Effects of silencing RPARP-AS1 on proliferation,migration and apoptosis of GBM U251 cells
Aim To investigate the effect of silencing long non-coding RNA RPARP-AS1 on the proliferation,migration and apoptosis of glioblastoma(GBM)U251 cells.Methods 80 tumor and paracancerous tissue samples were collected from patients diagnosed with GBM and GBM cell line U251 was routinely cultured.U251 cells were randomly separated into Control group,small interfering RNA negative control(si-NC)group,si-RPARP-AS1 group,si-RPARP-AS1+inhibitor NC group,and si-RPARP-AS1+miR-339-5p inhibitor group.qRT-PCR method was applied to detect the expression of RPARP-AS1,miR-339-5p,and murine double minute 2(MDM2)mRNA in tissues and cells.CCK-8 method was applied to detect the proliferation ability of cells in each group.Transwell method was applied to evaluate the migration and invasion abilities of U251 cell in each group.Flow cytometry was applied to detect cell apoptosis rate.Western blotting was applied to detect the expression of B cell lymphoblastoma-2(Bcl-2),Bcl-2 associated X protein(Bax),E-cadherin,N-cadherin,and MDM2 proteins.Double luciferase reporter gene experiment was applied to verify the relationship between RPARP-AS1 and miR-339-5p,miR-339-5p and MDM2 respectively.Results Compared with adjacent cancer tissues,RPARP-AS1 and MDM2 mRNA were highly expressed in GBM tumor tissues,while miR-339-5p was lowly expressed(P＜0.05).Compared with the Control group and the si-NC group,the expression of RPARP-AS1,MDM2 mRNA,and Bcl-2,N-cadherin,MDM2 proteins,and proliferation value,migration,invasion ability in U251 cells of the si-RPARP-AS1 group were obviously decreased(P＜0.05).And the expression of miR-339-5p,Bax,E-cadherin protein,and cell apoptosis rate were obviously increased(P＜0.05).Compared with the si-RPARP-AS1 group and the si-RPARP-AS1+inhibitor NC group,the expression of MDM2 mRNA,and Bcl-2,N-cadherin,and MDM2 proteins,and proliferation value,migration,cell invasion ability in the si-RPARP-AS1+miR-339-5p inhibitor group were obviously increased(P＜0.05),whereas the expression of miR-339-5p,Bax and E-cadherin proteins,and cell apoptosis rate were obviously decreased(P＜0.05).RPARP-AS1 targeted and negatively regulated miR-339-5p expression,while miR-339-5p targeted and negatively regulated MDM2 expression.Conclusion Silencing RPARP-AS1 has inhibitory effects on the proliferation,migration,and invasion of GBM U251 cells,and promotes cell apoptosis,which may be related to the upregulation of miR-339-5p and inhibition of MDM2 expression.
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