Department of Paediatrics, Suizhou Hospital, Hubei University of Medicine, Long Men Street 60th, Zeng Du District, Suizhou, 441300, Hubei, China.
Genes Genomics. 2021 Jul;43(7):815-827. doi: 10.1007/s13258-021-01103-1. Epub 2021 Apr 26.
Many long non-coding RNAs (lncRNAs) have been suggested to play critical roles in acute lung injury (ALI) pathogenesis, including lncRNA nuclear enriched abundant transcript 1 (NEAT1).
We aimed to further elucidate the functions and molecular mechanism of NEAT1 in ALI.
Human pulmonary alveolar epithelial cells (HPAEpiCs) stimulated by lipopolysaccharide (LPS) were served as a cellular model of ALI. Cell viability and cell apoptosis were determined by cell counting kit-8 (CCK-8) assay and flow cytometry, respectively. The expression of NEAT1, microRNA-424-5p (miR-424-5p), and mitogen-activated protein kinase 14 (MAPK14) was measured by quantitative real-time polymerase chain reaction (qRT-PCR) or western blot analysis. Caspase activity was determined by caspase activity kit. The inflammatory responses were evaluated using enzyme-linked immunosorbent assay (ELISA). The oxidative stress factors were analyzed by corresponding kits.
NEAT1 was upregulated in LPS-stimulated HPAEpiCs. NEAT1 knockdown weakened LPS-induced injury by inhibiting apoptosis, inflammation and oxidative stress in HPAEpiCs. Moreover, miR-424-5p was a direct target of NEAT1, and its knockdown reversed the effects caused by NEAT1 knockdown in LPS-induced HPAEpiCs. Furthermore, MAPK14 was a downstream target of miR-424-5p, and its overexpression attenuated the effects of miR-424-5p on reduction of LPS-induced injury in HPAEpiCs. Besides, NEAT1 acted as a sponge of miR-424-5p to regulate MAPK14 expression.
NEAT1 knockdown alleviated LPS-induced injury of HPAEpiCs by regulating miR-424-5p/MAPK14 axis, which provided a potential therapeutic target for the treatment of ALI.
许多长链非编码 RNA（lncRNA）被认为在急性肺损伤（ALI）发病机制中发挥关键作用，包括核丰富丰富转录物 1（NEAT1）lncRNA。
我们旨在进一步阐明 NEAT1 在 ALI 中的功能和分子机制。
用脂多糖（LPS）刺激的人肺泡上皮细胞（HPAEpiCs）作为 ALI 的细胞模型。通过细胞计数试剂盒-8（CCK-8）测定和流式细胞术分别测定细胞活力和细胞凋亡。通过定量实时聚合酶链反应（qRT-PCR）或蛋白质印迹分析测定 NEAT1、微 RNA-424-5p（miR-424-5p）和丝裂原活化蛋白激酶 14（MAPK14）的表达。通过 caspase 活性试剂盒测定半胱氨酸天冬氨酸蛋白酶活性。通过酶联免疫吸附试验（ELISA）评估炎症反应。通过相应的试剂盒分析氧化应激因素。
在 LPS 刺激的 HPAEpiCs 中上调 NEAT1。NEAT1 敲低通过抑制细胞凋亡、炎症和氧化应激来减弱 LPS 诱导的 HPAEpiCs 损伤。此外，miR-424-5p 是 NEAT1 的直接靶标，其敲低逆转了 LPS 诱导的 HPAEpiCs 中 NEAT1 敲低引起的作用。此外，MAPK14 是 miR-424-5p 的下游靶标，其过表达减弱了 miR-424-5p 对 LPS 诱导的 HPAEpiCs 损伤的作用。此外，NEAT1 作为 miR-424-5p 的海绵调节 MAPK14 表达。
NEAT1 敲低通过调节 miR-424-5p/MAPK14 轴缓解 LPS 诱导的 HPAEpiCs 损伤，为 ALI 的治疗提供了潜在的治疗靶点。
