ATPIF1/ATP合酶在异氟烷致秀丽线虫全身损伤中的机制研究

吸入麻醉药异氟烷导致术后认知功能障碍（POCD），与神经元线粒体凋亡密切相关，但其上游机制未明。申请者前期研究证实：①异氟烷可激活线粒体凋亡，造成秀丽线虫损伤；②异氟烷诱导神经细胞ATP酶抑制因子1（ATPIF1）表达上调，ATP合酶抑制剂可改善异氟烷的神经细胞凋亡，且不影响ATPIF1表达。最新研究报道：ATPIF1可抑制ATP合酶水解ATP的活性，加重线粒体膜电位降低。我们推断：异氟烷通过诱导ATPIF1表达上调，降低ATP合酶水解ATP的活性，导致线粒体膜电位持续下降，激活线粒体凋亡通路，最终造成秀丽线虫损伤。本研究采用异氟烷诱导秀丽线虫全身损伤创新模型和神经细胞凋亡模型，通过调控ATPIF1表达或应用ATP合酶抑制剂干预，研究ATPIF1与ATP合酶结合能力、线粒体形态和功能、细胞凋亡以及线虫行为学的变化，从而揭示异氟烷导致POCD和线虫损伤的机制，为防治吸入麻醉药毒性提供新靶点。

Currently, the study of inhalational anesthetic-induced neurotoxicity and subsequent cognitive impairment is of great research interest in anesthesiology. The inhalational anesthetic isoflurane has been reported to induce caspase activation and apoptosis, potentially leading to postoperative cognitive dysfunction and promotion of Alzheimer’s disease neuropathogenesis. However, the underlying mechanisms by which isoflurane induce apoptosis remains largely unknown. Our preliminary data has shown that: firstly, isoflurane can activate mitochondrial apoptotic pathway, leading systemic injury in Caenorhabditis elegans; secondly, isoflurane can induce ATPase inhibitory factor 1(ATPIF1) increasing in neuron, the inhibitor of ATP synthase can improve isoflurane-induced neuronal apoptosis, which doesn’t affect the increasing of ATPIF1.It is newly reported that the increasing expression of ATPIF1 can inhibit the activity of ATP synthase in hydrolysis of ATP and induce the decrease of mitochondrial membrane potential. Based on the results of our preliminary studies and the aforementioned findings, we hypothesize that isoflurane induces systemic injury in Caenorhabditis elegans by increasing expression of ATPIF1, which could inhibit the activity of ATP synthase in hydrolysis of ATP, lead to declining mitochondrial membrane potential and activate mitochondrial apoptotic pathway, which could be prevented by administration of the inhibitor of ATP synthase. . Both apoptotic neurons model and systemic injury Caenorhabditis elegans model induced by isoflurane are used to test the hypothesis that isoflurane induces systemic injury in Caenorhabditis elegans by increasing expression of ATPIF1, which could inhibit the activity of ATP synthase in hydrolysis of ATP, lead to declining mitochondrial membrane potential and activate mitochondrial apoptotic pathway, the potential protective role of the inhibitor of ATP synthase on this process. To test this hypothesis, we will apply gene overexpression, small interfering RNA, RNA interfering, co-Immunoprecipitation, western blot, immunofluorescence confocal microscopy, electron microscopy, and behavioral tests to study the novel mechanism underlying isoflurane-induced cellular apoptosis at multiple aspects of molecular signaling, mitochondrial morphology, mitochondrial function, cell function, and animal behavior. The results will reveal the molecular mechanism of isoflurane-induced neuronal apoptosis and systemic injury, provide a theoretical basis for the prevention of inhalational anesthetic neurotoxicity.