肝癌间质细胞中HGF／cMET系统的表达对肝癌细胞恶性生物学功能的影响

摘要

Objective To explore the effects of interstitial cells of liver cancer and normal liver cells co‐cultured on the biological function of liver cancer malignancy so as to understand the signal pathway involved by the interaction between these cells and confirm the role of interstitial cells in cancer progression in tumor microenvironment .Methods We co‐cultured interstitial cells or hepatocyte growth factor (HGF ) and human normal liver cell L‐02 ,and then detected the expressions of the tumor‐suppressing gene PTEN and the oncogene K‐RAS and changes of cell proliferation .The downstream signaling pathways were detected by Real‐time PCR and Western blot .Results The expression of PTEN was downregulated at the transcription level by 1 .15 times and translation level by 10 times (P<0 .05) ,while the transcription level and translation level of K‐RAS increased by 1 .4 times and more than 9 times , respectively ( P< 0 .05 ) in normal liver cells co‐cultured with liver cancer mesenchymal cells .The proliferation ability was increased by more than 2 times .ELISA experiment results showed that the medium from co‐culture contained HGF over 3 times more than the control group ( P<0 .05 ,1 085+108 vs .387+23) .At the same time ,cells in the experimental group expressed more than four times of cMET than the control group cells (P< 0 .05) .Exogenous HGF consistently promoted liver cell proliferation and viability (P<0 .05) .Conclusion Our study shows that liver cancer interstitial cells activate the HGF/cMET signaling pathway by secreting HGF and promote the proliferation of normal liver cells ,suggesting a new way to explore the molecular mechanism of tumor microenvironment in tumor development and treatment of hepatocellular carcinoma .%目的：探索肝癌间质细胞与正常肝细胞共培养对肝癌恶性生物学功能的影响，探讨其相互作用所涉及的信号通路，明确肝癌微环境中间质细胞在肿瘤进展中的作用。方法人肝癌间质细胞或人肝细胞生长因子（hepatocyte growth factor ，HGF）与肝正常细胞系L‐02共培养，检测肝正常细胞中抑癌基因PTEN及癌基因K‐RAS的表达及肝正常细胞的增殖变化；Real‐time PCR和Western blot检测下游相关信号通路的变化。结果肝癌间质细胞与正常肝细胞L‐02共培养使得抑癌基因 PT EN 的转录水平下调1．15倍，翻译水平下调10倍多（ P＜0．05），而原癌基因K‐RAS的转录水平上调1．4倍，翻译水平上调9倍以上（P＜0．05）；共培养后的肝细胞增殖能力较对照组增加2倍以上；ELISA实验结果显示，共培养后的培养液中含有较对照组3倍以上的HGF（P＜0．05，1085±108 vs ．387±23）；同时实验组细胞表达的cMET也较对照组细胞高出4倍以上（P＜0．05）；外源性HGF一致性促进了肝细胞的增殖能力和活力（P＜0．05）。结论肝癌间质细胞可能通过分泌 HGF激活 HGF／cMET 信号通路，促进正常肝细胞的增殖。这为探索肿瘤微环境对肿瘤发生、发展的分子机制及肝癌治疗提供新的途径。