miR-199b-5p靶向调控Sirt 1对氧糖剥夺/复糖复氧小鼠海马神经元HT22细胞增殖、凋亡的影响  

The Effect of miR-199b-5p Targeting Regulation of Sirt1 on the Proliferation and Apoptosis of OGD/R Mouse Hippocampal Neurons HT22 Cells

作　　者：王超奇 隋小芳[1] 陈尚君[1] WANG Chao-qi;SUI Xiao-fang;CHEN Shang-jun(Department of Geriatrics,the First Afiliated Hospital of Jiamusi University,Jiamusi 154000,China)

机构地区：[1]佳木斯大学附属第一医院老年病科,佳木斯154000

出　　处：《中国临床神经科学》2025年第1期19-28,共10页Chinese Journal of Clinical Neurosciences

摘　　要：目的研究miR-199b-5p对氧糖剥夺(OGD)/复糖复氧(R)后小鼠海马神经元HT22细胞增殖、凋亡的影响及机制.方法①收集2022年10月至2023年10月收治的24例急性缺血性脑卒中(AIS)患者作为AIS组,另选取同期24例行健康体检者作为对照组.使用qRT-PCR检测两组血清miR-199b-5p表达水平.②体外培养小鼠海马神经元HT22细胞并进行OGD/R处理,按细胞实验分组,分为对照组与OGD/R组,检测两组小鼠miR-199b-5p表达水平.③HT22细胞转染miR-199b-5p模拟物、抑制物及阴性对照后进行OGD/R处理,根据是否转染miRNA分为对照组、OGD/R+NC组、OGD/R+miR-199b-5p模拟组和OGD/R+miR-199b-5p抑制组,用CCK8、EdU实验检测细胞增殖率;流式细胞术检测细胞凋亡率;Western blot检测cleaved-caspase-3蛋白表达水平.④通过生物信息学预测软件MiR Walk筛选miR-199b-5p的下游靶基因Sirt1,检测miR-199b-5p对其表达的影响,在HT22细胞中转染Sirt 1过表达质粒,共转染miR-199b-5p后进行回复实验验证miR-199b-5p通过靶向Sirt1调控OGD/R后小鼠海马神经元HT22细胞增殖、凋亡.结果①AIS组血清miR-199b-5p表达较对照组升高(P<0.05);②与对照组比较,OGD/R组miR-199b-5p表达升高(P<0.05)、细胞增殖能力降低、细胞凋亡率及cleaved-caspase-3蛋白表达增高(均P<0.05);③与OGD/R+NC组比较,OGD/R+miR-199b-5p模拟组miR-199b-5p表达水平升高(P<0.05)、细胞增殖能力下降、细胞凋亡率及cleaved-caspase-3蛋白表达增高(均P<0.05),OGD/R+miR-199b-5p抑制组则呈相反表现(P<0.05);④生物信息学预测软件检测发现miR-199b-5p与Sirt1存在互补配对序列,miR-199b-5p负向调控Sirt1表达(P<0.05);OGD/R诱导的HT22细胞中,过表达Sirt 1可以逆转miR-199b-5p上调所引起的细胞增殖能力受损和凋亡增加.结论miRNA-199b-5p可能通过靶向调控Sirt 1影响OGD/R导致的神经元增殖和凋亡.Aim To investigate the effects and mechanisms of miR-199b-5p on the proliferation and apoptosis of mouse hippocampal HT22 neurons after oxygen-glucose deprivation/reoxygenation(OGD/R).Methods①Twenty-four patients with acute ischemic stroke(AIS)treated from October 2022 to October 2023 were enrolled as the AIS group,and 24 healthy individuals were selected as the control group.Serum miR-199b-5p expression was measured by qRT-PCR.②Mouse hippocampal neurons(HT22 cells)were cultured and subjected to OGD/R treatment,then divided into a control and an OGD/R group.miR-199b-5p expression was detected using qRT-PCR.③HT22 cells were transfected with miR-199b-5p mimics,inhibitors,and negative controls,followed by OGD/R treatment,forming the control,OGD/R+NC,OGD/R+miR-199b-5p mimics,and OGD/R+miR-199b-5p inhibitor groups.Cell proliferation was assessed by CCK8 and EdU assays,apoptosis rates were analyzed by flow cytometry,and cleaved-caspase-3 protein levels were detected by Western blot.④Bioinformatics(MiR Walk)predicted Sirt 1 as a target of miR-199b-5p,and its expression was evaluated.Rescue experiments with Sirt1 overexpression plasmids in HT22 cells were conducted to confirm the regulatory effect of miR-199b-5p on cell proliferation and apoptosis through Sirt1.Results①miR-199b-5p expression in AIS patients was significantly higher than that in the control(P<0.05);②Compared with the control group,the miR-199b-5p expression was increased in OGD/R group(P<0.05),cell proliferation capacity was decreased,and apoptosis rate and cleaved-caspase-3 expression were increased(all P<0.05);③The OGD/R+miR-199b-5p mimics group showed increased miR-199b-5p expression,reduced proliferation,and increased apoptosis and cleaved-caspase-3 levels,whereas the OGD/R+miR-199b-5p inhibitor group exhibited the opposite trends(P<0.05);④Bioinformatics revealed that miR-199b-5p has a complementary pairing sequence with Sirt 1,and miR-199b-5p negatively regulated Sirt 1(P<0.05).Sirt 1 overexpression could reverse proliferation impairm

关 键 词：miR-199b-5p 沉默信息调节因子 氧糖剥夺/复糖复氧 海马神经元 HT22细胞 增殖 凋亡 

分 类 号：R743[医药卫生—神经病学与精神病学] Q189[医药卫生—临床医学]