College of Animal Science, Inner Mongolia Key Laboratory of Animal Nutrition and Feed Science, Inner Mongolia Agricultural University, Hohhot, 010018, China.
College of Animal Science, Inner Mongolia Key Laboratory of Animal Nutrition and Feed Science, Inner Mongolia Agricultural University, Hohhot, 010018, China.
J Dairy Sci. 2021 Jan;104(1):337-345. doi: 10.3168/jds.2020-18246. Epub 2020 Nov 12.
Acetate is a short-chain fatty acid (SFA) that is the major substrate for de novo fatty acid synthesis. The mammalian target of rapamycin/eukaryotic initiation factor 4E (mTOR/eIF4E) signaling pathway is involved in fat synthesis. However, the effect and mechanism of acetate on fatty acid synthesis by the mTOR/eIF4E signaling pathway is unclear in bovine mammary epithelial cells (BMECs). The objectives of this study were to investigate the effect of acetate on cell viability, triacylglycerol (TG), and mRNA expression of the genes related to lipid synthesis. The mechanism of acetate regulation milk fat synthesis through the mTOR/eIF4E signaling pathway was assessed by blocking the mTOR signaling pathway and silencing eIF4E in BMECs. Third-passage BMECs were allocated to 6 treatments including 0, 4, 6, 8, 10, and 12 mM acetate to evaluate the effect of acetate on lipid synthesis; the optimum concentration in the first study was selected for the subsequent study. Subsequently, cells were randomly allocated to 4 treatments, 1 control group and 3 treated groups, consisting of acetate (6 mM), rapamycin (100 nM), and acetate + rapamycin to test the role of mTOR signaling pathway response to acetate in milk lipid synthesis. Finally, eIF4E was silenced by small interfering RNA (siRNA) to detect the role of eIF4E in milk lipid synthesis. Treatments included control, eIF4E siRNA, acetate (6 mM), and acetate+ eIF4E siRNA. Results showed that acetate increased TG accumulation and the relative expression of fatty acid synthase (FASN), acetyl-coenzyme A carboxylase α (ACACA), fatty acid-binding protein 3 (FABP3), sterol regulatory element binding protein 1 (SREBP1), peroxisome proliferator-activated receptor gamma (PPARG), mTOR, eIF4E, P70 ribosomal protein S6 kinase-1 (S6K1), and 4E-binding protein-1 (4EBP1) in a dose-dependent manner. Rapamycin effectively inhibited the positive effect of acetate on the relative expression of mTOR, eIF4E, S6K1, 4EBP1, FASN, ACACA, FABP3, stearoyl-CoA desaturase (SCD1), SREBP1, and PPARG. The upregulation of acetate on the relative expressions of FASN, ACACA, SCD1, and SREBP1 was suppressed when eIF4E was knocked down. It suggested that acetate regulated milk fat synthesis through mTOR/eIF4E signaling pathway in BMECs.
醋酸盐是一种短链脂肪酸 (SFA)，是从头合成脂肪酸的主要底物。哺乳动物雷帕霉素靶蛋白/真核起始因子 4E (mTOR/eIF4E) 信号通路参与脂肪合成。然而，在奶牛乳腺上皮细胞 (BMEC) 中，醋酸盐对 mTOR/eIF4E 信号通路的脂肪酸合成的影响和机制尚不清楚。本研究旨在探讨醋酸盐对细胞活力、三酰基甘油 (TG) 和与脂质合成相关基因的 mRNA 表达的影响。通过阻断 mTOR 信号通路和沉默 BMECs 中的 eIF4E，评估醋酸盐通过 mTOR/eIF4E 信号通路调节乳脂肪合成的机制。将第 3 代 BMEC 分配到 6 种处理组，包括 0、4、6、8、10 和 12 mM 醋酸盐，以评估醋酸盐对脂质合成的影响；在第一项研究中选择最佳浓度用于后续研究。随后，细胞被随机分配到 4 种处理组，1 个对照组和 3 个处理组，包括醋酸盐 (6 mM)、雷帕霉素 (100 nM) 和醋酸盐+雷帕霉素，以测试 mTOR 信号通路对醋酸盐在乳脂合成中的反应的作用。最后，通过小干扰 RNA (siRNA) 沉默 eIF4E，以检测 eIF4E 在乳脂合成中的作用。处理组包括对照组、eIF4E siRNA、醋酸盐 (6 mM) 和醋酸盐+eIF4E siRNA。结果表明，醋酸盐呈剂量依赖性增加 TG 积累和脂肪酸合成酶 (FASN)、乙酰辅酶 A 羧化酶 α (ACACA)、脂肪酸结合蛋白 3 (FABP3)、固醇调节元件结合蛋白 1 (SREBP1)、过氧化物酶体增殖物激活受体 γ (PPARG)、mTOR、eIF4E、p70 核糖体蛋白 S6 激酶 1 (S6K1) 和 4E 结合蛋白 1 (4EBP1) 的相对表达。雷帕霉素能有效抑制醋酸盐对 mTOR、eIF4E、S6K1、4EBP1、FASN、ACACA、FABP3、硬脂酰辅酶 A 去饱和酶 (SCD1)、SREBP1 和 PPARG 相对表达的正向作用。当沉默 eIF4E 时，醋酸盐对 FASN、ACACA、SCD1 和 SREBP1 相对表达的上调作用受到抑制。这表明醋酸盐通过 mTOR/eIF4E 信号通路调节 BMECs 中的乳脂肪合成。
