运用分子探针技术探寻异丙酚分子靶点及功能鉴定

Recent studies usually focused on particular receptors or downstream intracellular signal transduction pathways. However, the real targets of general anesthesia drugs are still unknown. Identification of molecular targets of small molecule compounds is one of the most challenging tasks in drug research. The appearance of chemical probe makes it possible to solve this biomedical challenge. Propofol is the most commonly drug used in general anesthesia, which has the characteristics of small molecule compounds. But its specific molecular target is not yet clear. Combined with the latest developments in chemical biology, we have successfully constructed the fluorescently labeled probe of propofol, and it was found co-localization on the cell membrane and mitochondria. Based on this work, we will construct epoxy agarose gel 6B labeled propofol as a small molecular probe, and then combined the use of affinity purification strategy with small molecules-proteomics to discover the specific molecular target of propofol. Finally, we will verify the specificity of this potential target at both cellular and animal levels, and to determine its exact role in the molecular mechanism of propofol. The advantages of this study is a systematic explore strategy to find the bona fide target of propofol. This study is expected to further reveal the anesthesia mechanism of propofol.

目前对全麻机制的研究，主要集中在某一受体或下游细胞内信号转导通路，其真正的靶点并未完全明确。小分子化合物分子靶点的确证是药物研究中最具挑战性的工作之一，化学探针的出现为攻克这一生物医学难题带来曙光。异丙酚为临床常用麻醉药物，符合小分子化合物特性，且其具体的分子靶点尚未明确。结合化学生物学最新进展，我们成功构建了荧光标记的活性异丙酚分子探针，并发现该分子与细胞膜表面及线粒体存在共定位。本研究拟在这一工作的基础上构建环氧琼脂糖凝胶6B耦连的特异性异丙酚分子探针，利用亲和纯化、小分子-蛋白质组学等最新研究策略，来发掘异丙酚作用的确切的、特异性的分子靶点，并在细胞和动物整体水平进一步验证该靶点的特异性，以确定其在异丙酚分子机制中的作用。本研究采用系统性挖掘策略，在蛋白质组学层面发掘异丙酚的分子靶点，有望进一步揭示异丙酚在全麻中的作用机制。

我们在神经系统中揭示了一条全新的APC/C-CDH1-SENP2-MEF2A信号通路的调控机制。同时在麻醉药物引起认知功能障碍中的作用及调控机制进一步研究：①核糖体蛋白S6参与异氟醚引起的发育大脑神经毒性。②异氟醚，而非地氟醚，可以损害幼年小鼠的学习和记忆，其机制是通过Akt/GSK3β信号通路，降低了Akt/GSK3β的磷酸化水平。③二十二碳六烯酸（docosahexaenoic acid，DHA）改善多次七氟醚暴露新生小鼠引起的突触损伤和长期记忆障碍。④ 丹参酮IIA（Tanshinone IIA） 改善七氟醚引起的发育小鼠神经毒性。⑤七氟醚作用于泛素蛋白酶体途径降低新生幼鼠突触后致密蛋白-95的水平。另外，我们还发现ERK MAP激酶激活Cdk5的159位丝氨酸磷酸化参与外周炎症引起的疼痛/超敏反应