Journal
BIOCHIMICA ET BIOPHYSICA ACTA-GENERAL SUBJECTS
Volume 1830, Issue 10, Pages 4611-4620Publisher
ELSEVIER
DOI: 10.1016/j.bbagen.2013.05.021
Keywords
Apoptosis; Liposome; Photosensitizer; Temocene
Categories
Funding
- Spanish Ministry of Economy and Competitiveness [CTQ2010-20870-C03]
- Comissionat per a Universitats i Recerca del Departament d'Innovacio Universitats i Empresa de la Generalitat de Catalunya i del Fons Social Europeu
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Background: The cell death pathway activated after photodynamic therapy (PDT) is controlled by a variety of parameters including the chemical structure of the photosensitizer, its subcellular localization, and the photodynamic damage induced. The present study aims to characterize a suitable m-THPPo liposomal formulation, to determine its subcellular localization in HeLa cells and to establish the cell death mechanisms that are activated after photodynamic treatments. Methods: Liposomes containing m-THPPo were prepared from a mixture of DPPC and DMPG at a 9:1 molar ratio. In order to procure the best encapsulation efficiency, the m-THPPo/lipid molar ratio was considered. HeLa cells were incubated with liposomal m-THPPo and the subcellular localization of m-THPPo was studied. Several assays such as TUNEL, annexin V/propidium iodide and Hoechst-33258 staining were performed after photodynamic treatments. The apoptotic initiation was assessed by cytochrome c and caspase-2 immunofluorescence. Results: m-THPPo encapsulated in liposomes showed a decrease of the fluorescence and singlet oxygen quantum yields, compared to those of m-THPPo dissolved in tetrahydrofuran. Liposomal m-THPPo showed colocalization with LysoTracker (R) and it induced photoinactivation of HeLa cells by an apoptotic mechanism. In apoptotic cells no relocalization of cytochrome c could be detected, but caspase-2 was positive immediately after. photosensitizing treatments. Conclusions: Photodynamic treatment with liposomal m-THPPo leads to a significant percentage of apoptotic morphology of HeLa cells. The activation of caspase-2, without the relocalization of cytochrome c, indicates a mitochondrial-independent apoptotic mechanism. General significance: These results provide a better understanding of the cell death mechanism induced after liposomal m-THPPo photodynamic treatment. (c) 2013 Elsevier B.V. All rights reserved.
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