Document Type : Original Article
Molecular Medicine Department, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran
Hepatitis and AIDS Department, Pasteur Institute of Iran, Tehran, Iran
National cell Bank of Iran, Pasteur Institute of Iran, Tehran, Iran
Introduction: Wnt and k-ras are two main signaling pathways activated in colon cancer. Many genes are upregulated downstream of these signaling pathways. The aim of this study was to assess the activity of Wnt and k-ras in HCT116 and SW480 cell lines by making two reporter constructs using promoters downstream of these pathways (fibroblast growth factor18 [FGF18] and urokinase plasminogen activator receptor [UPAR]).
Materials and Methods: UPARLacZ, FGF18LacZ, negative (pUCLacZ) and positive (CMVLacZ) control plasmids and pRc/CMV2CAT were constructed. Expressions of LacZ in both cell lines were studied by βgal staining and ELISA after normalization with CAT expression.
Results: In both cell lines, FGF18LacZ transfected cells stained more than UPARLacZ transfected ones. This difference was more prominent in SW480. Both constructs have the ability of expression in both cell lines. It was also proven that FGF18LacZ was significantly more active than UPARLacZ in both cell lines. Expression of FGF18LacZ in HCT116 and SW480 cell lines was respectively 1.34 and 4.4 times more than UPARLacZ.
Conclusion: Despite the fact that in HCT116 the Ras pathway is activated, FGF18LacZ is more active than UPARLacZ although the UPAR promoter is more active in HCT116 cell line than SW480 cell line. These findings are in accordance with previous studies that in all colon cancer cell lines Wnt signaling pathway is active even though there is no mutation in any part of it. Wnt is the main signaling pathway responsible for carcinogenesis in colon epithelial cells. These constructs can be used as reporters for studying the above mentioned signaling pathways in other cell lines.