联合抑制聚腺苷酸二磷酸核糖转移酶-1及DNA依赖蛋白激酶催化亚单位增加KG-1α细胞对依托泊苷的敏感性  

Study of combined inhibition of PARP-1 and DNA-PKcs in increasing sensitivity of KG-1αcells to etoposide

作　　者：庄英婷 张灵玉 苏雪静 ZHUANG Ying-ting;ZHANG Ling-yu;SU Xue-jing(School of Pharmacy of Fujian Medical University,Fuzhou 350122,Fujian Province,China)

机构地区：[1]福建医科大学药学院,福建福州350122

出　　处：《中国临床药理学杂志》2020年第20期3302-3305,共4页The Chinese Journal of Clinical Pharmacology

基　　金：福建省自然科学基金资助项目(2017J01823);福建医科大学苗圃科研基金资助项目(2015MP007)。

摘　　要：目的观察联合抑制聚腺苷酸二磷酸核糖转移酶-1[poly(ADP-ribose)polymerase-1,PARP-1]及DNA依赖蛋白激酶催化亚单位(catalytic sunbunit of the DNA-dependent protein kinase,DNA-PKcs)对依托泊苷(etoposide,VP-16)作用于急性髓系白血病KG-1α细胞的影响。方法分别用特异性抑制剂奥拉帕尼(Olaparib,OLA)和Nu7441(NU)抑制PARP-1、DNA-PKcs。将KG-1α细胞设置成对照组、模型组、PARP-1抑制组(PARP-1 inhibition,VP-16+OLA)、DNA-PKcs抑制组(DNA-PKcs inhibition,VP-16+NU)、联合抑制组(combined inhibition,VP-16+OLA+NU)。用四甲基偶氮唑蓝(MTT)法体外检测联合用药对KG-1α细胞增殖抑制的影响;用高内涵荧光成像检测分析不同处理组的γ-H2AX在DNA双链断裂断裂位点的募集情况;用蛋白质印迹法检测细胞凋亡相关蛋白Cleaved caspase-3和Cleaved PARP的表达情况。结果对照组、模型组、PARP-1抑制组、DNA-PKcs抑制组和联合抑制组的增殖抑制率分别为(1.16±1.45)%,(23.88±3.15)%,(28.12±2.28)%,(17.21±0.89)%和(60.72±4.38)%;γ-H2AX阳性细胞百分比分别为γ-H2 AX阳性细胞百分比分别为(1.24±0.07)%,(22.51±2.24)%,(24.55±3.29)%,(23.28±2.48)%和(40.55±1.61)%;Cleaved Caspase-3蛋白相对表达量分别为0.16±0.10,0.61±0.30,1.15±0.64,1.11±0.31和1.72±1.03;Cleaved PARP蛋白相对表达量分别为1.23±0.09,1.45±0.55,2.03±0.84,2.08±0.41和2.66±0.95。模型组分别与对照组、联合抑制组比较,差异均有统计学意义(均P<0.05)。结论联合抑制PARP-1及DNA-PKcs能够在体外增加KG-1α细胞对依托泊苷的敏感性。Objective To observe the effect of combined inhibition of poly(ADP-ribose)polymerase-1(PARP-1)and catalytic sunbunit of the DNA-dependent protein kinase(DNA-PKcs)on etoposide(VP-16)acting on KG-1αcells in acute myeloid leukemia(AML).Methods Olaparib(OLA)and Nu7441(NU)were used to inhibit PARP-1 and DNA-PKcs,respectively.KG-1αcells were assigned to the control group,model group,PARP-1 inhibition group(VP-16+OLA),DNA-PKcs inhibition group(VP-16+NU)and the combined inhibition group(VP-16+OLA+NU).The effect of combined medication on the inhibition of KG-1αcells proliferation was tested in vitro by using MTT assay;γ-H2AX recruitment at DNA double-strand break sites in different treatment groups was detected and analyzed by high-content fluorescence imaging;and the expression of apoptosis-related proteins Cleaved caspase-3 and Cleaved PARP was detected by using Western Blotting.Results The proliferation inhibition rates of KG-1αcells in the control group,model group and the combined inhibition group were(1.16±1.45)%,(23.88±3.15)%,(28.12±2.28)%,(17.21±0.89)%and(60.72±4.38)%,respectively;The percentage ofγ-H2AX positive cells were(1.24±0.07)%,(22.51±2.24)%,(24.55±3.29)%,(23.28±2.48)%and(40.55±1.61)%,respectively;The relative expression of cleaved Caspase-3 was 0.16±0.10,0.61±0.30,1.15±0.64,1.11±0.31 and 1.72±1.03,respectively;The relative expression of cleared PARP was 1.23±0.09,1.45±0.55,2.03±0.84,2.08±0.41 and 2.66±0.95,respectively.There were statistically significant differences between the model group and the control group/combination group(P<0.05).Conclusion The combined inhibition of PARP-1 and DNA-PKcs can increase the sensitivity of KG-1αcells to etoposide in vitro.

关 键 词：依托泊苷 白血病 DNA损伤修复 聚腺苷酸二磷酸核糖转移酶-1 DNA依赖蛋白激酶的催化亚单位 经典非同源性末端接合 替代性末端连接 

分 类 号：R97[医药卫生—药品]