黄芩苷通过上调miR-190表达缓解缺氧缺糖对神经细胞损伤的研究  

Baicalin alleviates damage of nerve cells caused by hypoxia and hypoglycemia by up-regulating expression of miR-190

作　　者：吴艳荣[1] 刘光伟[2] 刘全忠[2] 朱志强[1] 王琳琳[1] 郑颖颖[1] 李昕[3] WU Yan-rong;LIU Guang-wei;LIU Quan-zhong;ZHU Zhi-qiang;WANG Lin-lin;ZHENG Ying-ying;LI Xin(Department of Acupunture&Moxibustion,The Second Affiliated Hospital of Zhengzhou University,Zhengzhou 450003,China;Department of Gastroenterology,The First Affiliated Hospital of Henan University of Chinese Medicine,Zhengzhou 450004,China;Department of Neurology,The Second Affiliated Hospital of Zhengzhou University,Zhengzhou 450003,China)

机构地区：[1]郑州大学第二附属医院针灸科,河南郑州450003 [2]河南中医药大学第一附属医院消化内科,河南郑州450004 [3]郑州大学第二附属医院神经内科,河南郑州450003

出　　处：《中草药》2021年第10期3009-3017,共9页Chinese Traditional and Herbal Drugs

基　　金：国家中医药管理局国家中医临床研究基地业务建设科研专项(ZDZX20120660);河南省中医药科学研究专项(2018JDZX109)。

摘　　要：目的探讨黄芩苷对缺氧缺糖(oxygen-glucosedeprivation,OGD)诱导的神经元细胞增殖、凋亡、炎症、氧化应激的影响及其对miR-190的调控作用。方法采用OGD处理小鼠海马神经元HT22细胞建立细胞损伤模型,使用不同剂量(10、30mg/L)的黄芩苷处理HT22细胞,分别将miR-NC、miR-190mimics转染至HT22细胞后进行OGD处理(OGD+miR-NC组、OGD+miR-190组);分别将anti-miR-NC、anti-miR-190转染至HT22细胞后使用黄芩苷处理24h,随后进行OGD处理(OGD+黄芩苷+anti-miR-NC组、OGD+黄芩苷+anti-miR-190组);采用MTT法检测细胞增殖情况;采用流式细胞术检测细胞凋亡率;采用ELISA法检测白细胞介素-1β(interleukin-1β,IL-1β)、肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)的水平;采用2,4-二硝基苯肼显色法检测乳酸脱氢酶(lactate dehydrogenase,LDH)的活性;采用黄嘌呤氧化酶法检测超氧化物歧化酶(superoxide dismutase,SOD)的含量;采用qRT-PCR法检测miR-190的表达量;Western blotting法检测细胞周期蛋白1(Cyclin D1)、未剪切的半胱天冬酶-3(pro-cysteinyl aspartate-specific protease-3,pro-Caspase-3)、p21、活化的含半胱氨酸的天冬氨酸蛋白水解酶3(cleaved-cysteinylaspartate-specificprotease-3,cleaved-Caspase-3)蛋白表达量。结果OGD可明显降低HT22细胞活力及CyclinD1、pro-Caspase-3蛋白水平(P<0.05),升高细胞凋亡率及p21、cleavedCaspase-3蛋白水平(P<0.05),并可提高IL-1β、TNF-α的水平及LDH的活性(P<0.05),降低SOD的含量(P<0.05);黄芩苷可明显提高OGD诱导的HT22细胞活力及CyclinD1、pro-Caspase-3蛋白水平(P<0.05),降低细胞凋亡率及p21、cleaved-Caspase-3蛋白水平(P<0.05),并可降低IL-1β、TNF-α的水平及LDH的活性(P<0.05),升高SOD的含量(P<0.05);OGD可明显降低HT22细胞中miR-190的表达水平(P<0.05),而黄芩苷可明显升高miR-190的表达水平(P<0.05);miR-190过表达对OGD诱导的HT22细胞增殖、凋亡、炎症及氧化应激的作用与黄芩苷的作用相似;抑制miR-Objective To explore the effects of baicalin on the proliferation,apoptosis,inflammation,and oxidative stress of neuronal cells induced by oxygen-glucose deprivation(OGD)and its regulation on miR-190.Methods OGD was used to treat mouse hippocampal neuronal cells HT22 to establish a cell injury model,and different doses(10,30 mg/L)of baicalin was used to treat the cells.HT22 cells were transfected with miR-NC or miR-190 mimics and then treated with OGD(OGD+miR-NC group,OGD+mi R-190 group).HT22 cells were transfected with anti-miR-NC and anti-miR-190 and treated with baicalin for 24 h,followed by OGD treatment(OGD+baicalin high-dose group+anti-miR-NC group,OGD+baicalin high-dose group+anti-miR-190 group).The MTT method and flow cytometry were used to detect cell proliferation and apoptosis rate,respectively.The levels of IL-1βand TNF-αwere detected by ELISA.The 2,4-dinitrophenylhydrazine color method was performed to assess the activity of lactate dehydrogenase(LDH).Xanthine oxidase method was used to detect the content of SOD.The expression of miR-190 was determined by qRT-PCR.And the protein expression of CyclinD1,pro-Caspase-3,p21,and cleaved-Caspase-3 was detected by Western blot.Results OGD significantly reduced HT22 cell viability and the protein levels of CyclinD1 and pro-Caspase-3(P<0.05),increased cell apoptosis rate and the protein levels of p21 and cleaved-Caspase-3(P<0.05),as well as increased the levels of IL-1β,TNF-αand the activity of LDH(P<0.05),reduced the content of SOD(P<0.05).Baicalin could significantly increase OGD-induced HT22 cell viability and the protein levels of CyclinD1 and pro-Caspase-3(P<0.05),reduce cell apoptosis rate and the protein levels of p21 and Cleaved-caspase3(P<0.05),and reduce the levels of IL-1β,TNF-αand the activity of LDH(P<0.05),and increase the content of SOD(P<0.05).OGD significantly reduced the expression level of miR-190 in HT22 cells(P<0.05),while baicalin significantly increased the expression level of miR-190(P<0.05).The effects of miR-190 overexpression o

关 键 词：黄芩苷 miR-190 缺氧缺糖 神经元细胞 增殖 凋亡 

分 类 号：R285.5[医药卫生—中药学]