Snail-induced epithelial-mesenchymal transition in gastric carcinoma cells and generation of cancer stem cell characteristics.

Abstract

Biological changes in Snail-overexpressed SGC7901 cells were studied by establishing a pEGFP-C1-Snail carrier. The significance of Snail in epithelial-mesenchymal transition (EMT) as well as the invasion and metastatic capacity of gastric cancer cells was also discussed; moreover, we attempted to verify the probable cancer stem cell characteristics of Snail-overexpressed cells. A pEGFP-C1-Snail eukaryotic expression plasmid was constructed and pEGFP-C1(-) and pEGFP-C1-Snail plasmids were extracted and transfected into SGC7901 cells using Lipofectamine 2000. Stably expressed SGC7901-N [control group containing pEGFP-C1(-)] and SGC7901-S (test group containing pEGFP-C1-Snail) cells were screened using a G418 resistance medium. Snail, E-cadherin, b-catenin, vimentin, and fibronectin gene and protein expressions were detected by real-time quantitative PCR, western blot, and immunofluorescence. Cell invasion and metastasis were tested by scratch test, invasion assay, and an adhesion experiment. The positive rate of aldehyde dehydrogenase-1 (ALDH-1) expression was analyzed by flow cytometry. The results indicated the occurrence of EMT, accompanied by morphological changes in the cells and a weakening of the cell adhesion capacity. We also observed a decrease in the expression of epithelial markers E-cadherin and b-catenin and an increase in mesenchymal (Snail and vimentin) marker expression. Moreover, the cells showed increased invasiveness and metastatic capacity, and decreased proliferative ability. Moreover, the Snail-treated SGC7901 cells moved towards the scratch and produced fewer clones compared to the control cells. Owing to its capacity for self-renewal, SGC7901-S cells produced new clones and expressed ALDH-1. Therefore, we concluded that Snail overexpression induced EMT and endowed cells with tumor stem cell characteristics.