MiR-133b Affect the Proliferation and Drug Sensitivity in A549 Lung Cancer Stem Cells by Targeting PKM2
10.3779/j.issn.1009-3419.2017.06.02
- VernacularTitle:miR-133b靶向PKM2基因对肺癌A549干细胞增殖及药物敏感性的影响
- Author:
MI YONGHUA
1
;
HE MIAO
;
LIU BEIZHONG
Author Information
1. 重庆医科大学附属医院永川医院检验科
- Keywords:
MiR-133b;
A549;
PKM2;
DDP
- From:
Chinese Journal of Lung Cancer
2017;20(6):376-381
- CountryChina
- Language:Chinese
-
Abstract:
Background and objective It has been proven that miR-133b could inhibit cancer cell growth, the expression level of miR-133b was significant reduction in lung cancer tissue and serum of patients, and increase the radiation sensitivity of squamous cell carcinoma by targeting PKM2, but the exist mechanisms is not clear. The aim of this study is to explore the effect of miR-133b on proliferation in A549 lung cancer stem cells and drug sensitivity in DDP, and to explore the relationship between miR-133b and PKM2 gene, as well as the effect of cancer stem cells. Methods Using miRBase and miRNAMap database to sequence comparison miR-133b and PKM2 gene. Using im-mune magnetic separation method to select the CD133+/CD34+lung cancer stem cells from A549 cells, and using flow cytometry to detect the purity. The expression of miR-133b mRNA was detected by real-time fluorescence quantita-tive PCR (qRT-PCR). Cell proliferation was detected by CCK8 assay. 15 μg/mL DDP was treated to cells which was transfected with miR-133b, and apoptosis was detected by flow Cytometry at 0 h, 12 h, 24 h, 72 h. The expression of PKM2 protein was detected by Western blot. Results Gene binding site report that PKM2 gene may be the target gene of miR-133b; the results of flow cytometry showed that the purity of CD133+/CD34+ stem cells was (92.15±4.27)%. qRT-PCR results showed that compared with the control group, after overexpression of miR-133b, miR-133b was up-regulated and miR-133b was down regulated after miR-133b inhibition (P<0.05). Compared with the control group, cell proliferation of miR-133b mimics group was significantly decreased (P<0.05), PKM2 protein levels were signifi-cantly lower (P<0.05); and cell proliferation of the miR-133b inhibitor group and PKM2 level was increased (P<0.05). The apoptosis of miR-133b mimics group was significantly higher than that of control group (P<0.05) after DDP treat-ment with 12 h. The expression of PKM2 protein in miR-133b mimics+DDP group was significantly lower than that in control group (P<0.05). Conclusion Overexpression of miR-133b can inhibit the growth and proliferation of lung cancer stem cells by down regulating PKM2, and can enhance the sensitivity of lung cancer stem cells to DDP.
