Liposomal temocene (m-THPPo) photodynamic treatment induces cell death by mitochondria-independent apoptosis

• Published in: Biochimica et biophysica acta Vol. 1830; no. 10; pp. 4611 - 4620
• Main Authors: Soriano, Jorge, García-Díaz, María, Mora, Margarita, Sagristá, Maria Lluïsa, Nonell, Santi, Villanueva, Angeles, Stockert, Juan Carlos, Cañete, Magdalena
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.10.2013
• Subjects: Annexin A5 - metabolism; Apoptosis; Apoptosis - drug effects; Caspase 2 - metabolism; caspase-2; cell death; cells; chemical structure; cytochrome c; Cytochromes c - metabolism; encapsulation; fluorescence; Fluorescent Antibody Technique; HeLa Cells; Humans; In Situ Nick-End Labeling; Liposome; Liposomes; Mitochondria - drug effects; Photochemotherapy; Photosensitizer; Porphyrins - administration & dosage; Porphyrins - pharmacology; propidium; singlet oxygen; Temocene; tetrahydrofuran; therapeutics; DMPG; PBS; PDT; PCS; PS; DMSO; Liposome; m-THPPo; TB; Temocene; THF; Hoechst-33258; ROS; MLVs; PI; Photosensitizer; DPPC; Apoptosis; H-33258; dipalmitoylphosphatidylcholine; dimyristoylphosphatidylglycerol; photon correlation spectroscopy; dimethylsulfoxide; photodynamic therapy; toluidine blue; propidium iodide; reactive oxygen species; multilamellar vesicles; tetrahydrofuran; m-tetra (hydroxyphenyl) porphycene (temocene); phosphate buffered saline
• ISSN: 0304-4165; 0006-3002; 1872-8006
• DOI: 10.1016/j.bbagen.2013.05.021

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. 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. 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® 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. 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. These results provide a better understanding of the cell death mechanism induced after liposomal m-THPPo photodynamic treatment. •m-THPPo-lipo accumulates in lysosomes of HeLa cells.•The phototreatment, with m-THPPo-lipo produces independent-mitochondria apoptosis.•m-THPPo-DMSO is located in mitochondria of HeLa cells.•The phototreatment with m-THPPo-DMSO produces apoptosis mitochondria-dependent.•The vehiculization of a drug can determine different cell death routes.