摘要
目的探讨单用肝再生磷酸酶-3(PRL-3)抗体或联合培美曲塞(Pem)对人肺腺癌细胞A549凋亡的作用机制。方法将体外培养的A549细胞分为4组:PRL-3组(PRL-3抗体,164 ng·mL^(-1))、Pem组(Pem, 180 ng·mL^(-1))、联合组(PRL-3抗体+Pem)、溶剂对照组(二甲基亚砜)和空白组(不给予药物),4组均培养24 h。流式细胞技术检测细胞凋亡情况,蛋白质印迹法检测周期蛋白E(CyclinE)、周期蛋白依赖性激酶2(CDK2)、B淋巴细胞瘤-2蛋白(Bcl-2)和Bcl-2相关X蛋白(Bax)水平(灰度值),酶联免疫吸附法检测细胞超氧化物歧化酶(SOD)和丙二醛(MDA)水平,DCFH-DA荧光探针检测活性氧(ROS)水平。结果空白组、溶剂对照组、PRL-3组、Pem组和联合组的细胞总凋亡率分别为(3.87±1.27)%,(3.99±1.32)%,(17.54±2.59)%,(12.27±2.22)%和(32.51±3.77)%;上述5组Cyclin E蛋白表达水平分别为24.72±1.24,23.08±1.28,14.47±1.35,16.78±1.77和9.35±1.21;上述5组CDK2蛋白表达水平分别为39.01±1.87,39.21±1.98,32.35±1.23,28.35±1.17和17.37±1.08;上述5组Bcl-2蛋白表达水平分别为37.89±3.88,36.69±3.79,22.47±3.32,25.57±3.65和15.66±3.54;上述5组Bax蛋白表达水平分别为8.58±0.54,8.87±0.64,18.25±0.79,23.89±1.24和34.99±1.65;上述5组SOD分别为(6.21±0.71),(6.17±0.58),(9.23±1.47),(8.25±0.98)和(17.58±1.45)U·mL^(-1);上述5组ROS分别为(3.74±0.44),(3.68±0.38),(1.22±0.25),(1.87±0.29),和(0.90±0.12)MFI of DCF;上述5组MDA分别为(13.56±1.58),(13.24±1.35),(7.78±0.95),(9.25±1.04)和(4.32±0.52)μmol·L^(-1)。3个给药组与空白组和溶剂对照组比较,各指标差异均有统计学意义(均P<0.05);联合组与PRL-3组和Pem组比较,各指标差异均有统计学意义(均P<0.05)。结论联合组可能通过抑制A549细胞周期进程,促进细胞凋亡,降低氧化应激水平,表现抗肿瘤活性。
Objective To investigate the effect and mechanism of liver regeneration phosphatase-3(PRL-3) antibody alone or combined with pemetrexed(Pem) on apoptosis of human lung adenocarcinoma cell A549. Methods The A549 cells cultured in vitro were divided into 4 groups: PRL-3 group(164 ng·mL^(-1)PRL-3 antibody), Pem group(Pem, 180 ng·mL^(-1)), combined group PRL-3 antibody + Pem solvent group( dimethyl sulfoxide) and blank group( without drug treatment),all four groups were cultured for 24 h.Flow cytometry was used to detect cell apoptosis. Western blotting was used to detect the levels( gray value) of the cyclinE protein,cyclin-dependent kinases-2( CDK2) protein,B-cell lymphoma-2( Bcl-2) protein,Bcl-2 associated X( Bax) protein. Enzyme-linked immunosorbent assay was used to detect the levels of superoxide dismutase( SOD) and malondialdehyde( MDA). DCFH-DA by fluorescence probe was used to detect the levels of reactive oxygen species( ROS). Results The total apoptosis rate of blank group,solvent group,PRL-3 group,Pem group and combined group were( 3. 87 ± 1. 27) %,( 3. 99 ± 1. 32) %,( 17. 54 ± 2. 59) %,( 12. 27 ± 2. 22) %,( 32. 51 ±3. 77) %;Cyclin E protein levels in the above five groups were 24. 72 ± 1. 24,23. 08 ± 1. 28,14. 47 ± 1. 35,16. 78 ± 1. 77,9. 35 ± 1. 21;CDK2 protein levels in the above five groups were 39. 01 ± 1. 87,39. 21 ± 1. 98,32. 35 ± 1. 23,28. 35 ± 1. 17,17. 37 ± 1. 08;The Bcl-2 protein levels in the above five groups were37. 89 ± 3. 88,36. 69 ± 3. 79,22. 47 ± 3. 32,25. 57 ± 3. 65,15. 66 ± 3. 54;Bax protein levels in the above five groups were 8. 58 ± 0. 54,8. 87 ± 0. 64,18. 25 ± 0. 79,23. 89 ± 1. 24,34. 99 ± 1. 65;SOD in the above five groups were( 6. 21 ±0. 71),( 6. 17 ±0. 58),( 9. 23 ±1. 47),( 8. 25 ±0. 98),( 17. 58 ±1. 45) U· mL^(-1);ROS in the above five groups were( 3. 74 ± 0. 44),( 3. 68 ± 0. 38),( 1. 22 ± 0. 25),( 1. 87 ± 0. 29),( 0. 90 ± 0. 12) MFI of DCF;and MDA in the above five groups were( 13. 56 ± 1. 58),( 13. 24 ± 1. 35),( 7. 78 ± 0. 95),( 9. 25
作者
朱超男
王帅
郑贸根
王国臣
李作生
侯静朴
ZHU Chao-nan;WANG Shuai;ZHENG Mao-gen;WANG Guo-cheng;LI Zuo-sheng;HOU Jing-bu(Department of Thoracic Surgery,Affiliated Hospital of North China University of Technology,Tangshan 063000,Hebei Province,China)
出处
《中国临床药理学杂志》
CAS
CSCD
北大核心
2022年第1期10-13,共4页
The Chinese Journal of Clinical Pharmacology
基金
河北省医学研究重点计划基金资助项目(20130059)。
