mRNA profiling reveals divergent roles of PPARa and PPARß/d in regulating mouse liver gene expression (PPARa samples)

Little is known about the role of the transcription factor PPARß/d in liver. Here we set out to better elucidate the function of PPARß/d in liver by comparing the effect of PPARa and PPARß/d deletion using whole genome transcriptional profiling and analysis of plasma and liver metabolites. In fed state, the number of genes altered by PPARa and PPARß/d deletion was similar, whereas in fasted state the effect of PPARa deletion was much more pronounced, consistent with the pattern of gene expression of PPARa and PPARß/d. Minor overlap was found between PPARa- and PPARß/d-dependent gene regulation in liver. Pathways upregulated by PPARß/d deletion were connected to innate immunity. Pathways downregulated by PPARß/d deletion included lipoprotein metabolism and various pathways related to glucose utilization, which correlated with elevated plasma glucose and triglycerides and reduced plasma cholesterol in PPARß/d-/- mice. Downregulated genes that may underlie these metabolic alterations included Pklr, Fbp1, Apoa4, Vldlr, Lipg, and Pcsk9, which may represent novel PPARß/d target genes. In contrast to PPARa-/- mice, no changes in plasma FFA, plasma ß-hydroxybutyrate, liver triglycerides and liver glycogen were observed in PPARß/d-/- mice. Our data indicate a role for PPARß/d in hepatic glucose utilization and lipoprotein metabolism but not in the adaptive response to fasting. Keywords: Analysis of target gene regulation by using microarrays Pure-bred Sv129 PPARα -/- mice and corresponding wildtype mice were used. Male mice (n=4-5 per group) were either fed or fasted for 24 hours. At the end of the experiment, mice were anaesthetized with a mixture of isofluorane (1.5%), nitrous oxide (70%) and oxygen (30%). Blood was collected by orbital puncture, after which the mice were sacrificed by cervical dislocation. Livers were dissected, snap frozen in liquid nitrogen and kept at -80ºC until further analysis. For RNA analyses, tissue from the same part of the liver lobe was used.

目前学界对过氧化物酶体增殖物激活受体β/δ（PPARβ/δ）在肝脏中的作用所知甚少。本研究旨在通过全基因组转录谱分析，结合血浆与肝脏代谢物检测，对比敲除PPARα与PPARβ/δ所产生的效应，以更清晰地阐明PPARβ/δ在肝脏中的功能。在进食状态下，敲除PPARα与PPARβ/δ所改变的基因数量相近；而在禁食状态下，敲除PPARα的效应更为显著，这与PPARα和PPARβ/δ的基因表达模式相符。肝脏中受PPARα与PPARβ/δ依赖性调控的基因存在较少的重叠。PPARβ/δ敲除后上调的通路与固有免疫相关。PPARβ/δ敲除后下调的通路包括脂蛋白代谢以及多种与葡萄糖利用相关的通路，这与PPARβ/δ基因敲除（PPARβ/δ⁻/⁻）小鼠的血浆葡萄糖、甘油三酯水平升高，血浆胆固醇水平降低的表型相一致。可能介导上述代谢改变的下调基因包括Pklr、Fbp1、Apoa4、Vldlr、Lipg及Pcsk9，这些基因或为新型PPARβ/δ靶基因。与PPARα基因敲除（PPARα⁻/⁻）小鼠不同，PPARβ/δ⁻/⁻小鼠的血浆游离脂肪酸（FFA）、血浆β-羟丁酸、肝脏甘油三酯及肝糖原水平均无显著变化。本研究数据表明，PPARβ/δ在肝脏葡萄糖利用与脂蛋白代谢中发挥作用，但不参与机体对禁食的适应性应答。关键词：基于微阵列的靶基因调控分析。实验选用纯种Sv129背景的PPARα⁻/⁻小鼠及其同窝野生型小鼠。每组选取4~5只雄性小鼠，分别设置正常进食组与禁食24小时组。实验结束时，使用异氟烷（1.5%）、一氧化二氮（70%）与氧气（30%）的混合气体对小鼠进行麻醉。通过眼眶穿刺采集血液，随后采用颈椎脱臼法处死小鼠。摘取肝脏组织，经液氮快速冷冻后保存于-80℃冰箱，待后续分析使用。进行RNA分析时，取用肝叶同一部位的组织样本。