盐皮质激素受体激活后通过上调心房肌ROS进而调控Kv1.5/IKur而影响房颤易感性的研究

Kv1.5 potassium channel is an important therapeutic target of atrial fibrillation. Patients with hyperaldosteronemia have higher risk of atrial fibrillation than normal people. Our study showed that aldosterone could upregulate Kv1.5 expression by activating mineralocorticoid receptor (MR), but the underlying mechanism remains unknown. Our previous investigation indicated that NADPH oxidase (NOX)/reactive oxygen species (ROS) axis was closely related to the onset of atrial fibrillation. Our recent study showed that aldosterone increased the expression of NOX2, NOX4, Kv1.5, the production of reactive oxygen species and the intensity of IKur current and that antioxidants and Ang II type I receptor (AT1R) antagonists inhibited aldosterone-induced IKur augmentation. The above findings suggest that MR activation may initiate atrial fibrillation by upregulating Kv1.5 expression through activating NOX/ROS axis and that aldosterone/MR signaling may have crosstalk with Ang II/AT1R signaling in the regulation of Kv1.5 expression. In addition, we found that the expression of Kv1.5 peaked at 2 h and 24 h after aldosterone treatment, suggesting that there exist non-genomic and genomic pathway in aldosterone-induced Kv1.5 expression. The present study aims at clarifying how aldosterone/MR regulates Kv1.5/IKur and influences the onset of atrial fibrillation by modulating NOX/ROS axis. The elucidation of the mechanism underlying aldosterone-related Kv1.5/IKur regulation may provide new evidence for the utilization of MR antagonists in the treatment of atrial fibrillation in clinical practice.

Kv1.5是重要的房颤治疗靶点，临床上高醛固酮血症患者的房颤发病风险增加，醛固酮干预可增加心房Kv1.5通道的表达，但调控机制未明。我们报道NOX/ROS系统与房颤的发生密切相关。预实验显示醛固酮可上调心房肌NOX2、NOX4、ROS及Kv1.5的表达，并增强Kv1.5负载的IKur电流。抗氧化剂及AT1R阻断剂可抑制醛固酮介导的IKur增强。以上结果提示醛固酮/MR可通过激活NOX/ROS系统而上调Kv1.5进而促进房颤发生，同时醛固酮/MR可能与AngII/ AT1R交互作用，共同调控Kv1.5。醛固酮干预2小时及24小时后Kv1.5的表达分别达到峰值，提示醛固酮对Kv1.5调控可能涉及非基因组途径及基因组途径；本项目通过培养心房肌细胞及建立家兔房颤模型，深入探讨醛固酮/MR通过影响NOX/ROS-Kv1.5/IKur，进而影响房颤易感性的分子机制，为MR受体阻断剂治疗房颤提供新证据。

Kv1.5是重要的房颤治疗靶点，醛固酮（Aldo）干预可增加心房Kv1.5通道的表达，但调控机制未明。我们报道NOX/ROS系统与房颤的发生密切相关。预实验结果提示Aldo/盐皮质激素受体（MR）可通过激活NOX/ROS系统而上调Kv1.5进而促进房颤发生，同时Aldo/MR可能与血管紧张素II（AngII）/ 血管紧张素转换酶I型受体（AT1R）交互作用。醛固酮对Kv1.5调控可能涉及非基因组途径及基因组途径。本项目将探讨醛固酮/MR通过影响NOX/ROS-Kv1.5/IKur，进而影响房颤易感性的分子机制。通过培养乳鼠心房肌细胞，Aldo及AngII干预24h，可显著上调心房肌Kv1.5蛋白的表达，同时Aldo和AngII在诱导Kv1.5蛋白产生方面存在协同作用。螺内酯（Spiro）预处理，可抑制Aldo及AngII介导的Kv1.5蛋白表达，联合应用氯沙坦（Los）及Spiro可以抑制Aldo和Aldo诱导的Kv1.5蛋白表达。使用Aldo干预24h可上调心房肌NOX1、NOX2及NOX4的蛋白表达及ROS的含量，使用Spiro预处理可抑制这一上调作用。Src蛋白激酶家族抑制剂、NOX2抑制剂及NOX1/NOX4抑制剂均不同程度抑制Aldo介导的ROS产生及Kv1.5的蛋白表达。Aldo干预24h可以上调P-Src、P-Smad2/3及P-ERK1/2的蛋白表达，使用Spiro预处理可逆转上述作用。Aldo 干预 2h 显著增加心房肌细胞中 ROS 的产生，使用钙离子螯合剂 BAPTA 预处理 1h 后可明显抑制 Aldo 诱导的 ROS 产生。Aldo干预 2h 可显著增加心房肌细胞中Kv1.5的蛋白表达，使用Spiro预处理1h可显著抑制Aldo对Kv1.5的上调作用。与对照组相比，Aldo干预30min可显著上调结构蛋白caveolin-1。我们的研究结果表明AngII/ATR1以及Aldo/MR可通过NOX-ROS轴上调细胞Kv1.5及增加结构蛋白caveolin-1的表达而增加房颤的易感性。这为使用ACEI/ARB及MRA提高心房肌电稳定性以减少房颤的发生开辟了新的视野，也为使用ACEI/ARB及MRA类药物抑制NOX及下游的信号分子以抑制房颤的发生提供了新的治疗靶点，有潜在的转化应用价值。