Gu Wei, Zhang Ya-Ni, Li Bai, Han Jie, Cheng Bin-Bin, Ling Chang-Quan
Department of Traditional Chinese Medicine, Changhai Hospital, Second Military Medical University, Shanghai 200433, China.
Zhong Xi Yi Jie He Xue Bao. 2006 May;4(3):265-70.
To establish a multidrug-resistant cell line BEL-7402/5-FU of hepatocellular carcinoma (HCC).
BEL-7402/5-FU was induced by pulse therapy combined with continuous stepwise exposure to 5-fluorouracil in vitro. MTT assay was used to determine its multidrug resistance (MDR). Biological characteristics of the BEL-7402/5-FU cell line were observed including morphological changes, cell growth curve, population doubling time, plate cloning efficiency, adherence rate, cell cycle distribution, chromosome and tumorigenicity. Accumulation amount of adriamycin (ADM) in cytoplasm was measured by flow cytometry. The protein expression of thymidylate synthase (TS) was evaluated by immuno-cytochemical method.
The acquired MDR cell line of BEL-7402/5-FU was established successfully. The BEL-7402/5-FU cells showed cross-resistance to ADM, vincristine (VCR), methotrexate (MTX) and oxaliplatin (OHP), whereas still sensitive to hydroxycamptothecin (HCPT). The BEL-7402/5-FU cells tended to grow in clusters in vitro. It was found that the population doubling time of BEL-7402/5-FU cells was longer than that of its parental cells. The plate cloning efficiency and the adherence rate of BEL-7402/5-FU cells at the 2nd and 3rd hour were both lower than those of the parental cells. The distributing proportion of BEL-7402/5-FU cells in G(0)/G(1) phase was less than that of the parental cells, whereas the distributing proportion of BEL-7402/5-FU cells in S phase was higher than that of the parental cells. The accumulation amount of ADM in cytoplasm of BEL-7402/5-FU cells was significantly lower while the expression level of TS protein of which was highly up-regulated as compared with those of the parental cells.
Establishment of the human HCC cell line BEL-7402/5-FU might be beneficial to the studies of 5-Fluorouracil acquired MDR mechanisms and the selection of reversal modifiers.
建立人肝癌多药耐药细胞系BEL-7402/5-FU。
体外采用脉冲疗法联合持续逐步暴露于5-氟尿嘧啶诱导BEL-7402/5-FU。采用MTT法检测其多药耐药性（MDR）。观察BEL-7402/5-FU细胞系的生物学特性，包括形态变化、细胞生长曲线、群体倍增时间、平板克隆效率、贴壁率、细胞周期分布、染色体及致瘤性。采用流式细胞术检测阿霉素（ADM）在细胞质中的蓄积量。采用免疫细胞化学方法评估胸苷酸合成酶（TS）的蛋白表达。
成功建立了获得性多药耐药细胞系BEL-7402/5-FU。BEL-7402/5-FU细胞对ADM、长春新碱（VCR）、甲氨蝶呤（MTX）和奥沙利铂（OHP）表现出交叉耐药，而对羟基喜树碱（HCPT）仍敏感。BEL-7402/5-FU细胞在体外倾向于聚集成团生长。发现BEL-7402/5-FU细胞的群体倍增时间比其亲本细胞长。BEL-7402/5-FU细胞在第2小时和第3小时的平板克隆效率和贴壁率均低于亲本细胞。BEL-7402/5-FU细胞在G(0)/G(1)期的分布比例低于亲本细胞，而在S期的分布比例高于亲本细胞。与亲本细胞相比，BEL-7402/5-FU细胞细胞质中ADM的蓄积量显著降低，而TS蛋白的表达水平则高度上调。
人肝癌细胞系BEL-7402/5-FU的建立可能有助于5-氟尿嘧啶获得性多药耐药机制的研究及逆转剂的筛选。
