【摘要】 目的建立人肺腺癌耐药细胞模型 Anip973/NVB（去甲长春新碱）并鉴定其生物学特性。方法应用人肺腺癌细胞系 Anip973,采用 NVB 逐步增加剂量法,诱导建立耐药细胞模型Anip973/NVB,观察其生长规律;用 MTT 法鉴定抗药性;观察细胞形态和超微结构;流式细胞技术检测其细胞周期分布;高效液相色谱法测定细胞内 NVB 的浓度变化。结果经 MTT 法鉴定 Anip973/NVB 细胞较 Anip973细胞的 NVB 半数致死浓度(IC₅₀)增大21.81倍。两细胞系的倍增时间无明显差异。光镜及电镜下观察到,两细胞系结构变化较大。经流式细胞仪测定 Anip973/NVB 细胞,S 期细胞减少（P=0.035）而 G₀～G₁期细胞增多（P=0.014）;高效液相色谱法检测发现 Anip973细胞内药物浓度明显高于 Anip973/NVB。结论 Anip973/NVB 细胞是一个明确的多药耐药细胞模型,具有耐药细胞的基本生物学特性。更多还原

【Abstract】 Objective To establish a multidrug-resistant human lung adenocarcinoma cell line and to investigate its biologic characteristics.Methods NBV of the terminal concentration of 0.02mg/L was co- cultured with the human lung adenocarcinoma cells of the line Anip973.When the cells got a stable growth and generation the concentration of NVB was increased gradually till the 65th generation.Thus a drug- resistant line Anip973/NVB that could grow under the NVB of the concentration of 2.0mg/L was established.Anip973 and Anip973/NVB cells were co-cultured with 10 anti-cancer drugs:NVB,cisplastin, fluorouracil,gemcitabine,pacilitaxel,etopside,irinotecan,dacarbazine,ifosfamide,and pharmorubicin of different concentrations respectively.Forty-eight hours later MTT method was used to detect the 50% inhibition concentration(IC₅₀)values of different drugs.The doubling times of the Anip973 and Anip973/ NVB cells were calculated.Inverted microscopy and electron microscopy were used to observe the morphology of the cells.Flow cytometry was conducted to observe the cell cycle.The cells were cultured in the medium with NVB of the concentration of 2.0mg/L.High efficiency fluid chromatography was used to observe the drug concentration in the cells.Results Anip973/NVB cells showed resistance at different degrees to 8 drugs（all P<0.05）,except to gemcitabine and irinotecan.The doubling time of the Anip973 cells was 23.45h,not significantly different from that of the Anip973/NVB cells The percentage of the cells in the phase G₀～G₁ was 62.90% in the Anip973 cells,significantly higher than in the Anip973 cells （53.73%,P=0.014）,and the proportion of the cells in the phase S 28.32% in the Anip973/NVB cells , significantly lower than that in the Anip793 cells（38.25%,P=0.035）NVB could be detected in the Anip973 cells with a concentration of 0.22mg/L±0.05mg/L,however,could not be detected in the Anip973/NVB cells.MTT assay showed that the IC₅₀of Anip973/NVB cells was 21.81 times higher than Anip973 cells.Microscopy showed that the structure,especially the ultramicrostructure,of the Anip973/ NVB cells was more irregular than that of the Anip973 cells,and there were significant difference in ultramicrostructure.Conclusion A reliable multi-drug resistant human lung adenocarcinoma cell line Anip973/NVB has been successful established.更多还原