lncRNA NEAT1促进脓毒症诱导肝细胞损伤和炎症反应的机制研究

曹纪玲; 孙坚; 吴勇; 胡志青

doi:10.11659/jjssx.10E022092

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lncRNA NEAT1促进脓毒症诱导肝细胞损伤和炎症反应的机制研究

基础研究 | 更新时间：2023-12-13

- lncRNA NEAT1促进脓毒症诱导肝细胞损伤和炎症反应的机制研究
- Mechanism of lncRNA NEAT1 promoting hepatocyte injury and inflammatory reaction induced by sepsis
- 局解手术学杂志 2023年32卷第12期页码：1040-1047
- 作者机构：
  
  1.南京江北医院重症监护室，江苏南京 210048
  2.南京江北医院急诊医学，江苏南京 210048
  3.南京江北医院重症医学，江苏南京 210048
- 作者简介：
  
  曹纪玲，E-mail：t18992273@163.com
- 基金信息：
  
  江苏省卫生健康委员会科研课题项目(19PJ061)
- DOI：10.11659/jjssx.10E022092
  中图分类号： R459.7
- 纸质出版日期：2023-12-25，
  
  收稿日期：2022-10-19，
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曹纪玲,孙坚,吴勇等.lncRNA NEAT1促进脓毒症诱导肝细胞损伤和炎症反应的机制研究[J].局解手术学杂志,2023,32(12):1040-1047.

CAO Ji-ling,SUN Jian,WU Yong,et al.Mechanism of lncRNA NEAT1 promoting hepatocyte injury and inflammatory reaction induced by sepsis[J].JOURNAL OF REGIONAL ANATOMY AND OPERATIVE SURGERY,2023,32(12):1040-1047.
曹纪玲,孙坚,吴勇等.lncRNA NEAT1促进脓毒症诱导肝细胞损伤和炎症反应的机制研究[J].局解手术学杂志,2023,32(12):1040-1047. DOI： 10.11659/jjssx.10E022092.

CAO Ji-ling,SUN Jian,WU Yong,et al.Mechanism of lncRNA NEAT1 promoting hepatocyte injury and inflammatory reaction induced by sepsis[J].JOURNAL OF REGIONAL ANATOMY AND OPERATIVE SURGERY,2023,32(12):1040-1047. DOI： 10.11659/jjssx.10E022092.

摘要

目的

基于lncRNA NEAT1调控的miR-129-5p/TLR4信号轴探讨脓毒症诱导的肝细胞损伤和炎症反应机制。

方法

收集15例脓毒症患者外周血（脓毒症组）和15例健康者外周血（健康组）用于检测基因表达。体外实验使用小鼠正常肝细胞株NCTC1469，并用脂多糖（LPS）建立体外脓毒症模型（LPS组），将LPS刺激下转染mimic NC、mimic、siNC或siNEAT1质粒载体的细胞分别命名为LPS+mimic NC组、LPS+mimic组、LPS+siNC组或LPS+siNEAT1组，将LPS、siNEAT1、TLR4重组蛋白或LPS、mimic、TLR4重组蛋白联合处理的细胞命名为LPS+siNEAT1+TLR4组或LPS+mimic+TLR4组，而对照组不作处理。分别用qPCR和ELISA检测TNF-α、IL-6、IL-1β的mRNA和蛋白表达水平。TUNEL实验检测细胞的凋亡率。双荧光素酶报告基因分析法检测lncRNA NEAT1/miR-129-5p和miR-129-5p/TLR4的结合。Western blot检测细胞中TLR4的表达。

结果

在体内实验中，与健康组比较，脓毒症组外周血中TNF-α、IL-6、IL-1β、lncRNA NEAT1和TLR4的表达明显增加（

0.05），miR-129-5p表达减少（

0.05）。在体外实验中，与对照组比较，LPS组lncRNA NEAT1表达增加（

0.05）。与LPS组、LPS+siNC组比较，LPS+siNEAT1组的lncRNA NEAT1表达降低（

0.05），miR-129-5p表达增加（

0.05）。与对照组比较，LPS+siNC组细胞凋亡率及TLR4、TNF-α、IL-6、IL-1β的表达均上调（

0.05）。与LPS+siNC组比较，LPS+siNEAT1组细胞凋亡率及TLR4、TNF-α、IL-6、IL-1β的表达均下调（

0.05）。与LPS+siNEAT1组比较，LPS+siNEAT1+TLR4组细胞凋亡率及TLR4、TNF-α、IL-6、IL-1β的表达均上调（

0.05）。双荧光素酶报告基因实验表明lncRNA NEAT1与miR-129-5p存在相互作用，TLR4是miR-129-5p的靶基因。与对照组比较，LPS+mimic NC组TNF-α、IL-6、IL-1β的表达和细胞凋亡率均增加（

0.05）；与LPS+mimic NC组比较，LPS+mimic组TNF-α、IL-6、IL-1β的表达和细胞凋亡率均下调（

0.05）；与LPS+mimic组比较，LPS+mimic+TLR4组TNF-α、IL-6、IL-1β的表达和细胞凋亡率上调（

0.05）。

结论

lncRNA NEAT1与miR-129-5p存在相互作用，并直接靶向TLR4以促进LPS诱导的肝细胞损伤和炎症反应。本研究可为脓毒症引起的肝细胞损伤提供新的治疗靶点。

Abstract

Objective

To explore the mechanism of hepatocyte injury and inflammatory reaction induced by sepsis based on miR-129-5p/TLR4 signaling axis regulated by lncRNA NEAT1.

Methods

Peripheral blood from 15 sepsis patients （the sepsis group） and 15 healthy individuals （the healthy group） were collected for gene expression detection.The mouse normal liver cell line NCTC1469 was used in in vitro experiments， and an in vitro sepsis model （the LPS group） was established by lipopolysaccharide （LPS）. The cells stimulated with LPS and transfected with mimic NC，mimic，siNC or siNEAT1 plasmid vectors were named as the LPS+mimic NC group，the LPS+mimic group，the LPS+siNC group or the LPS+siNEAT1 group， respectively. The cells treated with a combination of LPS， siNEAT1， and TLR4 recombinant protein or LPS，mimic，TLR4 recombinant protein were named as the LPS+siNEAT1+TLR4 group or the LPS+mimic+TLR4 group， while the control group was not treated. The mRNA and protein expression levels of TNF-α， IL-6， and IL-1β were detected by qPCR and ELISA， respectively. The cell apoptosis rate was detected by TUNEL assay. The binding of lncRNA NEAT1/miR-129-5p and miR-129-5p/TLR4 was detected by dual-luciferase reporter gene analysis. The TLR4 expression in cells was detected by Western blot.

Results

In the in vivo experiment， compared with the healthy group， the expression of TNF-α， IL-6， IL-1β， lncRNA NEAT1， and TLR4 of peripheral blood in the sepsis group was significantly increased （

0.05）， while the expression of miR-129-5p was decreased （

0.05）. In the in vitro experiment，

compared with the control group， the expression of lncRNA NEAT1 in the LPS group was increased （

0.05）. Compared with the LPS group and the LPS+siNC group， the expression of lncRNA NEAT1 in the LPS+siNEAT1 group was decreased （

0.05）， and the expression of miR-129-5p was increased （

0.05）. Compared with the control group， the cell apoptosis rate and the expression of TLR4， TNF-α， IL-6， and IL-1β in the LPS+siNC group were up-regulated （

0.05）. Compared with the LPS+siNC group， the cell apoptosis rate and the expression of TLR4， TNF-α， IL-6， and IL-1β in the LPS+siNEAT1 group were down-regulated （

0.05）. Compared with the LPS+siNEAT1 group， the cell apoptosis rate and the expression of TLR4， TNF-α， IL-6， and IL-1β in the LPS+siNEAT1+TLR4 group were up-regulated （

0.05）. Dual-luciferase reporter gene assay revealed an interaction between lncRNA NEAT1 and miR-129-5p， and TLR4 was the target gene of miR-129-5p. Compared with the control group， the expression of TNF-α， IL-6， and IL-1β as well as cell apoptosis rate in the LPS+mimic NC group were increased （

0.05）. Compared with the LPS+mimic NC group， the expression of TNF-α， IL-6， and IL-1β as well as cell apoptosis rate in the LPS+mimic group were down-regulated （

0.05）. Compared with the LPS+mimic group， the expression of TNF-α， IL-6， and IL-1β as well as cell apoptosis rate in the LPS+mimic+TLR4 group were up-regulated （

0.05）.

Conclusion

lncRNA NEAT1 interacts with miR-129-5p and directly targets TLR4 to promote LPS-induced hepatocyte injury and inflammatory reaction. This study can provide a new therapeutic target for hepatocyte injury caused by sepsis.

关键词

lncRNA NEAT1miR-129-5p脓毒症肝细胞损伤炎症反应

Keywords

lncRNA NEAT1miR-129-5psepsishepatocyte injuryinflammatory reaction

references

Te Marvelde L，Whitfield A，Shepheard J，et al.Epidemiology of sepsis in cancer patients in Victoria，Australia：a population‐based study using linked data［J］.Aust N Z J Public Health，2020，44（1）：53-58.doi：10.1111/1753-6405.12935http://dx.doi.org/10.1111/1753-6405.12935.

Kim J，Kim K，Lee H，et al.Epidemiology of sepsis in Korea：a population-based study of incidence，mortality，cost and risk factors for death in sepsis［J］.Clin Exp Emerg Med，2019，6（1）：49-63.doi：10.15441/ceem.18.007http://dx.doi.org/10.15441/ceem.18.007.

Wang D，Yin Y，Yao Y.Advances in sepsis-associated liver dysfunction［J］.Burns Trauma，2014，2（3）：2321-3868.doi：10.4103/2321-3868.132689http://dx.doi.org/10.4103/2321-3868.132689.

Lin Y，Ding Y，Song S，et al.Expression patterns and prognostic value of miR-210，miR-494，and miR-205 in middle-aged and old patients with sepsis-induced acute kidney injury［J］.Bosn J Basic Med Sci，2019，19（3）：249-258.doi：10.17305/bjbms.2019.4131http://dx.doi.org/10.17305/bjbms.2019.4131.

Yang P，Xiong W，Chen X，et al.Overexpression of miR-129-5p mitigates sepsis-induced acute lung injury by targeting high mobility group box 1［J］.J Surg Res，2020，256（1）：23-30.doi：10.1016/j.jss.2020.05.101http://dx.doi.org/10.1016/j.jss.2020.05.101.

Fang J，Qiao F，Tu J，et al.High expression of long non-coding RNA NEAT1 indicates poor prognosis of human cancer［J］.Oncotarget，2017，8（28）：45918-49936.doi：10.18632/oncotarget.17439http://dx.doi.org/10.18632/oncotarget.17439.

江伟，杜斌.中国脓毒症流行病学现状［J］.医学研究生学报，2019，32（1）：5-8.doi：10.16571 /j.cnki.1008-8199.2019.01.002http://dx.doi.org/10.16571/j.cnki.1008-8199.2019.01.002.

赵广明，胡占升.脓毒症肝损伤的发病机制与治疗研究进展［J］.解放军医学杂志，2019，44（6）：515-520.doi：10.11855/j.issn.0577-7402.2019.06.10http://dx.doi.org/10.11855/j.issn.0577-7402.2019.06.10.

Lei F，Zhang H，Xie X.Comprehensive analysis of an lncRNA-miRNA-mRNA competing endogenous RNA network in pulpitis［J］.Peer J，2019，7：e7135.doi：10.7717/peerj.7135http://dx.doi.org/10.7717/peerj.7135.

马成龙，刘波，黄柯冰.lncRNA H19 对脂多糖诱导的脓毒症中microRNA-107表达的影响［J］.川北医学院学报，2018，33（6）：824-827.doi：10.3969 /j.issn.1005-3697.2018.06.005http://dx.doi.org/10.3969/j.issn.1005-3697.2018.06.005.

黄旦，刘健，纵瑞凯，等.miRNA与lncRNA在强直性脊柱炎中的研究进展［J］.中国免疫学杂志，2020，36（8）：1014-1028.doi：10.3969/j.issn.1000-484X.2020.08.025http://dx.doi.org/10.3969/j.issn.1000-484X.2020.08.025.

Jang JC，Li J，Gambini L，et al.Human resistin protects against endotoxic shock by blocking LPS-TLR4 interaction［J］.Proc Natl Acad Sci，2017，114（48）：E10399-E10408.doi：10.1073/pnas. 1716015114http://dx.doi.org/10.1073/pnas.1716015114.

Chen Y，Qiu J，Chen B，et al.RETRACTED：long non-coding RNA NEAT1 plays an important role in sepsis-induced acute kidney injury by targeting miR-204 and modulating the NF-κB pathway［J］.Int Immunopharmacol，2018，59：252-260.doi：10.1016/j.intimp. 2018.03.023http://dx.doi.org/10.1016/j.intimp.2018.03.023.

Ye J，Lin Y，Yu Y，et al.lncRNA NEAT1/microRNA-129-5p/SOCS2 axis regulates liver fibrosis in alcoholic steatohepatitis［J］.J Transl Med，2020，18（1）：1-12.doi：10.1186/s12967-020-02577-5http://dx.doi.org/10.1186/s12967-020-02577-5.

de Pádua Lúcio K，Rabelo ACS，Araújo CM，et al.Anti-inflammatory and antioxidant properties of black mulberry （Morus nigra L.） in a model of LPS-induced sepsis［J］.Oxid Med Cell Longev，2018，1（1）：1-13.doi：10.1155/2018/5048031http://dx.doi.org/10.1155/2018/5048031.

Dickson K，Lehmann C.Inflammatory response to different toxins in experimental sepsis models［J］.Int J Mol Sci，2019，20（18）：4341-4352.doi：10.3390/ijms20184341http://dx.doi.org/10.3390/ijms20184341.

Honda T，He Q，Wang F，et al.Acute and chronic remote ischemic conditioning attenuate septic cardiomyopathy，improve cardiac output，protect systemic organs，and improve mortality in a lipopolysaccharide-induced sepsis model［J］.Basic Res Cardiol，2019，114（3）：15.doi：10.1007/s00395-019-0724-3http://dx.doi.org/10.1007/s00395-019-0724-3.

Zhang Z，Wen H，Peng B，et al.Downregulated microRNA-129-5p by long non-coding RNA NEAT1 upregulates PEG3 expression to aggravate non-alcoholic steatohepatitis［J］.Front Genet，2021，11（1）：563265-563278.doi：10.3389/fgene.2020.563265http://dx.doi.org/10.3389/fgene.2020.563265.

Wei W，Sun Z，He S，et al.Protective role of dexmedetomidine against sevoflurane-induced postoperative cognitive dysfunction via the microRNA-129/TLR4 axis［J］.J Clin Neurosci，2021，92（1）：89-97.doi：10.1016/j.jocn.2021.07.057http://dx.doi.org/10.1016/j.jocn.2021.07.057.

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