The long noncoding RNA NKILA protects against myocardial ischaemic injury by enhancing myocardin expression via suppressing the NF-κB signalling pathway
The long noncoding RNA NKILA protects against myocardial ischaemic injury by enhancing myocardin expression via suppressing the NF-κB signalling pathway
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DOI：
10.1016/j.yexcr.2019.111774
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发表时间：
2020-02-15
影响因子：
3.7
通讯作者：
Xu，Jun-Mei
中科院分区：
文献类型：
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作者：
Liu，Qing;Liu，Zheng;Xu，Jun-Mei
lncRNA NKILA已被报道与NF-κB相互作用，并在多种人类疾病中发挥重要作用。然而，NKILA在心肌缺血损伤中的作用尚不清楚。方法建立心肌缺血损伤的细胞模型和动物模型。我们通过在H9c2细胞缺氧/再氧化（H/R）模型中过表达NKILA、沉默心肌素和使用NF-κB途径抑制剂证实了我们的发现。采用LAD结扎法建立缺血再灌注（I/R）损伤动物模型。通过尾静脉注射腺相关病毒（AAV）实现NKILA的过表达。Annexin-V/PI染色及流式细胞术检测细胞凋亡情况。采用elisa法检测炎症因子的分泌。采用TTC、HE、TUNEL染色观察心肌病理损伤。采用qRT-PCR或Western blotting检测NKILA、心肌素、NF-κB通路及凋亡相关蛋白的表达水平。结果sh /R和I/R处理可显著抑制NKILA的表达，激活NF-κB通路，导致心肌素丢失。过表达NKILA可抑制NF-κB通路，成功阻止H9c2细胞H/R刺激引起的细胞凋亡和炎症反应。沉默心肌素逆转NKILA的保护作用，导致H/R H9c2细胞严重损伤。NF-κB通路抑制剂BAY11-7028可减轻H9c2细胞H/R损伤，但对NKILA表达影响不大。在心肌I/R损伤动物模型中也证实了类似的结果，表明NKILA过表达可以抑制体内I/R引发的心肌损伤。结论nkila通过抑制NF-κB信号通路，抑制心肌细胞凋亡和炎症反应，增强心肌素的表达，从而改善心肌I/R损伤。
BackgroundThe lncRNA NKILA has been reported to interact with NF-κB and has an important role in various human diseases. However, the role of NKILA in myocardial ischaemic injury is still unknown.MethodsWe established cell and animal models of myocardial ischaemic injury. We confirmed our findings by overexpressing NKILA, silencing myocardin and using an NF-κB pathway inhibitor in a hypoxia/reoxygenation (H/R) model of H9c2 cells. An animal model of ischaemia-reperfusion (I/R) injury was established by LAD ligation. Overexpression of NKILA was achieved by adeno-associated virus (AAV) injection through the tail vein. Annexin-V/PI staining and flow cytometric analysis were performed to test cell apoptosis. ELISAs were used to determine the secretion of inflammatory factors. TTC, HE and TUNEL staining were performed to study myocardial pathological injury. qRT-PCR or Western blotting were used to test the expression levels of NKILA, myocardin, the NF-κB pathway and apoptosis-related proteins.ResultsH/R and I/R treatment significantly suppressed the expression of NKILA and activated the NF-κB pathway, resulting in the loss of myocardin. Overexpressing NKILA led to the suppressionof the NF-κB pathway and successfully prevented the cell apoptosis and inflammatory responses caused by H/R stimulation in H9c2 cells. Silencing myocardin reversed the protective effect of NKILA and led to severe injury in the H9c2 cells that underwent H/R. Furthermore, the NF-κB pathway inhibitor BAY11-7028 reduced the H/R injury in H9c2 cells with little effect on NKILA expression. Similar results were confirmed in an animal model of myocardial I/R injury and showed that overexpression of NKILA inhibited I/R-triggered myocardial injuryin vivo.ConclusionNKILA enhanced the expression of myocardin via inhibiting the NF-κB signalling pathway and preventing cell apoptosis and the inflammatory response of cardiomyocytes, thus ameliorating myocardial I/R injury.