解剖学报

解剖学报 ›› 2018, Vol. 49 ›› Issue (5): 624-629.doi: 10.16098/j.issn.0529-1356.2018.05.010

• 肿瘤生物学 • 上一篇    下一篇

沉默DLL3基因增强白血病K562/ADM细胞对阿霉素的敏感性

王世广1* 司旭艳2 王丽君1 仝雷3 李阳杰4   

  1. 1. 郑州工业应用技术学院医学院医学机能教研室，河南 新郑 451150; 2. 郑州工业应用技术学院临床基础教研室，河南 新郑 4511503;   3. 郑州工业应用技术学院医学形态学教研室，河南 新郑 4511503; 4. 郑州工业应用技术学院药学教研室，河南 新郑 451150
  • 收稿日期:2017-11-20 修回日期:2018-01-28 出版日期:2018-10-06 发布日期:2018-10-06
  • 通讯作者: 王世广 E-mail:wsgsxy@163.com

Enhancing effect of DLL3 gene silencing on the sensibility in human leukemia K562/ADM cells to adriamycin

WANG Shi-guang 1* SI Xu-yan2 WANG Li-jun1 TONG Lei3 LI Yang-jie4   

  1. 1. Department of Medical Function, Medical College, Zhengzhou University of Industrial Technology, He’nan Xinzheng 451150, China;  2. Department of Clinical Foundation, Medical College, Zhengzhou University of Industrial Technology, He’nan Xinzheng 451150, China; 3. Department of Medical Morphology, Medical College, Zhengzhou University of Industrial Technology, He’nan Xinzheng 451150, China;   4. Department of Pharmacy, Medical College, Zhengzhou University of Industrial Technology, He’nan Xinzheng 451150, China
  • Received:2017-11-20 Revised:2018-01-28 Online:2018-10-06 Published:2018-10-06
  • Contact: WANG Shi-guang E-mail:wsgsxy@163.com

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摘要：

目的 探讨沉默Delta-like ligand 3（DLL3）基因对白血病K562/ADM细胞对阿霉素（ADM）耐药性的影响及其分子机制。 方法 将干扰人DLL3基因表达的shRNA质粒和无义对照质粒转染K562/ADM细胞，采用RT-PCR法检测DLL3的mRNA表达水平，采用CCK-8法检测ADM对K562和K562/ADM细胞的毒性作用，采用流式细胞术检测细胞凋亡和细胞内ADM浓度，采用Western blotting方法检测DLL3、谷胱甘肽S转移酶-π（GST-π）和P-糖蛋白（P-gp）的蛋白表达水平。 结果 K562/ADM细胞DLL3的mRNA和蛋白表达水平均显著高于其亲代K562细胞（P<0.05）；ADM对K562和K562/ADM细胞的IC50 分别为1.08 mg/L和34.93 mg/L；沉默DLL3基因后，K562/ADM细胞的耐药倍数下降至13.12，反转倍数为2.47；尽管抑制DLL3基因表达未对K562/ADM细胞凋亡产生影响，但可下调P-gp和GST-π的蛋白表达水平（P<0.05），增加K562/ADM细胞内ADM的蓄积量（P<0.05），从而增强ADM诱导的K562/ADM细胞凋亡（P<0.05）。 结论 沉默DLL3基因可反转K562/ADM细胞对ADM的耐药性，这可能与下调P-gp和GST-π蛋白水平、从而减少K562/ADM细胞内阿霉素蓄积量有关。

关键词: DLL3基因, K562/ADM细胞, P-糖蛋白, 谷胱甘肽S转移酶-π, 反转录聚合酶链反应, 白血病, 人

Abstract:

Objective To investigate whether Delta-like ligand 3 (DLL) gene silencing could influence the sensibility of human leukemia K562/ADM cells to adriamycin (ADM) in vitro . Methods K562/ADM cells were stably transfected with specific shRNA interference plasmid vector targeting for DLL3. The mRNA expression level of DLL3 were measured by RT-PCR. CCK-8 assay was employed to detect the cytotoxic effect of ADM in K562 and K562/ADM cells. The apoptosis and intracellular ADM concentration were analyzed by flow cytometry. Western blotting was performed to determine the protein expression levels of DLL3, glutathione S transferases-π, (GST-π) and P-glycoprotein (P-gp). Results K562/ADM cells had a significantly higher mRNA and protein expression level of DLL3 than K562 cells (P<0.05). The IC50 value of K562 and K562/ADM cells to ADM were 1.08 and 34.93 mg/L respectively. After DLL3 gene silencing, the resistant factor of K562/ADM cells declined to 13.12 with a reversal fold of 2.47. DLL3 gene silencing had no effect on apoptosis of K562/ADM cells, but a significant effect on downregulating protein levels of P-gp and GST-π (P<0.05) and increasing intracellular ADM concentration (P<0.05), and resulting in an enhance of ADM-induced apoptosis (P<0.05). Conclusion DLL3 gene silencing may enhance the sensibility of K562/ADM cells to ADM by down-regulating P-gp and GST-π, and increasing intracellular ADM concentration.

Key words: DLL3 gene, K562/ADM cell, P-glycoprotein, Glutathione S transferases-π, RT-PCR, Leukemia, Human