长链非编码RNA TALNEC2通过EZH2/Nrf2/HO-1信号通路调控急性脑梗死损伤的研究  

LncRNA TALNEC2 regulates acute cerebral infarction injury through EZH2/Nrf2/HO-1 signaling pathway

作　　者：卢威 雷明 LU Wei;LEI Ming(Department of Intensive Care Medicine,Yangtze River Shipping General Hospital,Wuhan,Hubei 430000,China;Department of Neurology,Yangtze River Shipping General Hospital,Wuhan,Hubei 430000,China)

机构地区：[1]长江航运总医院重症医学科,湖北武汉430000 [2]长江航运总医院神经内科,湖北武汉430000

出　　处：《安徽医药》2024年第12期2483-2488,共6页Anhui Medical and Pharmaceutical Journal

基　　金：武汉市卫生健康委2020年度医学科研项目(WX20Z41)。

摘　　要：目的分析长链非编码RNA(lncRNA)TALNEC2在急性脑梗死体外模型缺氧-葡萄糖剥夺(OGD)模型中的分子调控机制。方法2021年1月至2022年6月,选用小鼠小胶质细胞系BV2建立OGD细胞模型,分为Blank组(未进行任何处理)、OGD组、OGD+si-NC组[转染阴性对照(si-NC)后进行OGD处理]、OGD+si-TALNEC2组(转染si-TALNEC2后进行OGD处理)。实时实时荧光定量逆转录聚合酶链反应(qRT-PCR)实验检测TALNEC2的表达;细胞计数试剂盒(CCK-8)、流式细胞术、酶联免疫吸附测定(ELISA)分别检测细胞活力、凋亡率及炎症因子白细胞介素(IL)-1β、IL-18和肿瘤坏死因子α(TNF-α)的变化;蛋白质印迹法检测核苷酸结合结构域富含亮氨酸重复序列和含热蛋白结构域受体3(NLRP3)、凋亡相关斑点样蛋白(ASC)、活化胱天蛋白酶1(cleaved caspase-1)、核因子E2相关因子2(Nrf2)、血红素氧化酶1(HO-1)、Zeste同源物增强子2(EZH2)蛋白表达。RNA pull-down实验及RNA结合蛋白免疫沉淀(RIP)实验证实TALNEC2与EZH2的结合。结果与Blank组[1.02±0.03、0.81±0.05、1.29±0.06、1.52±0.08、(4.72±1.09)%]相比,OGD组细胞TALNEC2表达(4.18±0.15)升高,在24 h(0.46±0.05)、48 h(0.61±0.05)、72 h(0.75±0.02)的活力下降,凋亡率(25.63±4.28)%升高(P<0.05);与OGD组相比,OGD+si-TALNEC2组细胞TALNEC2表达(1.16±0.09)减少,在24 h(0.65±0.07)、48 h(0.98±0.05)、72 h(1.13±0.07)的活力升高,凋亡率(7.25±1.93)%减少(P<0.05)。与Blank组[1.05±0.07、1.04±0.05、1.01±0.05、1.02±0.04、1.04±0.07、(47.21±5.01)ng/L、(38.26±4.13)ng/L、(48.22±7.15)ng/L、1.06±0.03、1.03±0.06]相比,OGD组细胞中NLRP3(2.69±0.15)、ASC(3.11±0.18)、cleaved caspase-1(2.51±0.12)、胞质Nrf2(2.91±0.13)、EZH2(3.12±0.18)蛋白表达及炎症因子TNF-α(157.22±13.18)ng/L、IL-1β(177.43±15.15)ng/L、IL-18(210.63±19.08)ng/L水平升高,胞核Nrf2(0.37±0.03)、HO-1(0.27±0.04)蛋白表达减少(P<0.05);与OGD组相比,OGD+si-TALNEC2组细胞中NLRP3(1.33±0.11)、ASCObjective To analyze the molecular regulatory mechanism of tumor-associated long non-coding RNA expressed on chromosome 2(TALNEC2)of long non-coding RNA(lncRNA)in the hypoxic-glucose deprivation model(OGD)of acute cerebral infarction in vitro.Methods The study was conducted from January 2021 to June 2022.Mouse microglia cell line BV2 was selected to establish the OGD cell model,they were divided into Blank group(without any treatment),OGD group,OGD+si-NC group(transfected with si-NC for OGD treatment),and OGD+si-TALNEC2 group(transfected with si-TALNEC2 for OGD treatment).Real-time quantitative fluorescent PCR(qRT-PCR)was used to detect the expression of TALNEC2.Cell count kit(CCK-8),flow cytometry and enzyme-linked immunosorbent assay(ELISA)were used to detect the changes of cell viability,apoptosis rate and inflammatory cytokines interleukin(IL)-1β,IL-18 and tumor necrosis factorα(TNF-α),respectively;Western blotting was used to detect proteins expression of NLR family Pyrindomain protein 3(NLRP3),apoptosis-associated speck-like protein(ASC),cleaved caspase-1,nuclear factor E2-related factor 2(Nrf2),heme oxidase 1(HO-1),and Zeste homolog enhancer 2(EZH2);The binding of TALNEC2 to EZH2 was confirmed by RNA pull-down assay and RNA binding protein immunoprecipitation(RIP)assay.Results Compared with Blank group[1.02±0.03,0.81±0.05,1.29±0.06,1.52±0.08,(4.72±1.09)%],the expression of TALNEC2 in OGD group was increased(4.18±0.15),the cell activity was decreased at 24 h(0.46±0.05),48 h(0.61±0.05),72 h(0.75±0.02),and the apoptosis rate was increased(25.63±4.28)%(P<0.05);Compared with OGD group,the expression of TALNEC2 in OGD+si-TALNEC2 group decreased(1.16±0.09),the cell activity was increased at 24 h(0.65±0.07),48 h(0.98±0.05)and 72 h(1.13±0.07),and the apoptosis rate was decreased(7.25±1.93)%(P<0.05).Compared with the Blank group[1.05±0.07,1.04±0.05,1.01±0.05,1.02±0.04,1.04±0.07,(47.21±5.01)ng/L,(38.26±4.13)ng/L,(48.22±7.15)ng/L,1.06±0.03,1.03±0.06],in the OGD group,the protein expressions

关 键 词：脑梗死 长链非编码RNA TALNEC2 缺氧-葡萄糖剥夺模型 小胶质细胞 炎症因子 

分 类 号：R743.33[医药卫生—神经病学与精神病学]