Lithium chloride induces mesenchymal‑to‑epithelial reverting transition in primary colon cancer cell cultures.
Epithelial‑to‑mesenchymal transition (EMT) confers stem cell‑like phenotype and more motile properties to carcinoma cells. During EMT, the expression of E‑cadherin decreases, resulting in loss of cell‑cell adhesion and increased migration. Expression of Twist1 and other pleiotropic transcription factors, such as Snail, is known to activate EMT. We established primary colon cancer cell cultures from samples of operated patients and validated cultures by cytogenetic and molecular biology approaches. Western blot assay, quantitative real‑time PCR and immunofluorescence were performed to investigate the expression of E‑cadherin, vimentin, β‑catenin, cytokeratin‑20 and ‑18, Twist1, Snail, CD44, cyclooxygenase‑2 (COX2), Sox2, Oct4 and Nanog. Moreover, cell differentiation was induced by incubation with LiCl‑containing medium for 10 days. We observed that these primary colorectal cancer (CRC) cells lost expression of the E‑cadherin epithelial marker, which was instead expressed in cancer and normal colon mucosa of the same patient, while overexpressed vimentin (mesenchymal marker), Twist1, Snail (EMT markers) and COX2. Cytokeratin‑18 was expressed both in tissues and cell cultures. Expression of stem cell markers, such as CD44, Oct4 and Nanog, were also observed. Following differentiation with the glycogen synthase kinase 3β (GSK3β) inhibitor LiCl, the cells began to express E‑cadherin and, at once, Twist1 and Snail expression was strongly downregulated, suggesting a MET‑reverting process. In conclusion, we established primary colon mesenchymal cancer cell cultures expressing mesenchymal and epithelial biomarkers together with high level of EMT transcription factors. We propose that they could represent a good model for studying EMT and its reverting mechanism, the mesenchymal‑to‑epithelial transition (MET). Our observation indicates that LiCl, a GSK3β inhibitor, induces MET in vitro, suggesting that LiCl and GSK3β could represent, respectively, interesting drug, and target for CRC therapy.
Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico Ⅱ, I‑80131 Naples, Italy.
Valeria
V
Costabile
Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico Ⅱ, I‑80131 Naples, Italy.
Francesca
F
Duraturo
Colorectal Surgical Oncology, Istituto Nazionale per lo Studio e la Cura dei Tumori, ̔Fondazione Giovanni Pascale̓ IRCCS, I‑80131 Naples, Italy.
Paolo
P
Delrio
Colorectal Surgical Oncology, Istituto Nazionale per lo Studio e la Cura dei Tumori, ̔Fondazione Giovanni Pascale̓ IRCCS, I‑80131 Naples, Italy.
Daniela
D
Rega
Colorectal Surgical Oncology, Istituto Nazionale per lo Studio e la Cura dei Tumori, ̔Fondazione Giovanni Pascale̓ IRCCS, I‑80131 Naples, Italy.
Ugo
U
Pace
Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico Ⅱ, I‑80131 Naples, Italy.
Raffaella
R
Liccardo
Endoscopy Unit, Istituto Nazionale per lo Studio e la Cura dei Tumori, ̔Fondazione Giovanni Pascale̓ IRCCS, I‑80131 Naples, Italy.
Giovanni Battista
GB
Rossi
Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico Ⅱ, I‑80131 Naples, Italy.
Rita
R
Genesio
Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico Ⅱ, I‑80131 Naples, Italy.
Lucio
L
Nitsch
Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico Ⅱ, I‑80131 Naples, Italy.
Paola
P
Izzo
Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico Ⅱ, I‑80131 Naples, Italy.
Marina
M
De Rosa
D000276:Adjuvants, Immunologic / Q000494:pharmacology*; D014408:Biomarkers, Tumor / Q000378:metabolism; D015153:Blotting, Western; D002454:Cell Differentiation / Q000187:drug effects; D003110:Colonic Neoplasms / Q000188:drug therapy* / Q000378:metabolism; D058750:Epithelial-Mesenchymal Transition / Q000187:drug effects; D005455:Fluorescent Antibody Technique; D006801:Humans; D018021:Lithium Chloride / Q000494:pharmacology*; D009363:Neoplasm Proteins / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D014407:Tumor Cells, Cultured
D000276:Adjuvants, Immunologic; D014408:Biomarkers, Tumor; D009363:Neoplasm Proteins; D018021:Lithium Chloride