Effect and mechanism of siRNA targeting α-enolase gene combined with paclitaxel on proliferation, invasion and apoptosis of hepatocellular carcinoma cell
10.3760/cma.j.cn112152-20191225-00840
- VernacularTitle:抑制α-烯醇化酶的表达联合紫杉醇对肝癌细胞增殖侵袭和凋亡的影响
- Author:
Hong CUI
1
;
Qinqin GAO
;
Hao ZHUANG
;
Tao HE
;
Baishun WAN
;
Xiaoqian WANG
;
Ling ZHANG
;
Tao HUANG
;
Feng HAN
Author Information
1. 郑州大学附属肿瘤医院 河南省肿瘤医院肝胆胰外科 450052
- Keywords:
Hepatocellular neoplasms;
Αlpha-enolase;
Paclitaxel;
Cell proliferation;
Cell invasion;
Apoptosis;
PI3K/Akt signaling pathway
- From:
Chinese Journal of Oncology
2021;43(12):1241-1247
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the effect of siRNA targeting inhibition of α-enolase (ENO1) combined with paclitaxel on the proliferation, invasion and apoptosis of hepatocellular carcinoma SK-HEP-1 cell and its mechanism.Methods:siRNA-ENO1 (siRNA-ENO1 group) and siRNA-negative control (siRNA-NC group) were transfected into SK-HEP-1 cells in vitro, the untransfected SK-HEP-1 cells were used as the control group, and the transfection effect was detected by real-time fluorescent quantitative polymerase chain reaction and western blotting. After SK-HEP-1 cells were treated with 0, 2.5, 5, 10, 20 and 40 μg/L paclitaxel for 48 hours, the cell survival rate was measured by 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2H tetrazolium bromide (MTT) method and the semi inhibitory concentration of paclitaxel was calculated. SK-HEP-1 cells transfected with siRNA-ENO1 or siRNA-NC were treated with 10 μg/L paclitaxel as paclitaxel+ siRNA-ENO1 group and paclitaxel+ siRNA-NC group. The proliferation, clonogenesis, invasion and apoptosis of siRNA-NC group, siRNA-ENO1 group, paclitaxel+ siRNA-ENO1 group and paclitaxel+ siRNA-NC group were detected by MTT, clonogenesis, Transwell chamber and flow cytometry respectively. The expression levels of the phosphorylation of phosphatidylinositol-3-kinase (p-PI3K), p-protein kinase B (Akt) and proliferating cell nuclear antigen (PCNA), matrix metalloproteinase 9 (MMP-9) and B lymphocytoma-2 gene (Bcl-2) were detected by western blotting. Results:Compared with the control group (1.00±0.00 and 0.69±0.04, respectively), the expression levels of ENO1 mRNA and protein (0.25±0.03 and 0.23±0.02, respectively) in siRNA-ENO1 group decreased significantly ( P<0.05), but there were no significant differences in the expression levels of ENO1 mRNA and protein in siRNA-NC group ( P>0.05). Compared without treatment group [(100.00±0.00)%, P<0.05], the survival rates of SK-HEP-1 cells treated with 2.5, 5, 10, 20 and 40 μg/L paclitaxel [(88.65±6.46)%, (72.36±6.08)%, (60.48±4.23)%, (38.52±3.56)% and (20.75±2.32)%, respectively] decreased significantly ( P<0.05), and the semi inhibitory concentration of paclitaxel was 13.26 μg/L. The cell survival rate and clone formation rate of siRNA-ENO1 group [(68.86±5.12)% and (18.12±2.25)%, respectively] were lower than those of siRNA-NC group [(100.00±0.00)% and (29.65±3.06)%, respectively, P<0.05]. The cell survival rate and clone formation rate of the paclitaxel+ siRNA-ENO1 group [(43.28±2.64)% and (8.72±0.52)%, respectively] were significantly different from those of the paclitaxel+ siRNA-NC group [(61.75±5.06)% and (13.48±2.16)%, respectively, P<0.05] and siRNA-ENO1 groups [(68.86±5.12)% and (18.12±2.25)%, respectively, P<0.05]. Cell invasion number in paclitaxel+ siRNA-ENO1 group (23.64±2.12) was lower than that in siRNA-ENO1 group and paclitaxel+ siRNA-NC group (42.16±2.75 and 37.35±2.42, respectively, P<0.05). The apoptosis rates of paclitaxel+ siRNA-NC group and siRNA-ENO1 group [(17.49±1.35)% and (15.29±1.50)%, respectively] were higher than that of siRNA-NC group [(7.21±0.70)%, P<0.05]. The apoptosis rate in the paclitaxel+ siRNA-ENO1 group [(24.59±2.40)%] was higher than those in the paclitaxel+ siRNA-NC group and siRNA-ENO1 group [(17.49±1.35)% and (15.29±1.50)%, respectively, P<0.05]. The expression levels of ENO1, PI3K/Akt signaling pathway related proteins including p-PI3K and p-Akt and the expression levels of PCNA, MMP-9 and Bcl-2 in siRNA-ENO1 group and paclitaxel+ siRNA-NC group were lower than those in siRNA-NC group ( P<0.05). The expression levels of ENO1, p-PI3K, p-Akt, PCNA, MMP-9 and Bcl-2 in paclitaxel+ siRNA-ENO1 group were lower than those in siRNA-ENO1 group or paclitaxel+ siRNA-NC group ( P<0.05). Conclusion:siRNA targeting inhibition of ENO1 expression can enhance the inhibitory effect of paclitaxel on proliferation, invasion and apoptosis of SK-HEP-1 cells, and its mechanism may be related to the inhibition of PI3K/AKT signaling pathway.
