Carcinogenesis Advance Access originally published online on June 22, 2009
Carcinogenesis 2009 30(9):1487-1496; doi:10.1093/carcin/bgp153
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Met receptor tyrosine kinase transactivation is involved in proteinase-activated receptor-2-mediated hepatocellular carcinoma cell invasion
Department of General, Visceral and Vascular Surgery, Research Center Lobeda Medical Faculty, Friedrich Schiller University Jena, 07747 Jena, Germany
1 Institute of Anatomy II
2 Institute of Pathology, Medical Faculty, Friedrich Schiller University Jena, 07743 Jena, Germany
3 Institute of Biochemistry, Charité, Humboldt University of Berlin, 10117 Berlin, Germany
4 Electron Microscopy Center
5 Institute of Molecular Cell Biology, Medical Faculty, Friedrich Schiller University Jena, 07743 Jena, Germany
6 Canadian Institutes of Health Research Proteinases and Inflammation Network, Inflammation Research Network, Department of Pharmacology and Therapeutics, University of Calgary, Calgary, Canada T2N 4N1
* To whom correspondence should be addressed. Research Laboratory, Department of General, Visceral and Vascular Surgery, Research Center Lobeda, Medical Faculty at the Friedrich Schiller University Jena, Erlanger Allee 101, D-07747 Jena, Germany. Tel: +49 9325870; Fax: +49 9325872; Email: roland.kaufmann{at}med.uni-jena.de
The expression of proteinase-activated receptor (PAR)2 in human hepatocellular carcinoma (HCC) was established by reverse transcription–polymerase chain reaction, confocal immunofluorescence and electron microscopy in permanent cell lines, primary HCC cell cultures and HCC tumor tissue. Stimulation of HCC cells with trypsin and the PAR2-selective activating peptide, 2-furoyl-LIGRLO-NH2, increased cell invasion across Matrigel. Both effects were blocked by a PAR2-selective pepducin antagonist peptide (pal-PAR2) and by PAR2 silencing with specific small interfering RNA (siRNA). PAR2-initiated HCC cell invasion was also blocked by inhibiting the hepatocyte growth factor receptor (Met receptor tyrosine kinase) with the receptor-targeted kinase inhibitors, SU 11274 and PHA 665752, or by downregulation of Met with specific siRNA. The involvement of Met in PAR2-mediated HCC invasive signaling was further supported by the finding that treatment of HCC cells with trypsin or the PAR2-selective agonist peptide, 2-furoyl-LIGRLO-NH2, stimulated Met activation-phosphorylation. In addition, Met-dependent stimulation of p42/p44 mitogen-activated protein Kinases was found to be critical for the PAR2–Met receptor tyrosine kinase-invasive signaling axis in HCC cells. Our study establishes an important link between the PAR2 and Met receptor tyrosine kinase signaling in promoting HCC cell invasion.
Abbreviations: HCC, hepatocellular carcinoma; HGF, hepatocyte growth factor; MAPK, mitogen-activated protein kinase; NAC, N-acetyl-L-cysteine; PAR, proteinase-activated receptor; PAR2-AP, proteinase-activated receptor2-activating peptide; ROS, reactive oxygen species; siRNA, small interfering RNA; TBS, Tris-buffered saline
Received January 1, 2009; revised May 21, 2009; accepted June 16, 2009.