miR-675通过靶向HIF-1α调控线粒体自噬在脑出血大鼠中神经保护作用  

miR-675 regulating neuroprotective effect of mitochondrial autophagy in rats with cerebral hemorrhage via targeting HIF-1α

作　　者：陈月阳 诸靖 王诗波 虞大为 陈涛 王忠祥 张烨 CHEN Yue-yang;ZHU Jing;WANG Shi-bo;YU DA-wei;CHEN Tao;WANG Zhong-xiang;ZHANG Ye(The 904 Hospital of the Joint Logistics Support Force of the Chinese People’s Liberation Army,Wuxi 214000,Jiangsu,China)

机构地区：[1]中国人民解放军联勤保障部队第九〇四医院,江苏无锡214000

出　　处：《生物医学工程与临床》2025年第2期135-141,共7页Biomedical Engineering and Clinical Medicine

基　　金：江苏省自然科学基金资助项目(BK20211044)。

摘　　要：目的探讨miR-675通过靶向缺氧诱导因子(HIF)-1α表达,进而调控线粒体自噬途径发挥脑出血大鼠的神经保护作用。方法选择健康8周龄雄性Wistar大鼠60只,平均体质量为275 g。随机分为假手术组、模型组、miR-675过表达组和miR-675低表达组,每组15只。采用Ⅶ型胶原酶定向注射法复制脑出血模型;miR-675过表达组和miR-675低表达组分别采用miR-675过表达和低表达慢病毒载体200 pmol/(kg·d)尾静脉注射3 d,假手术组和模型组采用等量0.9%氯化钠溶液(生理盐水)注射。实时荧光定量聚合酶链式反应(PCR)检测脑组织miR-675和HIF-1αmRNA表达量,改良神经功能损伤程度评分(mNSS)评估神经功能,Western blot检测人重组自噬效应蛋白(Beclin 1)、微管相关蛋白1轻链3(LC3)Ⅱ、磷脂酶和张力蛋白同源物诱导激酶(Pink-1)表达量,原位末端标记法检测细胞凋亡率。双荧光素酶报告基因实验探索miR-675和HIF-1αmRNA之间的靶向关系。结果假手术组、模型组、miR-675过表达组和miR-675低表达组miR-675表达量分别为1.003±0.004、1.003±0.004、1.502±0.103和0.412±0.004,即模型组低于假手术组,miR-675过表达组高于模型组,而miR-675低表达组低于模型组(F=12.326,P<0.001)。假手术组、模型组、miR-675过表达组和miR-675低表达组HIF-1αmRNA表达量分别为1.004±0.003、3.236±0.232、1.925±0.114和3.898±0.301,即模型组高于假手术组,miR-675过表达组低于模型组,而miR-675低表达组高于模型组(F=15.234,P<0.001)。假手术组、模型组、miR-675过表达组和miR-675低表达组mNSS分别为(0.5±0.1)分、(12.5±2.3)分、(7.5±1.2)分和(16.3±2.9)分,即模型组高于假手术组,miR-675过表达组低于模型组,而miR-675低表达组高于模型组(F=35.236,P<0.001)。假手术组、模型组、miR-675过表达组和miR-675低表达组Beclin 1蛋白表达量分别为1.003±0.003、2.756±0.326、1.759±0.125和3.126±0.421,即模型组高于假手术组,miR-675过�Objective To investigate the neuroprotective effect of miR-675 on cerebral hemorrhage rats via targeting expression of hypoxia-inducing factor(HIF-1α)and regulating mitochondrial autophagy pathway.Methods A total of 60 healthy 8-week-old male Wistar rats with mean body mass of 275 g were sacrificed.All of the rats were randomly divided into sham operation group,model group,miR-675 overexpression group and miR-675 low expression group,with 15 rats in each group.The cerebral hemorrhage model was replicated by typeⅦcollagenase directional injection;miR-675 overexpression group and low expression group were injected with miR-675 overexpression and low expression lentiviral vector 200 pmol/(kg·d)via tail vein for 3 days,respectively;sham operation group and model group were injected with the same amount of 0.9%sodium chloride solution(normal saline).The real-time fluorescence quantitative polymerase chain reaction(PCR)was used to detect expressions of miR-675 and HIF-1αmRNA in brain tissue,modified neurological severity score(mNSS)was used to evaluate neurological function.Western blot was used to detect the expressions of human recombinant autophagy effector protein(Beclin 1),microtubule-associated protein 1 light chain 3(LC3)II,phospholipase and tension protein homolog-induced kinase(Pink-1).The apoptosis rate was detected by in situ end labeling.The dual luciferase reporter gene assay was used to explore the targeting relationship between miR-675 and HIF-1αmRNA.Results The expression levels of miR-675 in sham operation group,model group,miR-675 overexpression group and miR-675 low expression group were 1.003±0.004,1.003±0.004,1.502±0.103 and 0.412±0.004,respectively.The expression level in model group was lower than that in sham operation group,the level in miR-675 overexpression group was higher than that in model group,and the level in miR-675 low expression group was lower than that in model group(F=12.326,P<0.001).The expression levels of HIF-1αmRNA in sham operation group,model group,miR-675 overexp

关 键 词：大鼠 脑出血 神经保护 miR-675 缺氧诱导因子-1Α 线粒体自噬 细胞凋亡 靶向调控 

分 类 号：R743.34[医药卫生—神经病学与精神病学] R363.2[医药卫生—临床医学] R-33