Inflammatory reactions play an important role in all stages of tumor development. A shift to the mesenchymal phenotype causes an increase of migratory capacity of tumor cells. Epithelial-mesenchymal transition (EMT) can also be caused by local inflammation. Searching for factors that can inhibit the transition of the cell population from the epithelial to the mesenchymal phenotype is very important for antitumor therapy. Interferon alfa (IFNα-2b) can be such a factor that has a direct effect on proliferation, differentiation and migration of tumor cells. The aim of this study was to compare the effect of IFNα-2b on the expression of EMT markers and on the basic cellular processes on 2D and 3D growth models.
2D cell culture was cultured in standard conditions with DMEM nutrient medium (Sigma, USA). The initial cell density was 2 × 104 cells/cm2. Concentrations of IFNα-2b were 103, 104 and 105 M/mL. For the initial generation of spheroids to DMEM nutrient medium 2% carboxymethylcellulose was added (Bio-Rad, USA). The spheroid culture was maintained for 7 days on an orbital shaker (PSU-10i, Biosan, Latvia). Every 24 h cells were counted in suspension and adhesion fractions using the routine method. Cell viability was evaluated by MTT assay. The Stemi2000 software AxioVisionRed 4.7 was used for image processing. The volume of spheroids was calculated by Bjerkvig formula. Markers were detected by applying IHC method with primary monoclonal antibodies: Ck (clone AE1/AE3, Dako, USA), vim (Clone V9, Dako, USA), EpCAM (Sigma, USA).
The change of the cell growth type caused a change of the expression of some EMT markers (CKs, EpCAM, Vim). IFNα-2b had a cytotoxic effect on tumor cells, and decreased the migration ability. IFNα-2b caused an increasing of Ck and EpCAM expression by 50.5% and 47.8%, respectively, compared with control in 2D cell culture. In 3D cell culture these increases were 33% and 34%, respectively, compared with the control. Expression of vim significantly showed no difference compared with the control.
IFNα-2b stimulated the differentiation and inhibited a migrational ability of tumor cells on early stages of breast cancer development.
Department of Biotechnical Problems of Diagnostics, Institute for Problems of Cryobiology and Cryomedicine, NAS of Ukraine
All authors have declared no conflicts of interest.