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Logo of neuroscibullNeuroscience Bulletin
 
Neurosci Bull. 2009 June; 25(3): 115–121.
Published online 2009 July 7. doi:  10.1007/s12264-009-8818-9
PMCID: PMC5552567

Language: English | Chinese

Relationship between transmembrane signal transduction pathway and DNA repair and the mechanism after global cerebral ischemia-reperfusion in rats

细胞信号通路与DNA损伤修复的关系及在大鼠全脑缺血再灌注中的作用

Abstract

Objective

To investigate the protein levels of phospho-ERK and phospho-APE/Ref-1 in hippocampal neurons after global cerebral ischemia reperfusion in rats, and observe the relationship between transmembrane signal transduction and repair of DNA damage. The role of ERK signal transduction pathway following global cerebral ischemia reperfusion in rats is further discussed.

Methods

Ninety healthy male SD rats were divided into 3 groups randomly: Sham group (S group), Ischemia reperfusion group (IR group) and Pd98059 pretreatment/ischemia reperfusion group (PD group). Global cerebral ischemia reperfusion model was established by four-vessel occlusion (4-VO) method, and reperfusion was performed 5 minutes following ischemia. Protein levels of phospho-ERK and phospho-APE/Ref-1 were detected using immunohistochemical method at 2 h, 6 h, 12 h, 24 h, 48 h and 72 h after reperfusion, and neuron apoptosis was observed by HE and TUNEL staining.

Results

In CA1 region of IR group, TUNEL positive cells began to appear at 6 h after IR, and reached the apex during 24 h to 48 h. However, TUNEL positive was most strongly exhibited in PD group. In IR group, phospho-ERK was obviously detected in CA3 region at 2 h after IR, and its level was gradually decreased from 6 h until totally absent at 48 h. Besides, phospho-ERK expression in PD group was weaker than that in IR group. For phospho-APE/Ref-1, its expression began to appear in CA1 region in IR group at 2 h after IR, with no obvious changes during 2 h to 12 h. Phospho-APE/Ref-1 expression began to decrease at 24 h and this decrease continued thereafter. Expression level of phospho-APE/Ref-1 in PD group was lower than that in IR group. Results showed the concurrence of decreased phospho-ERK expression level and increased neuron apoptosis after cerebral ischemia reperfusion, the former of which was consistent with the decrease of phospho-APE/Ref-1 expression. Also, the greater the inhibition of ERK phosphorylation was, the greater decrease of APE/Ref-1 expression occurred.

Conclusion

Activation of ERK signal transduction pathway increased the expression of phospho-APE/Ref-1, and thus faciliated the repair of DNA damage. So, activation of ERK signal transduction pathway may protect neurons from apoptosis after cerebral ischemia reperfusion.

Keywords: cerebral ischemia/reperfusion injury, ERK, APE/Ref-1, DNA repair, apoptosis

摘要

目的

观察全脑缺血再灌注时, 细胞外信号调节激酶(ERK)与无嘌呤/无嘧啶核酸内切酶/氧化还原因子-1(APE/Ref-1)蛋白在大鼠海马区神经元的表达。 将跨细胞膜信号转导与胞内DNA损伤修复相联系, 探讨ERK信号通路在大鼠全脑缺血再灌注中的作用。

方法

90只雄性SD大鼠随机分为3组: 假手术组(S组)、 全脑缺血再灌注组(IR组)、 pd98059抑制剂组(PD组)。 采用四血管阻断法建立大鼠全脑缺血再灌注模型, 脑缺血时间5 min, 再灌注后2、 6、 12、 24、 48、 72 h, 用免疫组化法分别检测磷酸化ERK和APE/Ref-1蛋白的表达, HE和TUNEL染色观察神经元凋亡。

结果

IR组CA1区TUNEL阳性细胞于缺血再灌注6 h后开始出现, 24 h至48 h达高峰;TUNEL阳性细胞在PD组表达最强。 IR组磷酸化ERK于再灌注2 h后, 在海马CA3区可见明显表达, 再灌注6 h后逐渐下降, 至再灌注后48 h未见, PD组表达弱于IR组。 IR组的APE/Ref-1蛋白于再灌注后2 h在海马CA1区可见表达, 2 h到12 h表达无明显变化, 再灌注后24 h明显减少, 之后表达逐渐下降, PD组表达弱于IR组。 磷酸化ERK表达下降的同时伴随有细胞凋亡的增加, APE/Ref-1在缺血再灌注后的表达变化与p-ERK一致, 并且随着ERK磷酸化的被抑制APE/Ref-1的表达更加减弱。

结论

缺血再灌注中ERK通路的激活能增加APE/Ref-1蛋白的表达, 对DNA的修复起支持作用, 并在大鼠脑缺血再灌注时发挥抗神经元凋亡的保护作用。

关键词: 脑缺血再灌注损伤, ERK, APE/Ref-1, DNA修复, 凋亡

References

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Articles from Neuroscience Bulletin are provided here courtesy of Springer