目的 探究异氟醚通过激活蛋白激酶样内质网激酶（protein kinase‑like endoplasmic reticulum kinase, PERK）/真核翻译起始因子2α（eukaryotic translation initiation factor 2α, eIF2α）通路诱导神经元凋亡和内质网应激（endoplasmic reticulum stress, ERS）在围手术期神经认知障碍（perioperative neurocognitive disorders, PND）中的作用机制。 方法 30只雄性SD大鼠按随机数字表法分为3组（每组10只）：对照组、异氟醚组和PERK抑制剂组（GSK组）。异氟醚组和GSK组大鼠放置于麻醉箱，通过吸入2%异氟醚4 h建立PND模型，GSK组大鼠在吸入异氟醚6 h前使用PERK抑制剂GSK2606414（150 mg/kg）灌胃；对照组大鼠进行同样处理，但不吸入异氟醚，给予等量生理盐水灌胃。吸入异氟醚4 h后进行水迷宫试验，记录逃避潜伏期、穿越平台次数和目标象限停留时间，分析大鼠神经功能。水迷宫实验结束后处死大鼠取海马组织，TUNEL染色检测海马组织神经元细胞凋亡率，Western blot法检测海马组织PERK、eIF2α磷酸化水平及78 kDa葡糖调节蛋白（78 kDa glucose‑regulated protein, GRP78）、C/EBP同源蛋白（C/EBP homologous protein, CHOP）水平，免疫组化染色检测海马组织B淋巴细胞瘤‑2（B‑cell lymphoma‑2, Bcl‑2）、B淋巴细胞瘤‑2‑Associated X（B‑cell lymphoma‑2‑Associated X, Bax）水平。 结果 异氟醚组逃避潜伏期长于GSK组及对照组（P<0.05），穿越平台次数和目标象限停留时间少于GSK组及对照组（P<0.05）。异氟醚组海马组织神经元细胞凋亡率高于GSK组及对照组（P<0.05），PERK、eIF2α磷酸化水平高于GSK组及对照组（P<0.05），GRP78、CHOP水平高于GSK组及对照组（P<0.05），Bax水平高于GSK组及对照组，Bcl‑2水平低于GSK组及对照组（P<0.05）。 结论 异氟醚可能通过激活PERK/eIF2α通路和ERS引起海马组织神经元细胞凋亡，抑制PERK/eIF2α通路的激活可通过抑制ERS缓解海马组织神经元细胞凋亡从而对PND模型大鼠的神经功能起到保护作用。
Abstract: Objective To explore isoflurane-induced neuronal apoptosis via RNA-activated protein kinase-like ER kinase (PERK)/eukaryotic translation initiation factor 2α (eIF2α) pathway And the mechanism of endoplasmic reticulum stress (ERS) in postoperative cognitive dysfunction (POCD). Methods Thirty male SD rats were randomly divided into three groups, namely the control group, the isoflurane group and the PERK inhibitor GSK2606414 group. The rats in isoflurane group and GSK2606414 group were placed in an anesthesia chamber, and a POCD model was established by inhaling 2% isoflurane for 4 h. Rats in the GSK2606414 group were orally administered with GSK2606414 (150 mg / kg) once 6 h before inhaling isoflurane. Four hours after inhalation of isoflurane, the neural function of the rats was analyzed using a water maze video tracking analysis system. TUNEL staining was used to detect hippocampal neuronal apoptosis. Western blot was used to detect the phosphorylation levels of eIF2α and PERK and the protein levels of glucose regulated protein 78 (GRP78) and C/EBP homologous protein (CHOP). Immunohistochemical staining was used to detect the levels of apoptosis-related proteins Bax and Bcl-2 in hippocampus. Results After modeling, the escape latency of the isoflurane group increased significantly and the number of crossing platforms and target quadrant residence time decreased significantly (P0.05). The escape latency of GSK2606414 group was significantly lower than that of isoflurane group, and the number of crossing platforms and target quadrant residence time was significantly higher than that of isoflurane group (P0.05). The p-PERK/PERK, p-eIF2α/eIF2α, GRP78 and CHOP in the hippocampus of the isoflurane group were significantly higher than those in the control group (P0.05). The p-PERK/PER, p-eIF2α/eIF2α, GRP78 and CHOP in the GSK2606414 group were significantly lower than those in the isoflurane group (P0.05). The apoptosis rate of hippocampus in the isoflurane group was significantly higher than that in the control group (P0.05). The apoptosis rate of GSK2606414 group was significantly lower than that of isoflurane group (P0.05). Bax in the hippocampus of the isoflurane group was significantly higher than that of the control group and Bcl-2 was significantly lower than the control group (P0.05). The Bax of GSK2606414 group was significantly lower than that of isoflurane group and Bcl-2 was significantly higher than that of isoflurane group (P0.05). Conclusion Isoflurane may induce hippocampal neuronal apoptosis by activating the PERK/eIF2α pathway and ERS, while inhibition of PERK/eIF2α pathway activation may attenuate hippocampal neuronal apoptosis by inhibiting ERS, thereby acting as a nerve in the POCD rat model. The function is protective.
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