Department of Endocrinology, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016, China.
Department of Endocrinology, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016, China; Laboratory of Lipids and Glucose Metabolism, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016, China.
Life Sci. 2015 Jun 15;131:23-9. doi: 10.1016/j.lfs.2015.03.003. Epub 2015 Mar 25.
It is well known that lipid accumulation and inflammation are two important steps in pathogenesis and progress of nonalcoholic fatty liver disease (NAFLD). However, fewer studies have explored the direct relationship between lipid accumulation and inflammation in early NAFLD. Tumor necrosis factor alpha (TNF-α) is one of the classical inflammatory cytokines. AMP-activated protein kinase (AMPK) is known as a critical regulator of energy homeostasis in metabolic processes. This study aims to investigate the role of TNF-α on lipid deposition of HepG2 cells and examine the modification of AMPK pathway.
TNF-α was added in HepG2 cells and lipid accumulation was analyzed by Oil Red O staining and quantitative test of triglyceride (TG). The expressions of phosphorylated AMPK and its pathway (including mTOR and SREBP-1) were determined. Furthermore, an AMPK agonist (metformin or AICAR) or antagonist (compound C) was co-administrated with TNF-α in HepG2 cells to investigate its effect on TNF-α induced lipid deposition.
A significant increment of TG content in HepG2 cells was observed after TNF-α treatment. Meanwhile, substantially suppressed AMPK and ACC phosphorylation, enhanced mTOR and p70S6K phosphorylation, and increased protein expression of FAS and SREBP-1 were found. Co-treatment with metformin or AICAR decreased the TNF-α-induced intracellular TG, accompanied by significantly enhanced AMPK and ACC phosphorylation, suppressed mTOR and p70S6K phosphorylation, and reduced SREBP-1 and FAS expressions. On the contrary, while co-incubated with compound C, AMPK and ACC phosphorylation were suppressed and the inhibitory effect of metformin on HepG2 cell lipid deposition was also attenuated.
Our results suggest that TNF-α directly induces lipid accumulation in HepG2 cells, at least in part, through the inhibition of AMPK/mTOR/SREBP-1 pathway.
众所周知，脂质堆积和炎症是非酒精性脂肪性肝病（NAFLD）发病和进展的两个重要步骤。然而，较少的研究探讨了早期 NAFLD 中脂质堆积和炎症之间的直接关系。肿瘤坏死因子-α（TNF-α）是经典的炎症细胞因子之一。AMP 激活的蛋白激酶（AMPK）被认为是代谢过程中能量平衡的关键调节剂。本研究旨在探讨 TNF-α对 HepG2 细胞脂质沉积的作用，并研究 AMPK 通路的修饰。
在 HepG2 细胞中加入 TNF-α，通过油红 O 染色和甘油三酯（TG）定量检测分析脂质堆积。测定磷酸化 AMPK 及其通路（包括 mTOR 和 SREBP-1）的表达。此外，在 HepG2 细胞中共同给予 AMPK 激动剂（二甲双胍或 AICAR）或拮抗剂（化合物 C）与 TNF-α共处理，以研究其对 TNF-α诱导的脂质沉积的影响。
TNF-α处理后 HepG2 细胞中的 TG 含量显著增加。同时，发现 AMPK 和 ACC 磷酸化显著受到抑制，mTOR 和 p70S6K 磷酸化增强，FAS 和 SREBP-1 蛋白表达增加。与二甲双胍或 AICAR 共同处理可降低 TNF-α诱导的细胞内 TG，同时显著增强 AMPK 和 ACC 磷酸化，抑制 mTOR 和 p70S6K 磷酸化，减少 SREBP-1 和 FAS 的表达。相反，当与化合物 C 共同孵育时，AMPK 和 ACC 磷酸化受到抑制，二甲双胍对 HepG2 细胞脂质沉积的抑制作用也减弱。
我们的结果表明，TNF-α通过抑制 AMPK/mTOR/SREBP-1 通路直接诱导 HepG2 细胞脂质堆积，至少部分是这样。
