Visualizing Autophagic Flux during Endothelial Injury with a Pathway-Inspired Tandem-Reaction Based Fluorogenic Probe

被引:15
作者
Lei, Yu [1 ]
Ren, Wenming [2 ]
Wang, Cheng-Kun [1 ]
Tao, Rong-Rong [1 ]
Xiang, Huai-Jiang [1 ]
Feng, Li-Li [3 ]
Gao, Yin-Ping [1 ]
Jiang, Quan [1 ]
Li, Xin [1 ]
Hu, Youhong [2 ]
Han, Feng [1 ,3 ]
机构
[1] Zhejiang Univ, Coll Pharmaceut Sci, Hangzhou 310058, Zhejiang, Peoples R China
[2] Chinese Acad Sci, State Key Lab Drug Res, Shanghai Inst Mat Med, Shanghai 201203, Peoples R China
[3] Nanjing Med Univ, Sch Pharm, Nanjing 211166, Jiangsu, Peoples R China
关键词
Fluorescent imaging; fluorescent probe; autophagy; endothelial injury; PEROXYNITRITE; FLUORESCENT; DESIGN; CELLS;
D O I
10.7150/thno.33867
中图分类号
R-3 [医学研究方法]; R3 [基础医学];
学科分类号
1001 ;
摘要
Autophagy is a dynamic and complicated catabolic process. Imaging autophagic flux can clearly advance knowledge of its pathophysiology significance. While the most common way autophagy is imaged relies on fluorescent protein-based probes, this method requires substantial genetic manipulation that severely restricts the application. Small fluorescent probes capable of tracking autophagic flux with good spatiotemporal resolution are highly demanable. Methods: In this study, we developed a small-molecule fluorogenic probe (AFG-1) that facilitates real-time imaging of autophagic flux in both intact cells and live mice. AFG-1 is inspired by the cascading nitrosative and acidic microenvironments evolving during autophagy. It operates over two sequential steps. In the first step, AFG-1 responds to the up-regulated peroxynitrite at the initiation of autophagy by its diphenylamino group being oxidatively dearylated to yield a daughter probe. In the second step, the daughter probe responds to the acidic autolysosomes at the late stage of autophagy by being protonated. Results: This pathway-dependent mechanism has been confirmed first by sequentially sensing ONOO- and acid in aqueous solution, and then by imaging autophagic flux in live cells. Furthermore, AFG-1 has been successfully applied to visualize autophagic flux in real-time in live mice following brain ischemic injury, justifying its robustness. Conclusion: Due to the specificity, easy operation, and the dynamic information yielded, AFG-1 should serve as a potential tool to explore the roles of autophagy under various pathological settings.
引用
收藏
页码:5672 / 5680
页数:9
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