Pulmonary vascular endothelial cell (PVEC) injury following acute lung injury or acute respiratory distress syndrome seriously affects disease development. Recently, accumulating evidence has suggested that long noncoding RNA (lncRNA) exerts significant effects in vascular endothelial cell injury. However, PRNCR1, a novel lncRNA, remains scarcely understood in terms of its functions in PVEC injury. Both in vivo and in vitro models of PVEC injury were constructed by lipopolysaccharide (LPS) administration. The relative expressions of PRNCR1, miR‐330‐5p, and TLR4 were detected by quantitative reverse transcription‐polymerase chain reaction, Western blot, and immunohistochemistry. Besides, gain and loss assays of PRNCR1/miR‐330‐5p were conducted to verify their effects on LPS‐induced PVEC injury. Cell Counting Kit‐8 assay used to measure cell viability and flow cytometry was used to detect apoptosis. Besides, the protein levels of caspase 3, nuclear factor‐κB (NF‐κB), and inflammatory cytokines (including tumor necrosis factor‐α, interleukin‐1β [IL‐1β], and IL‐6) were evaluated via Western blot and enzyme‐linked immunosorbent assay. Moreover, a dual‐luciferase activity experiment and RNA immunoprecipitation were applied to confirm the targeting relationship between PRNCR1 and miR‐330‐5p, miR‐330‐5p, and TLR4. PRNCR1 and TLR4 levels were significantly upregulated in LPS‐treated PVEC, both in vivo and in vitro, while miR‐330‐5p were downregulated. Inhibiting PRNCR1 or overexpressing miR‐330‐5p markedly attenuated LPS‐induced PVEC injury, expressions of TLR4, NF‐κB, and inflammatory cytokines. Mechanistically, PRNCR1 functioned as a competitive endogenous RNA by sponging miR‐330‐5p and then promoting TLR4 expression. PRNCR1 was upregulated in LPS‐induced PVEC and aggravated its injury via modulating the miR‐330‐5p/TLR4 axis.

中文翻译：

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下调lncRNA PRNCR1通过调节miR-330-5p / TLR4轴改善LPS诱导的肺血管内皮细胞损伤

急性肺损伤或急性呼吸窘迫综合征后的肺血管内皮细胞（PVEC）损伤严重影响疾病的发展。最近，越来越多的证据表明，长的非编码RNA（lncRNA）在血管内皮细胞损伤中发挥了重要作用。然而，就其在PVEC损伤中的功能而言，PRNCR1是一种新型的lncRNA，但对其了解仍很少。PVEC损伤的体内和体外模型都是通过脂多糖（LPS）给药构建的。通过定量逆转录聚合酶链反应，Western印迹和免疫组化检测PRNCR1，miR-330-5p和TLR4的相对表达。此外，进行了PRNCR1 / miR-330-5p的损益分析，以验证其对LPS诱发的PVEC损伤的影响。用于测量细胞活力的Cell Counting Kit-8检测法和流式细胞仪检测细胞凋亡。此外，通过Western blot和免疫印迹法评估了半胱天冬酶3，核因子-κB（NF-κB）和炎性细胞因子（包括肿瘤坏死因子-α，白介素-1β[IL-1β]和IL-6）的蛋白水平。酶联免疫吸附测定。此外，应用了双重荧光素酶活性实验和RNA免疫沉淀，以确认PRNCR1与miR-330-5p，miR-330-5p和TLR4之间的靶向关系。在体内和体外，经LPS处理的PVEC中PRNCR1和TLR4的水平均显着上调，而miR-330-5p的水平则下调。抑制PRNCR1或过表达miR-330-5p可以显着减轻LPS诱导的PVEC损伤，TLR4，NF-κB的表达和炎性细胞因子的表达。机械上，PRNCR1通过竞争miR-330-5p然后促进TLR4表达而起竞争性内源RNA的作用。PRNCR1在LPS诱导的PVEC中上调，并通过调节miR-330-5p / TLR4轴加剧了其损伤。