Defined as the dysfunction and/or cell death caused by toxic lipids accumulation in hepatocytes, hepatic lipotoxicity plays a pathological role in non-alcoholic fatty liver disease. The cellular and molecular mechanisms underlying lipotoxicity remain to be elucidated. In this study, using AML12 cells, a non-transformed murine hepatocyte cell line, exposed to palmitate (a 16-C saturated fatty acid) as an experimental model, we investigated the role and mechanisms of nicotinamide N-methyltransferase (NNMT), a methyltransferase catalyzing nicotinamide methylation and degradation, in hepatic lipotoxicity. We initially identified activating transcription factor 4 (ATF4) as a major transcription factor for hepatic NNMT expression. Here, we demonstrated that palmitate upregulates NNMT expression via activating ATF4 in a mechanistic target of rapamycin complex 1 (mTORC1)-dependent mechanism in that mTORC1 inhibition by both Torin1 and rapamycin attenuated ATF4 activation and NNMT upregulation. We further demonstrated that the mTORC1-dependent ATF4 activation is an integral signaling event of unfolded protein response (UPR) as both ATF4 activation and NNMT upregulation by tunicamycin, a well-documented endoplasmic reticulum (ER) stress inducer, are blunted when hepatocytes were pretreated with Torin1. Importantly, our data uncovered that NNMT upregulation contributes to palmitate-induced hepatotoxicity as NNMT inhibition, via either pharmacological (NNMT inhibitors) or genetic approach (siRNA transfection), provided protection against palmitate lipotoxicity. Our further mechanistic exploration identified protein kinase A (PKA) activation to contribute, at least, partially to the protective effect of NNMT inhibition against lipotoxicity. Collectively, our data demonstrated that NNMT upregulation by the mTORC1-ATF4 pathway activation contributes to the development of lipotoxicity in hepatocytes.

中文翻译：

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通过 mTORC1-ATF4 通路激活的烟酰胺 N-甲基转移酶 (NNMT) 上调有助于棕榈酸酯诱导的肝细胞脂毒性

肝脏脂毒性定义为肝细胞中有毒脂质积累引起的功能障碍和/或细胞死亡，在非酒精性脂肪肝疾病中起病理作用。脂毒性的细胞和分子机制仍有待阐明。在这项研究中，我们使用 AML12 细胞（一种未转化的鼠肝细胞系，暴露于棕榈酸酯（一种 16-C 饱和脂肪酸）作为实验模型），研究了烟酰胺 N-甲基转移酶 (NNMT) 的作用和机制。甲基转移酶催化烟酰胺甲基化和降解，在肝脂毒性中。我们最初将激活转录因子 4 (ATF4) 确定为肝脏 NNMT 表达的主要转录因子。这里，我们证明棕榈酸酯通过在雷帕霉素复合物 1 (mTORC1) 依赖机制的机械靶标中激活 ATF4 上调 NNMT 表达，因为 Torin1 和雷帕霉素对 mTORC1 的抑制减弱了 ATF4 活化和 NNMT 上调。我们进一步证明了 mTORC1 依赖性 ATF4 激活是未折叠蛋白反应 (UPR) 的一个完整信号事件，因为当肝细胞被预处理时，衣霉素（一种有据可查的内质网 (ER) 应激诱导剂）对 ATF4 激活和 NNMT 上调都减弱了与 Torin1。重要的是，我们的数据揭示了 NNMT 上调有助于棕榈酸酯诱导的肝毒性，因为 NNMT 抑制通过药理学（NNMT 抑制剂）或遗传方法（siRNA 转染）提供了对棕榈酸酯脂毒性的保护。我们进一步的机制探索确定了蛋白激酶 A (PKA) 激活至少部分有助于 NNMT 抑制对脂毒性的保护作用。总的来说，我们的数据表明 mTORC1-ATF4 通路激活引起的 NNMT 上调有助于肝细胞中脂毒性的发展。