/2019/12/23/mol.119.117887.short
4100  - http://molpharm.aspetjournals.org/content/early/2019/12/23/mol.119.117887.full
AB  - Peroxisome proliferator-activated receptors (PPARs) are members of the nuclear hormone receptor family playing pivotal roles in regulating glucose and lipid metabolism as well as inflammation. While characterizing potential PPARγ ligand activity of natural compounds in macrophages we investigated their influence on the expression of adipophilin (perilipin 2, PLIN2), a well-known PPARγ target. To confirm that a compound regulates PLIN2 expression via PPARγ we performed experiments using the widely used PPARγ antagonist 2-chloro-5-nitro-N-phenylbenzamide (GW9662). Surprisingly, instead of blocking up-regulation of PLIN2 expression in THP-1 macrophages, expression was concentration-dependently induced by GW9662 at concentrations and under conditions commonly used. We found that this unexpected up-regulation occurs in many human and murine macrophage cell models and also primary cells. Profiling expression of PPAR target genes showed up-regulation of several genes involved in lipid uptake, transport and storage, as well as fatty acid synthesis by GW9662. In line with this and with up-regulation of PLIN2 protein, GW9662 elevated lipogenesis and increased triglyceride levels. Finally, we identified PPARδ as a mediator of the substantial unexpected effects of GW9662. Our findings show that (i) the PPARγ antagonist GW9662 unexpectedly activates PPARδ-mediated signaling in macrophages, (ii) GW9662 significantly affects lipid metabolism in macrophages, (iii) careful validation of experimental conditions and results is required for experiments involving GW9662, and (iv) published studies in a context comparable to this work may have reported erroneous results if PPARγ independence was demonstrated using GW9662 only. In light of our findings, certain existing studies might require reinterpretation regarding the role of PPARγ.SIGNIFICANCE STATEMENT Peroxisome proliferator-activated receptors (PPARs) are targets for the treatment of various diseases as they are key regulators of inflammation as well as lipid and glucose metabolism. Hence, reliable tools to characterize the molecular effects of PPARs are indispensable. We describe profound and unexpected off-target effects of the PPARγ antagonist 2-chloro-5-nitro-N-phenylbenzamide (GW9662) involving PPARδ and in turn affecting macrophage lipid metabolism. Our results question certain existing studies using GW9662 and make better experimental design of future studies necessary.