An amphiphilic aggregate-induced emission polyurethane probe for in situ actin observation in living cells
An amphiphilic aggregate-induced emission polyurethane probe for in situ actin observation in living cells
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用于活细胞中原位肌动蛋白观察的两亲聚集体诱导发射聚氨酯探针
DOI：
10.1016/j.jcis.2020.08.113
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发表时间：
2021
影响因子：
9.9
通讯作者：
Massimiliano Galluzzi
中科院分区：
文献类型：
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作者：
Yuqing Niu;Bokai Zhang;Massimiliano Galluzzi
荧光探针或生物分子与肌动蛋白细胞骨架网络的特异性结合对于监测各种复杂的细胞活动，如细胞黏附、增殖、运动、内吞和细胞分裂等越来越重要。然而，提高细胞摄取和亚细胞分辨率仍然是细胞荧光探针成功和广泛应用的主要障碍。在这里，我们设计并合成了一种两亲性嵌段聚氨酯，它具有聚集诱导发射(AIE)的特殊光物理性质，可以用于活细胞成像，促进细胞结构的选择性可视化。以端异氰酸酯基聚乙二醇和聚己内酯为原料，与端羟基AIE染料进行两步聚合，制得AIE效应型聚氨酯(简称AIE-PU)。一系列表征技术证明了AIE-PU的成功合成。AIE-PU嵌段共聚物由于其线型嵌段分子的两亲性链段，可以在水溶液中自组装成球形纳米粒子，表现出相对稳定的光物理性质和良好的水分散性。细胞实验表明，AIE-PUS具有低毒和高亲和力的肌动蛋白网络。此外，通过低温和代谢抑制实验研究了AIE-PU纳米颗粒的摄取机制，表明AIE-PU纳米颗粒可以通过能量依赖的内吞作用很容易地内化到不同的活细胞中，并可以通过依赖于细胞骨架蛋白和小窝的运输途径从细胞外围转运到肌动蛋白网络。与肌动蛋白网络结合后，该探针的链间AIE机制显著增强，这对活细胞内肌动蛋白微丝网络的长期稳定荧光成像至关重要。最后，与商业肌动蛋白染料相比，该探针表现出更高的光稳定性，即使在较长的保留时间后，也没有明显的荧光猝灭。
The specific binding of fluorescent probes or biomolecules to the actin cytoskeleton network is increasingly important for monitoring various complex cellular activities such as cell adhesion, proliferation, locomotion, endocytosis, and cell division. However, improving cell uptake and subcellular resolution is still the main obstacle for successful and wide application of cellular fluorescent probes. Here, we designed and synthesized an amphiphilic block polyurethane with peculiar photophysical properties of aggregation induced emission (AIE), which can be used in living cell imaging to promote selective visualization of cell structures. The AIE effect polyurethane (abbreviated as AIE-PU) was prepared by two-step polymerization of diisocyanate terminated polyethylene glycol and polycaprolactone with hydroxyl terminated AIE dye. A series of characterization techniques proved the successful synthesis of AIE-PU. Due to the amphiphilic chain segment of its linear block molecule, AIE-PU block copolymers can self-assemble into spherical nanoparticles in aqueous solution, showing relatively stable photophysical properties and good water dispersion. Cellular experiments demonstrated that AIE-PUs have low toxicity and high actin network affinity. Moreover, the uptake mechanism was studied by low temperature and metabolic inhibition experiments, showing that AIE-PU nanoparticles could be easily internalized into different living cells through energy-dependent endocytosis, and can be transported from the cellular periphery to the actin network via clathrin- and caveolae-dependent transport pathway. Upon binding with the actin network, the inter-chain AIE mechanism of the probe was significantly enhanced, which is pivotal for the long-term stable fluorescence imaging of actin microfilament network in living cells. Finally, compared with commercial actin dyes, this probe showed higher photostability, even after a longer retention time, without significant fluorescence quenching.