Homer1b/c通过内质网功能调节由谷氨酸兴奋性毒性损伤诱发的小鼠海马神经元HT22细胞自噬  

Homer1b/c regulates autophagy in mouse hippocampal neuronal HT22 cells induced by glutamate excitotoxic injury through endoplasmic reticulum function

作　　者：张敏敏[1] 朱宣[1] 沈红健[1] 吕楠[1] 吴雄枫 徐小龙 吴涛[1] ZHANG Minmin;ZHU Xuan;SHEN Hongjian;LÜNan;WU Xiongfeng;XU Xiaolong;WU Tao(Neurovascular Center,The First Affiliated Hospital of Naval Medical University(Second Military Medical University),Shanghai 200433,China)

机构地区：[1]海军军医大学(第二军医大学)第一附属医院脑血管病中心,上海200433

出　　处：《海军军医大学学报》2024年第3期277-283,共7页Academic Journal of Naval Medical University

摘　　要：目的研究Homer1b/c蛋白在谷氨酸兴奋性毒性损伤诱发的细胞自噬中的作用及机制。方法选用小鼠海马神经元HT22细胞,通过500μmol/L L-谷氨酸处理建立细胞损伤模型。用siRNA慢病毒转染方式下调Homer1b/c表达,用10μmol/L钙离子螯合剂BAPTA-AM、10 mmol/L内质网应激抑制剂4-PBA分别抑制细胞内钙离子释放和内质网应激后,使用蛋白质印迹法检测细胞中Homer1b/c蛋白,自噬效应蛋白[beclin-1、微管相关蛋白1轻链3(LC3)]及内质网应激标志蛋白[C/EBP同源蛋白(CHOP)、葡萄糖调节蛋白78(GRP 78)]的表达水平。结果L-谷氨酸处理HT22细胞12 h后,细胞中beclin-1表达和LC3-Ⅱ/LC3-Ⅰ比值均较对照组升高(均P<0.05);与转染对照组相比,下调Homer1b/c表达可降低细胞中beclin-1表达和LC3-Ⅱ/LC3-Ⅰ比值(均P<0.05);抑制细胞内钙离子释放和内质网应激均能降低细胞中beclin-1表达和LC3-Ⅱ/LC3-Ⅰ比值(均P<0.05);下调Homer1b/c表达后,抑制细胞内钙离子释放和内质网应激未能进一步降低细胞中beclin-1表达和LC3-Ⅱ/LC3-Ⅰ比值。结论Homer1b/c能够调节谷氨酸兴奋性毒性损伤诱发的细胞自噬,其调节作用可能与内质网功能有关。Objective To study the role of Homer1b/c in cell autophagy induced by glutamate excitotoxic injury and its mechanism.Methods Mouse hippocampal neuronal HT22 cells were treated with 500μmol/L L-glutamate to establish cell injury model.The expression of Homer1b/c in HT22 cells was down-regulated by small interfering RNA(siRNA)lentivirus transfection.10μmol/L 1,2-bis(2-aminophenoxy)ethane-N,N,N’,N’-tetraacetic acid tetrakis(acetoxymethyl ester)(BAPTA-AM,a calcium ion chelating agent)and 10 mmol/L 4-phenylbutyric acid(4-PBA,an endoplasmic reticulum stress inhibitor)were used to inhibit intracellular calcium ion release and endoplasmic reticulum stress,respectively.Then,the expression levels of Homer1b/c,autophagy proteins(beclin-1 and microtubule-associated protein 1 light chain 3[LC3])and endoplasmic reticulum stress marker proteins(C/EBP homologous protein[CHOP]and glucose regulated protein 78[GRP-78])in cells were measured by Western blotting.Results Compared with the control group,the expression of beclin-1 and the ratio of LC3-Ⅱ/LC3-Ⅰwere significantly increased after the HT22 cells were treated with L-glutamate for 12 h(both P<0.05);down-regulation of Homer1b/c expression could reduce the expression of beclin-1 and the ratio of LC3-Ⅱ/LC3-Ⅰ(both P<0.05).Inhibition of intracellular calcium release and endoplasmic reticulum stress could reduce the beclin-1 expression and LC3-Ⅱ/LC3-Ⅰratio(both P<0.05).After down-regulation of Homer1b/c expression,inhibiting intracellular calcium release and endoplasmic reticulum stress failed to further reduce the beclin-1 expression and LC3-Ⅱ/LC3-Ⅰratio.Conclusion Homer1b/c can regulate cell autophagy induced by glutamate excitotoxic injury,and its regulatory effects may be related to the function of endoplasmic reticulum.

关 键 词：海马神经元 自噬 HOMER 钙稳态 内质网应激 兴奋性损伤 谷氨酸 

分 类 号：R741[医药卫生—神经病学与精神病学]