Departments of Clinical Pharmacy and Cardiology, Harbin Medical University Cancer Hospital, Institute of Metabolic Disease, Heilongjiang Academy of Medical Science, Key Laboratories of Education Ministry for Myocardial Ischemia Mechanism and Treatment, Harbin 150000, China.
Department of Physiology, Emory University School of Medicine, Atlanta, GA 30322, USA.
Oxid Med Cell Longev. 2018 Dec 2;2018:7560610. doi: 10.1155/2018/7560610. eCollection 2018.
Previous studies indicate that the epithelial sodium channel (ENaC) in the kidney is upregulated in diabetes mellitus. Here, we show that ENaC single-channel activity in distal nephron cells was significantly increased by palmitate, a free fatty acid which is elevated in diabetes mellitus. We also show that palmitate increased intracellular Ca and that after chelating intracellular Ca with BAPTA-AM, palmitate failed to affect ENaC activity. Treatment of the cells with 2-aminoethoxydiphenyl borate (2-APB, an inhibitor of IP receptors) abolished the elevation of both intracellular Ca and ENaC activity. Treatment of the cells with apocynin (an NADPH oxidase inhibitor), dithiothreitol/NaHS (reducing agents), or LY294002 (a phosphoinositide 3-kinase (PI3K) inhibitor) prevented palmitate-induced ENaC activity, whereas thimerosal (an oxidizing agent) mimicked the effects of palmitate on ENaC activity. However, these treatments did not alter the levels of intracellular Ca, indicating that elevation of reactive oxygen species (ROS) and activation of PI3K are downstream of the signaling cascade. Since we have shown that ROS stimulate ENaC by activating PI3K, these data together suggest that palmitate first elevates intracellular Ca, then activates an NADPH oxidase to elevate intracellular ROS and PI3K activity, and finally increases ENaC activity via the activated PI3K.
先前的研究表明，肾脏中的上皮钠离子通道（ENaC）在糖尿病中会被上调。在这里，我们发现棕榈酸（糖尿病中含量升高的游离脂肪酸）可显著增加远曲小管细胞中 ENaC 的单通道活性。我们还发现棕榈酸会增加细胞内 Ca 浓度，并且用 BAPTA-AM 螯合细胞内 Ca 后，棕榈酸无法影响 ENaC 活性。用 2-氨基乙氧基二苯硼酸钠（IP 受体抑制剂）处理细胞可消除细胞内 Ca 和 ENaC 活性的升高。用 apocynin（NADPH 氧化酶抑制剂）、dithiothreitol/NaHS（还原剂）或 LY294002（一种磷酸肌醇 3-激酶（PI3K）抑制剂）处理细胞可防止棕榈酸诱导的 ENaC 活性，而 thimerosal（一种氧化剂）模拟了棕榈酸对 ENaC 活性的影响。然而，这些处理并没有改变细胞内 Ca 的水平，表明活性氧（ROS）的升高和 PI3K 的激活是信号级联的下游事件。由于我们已经表明 ROS 通过激活 PI3K 来刺激 ENaC，这些数据共同表明，棕榈酸首先升高细胞内 Ca，然后激活 NADPH 氧化酶以升高细胞内 ROS 和 PI3K 活性，最后通过激活的 PI3K 增加 ENaC 活性。
