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目的:体外建立紫杉醇耐药人肺腺癌A549/TAX细胞系,并对其生物学特性进行研究。方法:采用逐步增加紫杉醇浓度结合低剂量持续诱导法,建立紫杉醇耐药人肺腺癌A549/TAX细胞系。MTT法描绘A549和A549/TAX细胞的生长曲线;流式细胞仪检测细胞周期并比较经紫杉醇诱导24h后A549和A549/TAX细胞平均凋亡率的差异;检测紫杉醇对A549/TAX细胞的半数抑制浓度、耐药系数(RI)及A549/TAX细胞对其他5种化疗药物(顺铂、长春新碱、表阿霉素、足叶乙甙及吉西他滨)的交叉耐药谱;RT-PCR半定量分析A549/TAX细胞凋亡基因(bcl-2)及耐药基因(MDR1、LRP、GST-π)的mRNA表达。结果:A549/TAX细胞生长较亲本细胞快,S期细胞增多[(51.61±0.48)%],G1期减少[(37.26±0.23)%],经紫杉醇(150μmol/L)诱导前后平均凋亡率差异与A549细胞相比差异有显著性。A549/TAX细胞对紫杉醇、长春新碱、表阿霉素及足叶乙甙的RI分别为21.3、12.91、5.88和4.79,而对顺铂(RI=1.06)和吉西他滨(RI=1.03)则无交叉耐药;A549/TAX细胞bcl-2、MDR1、LRPmRNA的表达均较亲本细胞显著增高(P<0.001),GST-πmRNA未见表达。结论:成功建立了紫杉醇耐药A549/TAX细胞系,该细胞抗药性能明显、稳定,推测其多药耐药性与bcl-2、MDR1和LRP高表达有关。
OBJECTIVE: To establish paclitaxel-resistant human lung adenocarcinoma A549 / TAX cell line in vitro and to study its biological characteristics. Methods: Paclitaxel-resistant human lung adenocarcinoma A549 / TAX cell line was established by gradually increasing paclitaxel concentration and low-dose continuous induction. MTT method was used to characterize the growth curve of A549 and A549 / TAX cells. The cell cycle was detected by flow cytometry and the apoptosis rates of A549 and A549 / TAX cells after paclitaxel induction were compared. The half inhibition of paclitaxel on A549 / TAX cells (RI) and the spectrum of drug resistance of A549 / TAX cells to the other 5 chemotherapeutic drugs (cisplatin, vincristine, epirubicin, etoposide and gemcitabine) The mRNA expressions of bcl-2 and MDR1, LRP and GST-π in A549 / TAX cells were analyzed. Results: The A549 / TAX cells grew faster than the parental cells, with an increase in S phase [(51.61 ± 0.48)%], a decrease in G1 phase (37.26 ± 0.23)%, and an average apoptotic rate after induction with paclitaxel (150μmol / L) The difference was significant compared with A549 cells. The RI of A549 / TAX cells to paclitaxel, vincristine, epirubicin and etoposide were 21.3, 12.91, 5.88 and 4.79, respectively, but not for cisplatin (RI = 1.06) and gemcitabine (RI = 1.03) The expression of bcl-2, MDR1 and LRPmRNA in A549 / TAX cells was significantly higher than that in parental cells (P <0.001), but no expression was found in GST-πmRNA. CONCLUSION: The paclitaxel-resistant A549 / TAX cell line was established successfully. The drug resistance of the A549 / TAX cell line was obvious and stable. It is speculated that multidrug resistance is related to the high expression of bcl-2, MDR1 and LRP.