高胰岛素血症是代谢组织，包括肝脏，胰岛素抵抗（IR）的发病关键风险因素。最近发现的代谢酶乙醇胺磷酸磷酸酯酶（ETNPPL）在肝脏组织中丰富表达，能将磷酸乙醇胺（PEA）转化为氨、无机磷酸盐和乙醛，但它是否在调节高胰岛素引起的肝细胞IR中起作用仍不明确。我们在人肝癌细胞系HepG2和小鼠肝细胞中建立了一个高胰岛素血症诱导的IR体外模型，通过高剂量胰岛素处理。然后，我们利用载体介导的过量表达来治疗ETNPPL以研究其对IR的影响而无需胰岛素刺激。为了探索ETNPPL介导高胰岛素血症引起的IR在HepG2中的潜在机制，我们使用RNA测序进行基因组范围的转录分析以识别ETNPPL的下游靶基因。结果表明，在HepG2和小鼠肝细胞的高胰岛素血症诱导下，ETNPPL的mRNA和蛋白质表达水平均显著上调。沉默ETNPPL后，高胰岛素血症诱导的IR得到了改善。在没有IR的正常情况下，过表达ETNPPL会促进IR、活性氧（ROS）产生和AKT失活。转录组分析表明，盐诱导激酶1（SIK1）在ETNPPL敲低的HepG2细胞中明显下调。此外，破坏SIK1可防止ETNPPL引起的ROS积累、损坏PI3K/AKT通路和IR。我们的研究揭示了ETNPPL通过SIK1/ROS介导的PI3K/AKT信号通路失活调节肝细胞中的高胰岛素血症引起的IR。针对ETNPPL可能是高胰岛素血症相关代谢紊乱，如2型糖尿病的潜在策略。©2023 Wiley Periodicals LLC。

Hyperinsulinemia is a critical risk factor for the pathogenesis of insulin resistance (IR) in metabolic tissues, including the liver. Ethanolamine phosphate phospholyase (ETNPPL), a newly discovered metabolic enzyme that converts phosphoethanolamine (PEA) to ammonia, inorganic phosphate, and acetaldehyde, is abundantly expressed in liver tissue. Whether it plays a role in the regulation of hyperinsulinemia-induced IR in hepatocytes remains elusive. Here, we established an in vitro hyperinsulinemia-induced IR model in the HepG2 human liver cancer cell line and primary mouse hepatocyte via a high dose of insulin treatment. Next, we overexpressed ETNPPL by using lentivirus-mediated ectopic to investigate the effects of ETNPPL per se on IR without insulin stimulation. To explore the underlying mechanism of ETNPPL mediating hyperinsulinemia-induced IR in HepG2, we performed genome-wide transcriptional analysis using RNA sequencing (RNA-seq) to identify the downstream target gene of ETNPPL. The results showed that ETNPPL expression levels in both mRNA and protein were significantly upregulated in hyperinsulinemia-induced IR in HepG2 and primary mouse hepatocytes. Upon silencing ETNPPL, hyperinsulinemia-induced IR was ameliorated. Under normal conditions without IR in hepatocytes, overexpressing ETNPPL promotes IR, reactive oxygen species (ROS) generation, and AKT inactivation. Transcriptome analysis revealed that salt-inducible kinase 1 (SIK1) is markedly downregulated in the ETNPPL knockdown HepG2 cells. Moreover, disrupting SIK1 prevents ETNPPL-induced ROS accumulation, damage to the PI3K/AKT pathway and IR. Our study reveals that ETNPPL mediates hyperinsulinemia-induced IR through the SIK1/ROS-mediated inactivation of the PI3K/AKT signaling pathway in hepatocyte cells. Targeting ETNPPL may present a potential strategy for hyperinsulinemia-associated metabolic disorders such as type 2 diabetes.© 2023 Wiley Periodicals LLC.