鸢尾素预处理激活Notch信号通路减轻大鼠全脑缺血-再灌注损伤

Irisin on reduce global cerebral ischemia-reperfusion injury by activating Notch signaling pathway

目的探讨鸢尾素预处理对全脑缺血-再灌注大鼠Notch信号通路的影响。方法成年雄性SD大鼠60只,体重300~350 g,按随机数字表法分为五组:假手术组(A组)、缺血-再灌注组(B组)、鸢尾素预处理组(C组)、DAPT(Notch信号通路选择性抑制剂)预处理组(D组)和鸢尾素复合DAPT预处理组(E组),每组12只。采用改良夹闭双侧颈总动脉合并低血压法建立大鼠全脑缺血-再灌注模型。假手术组仅分离双侧颈总动脉,不结扎。在大鼠全脑缺血-再灌注模型制备前30 min,C组静脉注射鸢尾素10μg/kg,D组腹腔注射DAPT 100 mg/kg,E组静脉注射鸢尾素10μg/kg后腹腔注射DAPT 100 mg/kg。A组和B组分别于相应时点静脉注射等体积生理盐水。再灌注24 h后采用神经功能缺失评分(NDS)法观察大鼠行为学变化,大脑干湿比重法观察大鼠脑含水量(BWC)变化,HE染色及原位末端标记(TUNEL)法观察大鼠脑组织海马CA1区坏死及凋亡情况,Western blot法检测Casepase-3、Notch1、NICD和Hes1蛋白相对含量。结果与A组比较,B组、C组、D组和E组NDS均明显升高,BWC均明显增加,脑组织海马CA1区形态学损伤和神经元凋亡程度明显加重,Caspase-3、NICD、Notch1和Hes1蛋白相对含量明显升高(P<0.05)。与B组比较,C组大鼠NDS明显降低,BWC明显减少,脑组织海马CA1区形态学损伤和神经元凋亡程度明显减轻,Caspase-3蛋白相对含量明显降低,NICD、Notch1和Hes1蛋白相对含量明显升高(P<0.05)。与C组比较,D组和E组大鼠NDS明显升高,海马CA1区形态学损伤和神经元凋亡程度明显加重,Caspase-3蛋白相对含量明显升高,NICD、Notch1和Hes1蛋白相对含量明显降低(P<0.05)。D组和E组各项指标差异无统计学意义。结论鸢尾素预处理可减轻大鼠全脑缺血-再灌注后海马神经元凋亡,降低脑水肿程度,其机制可能与激活Notch信号通路有关。

Objective To investigate the effect of irisin on Notch signaling pathway in rats with global cerebral ischemia-reperfusion.Methods A total of 60 healthy adult Sprague-Dawley rats were randomly divided into sham group(group A),global cerebral ischemia reperfusion group(group B),irisin-treated group(group C),DAPT(seletive inhibitor of Notch signaling pathway)-treated group(group D)and irisin combined with DAPT-treated group(group E),each group enrolled 12 rats.A rat model of global cerebral ischemia-reperfusion for group B was established by improved clipping of bilateral common carotid artery occlusion for 20 minutes and reperfusion for 24 hours combined with hypotension.In group A,only bilateral common carotid arteries separated without ligation.Thirty minutes before cerebral ischemia,rats in group C were injected with 10μg/kg irisin intravenously,rats in group D were intraperitoneally injected with DAPT 100 mg/kg,and rats in group E were injected with both irisin intravenously and DAPT intraperitoneally.Normal saline was intravenously injected at the corresponding time points in group A and group B.Twenty-four hours after reperfusion,neurological function was evaluated by neurological deficit score(NDS).The brain water content(BWC)was used to assess brain edema.HE staining and TdT-nediated dUTP nick end labeling(TUNEL)were used to observe the neuronal necrosis and apoptosis of hippocampal CA1 region in rats respectively.The protein expression of Caspase-3,Notch1,NICD,and Hes1 were detected by Western blot.Results Compared with group A,the NDS were higher,BWC were increased,the morphological damage were worse,the TUNEL positive cells increased,and the protein expression of Caspase-3,NICD,Notch1 and Hes1 up-regulated in other four groups(P<0.05).Compared with group B,the NDS were lower,the BWC were decreased,the morphological damage alleviated,the TUNEL positive cells reduced,the expression of Caspase-3 down-regulated,the protein expression of NICD,Notch1 and Hes1 up-regulated in group C(P<0.05).Compared with group C,the NDS were higher,the BWC were increased,the morphological damage were worse,the TUNEL positive cells increased,the expression of Caspase-3 up-regulated,and the protein expression of NICD,Notch1 and Hes1 down-regulated in groups D and E(P<0.05).Conclusion Irisin pretreatment can alleviate the morphological damage and cerebral edema after global cerebral ischemia-reperfusion.The mechanism may be related to the early activation of Notch signaling pathway.