人脐带间充质干细胞及其条件培养液对非小细胞肺癌多倍体A549细胞增殖、迁移和凋亡的影响  

Effects of human umbilical cord⁃derived mesenchymal stem cells and their conditioned medium on proliferation,migration and apoptosis of non⁃small cell lung cancer polyploid A549 cells

作　　者：欧阳明玥 王丽丽 邢思宁 赵松 刘硕 于卉影 Ouyang Mingyue;Wang Lili;Xing Sining;Zhao Song;Liu Shuo;Yu Huiying(Department of 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：《肿瘤研究与临床》2022年第1期8-14,共7页Cancer Research and Clinic

基　　金：辽宁省重点研发指导计划(2019JH8/10300082)。

摘　　要：目的探讨人脐带间充质干细胞(hUC‑MSC)及其条件培养液对人非小细胞肺癌多倍体A549细胞增殖、迁移和凋亡的影响。方法取对数生长期的A549细胞,采用1μmol/L多西他赛诱导24 h后,更换为含10%胎牛血清的DMEM/F12培养液继续培养3 d,建立多倍体A549细胞模型。分离与培养hUC‑MSC,制备hUC‑MSC条件培养液。正常培养的多倍体A549细胞作为对照组;条件培养液培养的多倍体A549细胞为条件培养液组;hUC‑MSC与多倍体A549细胞共培养,细胞总数比分别为2∶1(MSC 1组)和5∶1(MSC 2组)。各组细胞继续培养48 h或72 h。流式细胞术检测多倍体A549细胞增殖和凋亡情况,Transwell实验检测细胞迁移能力,蛋白质印迹法检测迁移和凋亡相关蛋白的表达情况。结果成功建立多倍体A549细胞模型,分离培养出hUC‑MSC。流式细胞术检测细胞增殖结果示,培养48 h时对照组、条件培养液组、MSC 1组、MSC 2组平均荧光强度分别为1695±305、2020±85、1259±35、1356±33,差异有统计学意义(F=14.00,P<0.05);72 h时分别为1052±77、1309±24、864±201、1103±237,差异无统计学意义(F=3.90,P>0.05)。Transwell实验结果显示,培养48 h时对照组、条件培养液组、MSC 1组、MSC 2组迁移细胞数分别为(52±9)个、(57±12)个、(68±8)个、(75±11)个,差异有统计学意义(F=32.16,P<0.05);各实验组迁移细胞数均高于对照组(均P<0.05)。对照组、条件培养液组、MSC 1组、MSC 2组凋亡细胞比例分别为(15.53±4.27)%、(13.77±1.75)%、(3.60±0.50)%、(2.33±0.06)%,差异有统计学意义(F=182.36,P<0.05);条件培养液组与对照组比较差异无统计学意义(P>0.05);MSC 1组和MSC 2组分别与对照组比较,差异均有统计学意义(均P<0.05)。蛋白质印迹法检测结果显示,与对照组比较,条件培养液组、MSC 1组和MSC 2组中迁移相关蛋白基质金属蛋白酶9(MMP‑9)表达上调,促凋亡蛋白bax表达下调,抗凋亡蛋白bcl‑xL表达上调。结论hUC�Objective To investigate the effects of human umbilical cord‑derived mesenchymal stem cells(hUC‑MSC)and their conditioned medium on proliferation,migration and apoptosis of human non‑small cell lung cancer(NSCLC)polyploid A549 cells.Methods A549 cells in logarithmic phase were selected.After induction treatment with 1μmol/L docetaxel for 24 h,DMEM/F12 medium with 10%fetal bovine serum was used to culture the cells for 3 d,finally the polyploid A549 cells model was successfully established.After finishing the separation and culture of hUC‑MSC,hUC‑MSC conditioned medium was prepared.Normally cultured polyploid A549 cells were treated as the control group,conditioned medium cultured polyploid A549 cells were treated as the conditioned medium group.hUC‑MSC was co‑cultured with polyploid A549 cells,and the ratio of the total number of cells was 2:1 and 5:1,respectively,which were recorded as MSC 1 group and MSC 2 group.Cells in each group were continually cultured for 48 h or 72 h.Proliferation and apoptosis of polyploid A549 cells in each group were detected by using flow cytometry,cell migration ability was detected by using Transwell assay,and the expressions of migration and apoptosis‑related proteins were detected by using Western blotting.Results Polyploid A549 cells model was successfully established and hUC‑MSC was cultured separately.The result of cell proliferation detected by flow cytometry showed that at 48 h,the mean fluorescence intensity of the control group,conditioned medium group,MSC 1 group and MSC 2 group was 1695±305,2020±85,1259±35 and 1356±33,respectively,and the difference was statistically significant(F=14.00,P<0.05);at 72 h,the mean fluorescence intensity of the control group,conditioned medium group,MSC 1 group and MSC 2 group was 1052±77,1309±24,864±201 and 1103±237,respectively,and the difference was statistically significant(F=3.90,P>0.05).The result of Transwell assay showed that at 48 h,the number of cell migration in the control group,conditioned medium group

关 键 词：间质干细胞 培养液 条件性 癌 非小细胞肺 细胞增殖 细胞凋亡 

分 类 号：R734.2[医药卫生—肿瘤]