Key Laboratory of Zoonosis Research, Ministry of Education, College of Veterinary Medicine, Jilin University, 5333 Xi'an Road, Changchun, 130062, Jilin, China.
College of Animal Science, Jilin University, 5333 Xi'an Road, Changchun, 130062, Jilin, China.
J Dairy Sci. 2019 Aug;102(8):7359-7370. doi: 10.3168/jds.2018-16015. Epub 2019 May 31.
Disruption of endoplasmic reticulum (ER) homeostasis, often termed ER stress, is intrinsically linked with perturbation of lipid metabolism in humans and mice. Whether ER homeostasis is affected in cows experiencing fatty liver is unknown. The aim of this study was to investigate the potential role of ER stress in hepatic lipid accumulation in calf hepatocytes and ER stress status in dairy cows with severe fatty liver. In vitro experiments were conducted in which hepatocytes were isolated from calves and treated with different concentrations of fatty acids, tauroursodeoxycholic acid (TUDCA; a canonical inhibitor of ER stress), or both. The increase in phosphorylation level of protein kinase RNA-like ER kinase (PERK) and inositol requiring protein-1α (IRE1α) proteins, and the cleavage of activating transcription factor-6 (ATF6) protein in response to increasing doses of fatty acids (which were reversed by TUDCA treatment) in primary hepatocytes underscored a mechanistic link between fatty acids and ER stress. In addition, fatty acid treatment increased the abundance of sterol regulatory element-binding protein 1c, acetyl-CoA carboxylase-α, fatty acid synthase, and diacylglycerol acyltransferase 1, and lipid accumulation in calf primary hepatocytes, whereas inhibition of ER stress by incubating with TUDCA significantly weakened these effects. Overall, results in vitro indicate that inhibition of ER stress in calf hepatocytes alleviates fatty acid-induced lipid accumulation by downregulating the expression of lipogenic genes. In vivo experiments, liver and blood samples were collected from cows diagnosed as healthy (n = 15) or with severe fatty liver (n = 15). The phosphorylation level of PERK and IRE1α, the cleavage of ATF6 protein, and the abundance of several unfolded protein response genes (78 kDa glucose-regulated protein, AMP-dependent transcription factor 4, and spliced X-box binding protein 1) were greater in liver of cows with severe fatty liver. The present in vivo study confirms the occurrence of ER stress in dairy cows with severe fatty liver. Considering the causative role of fatty acid-induced ER stress in hepatic lipid accumulation in calf hepatocytes, the existence of ER stress in the liver of severe fatty liver cows may presage its participation in fatty liver progression in dairy cows. However, the mechanistic relationship between ER stress and fatty liver in dairy cows remain to be determined.
内质网（ER）稳态的破坏，通常称为 ER 应激，与人类和小鼠中脂质代谢的扰动密切相关。在患有脂肪肝的奶牛中，ER 稳态是否受到影响尚不清楚。本研究旨在探讨 ER 应激在犊牛肝细胞中肝脂质积累中的潜在作用，以及在患有严重脂肪肝的奶牛中 ER 应激状态。在体外实验中，从犊牛中分离出肝细胞，并分别用不同浓度的脂肪酸、牛磺熊脱氧胆酸（TUDCA；一种经典的 ER 应激抑制剂）或两者处理。随着脂肪酸剂量的增加，磷酸化蛋白激酶 RNA 样内质网激酶（PERK）和需要肌醇的蛋白-1α（IRE1α）蛋白的水平增加，以及激活转录因子-6（ATF6）蛋白的切割（用 TUDCA 处理可逆转）在原代肝细胞中，强调了脂肪酸与 ER 应激之间的机制联系。此外，脂肪酸处理增加了固醇调节元件结合蛋白 1c、乙酰辅酶 A 羧化酶-α、脂肪酸合酶和二酰基甘油酰基转移酶 1 的丰度，并增加了犊牛原代肝细胞中的脂质积累，而用 TUDCA 孵育抑制 ER 应激显著减弱了这些作用。总之，体外结果表明，在犊牛肝细胞中抑制 ER 应激可通过下调生脂基因的表达来减轻脂肪酸诱导的脂质积累。在体内实验中，从诊断为健康（n=15）或严重脂肪肝（n=15）的奶牛中采集肝脏和血液样本。在患有严重脂肪肝的奶牛肝脏中，PERK 和 IRE1α 的磷酸化水平、ATF6 蛋白的切割以及几种未折叠蛋白反应基因（78 kDa 葡萄糖调节蛋白、AMP 依赖的转录因子 4 和剪接 X 盒结合蛋白 1）的丰度更高。本体内研究证实了严重脂肪肝奶牛中 ER 应激的发生。考虑到脂肪酸诱导的 ER 应激在犊牛肝细胞中肝脂质积累中的因果作用，严重脂肪肝奶牛肝脏中 ER 应激的存在可能预示着其参与奶牛脂肪肝的进展。然而，ER 应激与奶牛脂肪肝之间的机制关系仍有待确定。
