Induction of apoptosis in breast cancer cells in vitro by Fas ligand reverse signaling.

Abstract

The Fas-antigen is a cell surface receptor that transduces apoptotic signals into cells. The purpose of this study was to evaluate FasL expression in breast cancer and to elucidate the role of its signaling in different breast cancer cell lines. T47D and MCF7 cells were used and cultured in Dulbecco's modified Eagle's medium. FasL translocation to the membrane was achieved by culturing the cells in the presence of human interferon-γ (IFNγ). Translocation was detected by immunofluorescence. The ability of a Fas:Fc fusion protein to trigger apoptosis in these cells was investigated by cell death detection ELISA. After incubation with IFNγ for 4h and 18h, apoptosis was assessed in response to treatment with Fas:Fc. Immunofluorescence revealed that the used cell lines were positive for FasL which was increased and changed to more membrane-bound FasL expression after IFNγ stimulation. After stimulation with 50IU/ml IFNγ, Fas:Fc significantly increased MCF7 apoptosis (1.39 ± 0.06-fold, p = 0.0004) after 18h. After stimulation with 100IU/ml, Fas:Fc significantly increased apoptosis both after 4h (1.49 ± 0.15-fold, p = 0.018) and 18h (1.30 ± 0.06-fold, p = 0.013). In T47D cells this effect was seen after 4h of stimulation with 50IU/ml and addition of Fas:Fc (1.6 ± 0.08-fold, p = 0.03). Membrane-bound FasL expression could be induced by IFNγ in a breast cancer cell model. More importantly, in the presence of IFNγ the Fas:Fc fusion protein was able to transmit pro-apoptotic signals to T47D and MCF7 cells, significantly inducing apoptosis. The current findings support further in vivo studies regarding FasL activation as a potential target for therapeutic intervention in breast cancer.