异丙酚通过调控线粒体氧化磷酸化（OSPHOS）对神经发育及神经功能的影响及机制研究

Propofol is one of the most widely used anesthetics in clinical practice. However, it is still controversial that whether Propofol affects neurogenesis in children. In our previous study, we found that Propofol could improve the learning ability of neonatal mice. Meanwhile, the promotion of neural stem cells (NSCs) neuronal differentiation after Oropofol treatment was found, both in vivo and in vitro. Thus, whether propofol affects neural functions in neonatal mice and the underlying mechanisms remains a critical question. We have further observed an upregulation of mt-Atp8 in NSCs after Propofol exposure via RNA-seq screening, along with the activation of mitochondrial oxidative phosphorylation (OSPHOS) metabolic pathway. Therefore, in this project, we are going to use Nestin-GFP transgenic mice, cell tracking, patch clamp and flow-cytometry to clarify the effects of Propofol on the formation of neural circuits and the long-term cognitive functions in neonatal mice. In addition, RNA-sequence screening, gene knock-out and over-expression would be conducted to explore the specific molecular mechanisms of Propofol in promoting NSCs neuronal differentiation by activating OSPHOS and downstream Akt/mTOR pathway via mt-Atp8 upregulation. In this way, we hope to clarify the molecular mechanisms of Propofol in neurogenesis and further provide a new theoretical proof for the safely and reasonably perioperative application of Propofol in pediatrics.

异丙酚是临床应用最广泛的麻醉药物，而对于小儿神经发育的影响仍有争议，成为关注热点。课题组前期发现异丙酚能增强乳鼠的学习能力，在体内外均可促进神经干细胞(NSCs)向神经元分化，然而其是否影响神经功能以及具体机制如何是有待解决的关键科学问题。应用RNA-seq筛选我们发现，异丙酚致NSCs的mt-Atp8表达上调并激活线粒体氧化磷酸化（OSPHOS）途径。因此，本项目拟采用Nes-GFP转基因鼠、细胞示踪、膜片钳、流式细胞术等方法，明确异丙酚对乳鼠神经环路形成及远期神经功能的影响；利用RNA-seq、基因敲除及过表达技术，探讨异丙酚通过上调mt-Atp8激活OSPHOS及下游Akt/mTOR通路诱导NSCs向神经元分化的具体作用机制，从而阐明异丙酚对神经发育的作用地位和分子机制，为小儿围术期安全合理应用异丙酚提供理论依据。

为解决全身麻醉药物异丙酚对发育期大脑神经发育及认知功能的影响，本课题采用动物行为学、细胞生物学、生物信息学和分子生物学等技术，多维度展开对这一科学问题的探讨。研究发现，新生乳鼠接受异丙酚处理可以提高其空间认知功能、促进其海马神经发生。我们对异丙酚暴露后神经干细胞进行了bulk RNA-seq检测，结果提示与神经发生相关的基因转录水平明显上调。在此基础上，小鼠脑组织的单核细胞测序数据及代谢试验表明，异丙酚暴露后显著增强了神经干细胞中的氧化磷酸化水平。同时，通过qPCR、IF及基因调控等技术，我们确认了PGC-1α/NRF1/TFAM介导的线粒体氧化磷酸化在异丙酚促神经发生中的核心作用。上述研究明确了异丙酚通过调控线粒体能量代谢模式影响神经发育的分子机制，对于确保异丙酚用于小儿麻醉的安全性具有重大的社会意义。在该项目的资助下，我们共发表SCI论文6篇（其中3篇IF >5分），中文核心刊物1篇，培养数名硕士和博士研究生。