Thrombin conducts epithelial-mesenchymal transition via protease-activated receptor-1 in human gastric cancer

Abstract

Epithelial-mesenchymal transition (EMT) is thought to be a key step for cancer metastasis. Using an immunohistochemical approach with gastric carcinoma tissue, we found the expression of protease-activated receptor-1 (PAR 1), along with a metalloproteinase known to activate PAR 1, were associated with poorer prognosis, compared with expression-negative tumors, and activated PAR 1 promotes gastric cancer cell invasion and proliferation in vivo. In this study we observed EMT induction by the PAR 1 agonist α-thrombin, in human gastric cell lines stably expressing PAR 1. We investigated α-thrombin-induced changes in the cell forms of pcDNA 3-1-MKN45 (MKN45/Mock), pcDNA 3-1-PAR 1 transfected MKN45 (MKN45/PAR 1), and MKN74. Expression levels of epithelial and mesenchymal markers as well as the distribution of transcriptional factors of E-cadherin in the cytoplasm and nucleus were also noted in these cell lines. We observed α-thrombin-induced morphological changes in MKN45/PAR 1 and MKN74 cells. Western blotting and immunohistochemistry of these cells indicated a fall in the expression level of E-cadherin and an increase in fibronectin expression after 48 h. PAR1 activation also induced significant increases in nuclear levels of the Snail which is a repressor of E-cadherin gene expression. We found EMT in gastric cancer cell lines that underwent α-thrombininduced PAR 1 activation.