The Peroxisome Proliferator-Activated Receptor (PPAR)-γ Antagonist 2-Chloro-5-Nitro-N-Phenylbenzamide (GW9662) Triggers Perilipin 2 Expression via PPARδ and Induces Lipogenesis and Triglyceride Accumulation in Human THP-1 Macrophages
The Peroxisome Proliferator-Activated Receptor (PPAR)-γ Antagonist 2-Chloro-5-Nitro-N-Phenylbenzamide (GW9662) Triggers Perilipin 2 Expression via PPARδ and Induces Lipogenesis and Triglyceride Accumulation in Human THP-1 Macrophages
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DOI：
10.1124/mol.119.117887
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发表时间：
2020-03-01
影响因子：
3.6
通讯作者：
Lorkowski, Stefan
中科院分区：
文献类型：
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作者：
Schubert, Martin;Becher, Stefanie;Lorkowski, Stefan
过氧化物酶体增殖物激活受体(PPAR)是核激素受体家族的成员，在调节糖脂代谢和炎症等方面发挥着重要作用。在表征天然化合物在巨噬细胞中潜在的PPARγ配体活性的同时，我们研究了它们对脂联素[Perilipin 2(PLIN2)]--一个众所周知的PPAR伽马靶标--表达的影响。为了证实一种化合物通过PPAR伽马调节PLIN2的表达，我们使用广泛使用的PPAR伽马拮抗剂2-氯-5-硝基-N-苯基苯甲酰胺(GW9662)进行了实验。令人惊讶的是，GW9662没有阻止THP-1巨噬细胞PLIN2表达的上调，而是在常用的浓度和条件下以浓度依赖的方式诱导表达。我们发现，这种意想不到的上调发生在许多人和小鼠巨噬细胞模型中，也发生在原代细胞中。PPAR靶基因的表达谱显示，GW9662上调了几个与脂质吸收、运输和储存以及脂肪酸合成有关的基因。与此相一致的是，随着PLIN2蛋白的上调，GW9662提高了脂肪生成和甘油三酯水平。最后，我们确定PPAR Delta是GW9662重大意想不到的影响的中介。我们的发现表明：1)PPAR伽马拮抗剂GW9662出人意料地激活了巨噬细胞中PPAR Delta介导的信号转导，2)GW9662显著影响巨噬细胞的脂代谢，3)涉及GW9662的实验需要仔细验证实验条件和结果，4)如果仅使用GW9662来证明PPAR伽马独立性，那么在与本工作类似的背景下发表的研究可能报告了错误的结果。鉴于我们的发现，某些现有的研究可能需要重新解释PPAR伽玛的作用。信号统计PPAR是各种疾病的治疗靶点，因为它们是炎症以及脂和糖代谢的关键调节因子。因此，表征PPAR分子效应的可靠工具是必不可少的。我们描述了PPAR-γ拮抗剂2-氯-5-硝基-N-苯基苯甲酰胺(GW9662)涉及PPAR增量并进而影响巨噬细胞脂代谢的深刻和意想不到的非靶点效应。我们的结果质疑了使用GW9662进行的某些现有研究，并为未来的研究提供了更好的实验设计。
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 PPAR gamma ligand activity of natural compounds in macrophages, we investigated their influence on the expression of adipophilin [perilipin 2 (PLIN2)], a well-known PPAR gamma target. To confirm that a compound regulates PLIN2 expression via PPAR gamma, we performed experiments using the widely used PPAR gamma antagonist 2-chloro-5-nitro-N-phenylbenzannide (GW9662). Surprisingly, instead of blocking upregulation 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 upregulation occurs in many human and murine macrophage cell models and also primary cells. Profiling expression of PPAR target genes showed upregulation of several genes involved in lipid uptake, transport, and storage as well as fatty acid synthesis by GW9662. In line with this and with upregulation of PLIN2 protein, GW9662 elevated lipogenesis and increased triglyceride levels. Finally, we identified PPAR delta as a mediator of the substantial unexpected effects of GW9662. Our findings show that: 1) the PPAR gamma antagonist GW9662 unexpectedly activates PPAR delta-mediated signaling in macrophages, 2) GW9662 significantly affects lipid metabolism in macrophages, 3) careful validation of experimental conditions and results is required for experiments involving GW9662, and 4) published studies in a context comparable to this work may have reported erroneous results if PPAR gamma independence was demonstrated using GW9662 only. In light of our findings, certain existing studies might require reinterpretation regarding the role of PPAR gamma.SIGNIFICANCE STATEMENTPeroxisome 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 PPAR-gamma antagonist 2-chloro-5-nitro-N-phenylbenzannide (GW9662) involving PPAR delta and in turn affecting macrophage lipid metabolism. Our results question certain existing studies using GW9662 and make better experimental design of future studies necessary.