摘要
目的评价海马细胞外调节蛋白激酶1/2(ERK1/2)在外源性一氧化碳(CO)改善失血性休克复苏大鼠认知功能中的作用。方法清洁级健康雄性SD大鼠90只,9~10周龄,体重350~400g,采用随机数字表法分为5组(n=18):假手术组(S组)、失血性休克复苏组(H组)、CO组(CO组)、PD98059-CO组(PCO组)和PD98059组(P组)。失血性休克通过股静脉放血至MAP为25~35mmHg维持60min,将收集的血液在15min内回输体内达到初始血压水平作为复苏,必要时输注生理盐水,建立失血性休克复苏模型;CO组复苏结束后立即将大鼠置于含有1%CO混合空气的玻璃箱内3h;PCO组和PC组放血前30min时脑室内注射ERK1/2抑制剂PD98059(30μmol/L)30μl;S组只进行股静脉、股动脉穿刺置管和脑室内注射生理盐水。于复苏结束后3h时处死大鼠取海马,采用气相分析法测定海马CO含量;于复苏结束后第15天时采用Morris水迷宫实验评价认知功能,随后处死取海马,TUNEL联合活化caspase-3免疫荧光法测定海马CA1区细胞凋亡率;于复苏结束后6h时处死大鼠取海马,采用Westernblot法检测磷酸化ERK1/2(p-ERK1/2)、Bcl-2和Bax表达。结果与S组比较,H组、PCO组和P组逃避潜伏期延长,H组、CO组、PCO组和P组海马CO含量增加,神经细胞凋亡率升高,活化caspase-3和p-ERK1/2表达上调,Bcl-2/Bax比值下降(P<0.05);与H组比较,CO组逃避潜伏期缩短,海马CO含量增加,神经细胞凋亡率降低,活化caspase-3表达下调,p-ERK1/2表达上调,Bcl-2/Bax比值升高,P组逃避潜伏期延长,海马CO含量减少,神经细胞凋亡率升高,活化caspase-3表达上调,p-ERK1/2表达下调,Bcl-2/Bax比值降低(P<0.05);与CO组比较,PCO组逃避潜伏期延长,神经细胞凋亡率升高,活化caspase-3表达上调,p-ERK1/2表达下调,Bcl-2/Bax比值下降(P<0.05)。结论外源性CO改善失血性休克复苏大鼠的认知功能,与增加海马神经细胞ERK1/2磷酸化水平,抑制神经细胞凋亡有关。
Objective To evaluate the role of hippocampal extracellular signal-regulated kinase 1/2 (ERK1/2) in exogenous carbon monoxide(CO)-induced improvement of cognitive function in a rat model of hemorrhage shock and resuscitation. Methods Ninety clean-grade healthy male Sprague-Dawley rats, aged 9-10 weeks, weighing 350-400 g, were divided into 5 groups (n=18 each) using a random number table method: sham operation group (S group), hemorrhage shock and resuscitation group (H group), CO group, PD98059-CO group (PCO group) and PD98059 group (P group). Hemorrhagic shock was induced by withdrawing blood from the femoral vein until mean arterial pressure was reduced to 25-35 mmHg which was maintained for 60 min and resuscitated by infusing the blood withdrawn over 15 min until the initial blood pressure was achieved, and normal saline was infused when needed.Rats were exposed to air mixture containing 1% CO for 3 h in a glass box after the end of resuscitation in group CO.ERK1/2 inhibitor PD98059 (30 μmol/L) 30 μl was injected into the cerebral ventricle at 30 min before hemorrhage in PCO and PC groups.Right femoral artery and vein were only cannulated, and normal saline was injected into the cerebral ventricle in group S. Rats were sacrificed at 3 h after the end of resuscitation, brains were removed and hippocampi were isolated for determination of CO content (by gas chromatograph assay). Cognitive function was assessed by Morris water maze test at 15 days after the end of resuscitation, rats were then sacrificed and hippocampi were isolated for determination of cell apoptosis in hippocampal CA1 region by TUNEL and cleaved caspase-3 immunofluorescence, and the apoptosis rate was calculated.Rats were sacrificed at 6 h after the end of resuscitation, and hippocampi were isolated to detect the expression of phosphorylated ERK1/2 (p-ERK1/2), Bcl-2 and Bax by Western blot. Results Compared with group S, the escape latency was significantly prolonged in H, PCO and P groups, and the hippocampal CO content and apoptosis rate w
作者
孙文波
张立民
张冬雪
李睿
Sun Wenbo;Zhang Limin;Zhang Dongxue;Li Rui(Department of Anesthesiology,Cangzhou Central Hospital,Cangzhou 061001,China;Department of Gerontology,Cangzhou Central Hospital,Cangzhou 061001,China)
出处
《中华麻醉学杂志》
CAS
CSCD
北大核心
2018年第12期1500-1504,共5页
Chinese Journal of Anesthesiology
基金
国家自然科学基金(81701296).
关键词
细胞外信号调节MAP激酶类
一氧化碳
再灌注损伤
海马
细胞凋亡
认知障碍
Extracellular signal-regulated MAP kinases
Carbon monoxide
Reperfusion injury
Hippocampus
Apoptosis
Cognition disorders
