长链非编码RNA脑源性神经营养因子反义RNA靶向微小RNA-495-3p调控脂多糖诱导的大鼠肺泡巨噬细胞凋亡和炎症反应  

LncRNA BDNF-AS targets miR-495-3p to regulate LPS-induced apoptosis and inflammatory response in rat alveolar macrophages

作　　者：高炜 张莉 金培田 GAO Wei;ZHANG Li;JIN Peitian(Pediatric Internal Medicine,Zaozhuang Mining Group Central Hospital,Zaozhuang,Shandong 277000,China;Geriatrics,Zaozhuang Mining Group Central Hospital,Zaozhuang,Shandong 277000,China)

机构地区：[1]枣庄矿业集团中心医院小儿内科,山东枣庄277000 [2]枣庄矿业集团中心医院老年科,山东枣庄277000

出　　处：《安徽医药》2022年第6期1235-1239,共5页Anhui Medical and Pharmaceutical Journal

摘　　要：目的探讨长链非编码RNA(lncRNA)脑源性神经营养因子反义RNA(BDNF-AS)对脂多糖(LPS)诱导的大鼠肺泡巨噬细胞凋亡和炎症反应的影响及分子机制。方法该研究起止时间为2018年12月至2019年12月。用1 mg/L的LPS处理大鼠肺泡巨噬细胞NR8383细胞作为LPS组;正常培养的细胞作为Con组。将BDNF-AS小分子干扰RNA(si-BDNF-AS)及其阴性对照(si-NC)、微小RNA(miR)-495-3p及其阴性对照(miR-NC)转染至NR8383细胞中再用1 mg/L的LPS处理,记为LPS+siBDNF-AS组、LPS+si-NC组、LPS+miR-495-3p组、LPS+miR-NC组;将si-BDNF-AS分别与anti-miR-NC、anti-miR-495-3p共转染至NR8383细胞中再用1 mg/L的LPS处理,记为LPS+si-BDNF-AS+anti-miR-NC组、LPS+si-BDNF-AS+anti-miR-495-3p组。实时荧光定量PCR检测lncRNA BDNF-AS和miR-495-3p的表达水平;酶联免疫吸附测定(ELISA)法检测白细胞介素-1β(IL-1β)、肿瘤坏死因子-α(TNF-α)水平;流式细胞术检测细胞凋亡;蛋白质印迹法检测B细胞淋巴瘤-2(Bcl-2)、Bcl-2相关X蛋白(Bax)表达;荧光素酶报告实验检测BDNF-AS对miR-495-3p的靶向关系。结果LPS诱导的大鼠肺泡巨噬细胞中BDNF-AS高表达[(1.00±0.06)比(3.41±0.31)],miR-495-3p低表达[(1.02±0.07)比(0.47±0.04)],IL-1β[(22.14±2.25)ng/L比(513.20±41.22)ng/L]、TNF-α[(184.33±18.65)ng/L比(1125.65±110.36)ng/L]水平升高,细胞凋亡率[(8.11±0.81)%比(36.22±3.21)%]升高,Bax表达水平升高,Bcl-2表达水平降低(P<0.05)。抑制BDNF-AS表达或过表达miR-495-3p后,IL-1β[(526.14±46.87)ng/L比(132.41±12.58)ng/L、(536.14±51.02)ng/L比(175.98±18.41)ng/L]、TNF-α[(1203.33±185.22)ng/L比(354.26±21.58)ng/L、(1248.65±115.28)ng/L比(628.47±53.69)ng/L]水平降低,细胞凋亡率[(38.14±3.71)%比(12.03±1.25)%、(36.98±3.61)%比(14.87±1.42)%]降低,Bax表达水平降低,Bcl-2表达水平升高(P<0.05)。BDNF-AS靶向调控miR-495-3p,抑制miR-495-3p表达逆转了抑制lncRNA BDNF-AS表达对LPS诱导的大鼠肺泡巨噬细胞凋亡和炎症反应的抑制作用。�Objective To investigate the effect of lncRNA BDNF-AS on lipopolysaccharide(LPS)induced alveolar macrophage apoptosis and inflammatory response in rats and its molecular mechanism.Methods The study started and ended from December 2018 to December 2019.Rat alveolar macrophage NR8383 cells were treated with 1 mg/L LPS as the LPS group.Normal cultured cells served as the Con group.BDNF-AS small interfering RNA(si-BDNF-AS)and its negative control(si-NC),miR-495-3p oligonucleotide mimics(miR-495-3p mimics)and negative control mimic NC sequences(miR-NC)was transfected into NR8383 cells,which were then treated with 1 mg/L LPS,and recorded as LPS+si-NC group,LPS+si-BDNF-AS group,LPS+miR-NC group and LPS+miR-495-3p group;si-BDNF-AS was co-transfected with anti-miR-NC and anti-miR-495-3p into NR8383 cells which were then treated with 1 mg/L LPS and recorded as LPS+si-BDNF-AS+anti-miR-NC group and LPS+si-BDNF-AS+anti-miR-495-3p group.Real-time fluorescence quantitative PCR was used to detect the expressions of lncRNA BDNF-AS and miR-495-3p;enzyme-linked immunosorbent assay(ELISA)was used to detect interleukin-1β(IL-1β)and tumor necrosis factor-α(TNF-α)levels;flow cytometry was performed to detect apoptosis;Western blotting was employed to detect B-cell lymphoma/leukemia-2(Bcl-2),Bcl-2 related X protein(Bax)protein expressions;luciferase reporter assay was used to detect the targeting relationship of BDNF-AS to miR-495-3p.Results LPS-induced rat alveolar macrophages showed high expression of BDNF-AS[(1.00±0.06)vs.(3.41±0.31)],low expression of miR-495-3p[(1.02±0.07)vs.(0.47±0.04)],and low expression of IL-1β[(22.14±2.25)ng/L vs.(513.20±41.22)ng/L],TNF-α[(184.33±18.65)ng/L vs.(1125.65±110.36)ng/L],apoptosis rate[(8.11±0.81)%vs.(36.22±3.21)%],Bax expression were increased,the expression level of Bcl-2 was decreased,and the difference was statistically significant(P<0.05).After inhibiting BDNF-AS expression or overexpressing miR-495-3p,IL-1β[(526.14±46.87)ng/L vs.(132.41±12.58)ng/L,(536.14±51.02)ng/L vs.(175

关 键 词：巨噬细胞 肺泡 脑源性神经营养因子 RNA 反义 RNA 长链非编码 微小RNA-495-3p 凋亡 炎症反应 

分 类 号：R563.9[医药卫生—呼吸系统]