The effects of Ce6-mediated sono-photodynamic therapy on cell migration, apoptosis and autophagy in mouse mammary 4T1 cell line

Purpose: Sono-Photodynamic therapy (SPDT) is an alternative therapy which claims to enhance the anticancer effects by combining sonodynamic therapy (SDT) with photodynamic therapy (PDT). In the present study, we investigated the effects of chlorin e6 (Ce6) mediated SPDT on migration, apoptosis and autophagy in mouse mammary 4T1 cancer cells, and its underlying mechanisms. Materials: Cell migration was determined by wound healing assay. Apoptosis was analyzed using annexin V-PE/7-ADD staining as well as Hoechst 33342 staining. Changes of mitochondria membrane potential (MMP) was evaluated by flow cytometry. Formation of acidic vesicular organelles (AVOs) during autophagy was observed with fluorescence microscope by MDC staining. Immunofluorescence assays were performed to detect the co-localization of LC3 and Lamp2. Western blotting was employed to analyze the activity of the apoptosis related proteins Caspase-3, PARP, Bax and Bcl-2, as well as the autophagy associated processing of LC3-I to LC3-II and Beclin-1 expression. Results: Ce6 mediated SPDT further enhanced cell migration inhibition, significantly triggered cell apoptosis, nuclear condensation and MMP drop. Cleaved Caspase-3 and PARP increased dramatically after Ce6-SPDT, accompanied by decreased Bcl-2 expression, while the expression of Bax remained stable. Additionally, AVOs formation, co-localization of LC3 and Lamp2 occurred following Ce6-SPDT and simultaneously accompanied by LC3-II processing and increased Beclin-1 expression. Conclusions: Ce6-SPDT could enhance cell migration inhibition, and induce mitochondria-dependent apoptosis as well as autophagy in 4T1 cells. Crown Copyright (C) 2013 Published by Elsevier B.V. All rights reserved.