Involvement of PI3K and ERK1/2 pathways in hepatocyte growth factor-induced cholangiocarcinoma cell invasion.
OBJECTIVE
To investigate the role of hepatocyte growth factor (HGF) in cholangiocarcinoma (CCA) cell invasiveness and the mechanisms underlying such cellular responses.


METHODS
Effects of HGF on cell invasion and motility were investigated in two human CCA cell lines, HuCCA-1 and KKU-M213, using Transwell in vitro assay. Levels of proteins of interest and their phosphorylated forms were determined by Western blotting. Localization of E-cadherin was analyzed by immunofluorescence staining and visualized under confocal microscope. Activities of matrix degrading enzymes were determined by zymography.


RESULTS
Both CCA cell lines expressed higher Met levels than the H69 immortalized cholangiocyte cell line. HGF induced invasion and motility of the cell lines and altered E-cadherin from membrane to cytoplasm localization, but did not affect the levels of secreted matrix metalloproteinase (MMP)-2, MMP-9 and urokinase plasminogen activator, key matrix degrading enzymes involved in cell invasion. Concomitantly, HGF stimulated Akt and extracellular signal-regulated kinase (ERK)1/2 phosphorylation but with slightly different kinetic profiles in the two cell lines. Inhibition of the phosphoinositide 3-kinase (PI3K)/Akt pathway by the PI3K inhibitor, LY294002, markedly suppressed HGF-stimulated invasion of both CCA cell lines, and inhibition of the ERK pathway by U0126 suppressed HGF-induced invasion of the KKU-M213 cell line but had a moderate effect on HuCCA-1 cells.


CONCLUSIONS
These data indicate that HGF promotes CCA cell invasiveness through dys-localization of E-cadherin and induction of cell motility by distinct signaling pathways depending on cell line type.
Department of Biochemistry, Faculty of Science, Mahidol University, Bangkok 10400, Thailand.
Apaporn
A
Menakongka
Tuangporn
T
Suthiphongchai
D001650:Bile Duct Neoplasms / Q000378:metabolism* / Q000473:pathology; D001653:Bile Ducts, Intrahepatic; D015820:Cadherins / Q000378:metabolism; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects*; D018281:Cholangiocarcinoma / Q000378:metabolism* / Q000473:pathology; D017228:Hepatocyte Growth Factor / Q000494:pharmacology*; D006801:Humans; D020782:Matrix Metalloproteinases / Q000378:metabolism; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D019859:Proto-Oncogene Proteins c-met / Q000378:metabolism; D017978:Receptors, Growth Factor / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D014568:Urokinase-Type Plasminogen Activator / Q000378:metabolism
D015820:Cadherins; D017978:Receptors, Growth Factor; D017228:Hepatocyte Growth Factor; D019869:Phosphatidylinositol 3-Kinases; C491743:MET protein, human; D019859:Proto-Oncogene Proteins c-met; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D014568:Urokinase-Type Plasminogen Activator; D020782:Matrix Metalloproteinases