College of Animal Science and Veterinary Medicine, Heilongjiang Bayi Agricultural University, Daqing 163319, Heilongjiang, China.
College of Life Science and Technology, Heilongjiang Bayi Agricultural University, Daqing 163319, Heilongjiang, China.
J Anim Sci. 2021 Jul 1;99(7). doi: 10.1093/jas/skab184.
Fatty liver is a common metabolic disorder afflicting dairy cows during the periparturient period and is closely associated with endoplasmic reticulum (ER) stress. The onset of ER stress in humans and mice alters hepatic lipid metabolism, but it is unknown if such event contributes to fatty liver in dairy cows soon after parturition. ORAI calcium release-activated calcium modulator 1 (ORAI1) is a key component of the store-operated Ca2+ entry mechanism regulating cellular Ca2+ balance. The purpose of this study was to investigate the role of ORAI1 on hepatic lipidosis via ER stress in dairy cows. Liver tissue biopsies were collected from Holstein cows diagnosed as healthy (n = 6) or with hepatic lipidosis (n = 6). Protein and mRNA abundance of ER stress-related targets, lipogenic targets, or the transcription regulator SREBP1 and ORAI1 were greater in cows with lipidosis. In vitro, hepatocytes were isolated from four healthy female calves and used for culture with a 1.2 mM mixture of fatty acids (oleic, linoleic, palmitic, stearic, and palmitoleic acid) for various times (0, 3, 6, 9, or 12 h). As incubation time progressed, increases in concentration of Ca2+ and abundance of protein kinase RNA-like ER kinase (PERK), inositol-requiring protein 1α (IRE1α), and activating transcription factor-6 (ATF6) protein in response to exogenous fatty acids underscored a mechanistic link among Ca2+, fatty acids, and ER stress. In a subsequent study, hepatocytes were transfected with small interfering RNA (siORAI1) or the ORAI1 inhibitor BTP2 for 48 h or 2 h followed by a challenge with the 1.2 mM mixture of fatty acids for 6 h. Compared with control group, silencing or inhibition of ORAI1 led to decreased abundance of fatty acid synthesis (FASN, SREBP1, and ACACA) and ER stress-related proteins in bovine hepatocytes. Overall, data suggested that NEFA through ORAI1 regulate intracellular Ca2+ signaling, induce ER stress, and lead to lipidosis in isolated hepatocytes.
脂肪肝是围产期奶牛常见的代谢紊乱疾病，与内质网（ER）应激密切相关。人类和小鼠 ER 应激的发生会改变肝脏的脂质代谢，但尚不清楚这种事件是否会导致奶牛产后不久发生脂肪肝。钙释放激活钙调制器 1（ORAI1）是调节细胞内 Ca2+平衡的储存操作 Ca2+进入机制的关键组成部分。本研究旨在通过 ER 应激研究 ORAI1 在奶牛肝脏脂肪变性中的作用。从被诊断为健康（n=6）或患有肝脂肪变性（n=6）的荷斯坦奶牛中采集肝组织活检。脂肪变性牛的 ER 应激相关靶标、生脂靶标或转录调节因子 SREBP1 和 ORAI1 的蛋白和 mRNA 丰度更高。在体外，从 4 头健康雌性小牛中分离出肝细胞，并用于培养 1.2 mM 混合脂肪酸（油酸、亚油酸、棕榈酸、硬脂酸和棕榈油酸）不同时间（0、3、6、9 或 12 h）。随着孵育时间的延长，外源性脂肪酸引起的 Ca2+浓度增加和蛋白激酶 RNA 样内质网激酶（PERK）、肌醇需求蛋白 1α（IRE1α）和激活转录因子 6（ATF6）蛋白的丰度增加，突出了 Ca2+、脂肪酸和 ER 应激之间的机制联系。在随后的研究中，肝细胞用小干扰 RNA（siORAI1）或 ORAI1 抑制剂 BTP2 转染 48 h 或 2 h，然后用 1.2 mM 混合脂肪酸处理 6 h。与对照组相比，沉默或抑制 ORAI1 导致牛肝细胞中脂肪酸合成（FASN、SREBP1 和 ACACA）和 ER 应激相关蛋白的丰度降低。总体而言，数据表明 NEFA 通过 ORAI1 调节细胞内 Ca2+信号转导，诱导 ER 应激，并导致分离的肝细胞发生脂肪变性。
