Epithelial-mesenchymal transition stimulates human cancer cells to extend microtubule-based invasive protrusions and suppresses cell growth in collagen gel

• Published in: PloS one 2012-12, Vol.7 (12), p.e53209-e53209
• Main Authors: Oyanagi, Jun, Ogawa, Takashi, Sato, Hiroki, Higashi, Shouichi, Miyazaki, Kaoru
• Format: Article
• Language: English
• Subjects: Actin; Agar; Biology; Cancer; Cancer cells; Cancer treatment; Cell adhesion; Cell adhesion & migration; Cell Adhesion - drug effects; Cell Adhesion - physiology; Cell culture; Cell Culture Techniques; Cell Cycle - drug effects; Cell Cycle - physiology; Cell growth; Cell Line, Tumor; Cell Movement - drug effects; Cell Movement - physiology; Cell Proliferation - drug effects; Colchicine; Collagen; Collagen - metabolism; Cytoskeleton; Development and progression; Epithelial-Mesenchymal Transition - drug effects; Epithelial-Mesenchymal Transition - physiology; Fibroblasts; Fibronectin; Fibronectins; Gastric cancer; Gene expression; Growth; Growth factors; Heat shock proteins; Hsp90 protein; Humans; Infiltration; Inhibitors; Invasiveness; Kinases; Laminin; Laminin - metabolism; Medicine; Mesenchyme; Metastases; Metastasis; Microtubules; Microtubules - metabolism; Microtubules - pathology; Monomolecular films; Phosphoprotein phosphatase; Phosphorylation; Physiological aspects; Protein phosphatase; Proteins; Rodents; Science; Stem cells; Stomach cancer; Stroma; Studies; Transcription factors; Transforming Growth Factor beta - pharmacology; Transforming growth factors; Tumor cell lines; Tumors
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0053209

Epithelial-mesenchymal transition (EMT) is a crucial event in tumor invasion and metastasis. However, most of past EMT studies have been conducted in the conventional two-dimensional (2D) monolayer culture. Therefore, it remains unclear what invasive phenotypes are acquired by EMT-induced cancer cells. To address this point, we attempted to characterize EMT cells in more physiological, three-dimensional (3D) collagen gel culture. EMT was induced by treating three human carcinoma cell lines (A549, Panc-1 and MKN-1) with TGF-ß. The TGF-ß treatment stimulated these cells to overexpress the invasion markers laminin γ2 and MT1-MMP in 2D culture, in addition to the induction of well-known morphological change and EMT marker expression. EMT induction enhanced cell motility and adhesiveness to fibronectin and collagen in 2D culture. Although EMT cells showed comparable cell growth to control cells in 2D culture, their growth rates were extremely suppressed in soft agar and collagen gel cultures. Most characteristically, EMT-induced cancer cells commonly and markedly extended invasive protrusions in collagen gel. These protrusions were mainly supported by microtubules rather than actin cytoskeleton. Snail-introduced, stable EMT cells showed similar protrusions in 3D conditions without TGF-ß. Moreover, these protrusions were suppressed by colchicine or inhibitors of heat shock protein 90 (HSP-90) and protein phosphatase 2A. However, MMP inhibitors did not suppress the protrusion formation. These data suggest that EMT enhances tumor cell infiltration into interstitial stroma by extending microtubule-based protrusions and suppressing cell growth. The elevated cell adhesion to fibronectin and collagen and high cell motility also seem important for the tumor invasion.