MicroRNA-223 alleviates lipopolysaccharide-induced PC-12 cells apoptosis and autophagy by targeting RPH1 in spinal cord injury.
International journal of clinical and experimental pathology(2017)
Abstract
Spinal cord injury (SCI) is one of the most devastating diseases. MicroRNAs (miRNAs) are recognized as key regulators in SCI; however, the role of miR-223 in SCI remains unclear. Herein, our study aimed to explore the effect of miR-223 on lipopolysaccharide (LPS)-induced injury to PC-12 cells. PC-12 cells were treated with different concentrations of LPS, and then cell viability, apoptosis, apoptosis-related factors and autophagy-related factors were analyzed by CCK-8, flow cytometry and western blot. Subsequently, miR-223 mimic, miR-223 inhibitor, pEX-RPH1, sh-RPH1 and corresponding controls were transfected into PC-12 cells followed by 5 μg/ml of LPS treatment. Cell viability, apoptosis, apoptosis-related and autophagy-related factors were analyzed again. A target gene of miR-223 was validated by dual-luciferase assay. Besides, the main factors expressions of mTOR and NF-κB signal pathways were measured by western blot. LPS reduced cell viability but increased apoptotic cells rate, up-regulated Bax, cleaved-caspase-3, cleaved-caspase-9, LC-II and Beclin-1, and down-regulated Bcl-2 and p62 expressions in a dose-dependent way. Additionally, miR-223 overexpression promoted cell viability but inhibited apoptosis, and autophagy in LPS-stimulated PC-12 cells. RPH1 was a direct target of miR-223, and RPH1 exhibited contrary impacts to miR-223 on LPS-induced cell apoptosis and autophagy. Besides, the promoting effects of miR-223 suppression on cell apoptosis and autophagy were relieved by RPH1 silence. Furthermore, miR-223 blocked LPS-induced mTOR and NF-κB pathways by down-regulation of RPH1. MiR-223 improved cell viability but declined apoptosis and autophagy by targeting RPH1 and blocked mTOR and NF-κB pathways in LPS challenged PC-12 cells.
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