【作者】 汪进国；

【导师】 耿小平； 钱叶本； 朱立新；

【作者基本信息】 安徽医科大学 ， 外科学， 2002， 硕士

【摘要】 目的：探讨HGF/C-Met信号传导系统对肝细胞癌生长、侵袭和转移的影响及其作用机制。方法：将肝癌Hep3B细胞株按给予外源性HGF的剂量进行分组：不给予外源性HGF的为对照组，即A(0ng.ml^-1)组；给予外源性HGF的为实验组₁，并且按照外源性HGF的剂量将其分为B(5ng.ml^-1)，C(10ng.ml^-1)，D(15ng.ml^-1)，E(20ng.ml^-1)组。依照上述分组的方法将人正常肝细胞分为1，2，3，4，5组，其中1(0ng.ml^-1)组为对照组₂，2(5ng.ml^-1)，3(10ng.ml^-1)，4(15ng.ml^-1)，5(20ng.ml^-1)组为实验组₂。检测肝癌Hep3B细胞株细胞生长曲线和离散情况比较实验组₁和对照组₁肝癌Hep3B细胞株细胞的增长与离散状况。检测人正常肝细胞生长曲线比较实验组₂和对照组₂人正常肝细胞的增长情况，同时比较肝癌Hep3B细胞株和人正常肝细胞相对应组之间的细胞增长情况。分别用酶联免疫吸附实验（ELISA）检测肝癌Hep3B细胞培养上清液中血管内皮生长因子（VEGF）的含量；用免疫组织化学技术（ABC法）测定肝癌Hep3B细胞中增殖细胞核抗原（PCNA）的表达情况，并计算增殖细胞核抗原标记指数（PCNA—LI）。从整体水平上研究HGF/c-met信号传导系统对肝癌细胞生长、侵袭和转移的影响及其作用机制。结果：1．外源性HGF浓度在1～15ng.ml^-1时，肝癌Hep3B细胞增殖明显加快、细胞离散度增加并伴有细胞形态学的改变，同时HGF与细胞的增殖和离散度的增加呈剂量-效应关系，但是对照组₁肝癌Hep3B细胞株较B，C，D实验组₁细胞生长慢，并且与D组相比有显著性差异。当外源性HGF浓度为20ng.ml^-1时，肝癌Hep3B细胞增殖停止，细胞趋于凋亡。而HGF浓度在1～20ng.ml^-1时，人正常肝细胞的实验组₂和对照组₂细胞都处于持续性增长状态，其中以D组细胞增长最快，与其它组相比有显著性差异。2．肝癌Hep3B细胞培养上清液中，B，C，D实验组₁较对照组₁VEGF的OD值明显增高，并且有显著性安徽医科大学硕士学位论文差异；同时肝癌Hep3B细胞中，B，C，D组较对照组；PCNA十I也明显升高，其中D组 PCNA－LI最高，与对照组；和 B，C，E实验组；相比有显著性差异。而 E组中 VEGF的OD 值、PCNA－LI与对照组；个＊无明显差异。结论：HGF／c－met信号传导系统能促进肝癌细胞的生长、侵袭和转移，并受HGF浓度的影响，其作用机制可能与肝癌细胞的增殖、离散、VEGF及PCNA的表达有关。更多还原

【Abstract】 Objective: To explore the effects and mechanisms of hepatocyte growth factor (HGF)/C-Met transduction system on the growth, invasion and transmission of hepatocellular carcinoma.Methods: Hepatocarcinoma HepSB cell line was divided into five groups. In the groupj A ( control group ) HGF was not used. In the groupi B, C, D and E, HGF was used with different concentrations. Acorrding to the grouping of hepatocellular Hep3B cell line, normal hepatocytes were also divided into five groups. In the group2 (control group) HGF was not used. In the group2 2,3,4 and 5, HGF was used with different concentrations. After treated with HGF on different doses, the cell proliferation and scatter of hepatocarcinoma HepSB cells and normal hepatocytes were examined to compare the results with each group. Enzyme-Linked Immunosorbent Assay (ELISA) was used to detect the Vascular Endothelial Growth Factor (VEGF) concentration in the culture supernatant of hepatocarcinoma Hep3B cells. Immunohistochemistry technique was used to exam the Proliferating Cell Nuclear Antigen (PCNA) in hepatocarcinoma HepSB cells with Proliferating Cell Nuclear Antigen Label Index (PCNA-LI).Results: (1) The proliferation and scatter degree of hepatocarcinoma HepSB cell were significantly increased after treated with HGF in the concentrations of 1~15ng.ml-1,which presented a dose-dependent manner and cells modality alteration. When the concentration of HGF was 20ng.mr1, the cell proliferation stopped to apoptosis, while the proliferation of normal hepatocytes went on. (2) The Optical Density (OD) value of VEGF protein was significantly higher in the groups of B, C and D compared with the control one. In the groups of B, C and D, the PCNA-LI was significantly higher than that in the control. However, there is no difference between group E and the control with VEGF and PCNA-LI results.Conclusions: Our study proved that HGF/c-met signal transduction system could promote the proliferation, invasion and metastasis of hepatocyte, which was affected by the concentration of HGF. The mechanism is probably related to the proliferation, scatter, high expression of VEGF and PCNA.更多还原