Wang Lei, Mi Yuan, Zhang Xin-Fei, Li Xing, He Cai-Yi, Li Chao, Liu Liang
Department of Thoracic Surgery, the Fourth Hospital of Hebei Medical University, Shijiazhuang 050011, China.
Tumor Institute, the Fourth Hospital of Hebei Medical University, Shijiazhuang 050011, China.
Sichuan Da Xue Xue Bao Yi Xue Ban. 2022 May;53(3):452-456. doi: 10.12182/20220560201.
To investigate the regulatory role of extracellular vesicles (EVs) carrying ATP binding cassette transporter G2 (ABCG2) on the drug resistance of lung adenocarcinoma cells and the relevant molecular mechanisms.
A549 cells, human lung adenocarcinoma cells, were used to form cisplatin (or cis-Diaminedichloroplatinum, CDDP)-resistant lung adenocarcinoma cells, i.e., A549/CDDP cells. EVs from A549 and A549/CDDP cells were extracted by gradient centrifugation method and were hence named EVs and EVs , respectively. The A549 cells were treated with EVs and EVs for 48 hours, and the cells were named A549-EVs and A549-EVs cells, respectively. A549/ 2 cells were established by transfecting A549 cells with pCDNA3.1- 2 recombinant plasmids. On the other hand, A549 cells transfected with empty vectors were named A549/pCDNA3.1 cells. MTT assay was conducted to calculate the 24-hour cell drug resistance index for CDDP. The 2 gene expression in cells and EVs were assessed with real-time PCR. A549 and A549-EVs cells were transplanted subcutaneously into nude mice, which were labeled the control group and the experimental group accordingly. After tumor formation, 3 mg/kg CDDP was intraperitoneally injected once a week for two times. The 2 gene expression of subcutaneous transplanted tumor cells was examined by real-time PCR. The cell apoptosis rate of subcutaneous transplanted tumor cells was examined by flow cytometry.
Using the parental A549 cells as reference, the 24-h CDDP-resistance indexes of 549/CDDP, A549/ 2, A549/pCDNA3.1, A549-EVs , A549-EVs cells were 7.17, 10.06, 1.02, 1.19 and 5.40, respectively. When comparing the 2 gene expression levels in all cells and EVs, the findings were higher in A549/CDDP cells than those inA549 cells, higher in A549/ 2 cells than those in A549/pCDNA3.1 or A549 cells, higher in EVs than those in EVs , and higher in A549-EVs than those in A549-EVs cells ( <0.01) . The volume of transplanted tumor and the 2 gene expression level in the experimental group were higher than those in the control group, while the apoptosis rate was lower than that in the control group ( <0.01).
EVs carrying ABCG2 gene can regulate the drug resistance of lung adenocarcinoma cells.
探讨携带ATP结合盒转运蛋白G2（ABCG2）的细胞外囊泡（EVs）对肺腺癌细胞耐药性的调控作用及相关分子机制。
采用人肺腺癌细胞A549构建顺铂（或顺二氯二氨铂，CDDP）耐药的肺腺癌细胞，即A549/CDDP细胞。采用梯度离心法提取A549和A549/CDDP细胞的EVs，分别命名为EVs 和EVs 。将A549细胞分别用EVs 和EVs 处理48小时，处理后的细胞分别命名为A549-EVs 和A549-EVs 细胞。通过将A549细胞转染pCDNA3.1- 2重组质粒构建A549/ 2细胞。另一方面，将转染空载体的A549细胞命名为A549/pCDNA3.1细胞。采用MTT法计算24小时细胞对CDDP的耐药指数。采用实时荧光定量PCR检测细胞和EVs中 2基因的表达。将A549和A549-EVs 细胞皮下移植到裸鼠体内，分别标记为对照组和实验组。待肿瘤形成后，每周腹腔注射3 mg/kg CDDP，共注射2次。采用实时荧光定量PCR检测皮下移植瘤细胞中 2基因的表达。采用流式细胞术检测皮下移植瘤细胞的凋亡率。
以亲本A549细胞为参照，549/CDDP、A549/ 2、A549/pCDNA3.1、A549-EVs 、A549-EVs 细胞24小时对CDDP的耐药指数分别为7.17、10.06、1.02、1.19和5.40。比较所有细胞和EVs中 2基因的表达水平，结果显示A549/CDDP细胞高于A549细胞，A549/ 2细胞高于A549/pCDNA3.1细胞或A549细胞，EVs 高于EVs ，A549-EVs 高于A549-EVs 细胞（ <0.01）。实验组移植瘤体积和 2基因表达水平高于对照组，而凋亡率低于对照组（ <0.01）。
携带ABCG2基因的EVs可调控肺腺癌细胞的耐药性。
