Department of Gerontology, the First Affiliated Hospital of China Medical University, Shenyang, China.
APMIS. 2014 Oct;122(10):985-92. doi: 10.1111/apm.12242. Epub 2014 Mar 28.
The aim of this study was to explore if cell autophagy is activated by AngII before aging using human umbilical vascular endothelial cells (HUVECs). The ultrastructural analysis of HUVECs was performed to observe autophagosomes. The LC3-II/LC3-I ratio was determined by western blot assay. The β-gal staining was used to identify cell senescence. The flow cytometry was performed to evaluate cell apoptosis. The BH3 domain analog ABT737 or Beclin-1 knockdown by specific shRNA or valsartan was applied to investigate their effects on cell autophagy, senescence, and apoptosis induced by Ang II. Cell autophagy was initiated after Ang II treatment at 24 h. And cell senescence and apoptosis were observed in Ang II-treated cells at 48 h. The significant interaction of Beclin-1 and Bcl-2 was detected at 48 h after Ang II treatment. Beclin-1 was indispensable to Ang II-induced autophagy, and its BH3 domain was required for the interaction with Bcl-2 to attenuate autophagy. Pretreated with valsartan, cells were present with less autophagic, senescent, and apoptotic cells after Ang II stimulation. In conclusion, Ang II induced autophagy, senescence, and apoptosis of HUVECs progressively, and autophagy presented an early protective effect on vascular endothelial damage due to Ang II.
本研究旨在探讨血管紧张素 II（Ang II）是否在衰老前激活人脐静脉内皮细胞（HUVEC）的细胞自噬。通过超微结构分析观察自噬体，Western blot 检测 LC3-II/LC3-I 比值，β-半乳糖苷酶染色鉴定细胞衰老，流式细胞术评估细胞凋亡。用 BH3 结构域类似物 ABT737 或特异性 shRNA 敲低 Beclin-1 或缬沙坦处理，以研究其对 Ang II 诱导的细胞自噬、衰老和凋亡的影响。Ang II 处理 24 h 后开始发生细胞自噬，48 h 时观察到 Ang II 处理的细胞发生衰老和凋亡。Ang II 处理 48 h 后检测到 Beclin-1 和 Bcl-2 之间存在显著的相互作用。Beclin-1 是 Ang II 诱导自噬所必需的，其 BH3 结构域对于与 Bcl-2 的相互作用以减弱自噬也是必需的。Ang II 刺激前用缬沙坦预处理，细胞在 Ang II 刺激后自噬、衰老和凋亡的细胞数量减少。结论：Ang II 可渐进性诱导 HUVEC 自噬、衰老和凋亡，自噬对 Ang II 引起的血管内皮损伤具有早期保护作用。
