Abstract：
Carboxylesterase (CE), an enzyme widely present in organisms, is involved in various physiological and pathological processes. Changes in the levels of CEs in the liver may predict the presence of type 2 diabetes mellitus (T2DM). Here, a novel dicyanoisophorone (DCI)-based proximity-labeled far-red fluorescent probe DCI2F-Ac with endoplasmic reticulum targeting was proposed for real-time monitoring and imaging of the CEs activity. DCI2F-Ac featured very low cytotoxicity and biotoxicity and was highly selective and sensitive for CEs. Compared with traditional CEs probes, DCI2F-Ac was covalently anchored directly to CEs, thus effectively reducing the loss of in situ fluorescent signals due to diffusion. Through the \"on-off\" fluorescence signal readout, DCI2F-Ac was able to distinguish cell lines and screen for CEs inhibitors. In terms of endoplasmic reticulum (ER) stress, it was found that thapsigargin (Tg) induced upregulation of CEs levels but not tunicamycin (Tm), which was related to the calcium homeostasis of the ER. DCI2F-Ac could efficiently detect downregulated CEs in the livers of T2DM, and the therapeutic efficacy of metformin, acarbose, and a combination of these two drugs was assessed by tracking the fluctuation of CEs levels. The results showed that combining metformin and acarbose could restore CEs levels to near-normal levels with the best antidiabetic effect. Thus, the DCI2F-Ac probe provides a great opportunity to explore the untapped potential of CEs in liver metabolic disorders and drug efficacy assessment.
摘要：
羧酸酯酶(CE),一种广泛存在于生物体中的酶，参与各种生理和病理过程。肝脏中CE水平的变化可以预测2型糖尿病（T2DM）的存在。这里,提出了一种新型的基于双氰基异佛尔酮(DCI)的具有内质网靶向性的邻近标记远红荧光探针DCI2F-Ac,用于实时监测和成像CEs活性.DCI2F-Ac具有非常低的细胞毒性和生物毒性，对CEs具有高度选择性和敏感性。与传统CEs探头相比，DCI2F-Ac直接共价锚定在CEs上，从而有效地减少了由于扩散引起的原位荧光信号的损失。通过“开-关”荧光信号读出，DCI2F-Ac能够区分细胞系并筛选CEs抑制剂。在内质网（ER）应激方面，发现thapsigargin（Tg）诱导CEs水平上调，而不是衣霉素（Tm），这与ER的钙稳态有关。DCI2F-Ac可以有效检测T2DM肝脏中下调的CE，和二甲双胍的治疗效果，阿卡波糖,通过跟踪CEs水平的波动来评估这两种药物的组合。结果表明，二甲双胍和阿卡波糖联合使用可以使CEs水平恢复到接近正常水平，具有最佳的抗糖尿病作用。因此，DCI2F-Ac探针为探索CEs在肝脏代谢紊乱和药物疗效评估中的未开发潜力提供了一个很好的机会.
