Circ_0114428 knockdown inhibits ROCK2 expression to assuage lipopolysaccharide-induced human pulmonary alveolar epithelial cell injury through miR-574-5p

Abstract

BackgroundSepsis-induced acute lung injury (ALI) accounts for about 40% of ALI, accompanied by alveolar epithelial injury. The study aimed to reveal the role of circular RNA_0114428 (circ_0114428) in sepsis-induced ALI.MethodsHuman pulmonary alveolar epithelial cells (HPAEpiCs) were treated with lipopolysaccharide (LPS) to mimic a sepsis-induced ALI cell model. RNA expression of circ_0114428, miR-574-5p and Rho-associated coiled-coil containing protein kinase 2 (ROCK2) was detected by qRT-PCR. Protein expression was checked by Western blotting. Cell viability, proliferation and apoptosis were investigated by cell counting kit-8, 5-Ethynyl-29-deoxyuridine (EdU) and flow cytometry analysis, respectively. The levels of pro-inflammatory factors were detected by enzyme-linked immunosorbent assay (ELISA). Oxidative stress was analyzed by lipid peroxidation Malondialdehyde (MDA) and Superoxide Dismutase (SOD) activity detection assays. The interplay among circ_0114428, miR-574-5p and ROCK2 was identified by dual-luciferase reporter, RNA pull-down and RNA immunoprecipitation assays.ResultsCirc_0114428 and ROCK2 expression were significantly increased, but miR-574-5p was decreased in blood samples from sepsis patients and LPS-stimulated HPAEpiCs. LPS treatment led to decreased cell viability and proliferation and increased cell apoptosis, inflammation and oxidative stress; however, these effects were relieved after circ_0114428 knockdown. Besides, circ_0114428 acted as a miR-574-5p sponge and regulated LPS-treated HPAEpiC disorders through miR-574-5p. Meanwhile, ROCK2 was identified as a miR-574-5p target, and its silencing protected against LPS-induced cell injury. Importantly, circ_0114428 knockdown inhibited ROCK2 production by interacting with miR-574-5p.ConclusionCirc_0114428 knockdown protected against LPS-induced HPAEpiC injury through miR-574-5p/ROCK2 axis, providing a novel therapeutic target in sepsis-induced ALI.