间充质干细胞条件培养基增强肺癌多倍体肿瘤巨细胞化学治疗药物敏感性的研究  

Enhancement of chemosensitivity of lung cancer polyploid giant cancer cells by mesenchymal stem cell derived conditioned medium

作　　者：王丽丽 邢思宁 杨紫恩 欧阳明玥 赵松 刘硕 孙凌燕 于卉影 Wang Lili;Xing Sining;Yang Zien;Ouyang Mingyue;Zhao Song;Liu Shuo;Sun Lingyan;Yu Huiying(Basic Medical Laboratory,General Hospital of Northern Theater Command,Shenyang 110016,China)

机构地区：[1]北部战区总医院基础医学实验室,沈阳110016

出　　处：《中国临床实用医学》2022年第2期17-22,共6页China Clinical Practical Medicine

基　　金：辽宁省科学技术计划项目(2020JH2/10300160)。

摘　　要：目的探讨间充质干细胞条件培养基(MSCs-CM)对肺癌多倍体肿瘤巨细胞(PGCC)化学治疗药物敏感性的影响。方法使用1 μmol/L多西他赛处理A549细胞24 h(多西他赛A组), 撤除药物后继续在完全培养基中培养至第3天(多西他赛B组), 建立肺癌PGCC模型, 使用MSCs-CM培养PGCC 48 h(PGCC+MSCs-CM组), 以正常培养的A549细胞(A549组)或PGCC(PGCC组)作为对照, 流式细胞术检测细胞DNA含量, 细胞计数试剂盒(CCK-8)法检测A549细胞和PGCC对不同浓度多西他赛和顺铂的敏感性以及MSCs-CM对PGCC耐药性的影响, Western blot法检测相关蛋白表达。结果多西他赛B组细胞体积明显增大, 细胞核呈多核状态, PGCC亚群(DNA含量>4 N)比例均明显高于DMSO组和多西他赛A组(P<0.05), 成功建立肺癌PGCC模型, 将多西他赛B组作为PGCC细胞。CCK-8检测结果显示, 多西他赛对A549细胞和PGCC的IC50值分别为(0.20±0.04)μmol/L和(11.15±2.47)μmol/L, 差异有统计学意义(P<0.05), 顺铂对A549细胞和PGCC的IC50值分别为(14.87±3.72)μmol/L和(30.03±4.59)μmol/L, 差异有统计学意义(P<0.05), PGCC对多西他赛和顺铂的耐药指数分别为55.75倍和2.02倍。与PGCC组相比, PGCC+MSCs-CM+多西他赛组吸光度值明显下降(P<0.05), 且明显低于PGCC+多西他赛组和PGCC+MSCs-CM组(P<0.05)。与PGCC组相比, PGCC+MSCs-CM+顺铂组吸光度值明显下降(P<0.05), 且明显低于PGCC+顺铂组和PGCC+MSCs-CM组(P<0.05)。Western blot检测结果显示, 与A549组相比, PGCC组自噬微管相关蛋白轻链3A/B(LC3A/B)-Ⅰ/LC3A/B-Ⅱ、B淋巴细胞瘤-2基因(BCL-2)、多药耐药性相关蛋白1(MPR1)表达明显上调(P<0.05), 兔抗人选择性自噬接头蛋白1(SQSTM1/P62)、BCL-2相关X蛋白(BAX)表达明显下调(P<0.05);与PGCC组相比, PGCC+MSCs-CM组LC3A/B-Ⅰ/LC3A/B-Ⅱ、BCL-2、MRP1表达明显下调(P<0.05), SQSTM1/P62、BAX表达明显上调(P<0.05), 且MRP1表达水平显著低于A549组(P<0.05)。结论肺癌PGCC对化学治疗药物的耐药性Objective To investigate the effect of mesenchymal stem cell derived conditioned medium(MSCs-CM)on the chemosensitivity of lung cancer polyploid giant cancer cells(PGCC).Methods A549 cells were treated with 1μmol/L docetaxel for 24 hours(docetaxel A group).After removing the drug,the cells were cultured in complete medium until the third day(docetaxel B group).The lung cancer PGCC model was established.PGCC was cultured with MSCs-CM for 48 hours(PGCC+MSCs-CM group).Normally cultured A549 cells(A549 group)or PGCC(PGCC group)were used as control.The cell DNA content was detected by flow cytometry.The sensitivity of A549 cells and PGCC to different concentrations of docetaxel and cisplatin and the effect of MSCs-CM on PGCC drug resistance were detected by cell counting kit-8(CCK-8)method.The expression of related proteins was detected by Western blot.Results The cell volume of docetaxel B group was increased significantly,the nucleus was multinucleated,and the proportion of PGCC subsets(DNA content>4 N)was significantly higher than that of DMSO group and docetaxel A group(P<0.05).The PGCC model of lung cancer was successfully established,and docetaxel B group was used as the PGCC.The results of CCK-8 showed that the IC50 values of docetaxel on A549 cells and PGCC were(0.20±0.04)μmol/L and(11.15±2.47)μmol/L,and the differences were statistically significant(P<0.05).The drug resistance indexes of PGCC to docetaxel and cisplatin were 55.75 and 2.02 times,respectively.Compared with PGCC group,the absorbance value of PGCC+MSCs-CM+docetaxel group was decreased significantly(P<0.05),and was significantly lower than that of PGCC+docetaxel group and PGCC+MSCs-CM group(P<0.05).Compared with PGCC group,the absorbance value of PGCC+MSCs-CM+cisplatin group was decreased significantly(P<0.05),and was significantly lower than that of PGCC+cisplatin group and PGCC+MSCs-CM group(P<0.05).Western blot showed that compared with A549 group,the expressions of microtubule-associated protein light chain 3 A/B(LC3A/B)-Ⅰ/LC3A/B-Ⅱ,B-

关 键 词：间充质干细胞 肺癌 多倍体巨癌细胞 化学治疗药物敏感性 

分 类 号：R734.2[医药卫生—肿瘤]