摘要
目的观察乙醛脱氢酶(ALDH)在A549/DDP人肺腺癌耐顺铂细胞中的表达及在顺铂耐药性中的功能。方法流式细胞术检测肿瘤干细胞标志分子CD44、CD90、CD73、CD105、上皮细胞黏附分子(Ep CAM)、ATP结合盒式超家族G成员2(ABCG2)以及ALDH在A549细胞和A549/DDP细胞表达的阳性率;MTT法检测100μmol/L二乙氨基苯甲醛(DEAB)处理前后A549/DDP细胞的增殖能力和对顺铂的敏感性;实时定量PCR检测ALDH各亚型在A549细胞与A549/DDP细胞中的mRNA水平的表达;Western blot法检测实时定量PCR有明显差异结果的ALDH的亚型ALDH1A3和ALDH1B1在A549细胞与A549/DDP细胞中的蛋白水平的表达。结果与A549细胞相比,ABCG2和ALDH在A549/DDP细胞的阳性率增加,ALDH阳性率达97.7%,在A549细胞中仅仅为4.3%;CD44、CD90、CD73、CD105在二者间阳性率无明显差异,而Ep CAM在A549/DDP细胞阳性率降低;经100μmol/L DEAB处理后,A549/DDP细胞对顺铂的耐药性显著低于未处理前;与A549细胞相比,ALDH1A3和ALDH1B1的mRNA在A549/DDP细胞中表达水平增高;ALDH1B1蛋白表达升高,ALDH1A3表达降低。结论 ALDH可作为顺铂耐药人肺腺癌细胞的标志分子之一,其中ALDH1B1在顺铂耐药性中可能发挥一定作用,为临床上治疗肺腺癌耐药患者提供新的治疗靶点和策略。
Objective To investigate the expression and biological significance of acetaldehyde dehydrogenase (ALDH) in cisplatin-resistant human A549/DDP lung adenocarcinoma cell line. Methods The expressions of biomarkers of cancer stem cells (CSCs) including CD44, CD73, CD90, CD105, epithelial cell adhesion molecule (EpCAM), ATP-binding cassette sub-family G member 2 (ABCG2) and ALDH in human lung adenocarcinoma A549 cells and cisplatin-resistant A549/DDP cells were ascertained by flow cytometry. The proliferation and sensitivity of A549/DDP cells to cisplatin were evaluated by MTT assay before and after ALDH inhibitor diethylaminobenzaldehyde (DEAB) treatment. The differential expression of ALDH subtypes in cells was determined by real-time quantitative PCR, and was further confirmed by a Western blotting. Results The frequencies of ABCG2 and ALDH positive cells were higher in cisplatin-resistant A549/DDP cells than those in A549 cells, in particular the expression of ALDH which was 97.7% in A549/DDP cells, and only 4.3% in A549 cells. There was no significantly differential expression of CD44, CD90, CD73, CD105 between A549/DDP cells and A549 cells, whereas less EpCAM was found in A549/DDP cells compared with A549 cells. Importantly, the ALDH inhibitor DEAB (100 μmol/L) was able to significantly reduce cisplatin resistance in A549/DDP cells. Compared with A549 cells, the expressions of ALDH subtypes ALDH1A3 and ALDH1B1 mRNA significantly went up in A549/DDP cells. Interestedly, Western blotting revealed that ALDH1B1 protein expression was elevated but ALDH]A3 protein was reduced in A549/DDP cells in comparison with A549 cells. Conclusion ALDH can serve as a useful surface marker for cisplatin-resistant human lung adenocarcinoma cells. ALDH1B1 may play a role in the chemoresistance of these cells. Therefore, ALDH1B1 may be an important target for developing novel therapeutic agents and strategies for patients with cisplatin-resistant lung adenocarcinoma.
出处
《细胞与分子免疫学杂志》
CAS
CSCD
北大核心
2015年第5期625-629,共5页
Chinese Journal of Cellular and Molecular Immunology
基金
宁夏自然科学基金(NZ1247)
关键词
A549/DDP
顺铂耐药
乙醛脱氢酶
A549/DDP cells
cisplatin resistance
acetaldehyde dehydrogenase