Role Of Cytokeratin 19 In Metastasis Of Hepatocellular Carcinoma

Hepatocellular carcinoma (HCC) is among the most frequent cancers worldwide, especially in less developed and developing countries including China.Mechanisms for carcinogenesis and development of HCC is still far from clear. Current research is mainly focused on screening of novel tumor-specific markers and elucidating the molecules and mechanisms responsible for relapse and metastasis of HCC.Although a lot of molecules such as oncogenes, growth factors, adhesion molecules, have been shown to be related to relapse and metastasis of HCC, how they work and interact is poorly understood. Concerning the high heterogeneity and complex processes of HCC, It is urgent now to clarify the global > comprehensive change of signal transduction pathways during carcinogenesis, development, relapse and metastasis. The ever-improving proteomics technologies, mainly 2 dimensional electrophoresis and mass chromatography, may become an ideal platform for it.In this paper, co-operating with Prof. Xia from Shanghai Institute of Biological Chemistry, We screened out a differentially-expressed protein dot on 2D profile between two HCC cell lines (constructed by Medical College of Shanghai Fudan University )different only in metastatic capacity and identified that it is cytokeratin 19(CK19)with MS analysis, which implies the possible role of CK19 in metastasis60 paraffin-embedded HCC samples were screened for CK19 expression. 8 samples showed CK19-positive signals and their expressions were closely related to metastatic clinical indices. Moreover, CK19 is also expressed in two HCC cell lines: HepG-2 and SK-Hepl.To further elucidate the relation between CK19 expression and metastasis of HCC, CK19 cDNA was cloned from breast cancer cell line MCF-7 andpcDNA3-CK19 expression plasmid was constructed.Then Huh-7(HCC cell line without endogenous CK19 expression) was transfected with pcDNA3-CK19 and Huh-7 stably expressing CK19 was established. The migration capacity of this CK19-over-expressing Huh-7 was evaluated through Boyden Chamber test, which showed that CK19 increases dramatically the migration of Huh-7.The past researches of comparative proteomics on HCC are restricted to cell line-based study. But the state and inner pathways of the tumor cell in vivo is different from cell line in vitro very much .Moreover, tumor tissues, especially HCC, are highly heterogeneous compared to cell lines. So it is urgent to establish proteomics technologies suitable for HCC tissue.In this paper, we constructed the basic method of HCC tissue treatment for 2D analysis and compared influence of 4 different sample treatments of freshly-prepared HCC tissues on 2D profile .We found that, currently, hepatic cell pool based on Perocoll density-gradient separation is relatively the best method of HCC tissue treatment for 2D analysis.