Utilizing systems biology to reveal cellular responses to peroxisome proliferator-activated receptor γ ligand exposure.
Utilizing systems biology to reveal cellular responses to peroxisome proliferator-activated receptor γ ligand exposure.
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DOI：
10.1016/j.crtox.2021.03.003
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发表时间：
2021
影响因子：
3.3
通讯作者：
Volz DC
中科院分区：
文献类型：
--
作者：
Cheng V;Reddam A;Bhatia A;Hur M;Kirkwood JS;Volz DC
将人（HepG 2）细胞暴露于PPARγ配体以诱导系统水平效应。格列酮降低HepG 2细胞活力，而GW 9662没有影响。格列酮和GW 9662增加中性脂质作为浓度的函数。胆固醇生物合成转录受环格列酮和GW 9662的影响。阿格列酮改变脂质谱，但GW 9662与溶剂暴露的细胞相似。过氧化物酶体增殖物激活受体γ（Peroxisome Proliferator-Activated Receptor γ，PPARγ）是一种核受体，在被配体激活后，与类维生素A X受体（retinoid X receptor，RXR）异源二聚化，结合到过氧化物酶体增殖物激活受体应答元件（PPRES），并激活下游基因的转录。由于PPARγ在脂肪形成、脂肪酸储存和葡萄糖代谢中起着核心作用，因此PPARγ特异性药物（例如，噻唑烷二酮）已被开发用于治疗人类群体中的II型糖尿病和肥胖症。然而，据我们所知，没有先前的研究同时评估了在生物活性、非细胞毒性浓度下，PPARγ配体暴露对全基因组PPARγ结合的影响以及对人细胞内转录组和脂质组的影响。除了定量环格列酮的浓度依赖性作用外，（一种参照PPARγ激动剂）和GW 9662（一种参考的过氧化物酶体增殖物激活受体γ拮抗剂）对人肝癌（HepG 2）细胞活力、原位过氧化物酶体增殖物激活受体γ丰度和中性脂质的影响，将HepG 2细胞暴露于任一载体（0.1%DMSO）、环格列酮或GW 9662处理长达24小时，然后收获用于1）染色质免疫沉淀-测序（ChIP-seq）以鉴定整个基因组中的PPARγ结合区域，2）mRNA测序（mRNA-seq）以鉴定对转录组的潜在影响，3）脂质组学以鉴定脂质谱的潜在改变。暴露于环格列酮和GW 9662后，我们发现暴露2-8 h后，PPARγ水平无显著差异。虽然环格列酮和GW 9662导致中性脂质浓度依赖性增加，但基因组中PPARγ结合区域的大小和定位（通过ChIP-seq鉴定）不因治疗而变化。然而，mRNA-seq和脂质组学显示，HepG 2细胞暴露于环格列酮和GW 9662导致对转录组和脂质组的显著的治疗特异性作用。总体而言，我们的研究结果表明，人类细胞暴露于具有生物活性的非细胞毒性浓度的PPARγ配体会导致毒性，这可能是由PPARγ依赖性和PPARγ非依赖性机制共同驱动的。
Human (HepG2) cells were exposed to PPARγ ligands to induce systems-level effects. Ciglitazone decreases HepG2 cell viability while GW 9662 had no effect. Ciglitazone and GW 9662 increase neutral lipids as a function of concentration. Cholesterol biosynthesis transcripts are affected by ciglitazone and GW 9662. Ciglitazone alters lipid profiles but GW 9662 was similar to vehicle-exposed cells. Peroxisome proliferator-activated receptor γ (PPARγ) is a nuclear receptor that, upon activation by ligands, heterodimerizes with retinoid X receptor (RXR), binds to PPAR response elements (PPREs), and activates transcription of downstream genes. As PPARγ plays a central role in adipogenesis, fatty acid storage, and glucose metabolism, PPARγ-specific pharmaceuticals (e.g., thiazolidinediones) have been developed to treat Type II diabetes and obesity within human populations. However, to our knowledge, no prior studies have concurrently assessed the effects of PPARγ ligand exposure on genome-wide PPARγ binding as well as effects on the transcriptome and lipidome within human cells at biologically active, non-cytotoxic concentrations. In addition to quantifying concentration-dependent effects of ciglitazone (a reference PPARγ agonist) and GW 9662 (a reference PPARγ antagonist) on human hepatocarcinoma (HepG2) cell viability, PPARγ abundance in situ, and neutral lipids, HepG2 cells were exposed to either vehicle (0.1% DMSO), ciglitazone, or GW 9662 for up to 24 h, and then harvested for 1) chromatin immunoprecipitation-sequencing (ChIP-seq) to identify PPARγ-bound regions across the entire genome, 2) mRNA-sequencing (mRNA-seq) to identify potential impacts on the transcriptome, and 3) lipidomics to identify potential alterations in lipid profiles. Following exposure to ciglitazone and GW 9662, we found that PPARγ levels were not significantly different after 2–8 h of exposure. While ciglitazone and GW 9662 resulted in a concentration-dependent increase in neutral lipids, the magnitude and localization of PPARγ-bound regions across the genome (as identified by ChIP-seq) did not vary by treatment. However, mRNA-seq and lipidomics revealed that exposure of HepG2 cells to ciglitazone and GW 9662 resulted in significant, treatment-specific effects on the transcriptome and lipidome. Overall, our findings suggest that exposure of human cells to PPARγ ligands at biologically active, non-cytotoxic concentrations results in toxicity that may be driven by a combination of both PPARγ-dependent and PPARγ-independent mechanisms.
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