抑制 KLF6 可减少急性肺损伤中 II 型肺泡上皮细胞的炎症和凋亡。

Inhibition of KLF6 reduces the inflammation and apoptosis of type II alveolar epithelial cells in acute lung injury.

机构信息

Department of Anesthesiology, Qinghai University Affiliated Hospital, Xining, Qinghai, China.

Department of Critical Medicine, Yangpu Hospital, Tongji University School of Medicine, Shanghai, China;

出版信息

Allergol Immunopathol (Madr). 2022 Sep 1;50(5):138-147. doi: 10.15586/aei.v50i5.632. eCollection 2022.

DOI:10.15586/aei.v50i5.632

PMID:36086974

Abstract

BACKGROUND

The development of acute lung injury (ALI) into a severe stage leads to acute respiratory distress syndrome (ARDS). The morbidity and mortality of ALI and ARDS are very high. : This study is aimed to explore the effect of Krüppel-like factor 6 (KLF6) on lipopolysaccharide (LPS)-induced type II alveolar epithelial cells in ALI by interacting with cysteine-rich angiogenic inducer 61 (CYR61).

MATERIAL AND METHODS

ALI mice model and LPS-induced type II alveolar epithelial cells were conducted to simulate ALI and . The messenger RNA (mRNA) and protein expression of KLF6 in lung tissues were detected by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and Western blot analysis. Pathological changes in lung tissues were observed by hematoxylin and eosin (H&E) staining. The viability and KLF6 expression of A549 cells treated with different concentrations of LPS were detected by cell counting kit-8 (CCK-8) assay, RT-qPCR, and Western blot analysis. After indicated treatment, the viability and apoptosis of A549 cells were analyzed by CCK-8 and TUNEL assays, and the inflammation factors of A549 cells were detected by Enzyme-linked-immunosorbent serologic assay, RT-qPCR, and Western blot analysis. The combination of KLF6 and CYR61 was determined by chromatin immunoprecipitation (ChIP)-PCR and dual-luciferase reporter assay.

RESULTS

KLF6 expression was increased in lung tissues of ALI mice and LPS-induced A549 cells. Interference with KLF6 improved the viability, reduced the inflammatory damage, and promoted the apoptosis of LPS-induced A549 cells. In addition, KLF6 could bind to CYR61. Interference with KLF6 could decrease CYR61 expression in LPS-induced A549 cells. LPS also enhanced the TLR4/MYD88 signaling pathway, which was reversed by KLF6 interference. The above phenomena in LPS-induced A549 cells transfected with Si-KLF6 could be reversed by overexpression of CYR61.

CONCLUSION

Inhibition of KLF6 promoted the viability and reduced the inflammation and apoptosis of LPS-induced A549 cells, which was reversed by CYR61.

摘要

背景

急性肺损伤（ALI）发展为严重阶段会导致急性呼吸窘迫综合征（ARDS）。ALI 和 ARDS 的发病率和死亡率都非常高。本研究旨在探讨富含半胱氨酸的血管生成诱导因子 61（CYR61）与 Krüppel 样因子 6（KLF6）相互作用对脂多糖（LPS）诱导的 II 型肺泡上皮细胞在 ALI 中的作用。

材料和方法

采用脂多糖诱导的 ALI 小鼠模型和 II 型肺泡上皮细胞模拟 ALI。采用逆转录定量聚合酶链反应（RT-qPCR）和 Western blot 分析检测肺组织中 KLF6 的信使 RNA（mRNA）和蛋白表达。采用苏木精-伊红（H&E）染色观察肺组织的病理变化。用细胞计数试剂盒-8（CCK-8）法、RT-qPCR 和 Western blot 分析检测不同浓度 LPS 处理的 A549 细胞的活力和 KLF6 表达。经指示处理后，用 CCK-8 和 TUNEL 检测 A549 细胞的活力和凋亡，用酶联免疫吸附试验（ELISA）、RT-qPCR 和 Western blot 检测 A549 细胞的炎症因子。用染色质免疫沉淀（ChIP）-PCR 和双荧光素酶报告基因检测确定 KLF6 与 CYR61 的结合。

结果

ALI 小鼠肺组织和 LPS 诱导的 A549 细胞中 KLF6 表达增加。干扰 KLF6 可提高活力，减轻炎症损伤，促进 LPS 诱导的 A549 细胞凋亡。此外，KLF6 可以与 CYR61 结合。干扰 KLF6 可降低 LPS 诱导的 A549 细胞中 CYR61 的表达。LPS 还增强了 TLR4/MYD88 信号通路，干扰 KLF6 可使其逆转。在 LPS 诱导的 A549 细胞中转染 Si-KLF6 后，上述现象可被过表达的 CYR61 逆转。