miR-301a-3p通过靶向调节SNX27影响脊髓神经损伤修复  

miR-301a-3p modulates repair of spinal cord nerve injury through targeting SNX27

作　　者：史纪元[1] 姬乐[1] 易智[1] 刘时璋[1] 刘慧通[1] 王雄勋 Shi Jiyuan;Ji Le;Yi Zhi;Liu Shizhang;Liu Huitong;Wang Xiongxun(Department of Orthopedics,Shanxi Provincial People′s Hospital,Xi’an 710068,Shanxi,China;Department of Orthopedics,Xian International Medical Center Hospital,Xi’an 710100,Shanxi,China)

机构地区：[1]陕西省人民医院骨科,陕西西安710068 [2]西安市国际医学中心医院骨科,陕西西安710100

出　　处：《贵州医药》2021年第6期849-852,F0003,共5页Guizhou Medical Journal

基　　金：陕西省重点研发计划(S2017-ZDYF-YBXM-SF-0595);陕西省人民医院科技发展孵化项目(2020YXM-20)。

摘　　要：目的研究miR-301a-3p在脊髓损伤组织和神经细胞中的表达情况及其在神经损伤修复过程中的作用。方法建立SD大鼠脊髓损伤(SCI)模型,用qPCR和Western blot检测miR-301a-3p和分类连接蛋白(SNX27)的表达量;利用150μM H_(2)O_(2)诱导PC12神经细胞损伤;采用流式细胞术和Western blotting检测细胞凋亡以及凋亡相关蛋白和轴突相关蛋白的表达量,双荧光素酶报告基因分析验证miR-301a-3p对SNX27的相互作用关系。结果损伤脊髓组织中miR-301a-3p的表达下调,与SNX27呈现相反的表达模式。将miR-301a-3p mimic或miR-301a-3p inhibitor转染至PC12细胞中能够明显抑制或促进H_(2)O_(2)诱导的神经细胞凋亡,并促进或抑制神经轴突相关蛋白Gap-43和NF-200的表达。运用生物信息学分析、双荧光素酶报告基因实验、qPCR和Western blot证实SNX27是miR-301a-3p的靶基因。将腺病毒SNX27过表达载体(Ad-SNX27)转染至PC12细胞可拮抗miR-301a-3p对H_(2)O_(2)处理后细胞凋亡的抑制作用,以及神经轴突再生能力的促进作用。结论本研究表明miR-301a-3p可通过抑制SNX27的表达促进SCI后的神经功能修复,提示miR-301a-3p可能是脊髓损伤治疗的潜在靶点。Objective To explore the expression of miR-301a-3p in spinal cord injury(SCI)tissues and nerve cells and its role in the repair process of nerve injury.Methods The spinal cord injury(SCI)model of SD rats was established.qPCR and Western blotting were used to detect expression of miR-301a-3p and the Sorting nexin 27(SNX27).PC12 cells were treated with 150μM H_(2)O_(2) to establish injury model.Cell apoptosis and expression levels of apoptosis-related proteins and axon-related proteins were determined by Flow cytometry and Western blot(WB).Dual-luciferase reporter gene assay was used to verify the interaction between miR-301a-3p and SNX27.Results The expression of miR-301a-3p was down-regulated in injured spinal cord tissue,and SNX27 showed the opposite expression pattern.Transfection of miR-301a-3p mimic or miR-301a-3p inhibitor into PC12 cells can significantly inhibit or promote H_(2)O_(2)-induced cell apoptosis,and increase or decrease the expression levels of Gap-43 and NF-200.Bioinformatics analysis,dual-luciferase reporter gene assay,qPCR and Western blot assays showed that SNX27 was a target gene of miR-301a-3p.Over expression of SNX27 by transfection with adenovirus SNX27 over expression vector(Ad-SNX27)into PC12 cells antagonized the effects of miR-301a-3p on apoptosis and axonal regeneration.Conclusion This study demonstrates that miR-301a-3p can promote neurofunctional?repair after spinal cord injury through inhibiting the expression of SNX27,suggesting that miR-301a-3p may function as a potential target for the SCI treatment.

关 键 词：脊髓损伤(SCI) miR-301a-3p SNX27 细胞凋亡 轴突再生 

分 类 号：R745.4[医药卫生—神经病学与精神病学]