Effects of miR-21 on proliferation and apoptosis of WT cells via PTEN/Akt pathway.

Abstract

Micro ribonucleic acid (miR)-21 in the proliferation and apoptosis of Wilms' tumor (WT) cells was explored. SK-NEP-1 cells were transfected with miR-21 inhibitor to silence the expression of miR-21. Then, the effects of miR-21 silencing on the proliferation and apoptosis of WT SK-NEP-1 cells were detected through cell counting kit-8 (CCK-8), colony formation assay and flow cytometry. The targets of miR-21 were analyzed via TargetScan database. Fluorescence real-time quantitative polymerase chain reaction (RT-qPCR) assay and western blot analysis were conducted to detect the changes in messenger RNA (mRNA) and protein expression levels of gene of phosphate and tension homology deleted on chromosome ten (PTEN) after silencing miR-21. Whether miR-21 directly binds to PTEN was examined by activity detection via dual luciferase reporter gene assay. Western blotting was employed to detect the correlation of miR-21 with PTEN and protein kinase B (Akt). Compared with normal control (NC) group, miR-21 inhibitor group had significantly inhibited proliferation of SK-NEP-1 cells (P<0.05), notably reduced number of clones (P<0.05) and overtly raised proportion of apoptotic cells (P<0.05). The suppression of miR-21 expression upregulated the mRNA and protein expression levels of PTEN, and the results of activity detection via dual luciferase reporter gene assay indicated that miR-21 bound to PTEN 3′-untranslated region (UTR) to repress its expression (P<0.05). PTEN silencing increased phosphorylated Akt (p-Akt) level in SK-NEP-1 cells, but there was no changes in Akt protein level. After silencing both PTEN and miR-21, the decrease in p-Akt was reversed, thereby reversing the inhibitory effect of miR-21 on the proliferation of SK-NEP-1 cells (P<0.05). miR-21 affects the proliferation and apoptosis of WT SK-NEP-1 cells via the PTEN/Akt pathway.