纤维蛋白原通过激活EGFR信号通路抑制体外低糖低氧损伤神经元轴突再生  

Fibrinogen inhibits axonal regeneration of neurons subjected to in vitro low-glucose and low-oxygen damage by activating the EGFR signaling pathway

作　　者：曾雯 谭玲娟 周胜强 刘芳[2] Zeng Wen;Tan Lingjuan;Zhou Shengqiang;Liu Fang(Hunan University of Traditional Chinese Medicine,Changsha 410000,China;Affiliated Hospital of Hunan Academy of Traditional Chinese Medicine,Changsha 410006,China)

机构地区：[1]湖南中医药大学,长沙410000 [2]湖南省中医药研究院附属医院,长沙410006

出　　处：《中国组织化学与细胞化学杂志》2023年第5期451-457,473,共8页Chinese Journal of Histochemistry and Cytochemistry

基　　金：湖南省临床医疗技术创新引导项目(2021SK51010);湖南省中医药科研计划项目(C2022011);湖南省卫生健康委科研计划项目(D202319019427);中国中医科学院学部委员学术传承与传播专项项目(CI2022E014XB);长沙市自然科学基金(kq2208144);湖南省自然科学基金(2023JJ60458,2023JJ30367);湖南中医药大学研究生创新课题项目(2023CX17)。

摘　　要：目的探讨纤维蛋白原(fibrinogen,Fg)对体外低糖低氧(low glucose and oxygen,LGO)损伤神经元轴突再生的影响及机制。方法取出生后24 h内乳鼠皮层神经元进行原代培养,通过免疫荧光染色检测MAP-2鉴定神经元。将细胞分为3组,即LGO组、LGO+Fg组、LGO+Fg+PD168393组。3组均予以低糖低氧处理48 h后,LGO+Fg组加入Fg,LGO+Fg+PD168393组加入Fg、表皮生长因子受体(epidermal growth factor receptor,EGFR)抑制剂(PD168393),均处理细胞24 h。采用免疫荧光染色检测β-tubulin III(Tuj-1)表达,评价轴突生长;Western blot检测EGFR、p-EGFR在细胞中的表达;实时荧光定量PCR检测EGFR下游调控蛋白(MAP-1B,CRMP-2)mRNA表达。结果与LGO组比较,LGO+Fg组轴突生长明显受到抑制,轴突长度较短、数量减少,p-EGFR蛋白表达显著增加,MAP-1B、CRMP-2 mRNA表达下调;与LGO+Fg组比较,LGO+Fg+PD168393组轴突长度增加、数量明显增多,p-EGFR蛋白表达显著减少,MAP-1B、CRMP-2 mRNA表达上调。结论Fg可抑制低糖低氧损伤神经元的轴突再生能力,其机制与激活EGFR信号通路有关。Objective To investigate the influence and mechanisms of fibrinogen(Fg)on axonal regeneration of neurons under in vitro low glucose and oxygen(LGO)conditions.Methods Primary cultures of cortical neurons from neonatal rats within 24 h of birth were established,and immunofluorescence staining for MAP-2 was conducted to identify neurons.The cells were divided into three groups:LGO group,LGO+Fg group,and LGO+Fg+PD168393 group.All groups underwent low glucose and oxygen treatment for 48 h.In the LGO+Fg group,Fg was added,and in the LGO+Fg+PD168393 group,both Fg and the epidermal growth factor receptor(EGFR)inhibitor PD168393 were added.Cells were treated for an additional 24 h.β-tubulin III(Tuj-1)immunofluorescence staining was performed to assess axonal growth.Western blot analysis was conducted to measure the expression of EGFR and phosphorylated-EGFR(p-EGFR)in the cells.Real-time fluorescence quantitative PCR was used to detect mRNA expression of EGFR downstream regulatory proteins(MAP-1B,CRMP-2).Results In comparison to the LGO group,the LGO+Fg group exhibited significantly inhibited axonal growth,characterized by shorter lengths and reduced quantity of axons.The p-EGFR protein expression markedly increased,and the mRNA expression of MAP-1B and CRMP-2 decreased.When compared to the LGO+Fg group,the LGO+Fg+PD168393 group showed increased axonal length and a significant increase in the quantity of axons.The p-EGFR protein expression significantly decreased,and the mRNA expression of MAP-1B and CRMP-2 increased.Conclusion Fibrinogen can suppress the axonal regeneration capacity of neurons damaged under low glucose and oxygen conditions,and this mechanism is associated with the activation of the EGFR signaling pathway.

关 键 词：纤维蛋白原 皮层神经元 低糖低氧 轴突再生 表皮生长因子受体 

分 类 号：Q28[生物学—细胞生物学]