Abstract：
OBJECTIVE: Sepsis-associated encephalopathy (SAE) can lead to severe cerebral dysfunction as well as cognitive dysfunction, resulting in a significant disease burden. 3-Methyladenine (3-MA) has been confirmed to have anti-inflammatory effects on diseases characterized by enhanced autophagy. However, its role in SAE has not been clarified.
METHODS: An SAE mouse model was generated by intraperitoneal injection of lipopolysaccharide (LPS). Mice were given 5, 20, or 80 mg/kg 3-MA to determine the therapeutic dose. The mice in the different groups were given 20 mg/kg 3-MA or saline, and survival, body temperature, body weight and neurobehavioral scores were measured at different time points. The expression of autophagy-related proteins and inflammatory factors was detected by Western blotting, enzyme linked immunosorbent assay (ELISA) and real-time quantitative polymerase chain reaction (RT-qPCR) 12 h after LPS induction. Glial activation and neuronal injury in the hippocampus were detected by immunofluorescence staining and HE staining. The open Field test, novel object recognition (NOR) test, Y-maze test, and Morris water maze (MWM) test were performed to assess cognitive function.
RESULTS: Treatment with 20 or 80 mg/kg 3-MA reduced the increase in hippocampal TNF-α, IL-6, and IL-1β expression in SAE model mice, with 20 mg/kg 3-MA having the greatest therapeutic effect. Treatment with 20 mg/kg 3-MA effectively reduced the expression of hippocampal autophagy-related proteins and mortality, ameliorated hypothermia, decreased body weight and electroencephalography (EEG) performance, and attenuated the activation of neuroglia and neuronal damage. Moreover, it alleviated the cognitive dysfunction 2 weeks after LPS induction.
CONCLUSIONS: 3-MA reduced neuroglial activation and neuronal damage, attenuated neuroinflammation, and improved cognitive deficits during recovery period by inhibiting autophagy in SAE.
METHODS: An SAE mouse model was generated by intraperitoneal injection of lipopolysaccharide (LPS). Mice were given 5, 20, or 80 mg/kg 3-MA to determine the therapeutic dose. The mice in the different groups were given 20 mg/kg 3-MA or saline, and survival, body temperature, body weight and neurobehavioral scores were measured at different time points. The expression of autophagy-related proteins and inflammatory factors was detected by Western blotting, enzyme linked immunosorbent assay (ELISA) and real-time quantitative polymerase chain reaction (RT-qPCR) 12 h after LPS induction. Glial activation and neuronal injury in the hippocampus were detected by immunofluorescence staining and HE staining. The open Field test, novel object recognition (NOR) test, Y-maze test, and Morris water maze (MWM) test were performed to assess cognitive function.
RESULTS: Treatment with 20 or 80 mg/kg 3-MA reduced the increase in hippocampal TNF-α, IL-6, and IL-1β expression in SAE model mice, with 20 mg/kg 3-MA having the greatest therapeutic effect. Treatment with 20 mg/kg 3-MA effectively reduced the expression of hippocampal autophagy-related proteins and mortality, ameliorated hypothermia, decreased body weight and electroencephalography (EEG) performance, and attenuated the activation of neuroglia and neuronal damage. Moreover, it alleviated the cognitive dysfunction 2 weeks after LPS induction.
CONCLUSIONS: 3-MA reduced neuroglial activation and neuronal damage, attenuated neuroinflammation, and improved cognitive deficits during recovery period by inhibiting autophagy in SAE.
摘要：
目的：脓毒症相关性脑病(SAE)可导致严重的脑功能障碍和认知功能障碍,造成巨大的疾病负担。3-甲基腺嘌呤（3-MA）已被证实对以自噬增强为特征的疾病具有抗炎作用。然而,其在SAE中的作用尚未阐明。
方法：通过腹腔注射脂多糖（LPS）制备SAE小鼠模型。给予小鼠5、20或80mg/kg3-MA以确定治疗剂量。不同组的小鼠给予20mg/kg的3-MA或生理盐水,和生存，体温,在不同时间点测量体重和神经行为评分.免疫印迹法检测自噬相关蛋白和炎症因子的表达,酶联免疫吸附测定(ELISA)和实时定量聚合酶链反应(RT-qPCR)在LPS诱导后12h。免疫荧光染色和HE染色检测海马胶质细胞活化和神经元损伤。开放式现场测试,新型物体识别（NOR）测试，Y-迷宫测试,进行Morris水迷宫（MWM）测试以评估认知功能。
结果：用20或80mg/kg3-MA治疗可减少海马TNF-α的增加，IL-6和IL-1β在SAE模型小鼠中的表达,20mg/kg的3-MA具有最大的治疗效果。用20mg/kg的3-MA治疗可有效降低海马自噬相关蛋白的表达和死亡率。改善了体温过低，体重减轻和脑电图（EEG）表现，并减轻神经胶质细胞的激活和神经元损伤。此外，减轻LPS诱导后2周的认知功能障碍。
结论：3-MA减少神经胶质细胞活化和神经元损伤，减弱的神经炎症，并通过抑制SAE中的自噬改善恢复期的认知缺陷。
方法：通过腹腔注射脂多糖（LPS）制备SAE小鼠模型。给予小鼠5、20或80mg/kg3-MA以确定治疗剂量。不同组的小鼠给予20mg/kg的3-MA或生理盐水,和生存，体温,在不同时间点测量体重和神经行为评分.免疫印迹法检测自噬相关蛋白和炎症因子的表达,酶联免疫吸附测定(ELISA)和实时定量聚合酶链反应(RT-qPCR)在LPS诱导后12h。免疫荧光染色和HE染色检测海马胶质细胞活化和神经元损伤。开放式现场测试,新型物体识别（NOR）测试，Y-迷宫测试,进行Morris水迷宫（MWM）测试以评估认知功能。
结果：用20或80mg/kg3-MA治疗可减少海马TNF-α的增加，IL-6和IL-1β在SAE模型小鼠中的表达,20mg/kg的3-MA具有最大的治疗效果。用20mg/kg的3-MA治疗可有效降低海马自噬相关蛋白的表达和死亡率。改善了体温过低，体重减轻和脑电图（EEG）表现，并减轻神经胶质细胞的激活和神经元损伤。此外，减轻LPS诱导后2周的认知功能障碍。
结论：3-MA减少神经胶质细胞活化和神经元损伤，减弱的神经炎症，并通过抑制SAE中的自噬改善恢复期的认知缺陷。
