七氟烷致老龄小鼠认知功能减退时组蛋白乙酰化与NR2B-CREB-BDNF信号通路的关系

Relationship between histone acetylation and NR2B-CREB-BDNF signaling pathway during sevoflurane-induced cognitive decline in aged mice

目的评价七氟烷致老龄小鼠认知功能减退时组蛋白乙酰化与含2B亚基的NMDA受体(NR2B)-环腺苷酸反应元件结合蛋白(CREB)-脑源性神经营养因子(BDNF)信号通路的关系。方法SPF级雄性C57BL/6J小鼠48只,22月龄,体重32~40 g,采用随机数字表法分为4组(n=12):对照组(C组)、七氟烷组(S组)、二甲基亚砜+七氟烷组(DS组)和组蛋白去乙酰化酶抑制剂伏立诺它(SAHA)+七氟烷组(SS组)。C组吸入100%氧气2 h,S组、DS组、SS组吸入3.0%七氟烷2 h,七氟烷吸入结束后24 h行Morris水迷宫实验(共6 d)。于七氟烷麻醉前2 h以及每天水迷宫实验前2 h,SS组腹腔注射SAHA 50 mg/kg,DS组腹腔注射等容量二甲基亚砜。水迷宫实验后立即处死小鼠取海马组织,采用Western blot法检测胞核乙酰化-H3以及胞浆NR2B、磷酸化CREB(p-CREB)、BDNF的表达水平。采用RT-PCR法检测NR2B和BDNF的mRNA表达水平。结果与C组比较,S组、DS组和SS组逃避潜伏期延长,目标象限停留时间百分比降低,乙酰化-H3、NR2B、p-CREB、BDNF、NR2B mRNA和BDNF mRNA表达下调(P<0.05);与S组或DS组比较,SS组逃避潜伏期缩短,目标象限停留时间百分比升高,乙酰化-H3、NR2B、p-CREB、BDNF、NR2B mRNA和BDNF mRNA表达上调(P<0.05);S组与DS组上述指标比较差异无统计学意义(P>0.05)。结论组蛋白乙酰化可通过调控NR2B-CREB-BDNF信号通路表达,参与七氟烷致老龄小鼠认知功能减退的病理生理机制。

Objective To evaluate the relationship between histone acetylation and N-methyl-D-aspartic acid(NMDA)receptor subunit 2B(NR2B)-cAMP-response element binding protein(CREB)-brain-derived neurotrophic factor(BDNF)signaling pathway during sevoflurane-induced cognitive decline in aged mice.Methods Forty-eight SPF healthy male C57BL/6J mice,aged 22 months,weighing 32-40 g,were divided into 4 groups(n=12 each)using a random number table method:control group(group C),sevoflurane group(group S),dimethyl sulfoxide(DMSO)plus sevoflurane group(group DS)and histone deacetylase inhibitor vorinostat(SAHA)plus sevoflurane group(group SS).The 100%oxygen was inhaled for 2 h in group C.In S,DS and SS groups,3.0%sevoflurane was inhaled for 2 h,Morris Water Maze(MWM)test was performed at 24 h after the end of sevoflurane inhalation.SAHA 50 mg/kg was intraperitoneally injected in group SS,and the equal volume of DMSO was given instead in group DS at 2 h before sevoflurane inhalation and at 2 h before MWM test was performed every day.Animals were sacrificed after the MWM test,and hippocampi were isolated for determination of the expression of acetyl-H3 in the nucleus and NR2B,phosphorylated CREB(p-CREB)and BDNF in cytoplasm by Western blot.Real-time polymerase chain reaction was used to determine the expression of NR2B and BDNF mRNA.Results Compared with group C,the escape latency was significantly prolonged,and the percentage of time spent in target quadrant was decreased,and the expression of acetyl-H3,NR2B,p-CREB,BDNF,NR2B mRNA,and BDNF mRNA was down-regulated in S,DS and SS groups(P<0.05).Compared with group S or group DS,the escape latency was significantly shortened,the percentage of time spent in target quadrant was increased,the expression of acetyl-H3,NR2B,p-CREB,BDNF,NR2B mRNA,and BDNF mRNA was up-regulated in group SS(P<0.05).There was no significant difference in each parameter mentioned above between group S and group SD(P>0.05).Conclusion Histone acetylation is involved in the pathophysiological mechanism of cognitive decline induced by sevoflurane anesthesia by regulating NR2B-CREB-BDNF signaling pathway in aged mice.