D Gayathri Devi1, T R Cibin, Annie Abraham. 1. Department of Biochemistry, University of Kerala, Kariavattom, Thiruvananthapuram 695581, Kerala, India.
Abstract
BACKGROUND: Despite findings that photodynamic treatment with bis (3,5-diiodo-2,4,6-trihydroxyphenyl) squaraine initiated tumor regression in mice skin, queries regarding its mode of action - answers to which will be functional to design clinical trials on squaraine based photodynamic therapy - remain unanswered. Our investigation reveals the in vivo mechanism of action of the photosensitizer. METHODS: Skin tumor was induced in Swiss albino mice using 7,12-dimethyl benzanthacene. After the intraperitoneal administration of the dye in tumor induced mice, its concentration in subcellular fractions of the tumor tissue was determined fluorimetrically. Cytochrome c release from the mitochondrial membrane after the photodynamic treatment was analyzed. The observations stemming from this part lead to histopathological examination of tumor tissues. Apoptotic markers like caspase-3, Bcl-2 and Bax were also studied. RESULTS: Major portion of the dye accumulated in the mitochondria. Cytochrome c leakage from mitochondria after squaraine PDT suggests loss of mitochondrial membrane integrity, which was further confirmed by the results of histopathological analysis. The activity of caspase-3 was elevated, expression of Bcl-2 diminished and that of Bax increased - all these results show enhancement of apoptosis in the tumor region after the treatment. CONCLUSIONS: The results lead to the elucidation of mechanism of tumor destruction which proves to be mitochondria mediated apoptotic damage of tumor tissue. The study assumes significance since it defines the in vivo mode of action of a photosensitizer. Also, the query of how a squaraine based photosensitizer evokes tumor response is being dealt with here, for the first time.
BACKGROUND: Despite findings that photodynamic treatment with bis (3,5-diiodo-2,4,6-trihydroxyphenyl) squaraine initiated tumor regression in mice skin, queries regarding its mode of action - answers to which will be functional to design clinical trials on squaraine based photodynamic therapy - remain unanswered. Our investigation reveals the in vivo mechanism of action of the photosensitizer. METHODS:Skin tumor was induced in Swiss albino mice using 7,12-dimethyl benzanthacene. After the intraperitoneal administration of the dye in tumor induced mice, its concentration in subcellular fractions of the tumor tissue was determined fluorimetrically. Cytochrome c release from the mitochondrial membrane after the photodynamic treatment was analyzed. The observations stemming from this part lead to histopathological examination of tumor tissues. Apoptotic markers like caspase-3, Bcl-2 and Bax were also studied. RESULTS: Major portion of the dye accumulated in the mitochondria. Cytochrome c leakage from mitochondria after squaraine PDT suggests loss of mitochondrial membrane integrity, which was further confirmed by the results of histopathological analysis. The activity of caspase-3 was elevated, expression of Bcl-2 diminished and that of Bax increased - all these results show enhancement of apoptosis in the tumor region after the treatment. CONCLUSIONS: The results lead to the elucidation of mechanism of tumor destruction which proves to be mitochondria mediated apoptotic damage of tumor tissue. The study assumes significance since it defines the in vivo mode of action of a photosensitizer. Also, the query of how a squaraine based photosensitizer evokes tumor response is being dealt with here, for the first time.