Role of CIP2A in preoperative sleep deprivation-induced aggravation of postoperative cognitive dysfunction in aged mice
刘阳 1李慧敏 1巩红岩 1魏苗苗 1张高峰 1王明山 1郑芳 2王娟
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作者信息
1. 青岛市市立医院东院麻醉科，青岛 266071
2. 青岛市市立医院东院超声科，青岛 266071
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摘要
目的 评价抑癌因子蛋白磷酸酶2A(CIP2A)在术前睡眠剥夺加重老龄小鼠术后认知功能障碍中的作用。 方法 SPF级健康C57BL/6J小鼠110只，雌雄不拘，18～20月龄，体质量29～35 g。采用随机数字表法分为5组(n＝22)：假手术组(S组)、腹部手术组(O组)、术前睡眠剥夺＋腹部手术组(D＋O组)、CIP2A shRNA＋腹部手术组(CS＋O组)和CIP2A shRNA＋术前睡眠剥夺＋腹部手术组(CS＋D＋O组)。术前14 d时，S组、O组和CS＋O组海马注射对照shRNA慢病毒，D＋O组和CS＋D＋O组海马注射CIP2A shRNA慢病毒。术前连续3 d行睡眠剥夺。术后7～11 d采用Morris水迷宫实验评估认知功能。术后3 d时深麻醉下处死小鼠取海马，采用Western blot法测定CIP2A、高迁移率族蛋白B1(HMGB1)、离子化钙结合适配蛋白-1(Iba-1)、蛋白磷酸酶2A的α亚单位(PP2Aa)、蛋白磷酸酶2A的催化亚单位(PP2Ac)、磷酸化tau蛋白(p-tau) (S396)和p-tau (S404)表达，检测海马ROS、MDA和SOD的水平，免疫荧光染色法计数海马CA1区Iba-1阳性细胞。 结果 与S组比较，O组逃避潜伏期延长，穿越平台位置次数减少，目标象限停留时间缩短，海马CIP2A、Iba-1和HMGB1表达上调，PP2Ac表达下调，ROS、MDA水平和Iba-1阳性细胞计数升高，SOD活性降低(P<0.05)；与O组比较，D＋O组逃避潜伏期延长，穿越平台位置次数减少，目标象限停留时间缩短，海马CIP2A、Iba-1和HMGB1表达上调，PP2Ac表达下调，ROS、MDA水平和Iba-1阳性细胞计数升高，SOD水平降低，CS＋O组逃避潜伏期缩短，穿越平台位置次数增加，目标象限停留时间延长，海马CIP2A、Iba-1和HMGB1表达下调，PP2Ac表达上调，ROS、MDA水平和Iba-1阳性细胞计数降低，SOD活性升高(P<0.05)；与D＋O组比较，CS＋D＋O组逃避潜伏期缩短，穿越平台位置次数增加，目标象限停留时间延长，海马CIP2A、Iba-1和HMGB1表达下调，PP2Ac表达上调，ROS、MDA水平和Iba-1阳性细胞计数降低，SOD水平升高(P<0.05)。5组海马PP2Aa表达比较差异无统计学意义(P>0.05)。 结论 术前睡眠剥夺加重老龄小鼠术后认知功能障碍的机制与上调CIP2A的表达，促进氧化应激反应、神经炎症反应以及tau蛋白磷酸化有关。 Objective To evaluate the role of cancerous inhibitor of protein phosphatase 2A (CIP2A) in preoperative sleep deprivation (PSD)-induced aggravation of postoperative cognitive dysfunction (POCD) in aged mice. Methods One hundred and ten healthy C57BL/6J mice of either sex, aged 18-20 months, weighing 29-35 g, were divided into 5 groups (n=22 each) using a random number table method: sham operation group (S group), abdominal surgery group (O group), PSD + abdominal surgery group (D+ O group), CIP2A shRNA + abdominal surgery group (CS+ O group), and CIP2A shRNA+ PSD+ abdominal surgery group (CS+ D+ O group). At 14 days before surgery, control shRNA lentivirus was injected into the hippocampus in S, O and CS+ O groups, and CIP2A shRNA was injected into the hippocampus in D+ O and CS+ D+ O groups. PSD was carried out for 3 consecutive days prior to surgery. Cognitive function was assessed using the Morris water maze test at days 7-11 after surgery. The mice were sacrificed under deep anesthesia at day 3 after surgery, and hippocampal tissues were obtained to determine the expression of CIP2A, high mobility group box 1 (HMGB1), ionized calcium-binding adapter molecule 1 (Iba-1), alpha subunit of protein phosphatase 2A (PP2Aa), catalytic subunit of protein phosphatase 2A (PP2Ac), phosphorylated tau protein (p-tau) (S396), and p-tau (S404) (by Western blot), levels of reactive oxygen species (ROS), malondialdehyde (MDA), and superoxide dismutase (SOD), and count of Iba-1 positive cells in the hippocampal CA1 region (using immunofluorescence staining). Results Compared with S group, the escape latency was significantly prolonged, the frequency of crossing the platform was reduced, duration of stay in the target quadrant was shortened, the expression of CIP2A, Iba-1 and HMGB1 was up-regulated, PP2Ac expression was down-regulated, levels of ROS and MDA and count of Iba-1 positive cells were increased, and the activity of SOD was decreased in O group (P<0.05). Compared with O group, the escape latency was significantly prolonged, the frequency of crossing the platform was reduced, duration of stay in the target quadrant was shortened, the expression of CIP2A, Iba-1 and HMGB1 was up-regulated, PP2Ac expression was down-regulated, levels of ROS and MDA and count of Iba-1 positive cells were increased, and the activity of SOD was decreased in D+ O group, and the escape latency was significantly shortened, the frequency of crossing the platform was increased, duration of stay in the target quadrant was prolonged, the expression of CIP2A, Iba-1 and HMGB1 was down-regulated, PP2Ac expression was up-regulated, levels of ROS and MDA and count of Iba-1 positive cells were decreased, and the activity of SOD was increased in CS+ O group (P<0.05). Compared with D+ O group, the escape latency was significantly shortened, the frequency of crossing the platform was increased, duration of stay in the target quadrant was prolonged, the expression of CIP2A, Iba-1 and HMGB1 was down-regulated, PP2Ac expression was up-regulated, levels of ROS and MDA and count of Iba-1 positive cells were decreased, and the activity of SOD was increased in CS+ D+ O group (P<0.05). There was no significant difference in PP2Aa expression among the five groups (P>0.05). Conclusions The mechanism by which PSD aggravates POCD is related to up-regulating the expression of CIP2A and promoting oxidative stress responses, neuroinflammatory responses and phosphorylation of tau protein in aged mice.
Abstract
Objective To evaluate the role of cancerous inhibitor of protein phosphatase 2A (CIP2A) in preoperative sleep deprivation (PSD)-induced aggravation of postoperative cognitive dysfunction (POCD) in aged mice. Methods One hundred and ten healthy C57BL/6J mice of either sex, aged 18-20 months, weighing 29-35 g, were divided into 5 groups (n=22 each) using a random number table method: sham operation group (S group), abdominal surgery group (O group), PSD + abdominal surgery group (D+ O group), CIP2A shRNA + abdominal surgery group (CS+ O group), and CIP2A shRNA+ PSD+ abdominal surgery group (CS+ D+ O group). At 14 days before surgery, control shRNA lentivirus was injected into the hippocampus in S, O and CS+ O groups, and CIP2A shRNA was injected into the hippocampus in D+ O and CS+ D+ O groups. PSD was carried out for 3 consecutive days prior to surgery. Cognitive function was assessed using the Morris water maze test at days 7-11 after surgery. The mice were sacrificed under deep anesthesia at day 3 after surgery, and hippocampal tissues were obtained to determine the expression of CIP2A, high mobility group box 1 (HMGB1), ionized calcium-binding adapter molecule 1 (Iba-1), alpha subunit of protein phosphatase 2A (PP2Aa), catalytic subunit of protein phosphatase 2A (PP2Ac), phosphorylated tau protein (p-tau) (S396), and p-tau (S404) (by Western blot), levels of reactive oxygen species (ROS), malondialdehyde (MDA), and superoxide dismutase (SOD), and count of Iba-1 positive cells in the hippocampal CA1 region (using immunofluorescence staining). Results Compared with S group, the escape latency was significantly prolonged, the frequency of crossing the platform was reduced, duration of stay in the target quadrant was shortened, the expression of CIP2A, Iba-1 and HMGB1 was up-regulated, PP2Ac expression was down-regulated, levels of ROS and MDA and count of Iba-1 positive cells were increased, and the activity of SOD was decreased in O group (P<0.05). Compared with O group, the escape latency was significantly prolonged, the frequency of crossing the platform was reduced, duration of stay in the target quadrant was shortened, the expression of CIP2A, Iba-1 and HMGB1 was up-regulated, PP2Ac expression was down-regulated, levels of ROS and MDA and count of Iba-1 positive cells were increased, and the activity of SOD was decreased in D+ O group, and the escape latency was significantly shortened, the frequency of crossing the platform was increased, duration of stay in the target quadrant was prolonged, the expression of CIP2A, Iba-1 and HMGB1 was down-regulated, PP2Ac expression was up-regulated, levels of ROS and MDA and count of Iba-1 positive cells were decreased, and the activity of SOD was increased in CS+ O group (P<0.05). Compared with D+ O group, the escape latency was significantly shortened, the frequency of crossing the platform was increased, duration of stay in the target quadrant was prolonged, the expression of CIP2A, Iba-1 and HMGB1 was down-regulated, PP2Ac expression was up-regulated, levels of ROS and MDA and count of Iba-1 positive cells were decreased, and the activity of SOD was increased in CS+ D+ O group (P<0.05). There was no significant difference in PP2Aa expression among the five groups (P>0.05). Conclusions The mechanism by which PSD aggravates POCD is related to up-regulating the expression of CIP2A and promoting oxidative stress responses, neuroinflammatory responses and phosphorylation of tau protein in aged mice.
关键词
睡眠剥夺/术后认知并发症/抑癌因子蛋白磷酸酶2A
Key words
Sleep deprivation/Postoperative cognitive complications/Cancerous inhibitor of protein phosphatase 2A
万方数据