hypocrellin-b and Nasopharyngeal-Neoplasms

Hypocrellin B, a natural pigment from a traditional Chinese herb, has been attracting extensive attention. The present study aims to investigate whether hypocrellin B can enhance cell death induced by ultrasound sonification on nasopharyngeal carcinoma cells in vitro. The sonodynamic action of hypocrellin B was investigated on nasopharyngeal carcinoma cell line CNE2 cells as tumor model cells. In the experiments, the hypocrellin B concentration was kept constant at 2.5 microM and the cells were subject to ultrasound exposure for 15 s at an intensity of 0.65 W/cm(2). Cytotoxicity was investigated 24 h after ultrasound sonification. Apoptosis was evaluated using flow cytometry with annexin V-FITC and propidium iodine staining and nuclear staining with Hoechst 33258. Cell ultrastructure morphology was observed using transmission electron microscopy (TEM). No significant dark cytotoxicity of hypocrellin B in the CNE2 cells was observed at the concentration of 2.5 microM. The cell death rate induced by ultrasound sonification was significantly higher in the presence of hypocrellin B than in the absence of hypocrellin B. Flow cytometry showed that ultrasound exposure in the presence of hypocrellin B significantly increased the early and late apoptotic rate, 18.64% and 22.57%, respectively, compared with the controls. Nuclear condensation was observed in the nuclear staining and swollen mitochondria and more vacuolar and broken cell membrane were found in TEM after the treatment of hypocrellin B and ultrasound. Our findings demonstrated that the presence of hypocrellin B significantly enhanced the cytotoxicity of ultrasound radiation in CNE2 cells, suggesting that hypocrellin B is a novel sonosensitizer and hypocrellin B-mediated sonodynamic therapy is a potential therapeutic modality in the management of malignant tumors.

Topics: Carcinoma; Cell Death; Cell Line, Tumor; Flow Cytometry; Humans; Microscopy, Electron, Transmission; Molecular Structure; Nasopharyngeal Neoplasms; Perylene; Photosensitizing Agents; Quinones; Ultrasonic Therapy

The mitochondrion is an important target of ultrasound-induced cell death. This study aimed to investigate the mitochondrial damage in nasopharyngeal carcinoma (NPC) cells induced by ultrasound radiation in the presence of hypocrellin B (HB).. The NPC cell line CNE2 was used to investigate the effect of HB on ultrasonic action with an HB concentration of 2.5 mumol/L and ultrasound exposure for 15 seconds at an intensity of 0.65 W/cm(2). Cytotoxicity was investigated 24 hours after ultrasound exposure. Mitochondrial structure changes were observed by transmission electron microscopy. The mitochondrial membrane potential was evaluated by confocal laser-scanning microscopy with rhodamine 123 staining.. The mean death rates of the CNE2 cells +/- SD were 25.14% +/- 1.50% after ultrasound radiation alone and 76.72% +/- 1.13% after ultrasound radiation in the presence of HB. Transmission electron microscopy showed that slightly enlarging mitochondria were found in the ultrasound-treated cells. After treatment with ultrasound and HB together, some cells had seriously damaged mitochondria, namely, obvious swollen mitochondria and mitochondria in which cristae had almost completely disappeared. The mitochondrial membrane potential was more significantly collapsed when the CNE2 cells were exposed to HB for 5 hours and then ultrasound at 0.65 mW/cm(2) than with ultrasound radiation alone (P < .05).. Hypocrellin B significantly enhanced the cytotoxicity of ultrasound radiation in the CNE2 cells. The damage to the mitochondrial structure and function might be an important cause of death in the CNE2 cells induced by treatment with ultrasound radiation and HB together.

Topics: Apoptosis; Cell Line, Tumor; Humans; Mitochondria; Nasopharyngeal Neoplasms; Perylene; Quinones; Radiation Dosage; Radiation-Sensitizing Agents; Treatment Outcome; Ultrasonic Therapy

Novel photosensitizers Hypocrellin A (HA) and Hypocrellin B (HB), lipid soluble perylquinone derivatives of the genus Hypericum have a strong photodynamic effect on tumors and viruses. However, the mechanisms of tumor cell death induced by HA and HB are still unclear. In this study, we attempt to elucidate the photodynamic effects of HA and HB compounds in poorly differentiated (CNE2) and moderately differentiated (TW0-1) human nasopharyngeal carcinoma (NPC) cells as well as human mucosal colon (CCL-220.1) and bladder (SD) cells. Using these cell lines we investigated few hall marks of apoptotic commitments in a drug and light dose dependent manner. Tumor cells photoactivated with HA and HB showed cell size shrinkage and an increase in the sub-diploid DNA content. A loss of membrane phospholipid asymmetry associated with apoptosis was induced by all tumor cell lines as evidenced by the externalization of phosphatidylserine. Western blot analysis of poly (ADP-ribose) polymerase, a caspases substrate, showed the classical cleavage pattern (116 to 85kDa) associated with apoptosis in HA and HB-treated cell lysates. In addition, PARP cleavage was blocked by using tetrapepdide caspases inhibitors such as DEVD or z-VAD. These results demonstrate that tumor cell death induced by HB and HA is mediated by caspase proteases. This study also identifies both colon and bladder cells were more sensitive cell lines than NPC (CNE2 and TWO-1) cell lines.

Topics: Apoptosis; Carcinoma; Cell Culture Techniques; Cell Line, Tumor; Colon; Humans; Mucous Membrane; Nasopharyngeal Neoplasms; Perylene; Phenol; Photosensitizing Agents; Quinones; Urinary Bladder

It has been reported that novel photosensitizers Hypocrellin A and B, lipid soluble perylquinone derivatives of the genus Hypericum have a strong photodynamic effect on tumors and viruses. The molecular mechanisms of tumor cell death induction by Hypocrellin A and B are poorly understood. In this study, we have examined the photodynamic effects of Hypocrellin A and B compounds in poorly differentiated (CNE2) and moderately differentiated (TW0-1) human nasopharyngeal carcinoma (NPC) cells. Using these cell lines we investigated the role of the apoptotic pathway in photosensitized Hypocrellin A and B-mediated cell death. Tumor cells photoactivated with Hypocrellin A and B showed cell size shrinkage and an increase in the sub-diploid DNA content. A loss of membrane phospholipid asymmetry associated with apoptosis was induced by both tumor cell lines as evidenced by the externalization of phosphatidylserine (PS). A dose-dependent increase in caspases-3 protease activity inhibitable by the tetrapeptide inhibitor DEVD-CHO was also observed in both cell lines. Western blot analysis of poly (ADP-ribose) polymerase, a caspase substrate, showed the classical cleavage pattern (116 to 85 kDa) associated with apoptosis in Hypocrellin A and B-treated cell lysates. In addition, caspase inhibition blocked the externalization of membrane PS, indicating that the loss of membrane phospholipid asymmetry is a downstream event of caspases activation. These results demonstrate that tumor cell death induced by Hypocrellin A and B is mediated by caspase proteases. In conclusion, this study identifies both Hypocrellins (A and B) as potent and promising photosensitizers for the treatment of NPC.

Topics: Annexin A5; Apoptosis; Blotting, Western; Caspase 3; Caspase Inhibitors; Caspases; Cell Differentiation; DNA, Neoplasm; Dose-Response Relationship, Drug; Enzyme Inhibitors; Flow Cytometry; Humans; Nasopharyngeal Neoplasms; Perylene; Phenol; Phosphatidylserines; Photochemotherapy; Photosensitizing Agents; Poly(ADP-ribose) Polymerase Inhibitors; Poly(ADP-ribose) Polymerases; Quinones; Tumor Cells, Cultured

Potent photosensitizers hypocrellin A (HA), hypocrellin B (HB) and hypericin (HY) are lipid-soluble perylquinone derivatives of the genus Hypericum and have a strong photodynamic effect on tumors and viruses. However, the mechanisms of tumor cell death induced by HA, HB and HY are still unclear. Moreover, no reports have mentioned cell apoptosis induced by HA, HB and HY in human nasopharyngeal carcinoma (NPC) and other mucosal cells. In this study, we attempt to clarify the photodynamic effects of HA, HB and HY compounds in poorly differentiated (CNE2) and moderately differentiated (TW0-1) human NPC cells as well as human mucosal colon and bladder cells. Using these cell lines we investigated few hallmarks of apoptotic commitments in a drug dose dependent manner. Tumor cells photo-activated with HA, HB and HY showed cell size shrinkage and an increase in the sub-diploid DNA content. A loss of membrane phospholipid asymmetry associated with apoptosis was induced by all tumor cell lines as evidenced by the externalization of phosphatidylserine. Under apoptotic conditions, Western blot analysis of poly(ADP-ribose) polymerase, a caspases substrate, showed the classical cleavage pattern (116 to 85 kDa) associated with apoptosis in HA, HB and HY-treated cell lysates. In addition, 85 kDa cleaved product was blocked by the tetrapepdide caspase inhibitors such as DEVD-CHO or z-VAD-fmk. Both inhibitors protect tumor cells from apoptosis. These results demonstrate that tumor cell death induced by HA, HB and HY is mediated by caspase proteases. This study also identifies HB as a more potent and promising photosensitizer for the treatment of mucosal cancer cells.

Topics: Anthracenes; Antineoplastic Agents; Apoptosis; Caspase 3; Caspases; Cell Line; Cell Membrane; Cell Size; Cell Survival; DNA, Neoplasm; Enzyme Activation; Humans; Mucous Membrane; Nasopharyngeal Neoplasms; Perylene; Phenol; Phosphatidylserines; Photosensitizing Agents; Poly(ADP-ribose) Polymerases; Quinones; Tumor Cells, Cultured