Although long noncoding RNA nuclear paraspeckle assembly transcript 1 (NEAT1) was reported to be associated with acute lung injury (ALI), its specific mechanism has not been well studied. Mouse and cell ALI models were constructed by lipopolysaccharide (LPS). Cell viability was evaluated by 3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di- phenytetrazoliumromide assay. Cell death was evaluated by lactate dehydrogenase release (LDH) detection kit and flow cytometry. The levels of cytokines in lung tissues lysates were detected by quantitative real-time PCR (qRT-PCR) and enzyme-linked immunosorbent assay (ELISA). The expression of apoptosis-related markers was detected by Western blot. The relationship between NEAT1, miR-98-5p, and toll-like receptor 4 (TLR4) was determined by bioinformatics prediction, luciferase reporter assay, and RNA immunoprecipitation (RIP) assay. Rescue experiments were performed to determine the role of NEAT1/miR-98-5p/TLR4 in ALI. NEAT1 was significantly upregulated during ALI both in vitro and in vivo. NEAT1 knockdown efficiently attenuated LPS-induced ALI and reduced LPS-induced elevation of cytokines both in vitro and in vivo. NEAT1 negatively regulated miR-98-5p by directly sponging it, and TLR4 was a target of miR-98-5p. MiR-98-5p inhibition or TLR4 overexpression could obviously attenuate the protective effects of NEAT1 knockdown in LPS-treated A549 cells. Our study demonstrated that NEAT1 knockdown alleviated LPS-induced ALI by targeting the miR-98-5p/TLR4 axis.

中文翻译：

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LncRNA NEAT1通过调节miR-98-5p/TLR4轴加重脂多糖诱导的急性肺损伤

虽然据报道长非编码 RNA 核副啄木鸟组装转录物 1 (NEAT1) 与急性肺损伤 (ALI) 相关，但其具体机​​制尚未得到很好的研究。小鼠和细胞 ALI 模型由脂多糖 (LPS) 构建。细胞活力通过 3-(4,5)-二甲基噻唑 (-z-y1)-3,5-二苯基四唑溴化物测定法进行评估。通过乳酸脱氢酶释放 (LDH) 检测试剂盒和流式细胞术评估细胞死亡。通过定量实时PCR（qRT-PCR）和酶联免疫吸附试验（ELISA）检测肺组织裂解物中细胞因子的水平。Western blot检测凋亡相关标志物的表达。NEAT1、miR-98-5p 和 toll 样受体 4 (TLR4) 之间的关系通过生物信息学预测、荧光素酶报告基因测定、和 RNA 免疫沉淀 (RIP) 测定。进行救援实验以确定 NEAT1/miR-98-5p/TLR4 在 ALI 中的作用。NEAT1 在体外和体内 ALI 期间显着上调。NEAT1 敲低有效地减弱了 LPS 诱导的 ALI，并降低了 LPS 诱导的体外和体内细胞因子升高。NEAT1 通过直接擦拭 miR-98-5p 负调控 miR-98-5p，TLR4 是 miR-98-5p 的靶标。MiR-98-5p 抑制或 TLR4 过表达可明显减弱 NEAT1 敲低对 LPS 处理的 A549 细胞的保护作用。我们的研究表明，NEAT1 敲低通过靶向 miR-98-5p/TLR4 轴减轻了 LPS 诱导的 ALI。NEAT1 敲低有效地减弱了 LPS 诱导的 ALI，并降低了 LPS 诱导的体外和体内细胞因子升高。NEAT1 通过直接擦拭 miR-98-5p 负调控 miR-98-5p，TLR4 是 miR-98-5p 的靶标。MiR-98-5p 抑制或 TLR4 过表达可明显减弱 NEAT1 敲低对 LPS 处理的 A549 细胞的保护作用。我们的研究表明，NEAT1 敲低通过靶向 miR-98-5p/TLR4 轴减轻了 LPS 诱导的 ALI。NEAT1 敲低有效地减弱了 LPS 诱导的 ALI，并降低了 LPS 诱导的体外和体内细胞因子升高。NEAT1 通过直接擦拭 miR-98-5p 负调控 miR-98-5p，TLR4 是 miR-98-5p 的靶标。MiR-98-5p 抑制或 TLR4 过表达可明显减弱 NEAT1 敲低对 LPS 处理的 A549 细胞的保护作用。我们的研究表明，NEAT1 敲低通过靶向 miR-98-5p/TLR4 轴减轻了 LPS 诱导的 ALI。