Wen Yongqiang, Wang Yazhou, Loor Juan J, Zhao Chenxu, Wang Jianguo
College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi 712100, China.
Department of Animal Sciences, Division of Nutritional Sciences, University of Illinois, Urbana 61801, USA.
J Steroid Biochem Mol Biol. 2025 Oct;253:106813. doi: 10.1016/j.jsbmb.2025.106813. Epub 2025 Jun 13.
Excessive non-esterified fatty acids (NEFA) in the blood often lead to ketosis and fatty liver in peripartal cows. The integrity of mitochondrial structure and function plays a key role in the development of lipid metabolism imbalance and inflammation. However, the precise molecular mechanisms of NEFA's impact on these processes remain unclear. To explore how NEFA (0, 0.3, 0.6, 1.2, and 2.4 mM) influence lipid metabolism, inflammation, and mitochondrial function in perinatal cow hepatocytes. Bovine primary hepatocytes were isolated from five 1-day-old Holstein calves using collagenase IV perfusion, hepatocytes were treated with various concentrations of NEFA for 12 h to systematically simulate the metabolic changes in the negative energy balance (NEB) environment. Oxidative stress levels, lipid metabolism gene and protein expression, inflammatory pathway activation, and mitochondrial structure and function were assessed. Compared with the control group, 1.2 and 2.4 mM NEFA significantly increased lipid accumulation (TAG, T-CHO, LDL-c) and oxidative stress (MDA), while reducing antioxidant levels (GSH, SOD, T-AOC). NEFA upregulated lipogenic genes (SREBP-1c, ACC, FASN) and downregulated lipid oxidation genes (PPARα, CPT1A). Lipid transport was impaired, with altered APOE, LDLR, CD36, and L-FABP expression. NEFA activated the TLR4/MyD88/IRAK2/NF-κB pathway, increasing pro-inflammatory cytokines (IL-1β, IL-6, TNF-α). Mitochondrial function was impaired, as evidenced by increased ROS, reduced mitochondrial membrane potential, decreased mtDNA levels, and downregulated expression of oxidative phosphorylation genes. NEFA enhanced IRAK2 mitochondrial translocation, inhibited the mitochondrial biogenesis process, and increased apoptosis. NEFA alters lipid metabolism, inflammation, and mitochondrial function in bovine hepatocytes via the TLR4/MyD88/IRAK2 pathway, providing a new target for preventing metabolic diseases in perinatal cows.
血液中过量的非酯化脂肪酸（NEFA）常常导致围产期奶牛出现酮病和脂肪肝。线粒体结构和功能的完整性在脂质代谢失衡和炎症发展过程中起着关键作用。然而，NEFA对这些过程影响的确切分子机制仍不清楚。为了探究NEFA（0、0.3、0.6、1.2和2.4 mM）如何影响围产期奶牛肝细胞中的脂质代谢、炎症和线粒体功能。使用IV型胶原酶灌注法从五头1日龄的荷斯坦犊牛中分离出牛原代肝细胞，用不同浓度的NEFA处理肝细胞12小时，以系统模拟负能量平衡（NEB）环境下的代谢变化。评估氧化应激水平、脂质代谢基因和蛋白质表达、炎症途径激活以及线粒体结构和功能。与对照组相比，1.2和2.4 mM的NEFA显著增加脂质积累（TAG、T-CHO、LDL-c）和氧化应激（MDA），同时降低抗氧化水平（GSH、SOD、T-AOC）。NEFA上调脂肪生成基因（SREBP-1c、ACC、FASN）并下调脂质氧化基因（PPARα、CPT1A）。脂质转运受损，APOE、LDLR、CD36和L-FABP表达发生改变。NEFA激活TLR4/MyD88/IRAK2/NF-κB途径，增加促炎细胞因子（IL-1β、IL-6、TNF-α）。线粒体功能受损，表现为ROS增加、线粒体膜电位降低、mtDNA水平下降以及氧化磷酸化基因表达下调。NEFA增强IRAK2线粒体易位，抑制线粒体生物发生过程，并增加细胞凋亡。NEFA通过TLR4/MyD88/IRAK2途径改变牛肝细胞中的脂质代谢、炎症和线粒体功能，为预防围产期奶牛的代谢疾病提供了新靶点。
