长链非编码RNA NEAT1通过调控miR-122-5p/闭合蛋白轴保护尿毒症毒素诱导的肠上皮屏障损伤。
LncRNA NEAT1 protects uremic toxin-induced intestinal epithelial barrier injury by regulating miR-122-5p/Occludin axis.
作者信息
Han Meng, P Pathuama, Tian Jinhai, Wang Chen, Zhou Shengnan, Fu Lina, Wang Libin, Tian Na
机构信息
Department of Nephrology, General Hospital of Ningxia Medical University, Ningxia, China.
The Biochip Research Center, General Hospital of Ningxia Medical University, Ningxia, China.
出版信息
PLoS One. 2025 May 8;20(5):e0322989. doi: 10.1371/journal.pone.0322989. eCollection 2025.
BACKGROUND
Long non-coding RNA(LncRNA) has been reported to be associated with intestinal barrier damage. The aim of this study was to explore the mechanism of lncRNA Nuclear enriched abundant transcript 1 (NEAT1) in uremic toxin-induced intestinal epithelial barrier injury.
METHODS
Human colon cancer cells (Caco-2) were used to establish intestinal epithelial injury models with the urea treatment in different conditions. Cell Counting Kit-8 (CCK-8) and Western blot screening the best concentration and time. The expressions of lncRNA NEAT1 and miR-122-5p were measured by quantitative real-time polymerase chain reaction (qRT-PCR). Western blot and immunofluorescence were used to detect the expression of tight junction proteins Occludin, ZO-1 and Claudin-1. Sodium fluorescein was used to detect the paracellular permeability of intestinal epithelial injury models. The binding of miR-122-5p to lncRNA NEAT1 and Occludin was determined by bioinformatics analysis and dual luciferase reporter assay.
RESULTS
The best condition for the injury model was urea treatment in 144 mg/dl for 48 hours. With the increase of urea intervention time and concentration, the damage degree of intestinal epithelial cells is aggravated. Based on the qRT-PCR results, lncRNA NEAT1 was significantly down-regulated in the model group. Meanwhile, the tight junction proteins Occludin, ZO-1 and Claudin-1 were significantly reduced. The permeability of sodium fluorescein was significantly increased in the model group. Overexpression of lncRNA NEAT1 can alleviate the above performances. As the target gene of lncRNA NEAT1, miR-122-5p is significantly up-regulated in the model group. The dual luciferase reporter assay proved that miR-122-5p was targets to Occludin. The protective effect of overexpression lncRNA NEAT1 on intestinal epithelial barrier function is reversed by miR-122-5p mimics.
CONCLUSION
LncRNA NEAT1 protects uremic toxin-induced intestinal epithelial barrier injury by regulating miR-122-5p/Occludin axis.
背景
据报道，长链非编码RNA（LncRNA）与肠道屏障损伤有关。本研究旨在探讨长链非编码RNA核富集丰富转录本1（NEAT1）在尿毒症毒素诱导的肠上皮屏障损伤中的作用机制。
方法
采用人结肠癌细胞（Caco-2）在不同条件下用尿素处理建立肠上皮损伤模型。通过细胞计数试剂盒-8（CCK-8）和蛋白质免疫印迹法筛选最佳浓度和时间。采用定量实时聚合酶链反应（qRT-PCR）检测长链非编码RNA NEAT1和微小RNA-122-5p（miR-122-5p）的表达。用蛋白质免疫印迹法和免疫荧光法检测紧密连接蛋白闭合蛋白（Occludin）、紧密连接蛋白1（ZO-1）和闭合小环蛋白1（Claudin-1）的表达。用荧光素钠检测肠上皮损伤模型的细胞旁通透性。通过生物信息学分析和双荧光素酶报告基因检测确定miR-122-5p与长链非编码RNA NEAT1和Occludin的结合情况。
结果
损伤模型的最佳条件是144mg/dl尿素处理48小时。随着尿素干预时间和浓度的增加，肠上皮细胞的损伤程度加重。基于qRT-PCR结果，模型组长链非编码RNA NEAT1显著下调。同时，紧密连接蛋白Occludin、ZO-1和Claudin-1显著减少。模型组荧光素钠的通透性显著增加。长链非编码RNA NEAT1过表达可减轻上述表现。作为长链非编码RNA NEAT1的靶基因，miR-122-5p在模型组中显著上调。双荧光素酶报告基因检测证明miR-122-5p靶向Occludin。miR-122-5p模拟物可逆转长链非编码RNA NEAT1过表达对肠上皮屏障功能的保护作用。
结论
长链非编码RNA NEAT1通过调节miR-122-5p/Occludin轴保护尿毒症毒素诱导的肠上皮屏障损伤。
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