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Characterisation of mesenchymal colon tumour-derived cells in tumourspheres as a model for colorectal cancer progression.
International Journal of Oncology 2018 September 19
Cellular plasticity, the ability of cells to switch from an epitheial phenotype to a mesenchymal one and vice versa, plays a crucial role in tumour progression and metastases development. In 20-25% of patients with colon cancer and in 18% of patients with rectal cancer, metastases are present at the time of the first diagnosis. They are the first cause of colorectal cancer (CRC)-related mortality, defining stage IV CRC, which is characterized by a relatively short overall survival. We previously isolated two primary colon adenocarcinoma cell cultures that had undergone epithelial-mesenchymal transition (EMT), one with a high microsatellite instability phenotype (T88) and one with a chromosomal instability phenotype (T93). The aim of this study was to establish a model with which to study EMT, stemness features and cell plasticity in cancer progression and to examine the effects of incubation with lithium chloride (LiCl), a specific glycogen synthase kinase 3 β (GSK-3β) inhibitor, on these cellular processes. Indeed, GSK3β is an important regulator of cell survival, which promotes tumourigenesis in colon cells by facilitating the crosstalk between colorectal cancer pathways. Thus, we further characterized our system of adherent primary mesenchymal colon cancer cells and their paired tumourspheres by examining the expression and localisation of a panel of markers, including E- and N‑cadherin, CD133, CD44v6, aldehyde dehydrogenase 1 (ALDH1) and leucine-rich repeat‑containing G-protein coupled receptor 5 (LGR5). We also characterised the molecular features of these tumourspheres and examined their response to LiCl. Furthermore, we explored the effects of LiCl on cell motility and plasticity. We demonstrated that LiCl reduced cell migration, stemness features and cell plasticity. We also observed the atypical nuclear localisation of membrane proteins, including N‑cadherin, CD133 and CD44v6 in mesenchymal tumour cells. Of note, CD133 and CD44v6 appeared to localise at the plasma membrane in cells with a more epithelial phenotype, suggesting that the cytoplasmic/nuclear localisation of these proteins could favour and characterize cell plasticity in colorectal cancer progression.
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