Targeting MicroRNA-7-EGFR Pathway To Resist Osteoblast Damage Induced By Dexamethasone And Its Mechanism

Aims: Dexamethasone(Dex)can induce potent cytotoxicity in cultured human osteoblasts.The aim of this study was to test the potential role of micro RNA-7(mi R-7),which targets the epidermal growth factor receptor(EGFR),in Dex-treated human osteoblasts.Materials: Human osteoblastic cells(OB-6 and h FOB1.19 lines)and primary human osteoblasts were treated with Dex.mi R-7 expression was genetically decreased with a lentiviral antagomi R-7 construct(LV-antagomi R-7)or increased with a lentiviral mi R-7 construct(LV-mi R-7).AG1478 and PD153035 were used to inhibit EGFR.The CRISPR/Cas9 method was utilized for complete EGFR knockout(EGFR-KO).Cell viability,death,and apoptosis were tested by MTT,ELISA,and trypan blue staining assays,respectively.Expression of mi R-7 and EGFR signaling components was tested by quantitative real-time polymerase chain reaction(q PCR)and immunoblotting.Results: In OB-6,h FOB1.19,and primary human osteoblasts,mi R-7 depletion by LV-antagomi R-7 increased EGFR expression and downstream Akt activation,protecting cells from Dex-induced viability reduction,cell death,and apoptosis.In contrast,forced overexpression of mi R-7 by LV-mi R-7 inhibited EGFR expression and Akt activation,potentiating Dex-induced cytotoxicity in OB-6,h FOB1.19,and primary human osteoblasts.EGFR is the primary target of mi R-7 in human osteoblasts.Luciferase activity of the EGFR 3-untranslated region was enhanced by LV-antagomi R-7,but decreased by LV-mi R-7 in OB-6 cells.Further,LV-antagomi R-7–induced osteoblast cytoprotection against Dex was abolished by the EGFR inhibitors AG1478 and PD153035.Moreover,neither LV-antagomi R-7 nor LV-mi R-7 was functional in EGFR-KO OB-6 cells.We also showed that mi R-7 is upregulated in the necrotic femoral head tissues of Dex-administered patients,correlating with EGFR downregulation.Conclusion: mi R-7 inhibition protects human osteoblasts from Dex via activation of EGFR signaling.