Journal of Cancer Prevention

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pISSN 2288-3649
Fig. 1.

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Fig. 1. A549RT-eto cells exhibit CSC-like phenotypes. (A) Cell migration was measured by a wound-healing assay after treatment for 24 hours. The cell wound closure rate was measured at 24 hours and was calculated according to the equation described in Materials and Methods. The experiments were performed in triplicate (***P < 0.001, A549RT-eto vs. the A549 cell). (B) An invasion assay was performed using 24-well chambers coated with Matrigel after 24 hours. The invading cells in the membrane were stained with hematoxylin-eosin. (C) The expression of E-cadherin and vimentin was detected by immunofluorescence using an Alexa Fluor 488-conjugated secondary antibody (green; outside part). DAPI was added for nuclear staining (blue; inside part) (magnificaiton ×400). (D) Lysates from A549 and A549RT-eto cells were prepared and separated on an 8% SDS-PAGE gel. The expression of E-cadherin, N-cadherin and vimentin was detected by immunoblotting. Densitometric analysis was performed on all blots and the integrated optical density of each band was normalized with corresponding β-actin (***P < 0.001, A549RT-eto vs. the A549 cells). (E) The spheroid size and the number of A549 and A549RT-eto cells were evaluated after 2, 4 and 6 days. A spheroid greater than 50 µm in size was counted as the criterion for evaluating sphere formation. Scale bars indicate 50 µm. The assay was carried out from three independent experiments (***P < 0.001, A549RT-eto vs. A549 cells). (F) The expressions of Oct4, Bmi1, Nanog, Sox2 and Klf4 was detected by immunoblotting after 24 hours. Densitometric analysis was performed on all blots and the integrated optical density of each band was normalized with corresponding β-actin (**P < 0.05 and ***P < 0.001, A549RT-eto vs. the A549 cells). A549RT-eto, A549 lung cancer cells resistant to etoposide; CSC, cancer stem cell; DAPI, diamidino-2-phenylindole; SDS, sodium dodecyl sulfate; PAGE, polyacrylamide gel electrophoresis.
J Cancer Prev 2022;27:112~121 https://doi.org/10.15430/JCP.2022.27.2.112
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