细胞外信号调节激酶信号转导通路活化在新生大鼠缺氧缺血性脑损伤中的作用

目的 观察细胞外信号调节激酶(extracellular-signal regulated kinase,ERK)在新生大鼠缺氧缺血性脑损伤中的作用.方法 Wistar大鼠随机分为假手术组、缺氧缺血组和PD98059(ERK抑制剂)组,通过结扎右侧颈总动脉,恢复2 h后,吸入含8%氧气的氧氮混合气体2 h的方法 建立新生大鼠缺氧缺血性脑损伤模型,PD98059组结扎右侧颈总动脉前10 min股静脉注射PD980592 mg/kg.各组新生大鼠于模型制作后6 h剥离结扎侧海马及皮层脑组织,分别测定脑组织丙二醛(malondialdehyde,MDA)含量、超氧化物歧化酶(superoxide dismutase,SOD)活性和细胞凋亡水平.Western印迹法检测ERK1和ERK2,Bcl-2和Bax蛋白的表达.组间差异比较采用方差分析及q检验.结果 缺氧缺血组细胞凋亡率明显高于假手术组(18.80±1.37)%和(3.53±0.34)%](q=6.06,P＜0.01),PD98059组细胞凋亡率(15.53±0.64)%]低于缺氧缺血组(q=3.87,P＜0.01).缺氧缺血组MDA含量为(342.9±10.8)μmol/L,明显高于假手术组(181.5±17.0)μmol/L](q=6.35,P＜0.01)和PD98059组(252.0±17.1)μmol/L](q=5.28,P＜0.01),而SOD活性(34.8±4.3)U/ml]明显低于假手术组(63.4±4.3)U/ml](q=4.99,P＜0.01)和PD98059组(51.5±3.8)U/ml](q=4.17,P＜0.01).3组总ERK1和ERK2蛋白表达差异无统计学意义.缺氧缺血组磷酸化ERK1和磷酸化ERK2蛋白表达明显高于假手术组(q分别=3.82和4.08,P均＜0.01)和PD98059组(q分别=4.79和5.12,P均＜0.01).缺氧缺血后Bcl-2和Bax蛋白表达明显高于假手术组(q分别=3.55和3.42,P均＜0.01);PD98059组较缺氧缺血组抗凋亡蛋白Bcl-2表达增加,促凋亡蛋白Bax表达降低(q分别=3.71和5.86,P均＜0.01).结论 ERK激活通过调节凋亡蛋白表达参与了新生大鼠缺氧缺血性脑损伤的发生.

Abstract：

Objective To investigate the effects of extracellular-signal regulated kinase (ERK) on hypoxic-ischemic brain damage of neonatal rats.Methods The hypoxic-ischemic brain damage model of neonatal Wistar rats was established as following:first the right common carotid artery of the rats was ligated;2 h after operation,the rats began to inhale 8%-oxygen oxygen-nitrogen gas mixture lasting for 2 h.Rats were randomly divided into three groups:sham-group,hypoxic-ischemic group and ERK inhibitor PD98059 group (the rats were injected PD98059 2 mg/kg 10 min before the ligation).Six hours after the models were done,hippocampi and cortex of the ligation side of rats in the three groups were collected,and the levels of malondialdehyde (MDA) and superoxide dismutase (SOD) were measured.Apoptosis of neuron was assessed by TUNEL staining.The expression of ERK1,ERK 2,Bcl-2 and Bax were examined by Western blot.The differences among the groups were analyzed with ANOVA and q test.Results Compared with the sham-group,the MDA level (342.9± 10.8) μmol/L vs (181.5± 17.0) μmol/L,q= 6.35,P＜0.01) and the apoptosis rate of neuron (18.80±1.37)% vs (3.53±0.34)%,q=6.06,P＜0.01) of hypoxic-ischemic group was higher,and SOD level was lower (34.8±4.3) U/ml vs (63.4±4.3) U/ml,q=4.99,P＜0.01].While the apoptosis rate of neuron (15.53±0.64) %] and MDA level (252.0± 17.1) μmol/L] of PD98059 group were lower than those of hypoxic-ischemic group(q=3.87 and 5.28,P＜0.01respectively),the SOD level (51.5 ± 3.8) U/ml] was higher than that of hypoxic-ischemic group (q=4.17,P＜0.01).There were no differences of ERK1 and ERK2 expressions among the three groups.The phosphorylated ERK1 and ERK2 levels of hypoxic-ischemic group were higher than those of sham-group (q=3.82 and 4.08,P＜0.01) and PD98059 group (q=4.79 and 5.12,P＜0.01).The expression of Bcl-2 and Bax of hypoxic-ischemic group were higher than those of sham-group (q=3.55 and 3.42,P＜0.01).Compared with hypoxic-ischemic group,Bcl-2 expression (q=3.71,P＜0.01) of PD98059 group was higher,and Bax expression (q=5.86,P ＜ 0.01) was lower.Conclusions ERK is involved in hypoxic-ischemic brain damage of neonatal rats through regulating the expression of apoptosis protein.

Effects of extracellular-signal regulated kinase signaling pathway activation on hypoxic-ischemic brain damage of neonatal rats

Objective To investigate the effects of extracellular-signal regulated kinase (ERK) on hypoxic-ischemic brain damage of neonatal rats.Methods The hypoxic-ischemic brain damage model of neonatal Wistar rats was established as following:first the right common carotid artery of the rats was ligated;2 h after operation,the rats began to inhale 8%-oxygen oxygen-nitrogen gas mixture lasting for 2 h.Rats were randomly divided into three groups:sham-group,hypoxic-ischemic group and ERK inhibitor PD98059 group (the rats were injected PD98059 2 mg/kg 10 min before the ligation).Six hours after the models were done,hippocampi and cortex of the ligation side of rats in the three groups were collected,and the levels of malondialdehyde (MDA) and superoxide dismutase (SOD) were measured.Apoptosis of neuron was assessed by TUNEL staining.The expression of ERK1,ERK 2,Bcl-2 and Bax were examined by Western blot.The differences among the groups were analyzed with ANOVA and q test.Results Compared with the sham-group,the MDA level (342.9± 10.8) μmol/L vs (181.5± 17.0) μmol/L,q= 6.35,P＜0.01) and the apoptosis rate of neuron (18.80±1.37)% vs (3.53±0.34)%,q=6.06,P＜0.01) of hypoxic-ischemic group was higher,and SOD level was lower (34.8±4.3) U/ml vs (63.4±4.3) U/ml,q=4.99,P＜0.01].While the apoptosis rate of neuron (15.53±0.64) %] and MDA level (252.0± 17.1) μmol/L] of PD98059 group were lower than those of hypoxic-ischemic group(q=3.87 and 5.28,P＜0.01respectively),the SOD level (51.5 ± 3.8) U/ml] was higher than that of hypoxic-ischemic group (q=4.17,P＜0.01).There were no differences of ERK1 and ERK2 expressions among the three groups.The phosphorylated ERK1 and ERK2 levels of hypoxic-ischemic group were higher than those of sham-group (q=3.82 and 4.08,P＜0.01) and PD98059 group (q=4.79 and 5.12,P＜0.01).The expression of Bcl-2 and Bax of hypoxic-ischemic group were higher than those of sham-group (q=3.55 and 3.42,P＜0.01).Compared with hypoxic-ischemic group,Bcl-2 expression (q=3.71,P＜0.01) of PD98059 group was higher,and Bax expression (q=5.86,P ＜ 0.01) was lower.Conclusions ERK is involved in hypoxic-ischemic brain damage of neonatal rats through regulating the expression of apoptosis protein.