ABSTRACT

Background:

Signal transduction of Angiotensin II (Ang II) induced autophagy and its role in Ang II–induced dysfunction of HUVECs are still unclear.

Methods:

HUVECs are stimulated with different doses of Ang II (10-9-10-5 mol/L) for different time (6–48 hours). Autophagy-related protein markers: LC3, Beclin-1 and SQSTM1/p62 are measured by western blot.

Results:

Incubation with Ang II increases autophagic flux (Beclin-1, autophagosomes formation, and degradation of SQSTM1/p62, LC3-I). Increased autophagic levels are inhibited by pretreatment with Ang II type 1 receptor (AT1) blocker (Candesartan), NADPH Oxidase inhibitor (apocycin), mitochondrial K_ATP channels inhibitor (5-hydroxydecanoate, 5HD). 3-Methyladenine (inhibitors of autophagy) and rapamycin (activator of autophagy) respectively inhibits or activates Ang II–induced autophagy levels. Ang II decreases phosphorylation of endothelial nitric oxide synthase (eNOS) and NO production in HUVECs. L-NAME (NOS inhibitor) totally mimics the actions of Ang II on eNOS, NO production and autophagy levels. Rapamycin further decreases NO production combined with Ang II. Silence Atg5 completely reverses Ang II-activated autophagy levels.

Conclusions:

Our results demonstrate that Ang II stimulation increases autophagy levels via AT1 receptor, NADPH oxidase, mitochondrial K_ATP channel, eNOS, Atg5 signal pathway in HUVECs, and activation of autophagy contributes to Ang II induced dysfunction of HUVECs.

中文翻译：

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自噬导致血管紧张素 II 诱导的 HUVEC 功能障碍

摘要

背景：

血管紧张素 II (Ang II) 诱导的自噬的信号转导及其在 Ang II 诱导的 HUVEC 功能障碍中的作用仍不清楚。

方法：

HUVEC 用不同剂量的 Ang II (10-9-10-5 mol/L) 刺激不同时间（6-48 小时）。自噬相关蛋白标志物：LC3、Beclin-1 和 SQSTM1/p62 通过蛋白质印迹法测量。

结果：

与 Ang II 孵育会增加自噬通量（Beclin-1、自噬体形成和 SQSTM1/p62、LC3-I 的降解）。使用 Ang II 1 型受体 (AT1) 阻滞剂（坎地沙坦）、NADPH 氧化酶抑制剂（夹心霉素）、线粒体 K _ATP预处理可抑制自噬水平升高通道抑制剂（5-羟基癸酸酯，5HD）。3-甲基腺嘌呤（自噬抑制剂）和雷帕霉素（自噬激活剂）分别抑制或激活 Ang II 诱导的自噬水平。Ang II 降低内皮一氧化氮合酶 (eNOS) 的磷酸化和 HUVEC 中 NO 的产生。L-NAME（NOS 抑制剂）完全模拟了 Ang II 对 eNOS、NO 产生和自噬水平的作用。雷帕霉素与 Ang II 结合进一步降低 NO 的产生。Silence Atg5 完全逆转 Ang II 激活的自噬水平。

结论：

我们的结果表明，Ang II 刺激通过 AT1 受体、NADPH 氧化酶、线粒体 K _ATP通道、eNOS、HUVEC 中的 Atg5 信号通路增加自噬水平，并且自噬的激活有助于 Ang II 诱导的 HUVEC 功能障碍。