Down-regulation of Ech1 decreases the adhesion ability of mouse hepatocarcinoma Hca-F cells.
10.3760/cma.j.issn.1007-3418.2012.08.013
- Author:
Jun ZHANG
1
;
Jian-wu TANG
;
Ming-zhong SUN
;
Shu-qing LIU
;
Mei-ying SONG
;
Bo WANG
;
Bo SONG
;
Yu-hong HUANG
Author Information
1. Department of Pathology, Dalian Medical University, Dalian, China.
- Publication Type:Journal Article
- MeSH:
Animals;
Carbon-Carbon Double Bond Isomerases;
genetics;
metabolism;
Carcinoma, Hepatocellular;
enzymology;
genetics;
pathology;
Cell Adhesion;
Cell Line, Tumor;
Down-Regulation;
Electrophoresis, Gel, Two-Dimensional;
Gene Expression Regulation, Neoplastic;
Liver Neoplasms;
enzymology;
genetics;
pathology;
Lymph Nodes;
pathology;
Lymphatic Metastasis;
Mice;
Plasmids;
RNA, Messenger;
genetics;
metabolism;
RNA, Small Interfering;
genetics;
pharmacology;
Transfection
- From:
Chinese Journal of Hepatology
2012;20(8):605-610
- CountryChina
- Language:Chinese
-
Abstract:
To examine the differential expression pattern of Ech1 protein in mouse Hca-F and Hca-P hepatocarcinoma cell lines with high and low rates of lymphatic metastasis, respectively, and to investigate the relationships between Ech1 expression and adhesion of Hca-F cells. Fluorescence two-dimensional difference in-gel electrophoresis (2D DIGE) and mass spectrometry were used to detect Ech1 expression. Ech1 gene silencing was achieved by stable transfection of Hca-F cells with a plasmid vector harboring short hairpin RNA (shRNA) targeting Ech1, pGPU6/GFP/Neo-shRNA-Ech1. Ech1 mRNA and protein expressions were detected by real-time quantitative polymerase chain reaction (qRT-PCR) and Western blotting analysis, respectively. Adhesive properties of cells were assessed by hematoxylin-eosin staining and fluorimetric detection of extracellular matrix (ECM) proteins. Endogenous Ech1 protein level was remarkably higher in the highly metastatic Hca-F cell line than in the Hca-P cell line (2.7-fold by 2D DIGE; 1.5-fold by Western blotting). shRNA-induced silencing of Ech1 significantly reduced the adhesion ability of Hca-F cells, as evidenced by decreased absorbance values of fibronectin and collagen I (Hca-F cells vs. pGPU6/GFP/Neo-shRNA-Ech1 cells: 1.42+/-0.26 vs. 1.01+/-0.27 and 1.14+/-0.07 vs. 0.90+/-0.09, respectively; P less than 0.05). Down-regulation of Ech1 can inhibit the adhesive capacity of metastatic Hca-F cells.
