长链非编码RNA alpha-2-巨球蛋白反义RNA 1靶向微小RNA-106b-5p调控氧化型低密度脂蛋白诱导的人脑微血管内皮细胞损伤

李薇; 王丽; 汪志华; 刘庆春; 韩荣胜

doi:10.16098/j.issn.0529-1356.2023.03.010

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解剖学报 ›› 2023, Vol. 54 ›› Issue (3) : 319-327. DOI: 10.16098/j.issn.0529-1356.2023.03.010

细胞和分子生物学

长链非编码RNA alpha-2-巨球蛋白反义RNA 1靶向微小RNA-106b-5p调控氧化型低密度脂蛋白诱导的人脑微血管内皮细胞损伤

李薇^1* 王丽² 汪志华³刘庆春¹韩荣胜4

作者信息 +

Long non-coding RNA alpha-2-macroglobulin antisense RNA 1 regulating oxidized low density lipoprotein-induced human brain microvascular endothelial cell damage by targeting microRNA-106b-5p

LI Wei^1* WANG Li² WANG Zhi-hua³ LIU Qing-chun¹ HAN Rong-sheng⁴

Author information +

文章历史 +

摘要

目的探讨长链非编码RNA (lncRNA）alpha-2-巨球蛋白反义RNA 1（A2M-AS1）靶向微小RNA（miR）-106b-5p对氧化型低密度脂蛋白（ox-LDL）诱导的人脑微血管内皮细胞损伤的影响。方法用ox-LDL诱导人脑微血管内皮细胞设为ox-LDL组，正常培养细胞为对照(Ctrl)组；A2M-AS1过表达（pcDNA-A2M-AS1组）、空载体（pcDNA组）、miR-106b-5p抑制剂（anti-miR-106b-5p组）、阴性对照（anti-miR-NC组）、pcDNA-A2M-AS1与对照mimic NC（miR-NC组）、pcDNA-A2M-AS1与miR-106b-5p模拟物（miR-106b-5p mimics组）转染细胞后加ox-LDL处理，n=9；Real-time PCR检测A2M-AS1与miR-106b-5p表达；试剂盒检测丙二醛（MDA）、超氧化物歧化酶（SOD）和过氧化氢酶（CAT）水平；流式细胞术及TUNEL法检测细胞凋亡；双荧光素酶报告基因实验检测A2M-AS1与miR-106b-5p靶向关系；Western blotting检测Bcl-2和Bax蛋白表达量。结果与Ctrl组比较，ox-LDL组A2M-AS1表达水平、SOD和CAT活性、Bcl-2蛋白水平降低，miR-106b-5p表达水平、MDA水平、凋亡率、Bax蛋白水平升高（P＜0.05）；过表达A2M-AS1或干扰miR-106b-5p降低ox-LDL诱导细胞后MDA水平、凋亡率与Bax蛋白水平，升高SOD、CAT活性和Bcl-2蛋白水平（P＜0.05）；A2M-AS1靶向miR-106b-5p；上调miR-106b-5p逆转过表达lncRNA A2M-AS1对ox-LDL诱导的人脑微血管内皮细胞损伤的作用。结论 A2M-AS1通过靶向miR-106b-5p减轻ox-LDL诱导的人脑微血管内皮细胞损伤。

Abstract

Objective To investigate the effect of long non-coding RNA(lncRNA) alpha-2-macroglobulin antisense RNA 1 (A2M-AS1) targeting microRNA (miR）-106b-5p on oxidized low-density lipoprotein (ox-LDL)-induced injury of human brain microvascular endothelial cells. Methods Human brain microvascular endothelial cells (ox-LDL group) were induced by ox-LDL, normal cultured cells were control group (Ctrl); A2M-AS1 overexpression (pcDNA-A2M-AS1 group), empty vector (pcDNA group), miR-106b-5p inhibitor (anti-miR-106b-5p group), negative control (anti-miR-NC group), pcDNA-A2M-AS1 with control mimic NC (miR-NC group), pcDNA-A2M-AS1 with miR-106b-5p mimic (miR-106b-5p mimics group) were transfected into cells and treated with ox-LDL, n=9. Real-time PCR was used to detect the expression levels of A2M-AS1 and miR-106b-5p; Kits were used to detect malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT)); Flow cytometry and TUNEL detected apoptosis; Dual luciferase reporter gene assay detected A2M-AS1 and miR-106b-5p targeting; Western blotting detected Bcl-2 and Bax protein expression. Results Compared with the Ctrl group, the expression level of A2M-AS1 in the ox-LDL group decreased, and the activity of SOD and CAT and the protein level of Bcl-2 decreased (P＜0.05), while the expression level of miR-106b-5p and the level of MDA increased (P<0.05), and the rate of apoptosis and the protein level of Bax increased (P<0.05). Overexpressing A2M-AS1 or interfering with miR-106b-5p decreased the MDA level, apoptosis rate and Bax protein level after ox-LDL-induced cells, and increased SOD, CAT activity and Bcl-2 protein level (P＜0.05). A2M-AS1 targeted miR-106b-5p; up-regulation of miR-106b-5p reversed the effect of overexpressed lncRNA A2M-AS1 on ox-LDL-induced injury of human brain microvascular endothelial cells (P<0.05). Conclusion A2M-AS1 attenuates ox-LDL-induced injury of human brain microvascular endothelial cells by targeting miR-106b-5p.

导出引用

李薇王丽汪志华刘庆春韩荣胜. 长链非编码RNA alpha-2-巨球蛋白反义RNA 1靶向微小RNA-106b-5p调控氧化型低密度脂蛋白诱导的人脑微血管内皮细胞损伤[J]. 解剖学报. 2023, 54(3): 319-327 https://doi.org/10.16098/j.issn.0529-1356.2023.03.010

LI Wei WANG Li WANG Zhi-hua LIU Qing-chun HAN Rong-sheng.

Long non-coding RNA alpha-2-macroglobulin antisense RNA 1 regulating oxidized low density lipoprotein-induced human brain microvascular endothelial cell damage by targeting microRNA-106b-5p

[J]. Acta Anatomica Sinica. 2023, 54(3): 319-327 https://doi.org/10.16098/j.issn.0529-1356.2023.03.010

中图分类号： R657.51

参考文献

［1］He ShB, Jiang ChM, Yang Q, et al. TSG alleviates TNF-αInduced human brain microvascular endothelial cells damage by inhibiting NF-κB signaling pathway［J］. Journal of Shaoyang University (Natural Science Edition), 2018, 15(1): 79-84. (in Chinese)

何邵波, 蒋传命, 杨秦, 等. 二苯乙烯苷通过抑制NF-κB信号通路减轻TNF-α诱导的人脑微血管内皮细胞损伤［J］. 邵阳学院学报(自然科学版), 2018, 15(1): 79-84.

［2］Yang WY, Yang L. Analysis of radix hedysari on oxLDL induced human vascular endothelial cell injury model of micro oxidation protection mechanism ［J］. Journal of Sichuan Traditional Chinese Medicine, 2018, 36(1): 60-63. (in Chinese)

杨为亚, 杨林. 红芪多糖对氧化低密度脂蛋白诱导人脑微血管内皮细胞氧化损伤模型保护作用机制分析［J］. 四川中医, 2018, 36(1): 60-63.

［3］Yang MZ, Wang L. AMPK activation repairs the damage of human brain microvascular endothelial cells induced by high-glucose via regulating oxidative stress ［J］. Chinese Journal of Histochemistry and Cytochemistry, 2020, 29(1): 1-7. (in Chinese)

杨梦珍, 汪琳. AMPK激活通过调控氧化应激修复高糖诱导的人脑微血管内皮细胞损伤［J］. 中国组织化学与细胞化学杂志, 2020, 29(1): 1-7.

［4］Zhang M, Tang M, Wu Q, et al. LncRNA DANCR attenuates brain microvascular endothelial cell damage induced by oxygen-glucose deprivation through regulating of miR-33a-5p/XBP1s ［J］. Aging (Albany NY), 2020, 12(2): 1778-1791.

［5］Song XL, Zhang FF, Wang WJ, et al. LncRNA A2M-AS1 lessens the injury of cardiomyocytes caused by hypoxia and reoxygenation via regulating IL1R2［J］. Genes Genomics, 2020, 42(12): 1431-1441.

［6］Li P, Shen M, Gao F, et al. An antagomir to microRNA-106b-5p ameliorates cerebral ischemia and reperfusion injury in rats via inhibiting apoptosis and oxidative stress ［J］. Mol Neurobiol, 2017, 54(4): 2901-2921.

［7］Yang J, Wei DF, Li XD, et al. Differential protein study of protective effect of Hedysari Radix Polysaccharides on brain microvascular endothelial cells of oxidative damage ［J］. Pharmacology and Clinics of Chinese Materia Medica, 2016, 32(2): 113-118. (in Chinese)

杨杰, 卫东锋, 李晓东, 等. 红芪多糖对脑微血管内皮细胞氧化损伤保护作用的差异蛋白质研究［J］. 中药药理与临床, 2016, 32(2): 113-118.

［8］Dong B, Zhou B, Sun Z, et al. LncRNA-FENDRR mediates VEGFA to promote the apoptosis of brain microvascular endothelial cells via regulating miR-126 in mice with hypertensive intracerebral hemorrhage［J］. Microcirculation, 2018, 25(8): e12499-e12509.

［9］Xin JW, Jiang YG. Long noncoding RNA MALAT1 inhibits apoptosis induced by oxygen-glucose deprivation and reoxygenation in human brain microvascular endothelial cells ［J］. Exp Ther Med, 2017, 13(4): 1225-1234.

［10］Gao M, Fu J, Wang Y. The lncRNA FAL1 protects against hypoxia-reoxygenation-induced brain endothelial damages through regulating PAK1 ［J］. J Bioenerg Biomembr, 2020, 52(1): 17-25.

［11］He F, Huang L, Xu Q, et al. Microarray profiling of differentially expressed lncRNAs and mRNAs in lung adenocarcinomas and bioinformatics analysis［J］. Cancer Med, 2020, 9(20): 7717-7728.

［12］Giulietti M, Righetti A, Principato G, et al. LncRNA co-expression network analysis reveals novel biomarkers for pancreatic cancer ［J］. Carcinogenesis, 2018, 39(8): 1016-1025.

［13］Su HY, Tian RY, Zhu CX, et al. The effect of lentivirus-mediated PDCD4 siRNA on the oxidative damage of vascular endothelial cells induced by high glucose and its mechanism ［J］. Journal of Inner Mongolia Medical University, 2018, 40(6): 614-617. (in Chinese)

苏海燕, 田瑞媛, 朱彩霞, 等. 慢病毒介导PDCD4 siRNA对高糖诱导的血管内皮细胞氧化损伤的影响及机制［J］. 内蒙古医学院学报, 2018, 40(6): 614-617.

［14］Yan G, Zhao H, Hong X. LncRNA MACC1-AS1 attenuates microvascular endothelial cell injury and promotes angiogenesis under hypoxic conditions via modulating miR-6867-5p/TWIST1 in human brain microvascular endothelial cells ［J］. Ann Transl Med, 2020, 8(14): 876-886.

［15］Wu Y, Liu Y, Liu QT, et al. A preliminary study on the protective effect and mechanism of paraoxonase 3 gene overexpression on hepatic injury in acutely poisoned mice［J］. Acta Anatomica Sinica, 2019, 50(2): 179-184. (in Chinese)

伍洋, 刘英, 刘济滔, 等. 对氧磷酶3基因过表达对急性中毒小鼠肝损伤的保护作用及机制［J］. 解剖学报, 2019, 50(2): 179-184.

［16］Wu H, Wang J, Ma Z. Long noncoding RNA HOXA-AS2 mediates microRNA-106b-5p to repress sepsis-engendered acute kidney injury ［J］. J Biochem Mol Toxicol, 2020, 34(4): e22453-e22463.

［17］Xu G, Zhang W, Wang Z, et al. Matrine regulates H2O2-induced oxidative stress through long non-coding RNA HOTAIR/miR-106b-5p axis via AKT and STAT3 pathways ［J］. Biosci Rep, 2020, 40(5): 1-12.

［18］Pan H, Zhao F, Yang Y, et al. Overexpression of long non-coding RNA SNHG16 against cerebral ischemia-reperfusion injury through miR-106b-5p/LIMK1 axis ［J］. Life Sci, 2020, 254(1): 117778-117788.