中文摘要：

目的观察HGF／Met通路对肝癌细胞株MHCC97-H干细胞样表型的影响，探讨其可能的作用机制。方法体外培养肝癌细胞株Huh7和MHCC97-H，采用细胞免疫荧光染色技术筛选高表达Met的肝癌细胞株MHCC97-H，将其分为4组：空白组（不做处理）、HGF刺激组（50μg/L HGF）、抑制组（50μg／LHGF＋1mol／LHGF抑制剂PHA665752）和表皮生长因子（EGF）／成纤维细胞生长因子（FGF）刺激组（50μg/LHGF＋20μg/LEGF＋20μg/LFGF）。Westernblot检测前3组MHCC97-H细胞中Met和磷酸化Met（p-Met）蛋白的表达。培养24h后光镜下观察前3组细胞形态学变化。克隆球形成实验检测4组细胞克隆球形成能力。荧光实时定量PCR检测空白组和HGF刺激组不同时间点（2、4、8、16、24h）MHCC97-H细胞中多能干细胞相关基因表达变化。计量资料采用x±s表示，多组间比较采用单因素方差分析和重复测量的方差分析，两两比较采用LSD—t检验。结果细胞免疫荧光染色检测结果显示：肝癌细胞株MHCC97-H与Huh7细胞比较．前者高表达Met，将其作为后续研究对象。Westernblot检测结果显示：空白组、HGF刺激组和抑制组细胞中Met蛋白的相对表达量分别为0．44±0．04、0．37±0．03和0．41±0．04，3组比较，差异无统计学意义（F=2．31，P〉0．05）。而3组细胞中p-Met蛋白的相对表达量分别为0．020±0．010、0．070±0．020和0．010±0．000，3组比较，差异有统计学意义（F=34．11，P〈0．05），其中HGF刺激组p-Met蛋白的表达水平高于空白组和抑制组（t=3．87，5．20，P〈0．05）。HGF刺激组MHCC97-H细胞形态呈现长梭状问质样改变，而抑制组和空白组细胞形态相似，呈上皮样。克隆球形成实验结果表明：空白组、HGF刺激组、抑制组和EGF／FGF刺激组克隆球数目分别为0、（35．0±6．3）个、（4．3±1．5）个和（54．3±2．5）个，4组比较，差异有统计学意义（F：511

英文摘要：

Objective To investigate the effects of HGF/Met signaling pathway on stem-like phenotype of hepatocellular carcinoma cell line MHCC97-H. Methods The hepatocellular carcinoma cell lines Huh7 and MHCC97-H were cultured in vitro. Cell lines with high expression of Met were selected and divided into the blank group （cells untreated）, HGF stimulation group （cells treated with 50 μg/L of HGF）, HGF inhibition group （cells treated with 50 μg/L of HGF ＋ 1 mol/L of PHA665752） and epidermal growth factor/fibroblast growth factor （EGF/FGF） stimulation group （ cells treated with 50 μg/L of HGF ＋ 20 μg/L of EGF ＋ 20 μg/L of FGF）. The protein expressions of Met and phospho-Met （p-Met） in the blank group, the HGF stimulation group and the HGF inhibition group were detected by Western blot. The cell morphological alteration of the blank group, the HGF stimulation group and the HGF inhibition group was observed under the light microscope at 24 hours after the treatment. Different sphere-formation ability was detected by sphere-formation assay under serum-free condition. The expression change of induced pluripotent stem cells related genes was analyzed by Real-Time polymerase chain reaction when MHCC97-H cells had been treated with HGF for different hours （2, 4, 8, 16, 24 hours）. The measurement data were presented as x ± s. All data were analyzed using the one-way analysis of variance, repeated measurement and LSD-t test. Results The results of immunofluoreseence showed that MHCC97-H cells possessed high expression of Met when compared with Huh7 cells. In the blank group, HGF stimulation group and the HGF inhibition group, the protein expressions of Met were 0.44 ± 0.04, 0.37 ± 0.03, 0.41±0.04, with no significant difference between the 3 groups （ F = 2. 31, P 〉 0.05 ） , and the protein expressions of p-Met in the 3 groups were 0. 020 ± 0. 010, 0. 070 ± 0. 020, 0. 010± 0. 000, with significant difference between the 3 groups （ F = 34. 11, P 〈 0.05 ）. The protein expression