Basic & Clinical Medicine ›› 2022, Vol. 42 ›› Issue (1): 106-113.

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Silencing eEF2K combined with sodium tanshinone IIA sulfonate synergistically inhibit proliferation of human lung adenocarcinoma cell line A549

  

  • Received:2020-11-09 Revised:2021-04-12 Online:2022-01-05 Published:2022-01-05

Abstract: Objective: To investigate the effects of eukaryotic elongation factor 2 kinase (eEF-2K) gene transfection and sodium tanshinone IIA sulfonate (STS) on proliferation, invasion and migration of lung adenocarcinoma A549 cells and its mechanism. Methods: After treated with 0, 1.25, 2.5, 5, 10 and 20 μg/mL STS for 48 h, the survival rate of A549 cells was detected by cell counting kit-8 (CCK-8) method to screen the concentration of STS. After siRNA targeting eEF-2K gene (siRNA-eEF-2K) was transfected into A549 cells, the expression levels of eEF-2K protein and mRNA in A549 cells were detected by Western blot and real-time quantitative PCR. A549 cells were divided into siRNA-NC group (transfected with siRNA-NC), siRNA-eEF-2K group (transfected with siRNA-eEF-2K), siRNA-NC + STS group (treated with 10 μg/mL STS after siRNA-NC) and siRNA-eEF-2K + STS group (treated with 10 μg/mL STS after siRNA-eEF-2K), the survival rate, clone formation rate, number of transmembrane cells, migration rate and apoptosis rate of A549 cells were detected by CCK-8 method, clone formation test, Transwell chamber test, scratch test and flow cytometry, the protein expression levels of protein kinase B (AKT), phosphorylation (p)-AKT, Ki67, matrix metalloproteinase-2 (MMP-2) and B-lymphoma-2 genes (Bcl-2) were detected by Western blot. Results: Compared with 0 μg/mL STS, the survival rate of A549 cells was significantly decreased after treatment with 1.25, 2.5, 5, 10 and 20 μg/mL STS (P < 0.05), and it was in a concentration dependent manner; the 50% inhibitory concentration of STS on A549 cells was 10.39 μg/mL. Transfection of siRNA-eEF-2K can successfully inhibit the expression of eEF-2K protein and mRNA in A549 cells. Compared with those in siRNA-NC group, the cell survival rate, clone formation rate, number of transmembrane cells, migration rate and p-AKT, Ki67, MMP-2 and Bcl-2 in proteins siRNA-eEF-2K + STS group were significantly lower, while the apoptosis rate was significantly higher (P < 0.05), the changes of the above indexes in siRNA-eEF-2K + STS group were significantly greater than those in siRNA-NC + STS group and siRNA-eEF-2K group. Conclusions: The eEF-2K gene silencing combined with STS can synergistically inhibit the proliferation, invasion and migration of A549 cells and promote cell apoptosis. The mechanism may be related to the inhibition of protein expression of p-AKT, Ki67, MMP-2 and Bcl-2.

Key words: lung adenocarcinoma, eukaryotic elongation factor 2 kinase, sodium tanshinone IIA sulfonate, cell proliferation, cell invasion, apoptosis