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  • Progeny from irradiated colorectal cancer cells acquire an EMT-like phenotype and activate Wnt/β-catenin pathway.

Progeny from irradiated colorectal cancer cells acquire an EMT-like phenotype and activate Wnt/β-catenin pathway.

Journal of cellular biochemistry (2014-08-12)
Lilian Gonçalves dos Reis Bastos, Priscila Guimarães de Marcondes, Julio Cesar Madureira de-Freitas-Junior, Fernanda Leve, André Luiz Mencalha, Waldemir Fernandes de Souza, Wallace Martins de Araujo, Marcelo Neves Tanaka, Eliana Saul Furquim Werneck Abdelhay, José Andrés Morgado-Díaz
ABSTRACT

Radiotherapy remains a major approach to adjuvant therapy for patients with advanced colorectal cancer, however, the fractionation schedules frequently allow for the repopulation of surviving tumors cells, neoplastic progression, and subsequent metastasis. The aim of the present study was to analyze the transgenerational effects induced by radiation and evaluate whether it could increase the malignant features on the progeny derived from irradiated parental colorectal cancer cells, Caco-2, HT-29, and HCT-116. The progeny of these cells displayed a differential radioresistance as seen by clonogenic and caspase activation assay and had a direct correlation with survivin expression as observed by immunoblotting. Immunofluorescence showed that the most radioresistant progenies had an aberrant morphology, disturbance of the cell-cell adhesion contacts, disorganization of the actin cytoskeleton, and vimentin filaments. Only the progeny derived from intermediary radioresistant cells, HT-29, reduced the E-cadherin expression and overexpressed β-catenin and vimentin with increased cell migration, invasion, and metalloprotease activation as seen by immunoblotting, wound healing, invasion, and metalloprotease activity assay. We also observed that this most aggressive progeny increased the Wnt/β-catenin-dependent TCF/LEF activity and underwent an upregulation of mesenchymal markers and downregulation of E-cadherin, as determined by qRT-PCR. Our results showed that the intermediate radioresistant cells can generate more aggressive cellular progeny with the EMT-like phenotype. The Wnt/β-catenin pathway may constitute an important target for new adjuvant treatment schedules with radiotherapy, with the goal of reducing the migratory and invasive potential of the remaining cells after treatment.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Gelatin from porcine skin, Type A, lyophilized powder, γ-irradiated, BioXtra, suitable for cell culture
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Monoclonal Anti-Vimentin−Cy3 antibody produced in mouse, clone V9, purified immunoglobulin, buffered aqueous solution
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4′,6-Diamidino-2-phenylindole dihydrochloride, suitable for fluorescence, BioReagent, ≥95.0% (HPLC)
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Protease Inhibitor Cocktail, for use with mammalian cell and tissue extracts, DMSO solution
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Monoclonal Anti-Vimentin antibody produced in mouse, clone V9, ascites fluid