부산대학교 도서관

目的 通过观察不同剂量艾司氯胺酮对高迁移率族蛋白1(high mobility group box-1,HMGB1)/丝裂原活化蛋白激酶(mitogen activated protein kinase,MAPK)轴的调控,探究艾司氯胺酮改善小鼠POCD的作用机制.方法 选取健康的18 个月雄性小鼠随机进行分组,对照组(C组)、脓毒症组(LPS组)、LPS+低剂量艾司氯胺酮组 5 mg/kg(LPS+S1)、LPS组+中等剂量艾司氯胺酮组10 mg/kg(LPS+S2)、LPS组+高剂量艾司氯胺酮组15 mg/kg(LPS+S3),LPS+HMGB1 抑制剂(LPS+甘草酸组)、LPS+中等剂量艾司氯胺酮干预组+HMGB1 抑制剂(LPS+S2+甘草酸组)、LPS+中等剂量艾司氯胺酮干预组+MAPK信号通路抑制剂组(LPS+S2+PD98095 组)、LPS+中等剂量艾司氯胺酮干预组+HMGB1 抑制剂+MAPK信号通路激活剂(LPS+S2+甘草酸+ANS组).(艾司氯胺酮干预组是在模型建立后给予第一部分所研究的最佳艾司氯胺酮剂量进行处理).LPS组是通过腹腔注射LPS 5 mg/kg,HMGB1 抑制剂组给予腹腔注射甘草酸剂量为40 mg/kg,对照组注射同等剂量生理盐水.不同药物处理后进行旷场试验和水迷宫实验,之后立即处死小鼠并取其海马组织,采用ELISA测定海马组织白细胞介素-6(interleukin-6,IL-6)、白细胞介素-1(interleukin-1,IL-1)、肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α).Western Blot检测海马组织HMGB1 和MAPK信号通路相关蛋白(p38 MAPK、ERK1/ERK2、JNK)表达水平.结果 与对照组小鼠对比,LPS组脓毒症模型小鼠HMGB1、p38 MAPK、ERK1/ERK2、JNK表达增加,差异有统计学意义(P<0.05).与模型小鼠对比,抑制HMGB1 表达后,LPS+S2+甘草酸组脓毒症模型小鼠炎症因子表达降低,差异有统计学意义(P<0.05);与模型小鼠对比,抑制HMGB1 表达后,LPS+S2+甘草酸组脓毒症模型小鼠各组小鼠各时点逃避潜伏期时间减少、穿越平台次数增加、总行驶距离增加、中央区域停留时间增加、目标象限时间增加,差异有统计学意义(P<0.05);与模型小鼠对比,抑制MAPK信号通路后,LPS+S2+PD98095 组模型小鼠炎症因子表达降低,差异有统计学意义(P<0.05);与模型小鼠对比,逃避潜伏期时间等上述指标优于模型组,差异有统计学意义(P<0.05);与模型小鼠对比,抑制HMGB1 表达同时激活MAPK信号通路后,LPS+S2+甘草酸+ANS组模型小鼠炎症因子表达降低,差异有统计学意义(P<0.05);与模型小鼠对比,逃避潜伏期时间等上述指标优于模型组,差异有统计学意义(P<0.05).结论 艾司氯胺酮能够一定程度改善脓毒症小鼠炎症因子表达,其机制主要是艾司氯胺酮能够直接抑制HMGB1 与MAPK通路的表达或是激活,更深层次的机制可能艾司氯胺酮能够通过抑制HMGB1 进而抑制MAPK信号通路的激活,进而改善脓毒症小鼠模型的症状.
Objective By observing the regulation of high mobility group box-1(HMGB1)/mitogen activated protein kinase(MAPK)axis at different doses of esketamine,to explore the mechanism of esketamine improving POCD in mice.Methods Healthy 18-month male mice were randomly divided into different groups:control group(group C),sepsis group(LPS group),and LPS+low-dose esketamine group 5 mg/kg(LPS+S1),LPS+medium-dose esketamine group 10 mg/kg(LPS+S2),LPS+high-dose esketamine group 15 mg/kg(LPS+S3),LPS+HMGB1 inhibitor(LPS+glycyrrhizin group),LPS+medium-dose esketamine in-tervention group+HMGB1 inhibitor(LPS+S2+glycyrrhizin group),LPS+medium-dose esketamine intervention group+MAPK signa-ling pathway inhibitor group(LPS+S2+PD98095 group),and LPS+medium-dose esketamine dry Pre-group+HMGB1 inhibitor+ MAPK signaling pathway activator(LPS+S2+glycyrrhizic acid+ANS group).The Esketamine intervention group was treated with the optimal dose of Esketamine studied in Part I after the model was established.The LPS group was intraperitoneally injected with LPS 5 mg/kg,the HMGB1 inhibitor group was intraperitoneally injected with glycyrrhizic acid at a dose of 40 mg/kg,and the control group was injected with the same dose of normal saline.Open field test and water maze test were performed after treatment with different drugs,and then the mice were killed immediately and their hippocampal tissues were taken,and the interleukin-6(IL-6),inter-leukin-1(IL-1)and tumor necrosis factor-α(TNF-α)were determined by ELISA.The expression levels of HMGB1 and MAPK signaling pathway related proteins(p38 MAPK,ERK1/ERK2,JNK)in hippocampus were detected by Western Blot.Results Compared with the control group,the expression of HMGB1,p38 MAPK,ERK1/ERK2 and JNK in the LPS group was increased,and the difference was statistically significant(P<0.05).Compared with the model mice,after inhibiting the expression of HMGB1,the LPS+S2+GA group had a significant reduction in the expression of inflammatory factors(P<0.05);compared with the model mice,after inhibiting the expression of HMGB1,the escape latency time was decreased,the number of crossing the platform was in-creased,the total travel distance was increased,the residence time in the central area was increased,and the target quadrant time was increased at each time point in the LPS+S2+GA group of sepsis model mice(P<0.05).Compared with the model mice,after in-hibiting the MAPK signaling pathway,the expression of inflammatory factors in LPS+S2+PD98095 group was decreased,and the difference was statistically significant(P<0.05).Compared with the model mice,the escape latency time and other indicators of the model group were significantly different(P<0.05).Compared with the model mice,after inhibiting the expression of HMGB1 and ac-tivating the MAPK signaling pathway,the LPS+S2+GA+ANS group had a significant reduction in the expression of inflammatory factors(P<0.05);compared with the model group,the escape latency time and other indicators of the model group were better than those of the model group,and the differences were statistically significant(P<0.05).Conclusion Esketamine can improve the expression of inflammatory factors in sepsis mice to a certain extent,and the mechanism is mainly that esketamine can directly inhibit the expres-sion or activation of HMGB1 and MAPK pathways.A deeper mechanism may be that esketamine can inhibit the activation of MAPK signaling pathways by inhibiting HMGB1,thus improving the symptoms of sepsis mouse models