Effect of Bmi-1 on Multidrug Resistance in K562/ADR Cells and Its Mechanisms.
10.19746/j.cnki.issn.1009-2137.2020.03.008
- Author:
Bao-Xia ZHAO
1
;
Si-Qi LIU
2
;
Si-Cong DONG
1
;
Ru-Nan JING
1
;
Xiu-Xiang MENG
3
Author Information
1. Department of Clinical Hematology, Laboratorial Medicine College of Dalian Medical University, Dalian 116044, Liaoning Province, China.
2. Department of Laboratorial Medicine, Municipal Hospital Affiliated to Xuzhou Medical College，Xuzhou 221005, Jiangsu Province, China.
3. Department of Clinical Hematology, Laboratorial Medicine College of Dalian Medical University, Dalian 116044, Liaoning Province, China,E-mail: xiuxiang_meng@sina.com.
- Publication Type:Journal Article
- MeSH:
Doxorubicin;
Drug Resistance, Multiple;
Drug Resistance, Neoplasm;
Humans;
K562 Cells;
Mitogen-Activated Protein Kinase 7
- From:
Journal of Experimental Hematology
2020;28(3):758-766
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVE:To investigate the effect of Bmi-1 gene silencing on drug resistance of leukemia cell K562/ADR and to explore its possible mechanism.
METHODS:After two sequences of Bmi-1-siRNA were transfected into drug-resistant K562/ADR cells, the mRNA and protein expressions of Bmi-1 gene were detected. After Bmi-1 gene silencing the expression of P-gp and MDR1 were detected and the accumulation of doxorubicin in K562/ADR cells were detected by flow cytometry to determine the effect of Bmi-1 gene silencing on drug resistance of K562/ADR cells. The protein expression of NF-κB was analyzed after Bmi-1 gene silencing. Then after K562/ADR cells were treated with NF-κB inhibitor PDTC, the protein expression of P-gp and its functional changes were analyzed to determine the effect of NF-κB on drug resistance of leukemia cells. The protein expressions of PTEN, AKT and p-AKT after Bmi-1 gene silencing were detected and the effect of Bmi-1 gene silencing on PTEN/PI3K/AKT signaling pathway in drug-resistant cells was determined. After K562/ADR cells were treated with PI3K/AKT pathway inhibitor LY294002, the protein expressions of NF-κB and P-gp were analyzed to determine the regulation of AKT on the expression of NF-κB and P-gp. The protein expressions of AKT, p-AKT, NF-κB and P-gp were detected after the Bmi-1-siRNA transfected cells were treated by PTEN inhibitor BPV. Above-mentioned expression of mRNA was detected by RT-PCR, and the protein expression was detected by Western blot.
RESULTS:The expression of Bmi-1 gene in K562/ADR cells decreased at both mRNA and protein levels and the doxorubicin accumulation increased after Bmi-1 gene silencing. The expression of MDR1/P-gp in Bmi-1-siRNA transfected cells was lower than that in K562/ADR cells (P＜0.05). After Bmi-1 gene silencing, the activity of NF-κB decreased. The activity of NF-κB and P-gp expression was inhibited and the function of P-gp in K562/ADR cells was reduced by using NF-κB inhibitor (PDTC). The protein expression of PTEN increased while the protein expression of p-AKT decreased after Bmi-1 gene silencing (P＜0.05). The protein expressions of p-AKT, P-gp and the activity of NF-κB were inhibited significantly by using PI3K/AKT inhibitor LY294002 (P＜0.05). After the Bmi-1-siRNA transfected cells were treated by PTEN inhibitor BPV, the activity of NF-κB and the protein expressions of P-gp were restored.
CONCLUSION:Bmi-1 plays a key role in MDR-mediated multidrug resistance in K562/ADR cells, which may be mediated by activating PTEN/AKT pathway to regulate NF-κB.
