p38 MAPK在沙土鼠前脑缺血再灌注损伤及缺血预处理中的作用

Role of p38 mitogen-activated protein kinase in forebrain ischemia-reperfusion injury and ischemic preconditioning in gerbils

目的 研究p38 MAPK在沙土鼠前脑缺血再灌注损伤及缺血预处理中的作用。方法 雄性蒙古沙土鼠384只，体重50-80 g，随机分为6组，每组64只。假手术组（SH组）：仅游离双侧颈总动脉但不阻断;缺血再灌注组（I／R组）：夹闭双侧颈总动脉，前脑缺血5min后恢复灌注;缺血预处理组（IP组）：前脑缺血3 min后恢复灌注，24 h后再行前脑缺血5 min;P组：于前脑缺血前20 min侧脑室内注射0．8μg p38 MAPK特异性激动剂P79350;SB组：于前脑缺血前20 min侧脑室内注射0．4μg p38 MAPK特异性抑制剂SB202190;溶剂对照组（VE组）：于前脑缺血前20min侧脑室内注射1％二甲基亚砜4μl。各组于再灌注15min、2h、4h、6h分别取8只沙土鼠，测定海马CA1区p-p38 MAPK的表达，再灌注1、3、5、7d分别取8只沙土鼠，采用开阔法观察行为学，然后测定海马CA1区存活神经元计数、凋亡神经元计数及p-p38 MAPK、HSP27、Bcl-2、Bax的表达。结果 I／R组再灌注期p-p38 MAPK表达上调，IP组及SB组再灌注各时点p-p38 MAPK表达水平低于I／R组，P组再灌注各时点高于I／R组、IP组及SB组（P〈0．05）;IP组、SB组较I／R组及vE组沙土鼠探索活动减少，CA1区再灌注期凋亡神经元数减少，HSP27、Bax表达下调，存活神经元数增加，Bcl-2表达上调（P〈0．05）;P组再灌注1 d探索活动增加，再灌注各时点p38 MAPK及HSP27表达均较I／R组上调（P〈0．05）。结论 沙土鼠脑缺血再灌注损伤及神经元凋亡与p38 MAPK的激活有关;缺血预处理可通过抑制p38 MAPK的激活，下调HSP27及Bax的表达、上调Bcl-2的表达。

Objective To investigate the role of p38 mitogen-activated protein kinase （p38 MAPK） in forebrain ischemia-reperfusion injury （I/R） in gerbils and the mechanism of ischemic preconditioning （IP）. Methods Three hundred and eighty-four male gerbils weighing 50-80 g were randomly divided into 6 groups （ n = 64each）： group Ⅰ sham operation; group Ⅱ I/R; group Ⅲ IP＋ I/R; group Ⅳ SB202190 （a specific p38 antagonist） ＋ I/R; group V P79350 （a specific p38 agonist） ＋ I/R and group Ⅵ 1% DMSO ＋ I/R （solvent control）. Forebrain ischemia was produced by occlusion of bilateral common carotid arteries for 5 min and confirmed by isoelectricity of EEG. In ischemic preconditioning group 3 min brain ischemia was induced 24 h before I/R. In group Ⅳ, Ⅴand Ⅵ SB202190 0.4μg in 4μl, P79350 0.8μg in 4 μl and 1% DMSO 4μl were injected into lateral ventricle of the brain 20 min before I/R respectively. The changes of animal behavior in the open field were evaluated. The animals were sacrificed at 15 min, 2 h, 4 h, 6 h, 1 d, 3 d, 5 d and 7 d after reperfusionwas started. Apoptosis in neurons and the expression of p-p38 MAPK, HSP27, Bcl-2 and Bax in hippocampal CA1 region were detected. Results The motor activity, the number of apoptotic neurons and expression of HSP27 and Bax were significantly decreased, while the number of living neurons and Bcl-2 expression were significantly increased in ischemic preconditioning group （ group Ⅲ ） and SB202190 group （ group Ⅳ ） as compared with I/R group and solvent control group （ Ⅵ ）. p-p38 MAPK expression was significantly higher in I/R group than in ischemic preconditioning group and SB202190 group. The expression of p-p38 MAPK and HSP27 was significantly higher in P79350 group than in I/R group. Conclusion p38 MAPK is involved in the cerebral I/R injury and apoptosis in neurons in gerbils. IP can reduce p38 MAPK activation to down-regnlate HSP27 and Bax expression and up-regulate Bcl-2 expression.