CX3CR1通过调节Ca2+内流介导神经元凋亡影响缺血性卒中小鼠预后

基础医学与临床 ›› 2020, Vol. 40 ›› Issue (1): 1-8.

• 研究论文 • 下一篇

CX3CR1通过调节Ca²⁺内流介导神经元凋亡影响缺血性卒中小鼠预后

刘畅^#, 张申^#, 周菲惠, 夏阳, 李智高, 王进昆, 汤志伟^*

昆明医科大学第一附属医院神经外一科, 云南昆明 650031

收稿日期:2018-11-05 修回日期:2019-04-22 出版日期:2020-01-05 发布日期:2019-12-27
通讯作者: ^*tangzhiwei7755@hotmail.com
作者简介:^# 对本文有相同贡献
基金资助:
国家自然科学基金(81760220);云南省基金项目(2016NS058,2017NS057,2018NS0140,2017FE467-135,2017FB111)

CX3CR1 mediates neuronal apoptosis by regulating Ca²⁺ influx to affect the prognosis of ischemic stroke mice

LIU Chang^#, ZHANG Shen^#, ZHOU Fei-hui, XIA Yang, LI Zhi-gao, WANG Jin-kun, TANG Zhi-wei^*

Department of Neurosurgery, the First Affiliated Hospital of Kunming Medical University, Kunming 650031, China

Received:2018-11-05 Revised:2019-04-22 Online:2020-01-05 Published:2019-12-27
Contact: ^*tangzhiwei7755@hotmail.com

摘要/Abstract

摘要： 目的观察脑缺血后趋化因子受体1(CX3CR1)在神经元上的表达变化,并探讨其作用及机制。方法将野生型C57/BL6小鼠和CX3CR1基因敲除小鼠,分别设置对照组(假手术组)和小鼠中动脉永久闭塞(pMCAO)模型,用磁共振成像(MRI)检测30 min后脑缺血范围和24 h后梗死面积;免疫荧光三重染色法检测凋亡;Western blot检测CX3CR1蛋白的表达。在体外,培养原代神经元,建立氧糖剥离(OGD)细胞缺血模型;用MTT法检测细胞存活率;免疫荧光检测神经元CX3CR1的表达;激光共聚焦显微镜观察神经元内Ca²⁺浓度变化。结果在野生型小鼠中,与对照组和健侧相比,pMACO后,患侧CX3CR1表达显著升高(P＜0.05),且CX3CR1与凋亡蛋白caspase-3在神经元上共表达;与野生型小鼠相比,CX3CR1基因敲除小鼠pMACO 30 min后缺血损伤面积相似,但24 h后CX3CR1基因敲除小鼠梗死面积小于对照组(P＜0.05);在体外,原代神经元OGD后CX3CR1表达显著升高(P＜0.05),敲除神经元上CX3CR1,可减轻谷氨酸介导的兴奋性损伤,细胞存活率显著上升,同时观察到,敲除CX3CR1可降低谷氨酸介导的Ca²⁺ 内流到神经元的速度和总量。结论缺血可诱导神经元CX3CR1的表达,且神经元CX3CR1可以通过调节Ca²⁺内流来介导神经元的凋亡。

关键词: 缺血, CX3CR1, 神经元, 凋亡, 钙离子

Abstract: Objective To observe the changes of chemokine receptor 1(CX3CR1) expression in neurons after cerebral ischemia, and to explore its function mechanism. Methods Wild-type C57/BL6 mice and CX3CR1 gene knockout mice were set up as control group (sham operation group) and permanent middle cerebral artery occlusion(pMACO) model, respectively. Cerebral ischemia range and infarction area were detected after 30min by MRI, and apoptosis was detected by immunofluorescence triple staining.Western blot was used to detect the expression of CX3CR1 protein.In vitro, primary neurons were cultured to establish an oxysaccharide dissection (OGD) cell ischemia model. Cell survival rate was detected by MTT colorimetry, expression of CX3CR1 on neurons was detected by immunofluorescence, and the changes of Ca²⁺ concentration in neurons was observed by laser confocal microscopy. Results Compared with the control group, the expression of CX3CR1 in affected side of pMACO significantly increased in wild-type mice (P＜0.05), and CX3CR1 was co-expressed with the apoptotic protein caspase-3 in neurons.Compared with wild-type ones, the ischemic injury area of CX3CR1 gene knockout mice was similar 30 min after pMACO, but the infarct area of CX3CR1 gene knockout mice 24 h after pMACO was smaller than that of the control group (P＜0.05).In vitro, the expression of CX3CR1 significantly increased after OGD of primary neurons (P＜0.05). Knockout of CX3CR1 on neurons can reduce the excitatory injury mediated by glutamate and significantly increase the cell survival rate. Meanwhile, the knockout of CX3CR1 reduces the speed and quantity of glutamate-mediated Ca²⁺ flowing into neurons. Conclusions Ischemia can induce the expression of CX3CR1 on neurons, and CX3CR1 on neurons mediates the apoptosis of neurons by regulating Ca²⁺ internal flow.

Key words: ischemia, CX3CR1, neurons, apoptosis, calcium ion

中图分类号:

R743.33

刘畅, 张申, 周菲惠, 夏阳, 李智高, 王进昆, 汤志伟. CX3CR1通过调节Ca²⁺内流介导神经元凋亡影响缺血性卒中小鼠预后[J]. 基础医学与临床, 2020, 40(1): 1-8.

LIU Chang, ZHANG Shen, ZHOU Fei-hui, XIA Yang, LI Zhi-gao, WANG Jin-kun, TANG Zhi-wei. CX3CR1 mediates neuronal apoptosis by regulating Ca²⁺ influx to affect the prognosis of ischemic stroke mice[J]. Basic & Clinical Medicine, 2020, 40(1): 1-8.

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CX3CR1通过调节Ca²⁺内流介导神经元凋亡影响缺血性卒中小鼠预后

CX3CR1 mediates neuronal apoptosis by regulating Ca²⁺ influx to affect the prognosis of ischemic stroke mice

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