蟾酥是中國的傳統中藥，在中國南方用來治療癌症已有數百年的歷史，但蟾酥的抗癌機制至今尚不明瞭。在台灣，肝癌是最常罹患的惡性腫瘤之一，給予化學療法對肝癌的治療的並無顯著療效。在台灣的蟾蜍所得到的蟾酥，其主要成分為bufotalin，我們是第一個對bufotalin的抗肝癌作用機轉進行研究，並設計了一系列的實驗來探討bufotalin對於肝癌的抗癌機制，實驗結果顯示以bufotalin處理Hep 3B細胞6天，其抑制細胞50 %生長的濃度為0.24 mM，而以bufotalin處理Hep 3B細胞14天，其抑制細胞在軟凝膠中50 %細胞群落形成的濃度為0.12 mM。以bufotalin處理Hep 3B細胞後，可以觀察到細胞膜內面的磷脂絲胺酸翻轉至膜外、sub-G1期的DNA含量增加 及DNA片段和凋亡小體的形成，顯示bufotalin是經由細胞凋亡的過程抑制Hep 3B細胞的生長。Bufotalin引起細胞凋亡的過程中會造成粒線體膜電位喪失，其分子機制是經由活化caspase-8，使粒線體上tBid和Bak的表現增加，而粒線體上Bcl-2和Bcl-XL的表現減少，並造成XIAP蛋白質含量的減少，接著引發caspase-9和caspase-3的活化，進而切割核內的poly (ADP-ribose) polymerase和DFF45，最後引發DNA的斷裂。這些結果顯示出bufotalin會透過caspase-8的活化而引發細胞凋亡，有助於作為抗癌藥物的研發。

　　Ch’an Su, a traditional Chinese medicine, has been used in cancer therapy in south area of China for hundreds of years. The antitumor mechanism of Chan Su is still unclear. Human hepatocellular carcinoma (HCC) is one of the most common malignant neoplasms in Taiwan with poor response to chemotherapy. Bufotalin is one of major constituents of Chan Su. In the present study, we are the first to explore the antitumor mechanism of bufotalin in HCC. The inhibition concentrations (IC50) of bufotalin on HCC Hep 3B cell growth was 0.24 mM at day 6 of post-treatment. At day 14 post-treatment, the IC50 of colony formation in soft agar was 0.12 mM. Bufotalin inhibited the growth of Hep 3B cells via apoptosis, following by phosphatidylserine exposure, increased in sub-G1 content, and DNA fragmentation. The molecular basis of bufotalin triggered apoptosis seems to associate with the activation of caspase-8, the loss of mitochondrial membrane potential, decreased the expression of XIAP, up expressions of mitochondrial tBid and Bak, and down expressions of mitochondrial Bcl-2 and Bcl-XL, than activation of caspase-9 and -3, which, in turn, increased nuclear cleavages of poly (ADP-ribose) polymerase and DFF45, and DNA fragmentation. These findings indicate that bufotalin is an anticancer agent that triggered apoptosis via the activation of caspase-8.

Adams, J. M., and Cory, S. (1998). The Bcl-2 protein family: arbiters of cell survival. Science 281, 1322-1326.
Arii, S., Yamaoka, Y., Futagawa, S., Inoue, K., Kobayashi, K., Kojiro, M., Makuuchi, M., Nakamura, Y., Okita, K., and Yamada, R. (2000). Results of surgical and nonsurgical treatment for small-sized hepatocellular carcinomas: a retrospective and nationwide survey in Japan. The Liver Cancer Study Group of Japan. Hepatology 32, 1224-1229.
Ashkenazi, A., and Dixit, V. M. (1998). Death receptors: signaling and modulation. Science 281, 1305-1308.
Barnett, D. K., Kimura, J., and Bavister, B. D. (1996). Translocation of active mitochondria during hamster preimplantation embryo development studied by confocal laser scanning microscopy. Dev Dyn 205, 64-72.
Bismuth, H., and Majno, P. E. (2000). Hepatobiliary surgery. J Hepatol 32, 208-224.
Block, T. M., Mehta, A. S., Fimmel, C. J., and Jordan, R. (2003). Molecular viral oncology of hepatocellular carcinoma. Oncogene 22, 5093-5107.
Boatright, K. M., and Salvesen, G. S. (2003). Mechanisms of caspase activation. Curr Opin Cell Biol 15, 725-731.
Bodmer, J. L., Holler, N., Reynard, S., Vinciguerra, P., Schneider, P., Juo, P., Blenis, J., and Tschopp, J. (2000). TRAIL receptor-2 signals apoptosis through FADD and caspase-8. Nat Cell Biol 2, 241-243.
Boise, L. H., Gonzalez-Garcia, M., Postema, C. E., Ding, L., Lindsten, T., Turka, L. A., Mao, X., Nunez, G., and Thompson, C. B. (1993). bcl-x, a bcl-2-related gene that functions as a dominant regulator of apoptotic cell death. Cell 74, 597-608.
Borner, C. (2003). The Bcl-2 protein family: sensors and checkpoints for life-or-death decisions. Mol Immunol 39, 615-647.
Bruno, S., Silini, E., Crosignani, A., Borzio, F., Leandro, G., Bono, F., Asti, M., Rossi, S., Larghi, A., Cerino, A., et al. (1997). Hepatitis C virus genotypes and risk of hepatocellular carcinoma in cirrhosis: a prospective study. Hepatology 25, 754-758.
Cain, K., Bratton, S. B., and Cohen, G. M. (2002). The Apaf-1 apoptosome: a large caspase-activating complex. Biochimie 84, 203-214.
Cande, C., Cohen, I., Daugas, E., Ravagnan, L., Larochette, N., Zamzami, N., and Kroemer, G. (2002). Apoptosis-inducing factor (AIF): a novel caspase-independent death effector released from mitochondria. Biochimie 84, 215-222.
Chapman, R. S., Chresta, C. M., Herberg, A. A., Beere, H. M., Heer, S., Whetton, A. D., Hickman, J. A., and Dive, C. (1995). Further characterisation of the in situ terminal deoxynucleotidyl transferase (TdT) assay for the flow cytometric analysis of apoptosis in drug resistant and drug sensitive leukaemic cells. Cytometry 20, 245-256.
Chen, K. K., and Kovarikova, A. (1967). Pharmacology and toxicology of toad venom. J Pharm Sci 56, 1535-1541.
Chittenden, T., Harrington, E. A., O'Connor, R., Flemington, C., Lutz, R. J., Evan, G. I., and Guild, B. C. (1995). Induction of apoptosis by the Bcl-2 homologue Bak. Nature 374, 733-736.
Coppola, D., Saunders, B., Fu, L., Mao, W., and Nicosia, S. V. (1999). The insulin-like growth factor 1 receptor induces transformation and tumorigenicity of ovarian mesothelial cells and down-regulates their Fas-receptor expression. Cancer Res 59, 3264-3270.
D'Amours, D., Desnoyers, S., D'Silva, I., and Poirier, G. G. (1999). Poly(ADP-ribosyl)ation reactions in the regulation of nuclear functions. Biochem J 342 ( Pt 2), 249-268.
Degli Esposti, M., and Dive, C. (2003). Mitochondrial membrane permeabilisation by Bax/Bak. Biochem Biophys Res Commun 304, 455-461.
Deveraux, Q. L., Takahashi, R., Salvesen, G. S., and Reed, J. C. (1997). X-linked IAP is a direct inhibitor of cell-death proteases. Nature 388, 300-304.
Di Bisceglie, A. M. (1997). Hepatitis C and hepatocellular carcinoma. Hepatology 26, 34S-38S.
Du, C., Fang, M., Li, Y., Li, L., and Wang, X. (2000). Smac, a mitochondrial protein that promotes cytochrome c-dependent caspase activation by eliminating IAP inhibition. Cell 102, 33-42.
Earnshaw, W. C., Martins, L. M., and Kaufmann, S. H. (1999). Mammalian caspases: structure, activation, substrates, and functions during apoptosis. Annu Rev Biochem 68, 383-424.
Enari, M., Sakahira, H., Yokoyama, H., Okawa, K., Iwamatsu, A., and Nagata, S. (1998). A caspase-activated DNase that degrades DNA during apoptosis, and its inhibitor ICAD. Nature 391, 43-50.
Eskes, R., Desagher, S., Antonsson, B., and Martinou, J. C. (2000). Bid induces the oligomerization and insertion of Bax into the outer mitochondrial membrane. Mol Cell Biol 20, 929-935.
Farrow, S. N., White, J. H., Martinou, I., Raven, T., Pun, K. T., Grinham, C. J., Martinou, J. C., and Brown, R. (1995). Cloning of a bcl-2 homologue by interaction with adenovirus E1B 19K. Nature 374, 731-733.
Feitelson, M. A. (1999). Hepatitis B virus in hepatocarcinogenesis. J Cell Physiol 181, 188-202.
Ferri, K. F., and Kroemer, G. (2001). Organelle-specific initiation of cell death pathways. Nat Cell Biol 3, E255-263.
Furuya, T., Kamada, T., Murakami, T., Kurose, A., and Sasaki, K. (1997). Laser scanning cytometry allows detection of cell death with morphological features of apoptosis in cells stained with PI. Cytometry 29, 173-177.
Garcea, G., Lloyd, T. D., Aylott, C., Maddern, G., and Berry, D. P. (2003). The emergent role of focal liver ablation techniques in the treatment of primary and secondary liver tumours. Eur J Cancer 39, 2150-2164.
Giuliano, M., Lauricella, M., Calvaruso, G., Carabillo, M., Emanuele, S., Vento, R., and Tesoriere, G. (1999). The apoptotic effects and synergistic interaction of sodium butyrate and MG132 in human retinoblastoma Y79 cells. Cancer Res 59, 5586-5595.
Giuliano, M., Vento, R., Lauricella, M., Calvaruso, G., Carabillo, M., and Tesoriere, G. (1996). Role of insulin-like growth factors in autocrine growth of human retinoblastoma Y79 cells. Eur J Biochem 236, 523-532.
Green, D. R. (2000). Apoptotic pathways: paper wraps stone blunts scissors. Cell 102, 1-4.
Green, D. R., and Kroemer, G. (2004). The pathophysiology of mitochondrial cell death. Science 305, 626-629.
Griffiths, G. J., Corfe, B. M., Savory, P., Leech, S., Esposti, M. D., Hickman, J. A., and Dive, C. (2001). Cellular damage signals promote sequential changes at the N-terminus and BH-1 domain of the pro-apoptotic protein Bak. Oncogene 20, 7668-7676.
Gross, A., Jockel, J., Wei, M. C., and Korsmeyer, S. J. (1998). Enforced dimerization of BAX results in its translocation, mitochondrial dysfunction and apoptosis. Embo J 17, 3878-3885.
Gu, J., Dong, R. P., Zhang, C., McLaughlin, D. F., Wu, M. X., and Schlossman, S. F. (1999). Functional interaction of DFF35 and DFF45 with caspase-activated DNA fragmentation nuclease DFF40. J Biol Chem 274, 20759-20762.
Hengartner, M. O. (2000). The biochemistry of apoptosis. Nature 407, 770-776.
Herr, I., and Debatin, K. M. (2001). Cellular stress response and apoptosis in cancer therapy. Blood 98, 2603-2614.
Hofer-Warbinek, R., Schmid, J. A., Stehlik, C., Binder, B. R., Lipp, J., and de Martin, R. (2000). Activation of NF-kappa B by XIAP, the X chromosome-linked inhibitor of apoptosis, in endothelial cells involves TAK1. J Biol Chem 275, 22064-22068.
Hong, S. J., Dawson, T. M., and Dawson, V. L. (2004). Nuclear and mitochondrial conversations in cell death: PARP-1 and AIF signaling. Trends Pharmacol Sci 25, 259-264.
Inohara, N., Koseki, T., Chen, S., Benedict, M. A., and Nunez, G. (1999). Identification of regulatory and catalytic domains in the apoptosis nuclease DFF40/CAD. J Biol Chem 274, 270-274.
Ishikawa, M., Yogita, S., Miyake, H., Fukuda, Y., Harada, M., Wada, D., and Tashiro, S. (2002). Clarification of risk factors for hepatectomy in patients with hepatocellular carcinoma. Hepatogastroenterology 49, 1625-1631.
Jacotot, E., Costantini, P., Laboureau, E., Zamzami, N., Susin, S. A., and Kroemer, G. (1999). Mitochondrial membrane permeabilization during the apoptotic process. Ann N Y Acad Sci 887, 18-30.
Jemmerson, R., LaPlante, B., and Treeful, A. (2002). Release of intact, monomeric cytochrome c from apoptotic and necrotic cells. Cell Death Differ 9, 538-548.
Jiang, X., and Wang, X. (2000). Cytochrome c promotes caspase-9 activation by inducing nucleotide binding to Apaf-1. J Biol Chem 275, 31199-31203.
Jonas, S., Steinmuller, T., Tullius, S. G., Thelen, A., Settmacher, U., Berg, T., Radtke, C., and Neuhaus, P. (2003). Increased mortality after liver transplantation for hepatocellular carcinoma in hepatitis B-associated cirrhosis. Transpl Int 16, 33-36.
Juan, G., Cavazzoni, M., Saez, G. T., and O'Connor, J. E. (1994). A fast kinetic method for assessing mitochondrial membrane potential in isolated hepatocytes with rhodamine 123 and flow cytometry. Cytometry 15, 335-342.
Juo, P., Kuo, C. J., Yuan, J., and Blenis, J. (1998). Essential requirement for caspase-8/FLICE in the initiation of the Fas-induced apoptotic cascade. Curr Biol 8, 1001-1008.
Kaufmann, S. H., Desnoyers, S., Ottaviano, Y., Davidson, N. E., and Poirier, G. G. (1993). Specific proteolytic cleavage of poly(ADP-ribose) polymerase: an early marker of chemotherapy-induced apoptosis. Cancer Res 53, 3976-3985.
Kawasaki, M., Sasaki, K., Satoh, T., Kurose, A., Kamada, T., Furuya, T., Murakami, T., and Todoroki, T. (1997). Laser scanning cytometry (LCS) allows detailed analysis of the cell cycle in PI stained human fibroblasts (TIG-7). Cell Prolif 30, 139-147.
Kelekar, A., and Thompson, C. B. (1998). Bcl-2-family proteins: the role of the BH3 domain in apoptosis. Trends Cell Biol 8, 324-330.
Kemeny, N., Huang, Y., Cohen, A. M., Shi, W., Conti, J. A., Brennan, M. F., Bertino, J. R., Turnbull, A. D., Sullivan, D., Stockman, J., et al. (1999). Hepatic arterial infusion of chemotherapy after resection of hepatic metastases from colorectal cancer. N Engl J Med 341, 2039-2048.
Kischkel, F. C., Lawrence, D. A., Chuntharapai, A., Schow, P., Kim, K. J., and Ashkenazi, A. (2000). Apo2L/TRAIL-dependent recruitment of endogenous FADD and caspase-8 to death receptors 4 and 5. Immunity 12, 611-620.
Knowles, B. B., Howe, C. C., and Aden, D. P. (1980). Human hepatocellular carcinoma cell lines secrete the major plasma proteins and hepatitis B surface antigen. Science 209, 497-499.
Korsmeyer, S. J., Wei, M. C., Saito, M., Weiler, S., Oh, K. J., and Schlesinger, P. H. (2000). Pro-apoptotic cascade activates BID, which oligomerizes BAK or BAX into pores that result in the release of cytochrome c. Cell Death Differ 7, 1166-1173.
Krammer, P. H. (2000). CD95's deadly mission in the immune system. Nature 407, 789-795.
Kroemer, G., and Reed, J. C. (2000). Mitochondrial control of cell death. Nat Med 6, 513-519.
Krysko, O., De Ridder, L., and Cornelissen, M. (2004). Phosphatidylserine exposure during early primary necrosis (oncosis) in JB6 cells as evidenced by immunogold labeling technique. Apoptosis 9, 495-500.
Kunji, E. R. (2004). The role and structure of mitochondrial carriers. FEBS Lett 564, 239-244.
Lassus, P., Opitz-Araya, X., and Lazebnik, Y. (2002). Requirement for caspase-2 in stress-induced apoptosis before mitochondrial permeabilization. Science 297, 1352-1354.
Lautier, D., Lagueux, J., Thibodeau, J., Menard, L., and Poirier, G. G. (1993). Molecular and biochemical features of poly (ADP-ribose) metabolism. Mol Cell Biochem 122, 171-193.
Leon, J., Acuna-Castroviejo, D., Sainz, R. M., Mayo, J. C., Tan, D. X., and Reiter, R. J. (2004). Melatonin and mitochondrial function. Life Sci 75, 765-790.
Li, L. Y., Luo, X., and Wang, X. (2001). Endonuclease G is an apoptotic DNase when released from mitochondria. Nature 412, 95-99.
Li, P., Nijhawan, D., Budihardjo, I., Srinivasula, S. M., Ahmad, M., Alnemri, E. S., and Wang, X. (1997). Cytochrome c and dATP-dependent formation of Apaf-1/caspase-9 complex initiates an apoptotic protease cascade. Cell 91, 479-489.
Liu, X., Kim, C. N., Yang, J., Jemmerson, R., and Wang, X. (1996). Induction of apoptotic program in cell-free extracts: requirement for dATP and cytochrome c. Cell 86, 147-157.
Liu, X., Li, P., Widlak, P., Zou, H., Luo, X., Garrard, W. T., and Wang, X. (1998). The 40-kDa subunit of DNA fragmentation factor induces DNA fragmentation and chromatin condensation during apoptosis. Proc Natl Acad Sci U S A 95, 8461-8466.
Liu, X., Zou, H., Slaughter, C., and Wang, X. (1997). DFF, a heterodimeric protein that functions downstream of caspase-3 to trigger DNA fragmentation during apoptosis. Cell 89, 175-184.
Llovet, J. M., Burroughs, A., and Bruix, J. (2003). Hepatocellular carcinoma. Lancet 362, 1907-1917.
Llovet, J. M., Fuster, J., and Bruix, J. (1999). Intention-to-treat analysis of surgical treatment for early hepatocellular carcinoma: resection versus transplantation. Hepatology 30, 1434-1440.
Lo, C. M., Ngan, H., Tso, W. K., Liu, C. L., Lam, C. M., Poon, R. T., Fan, S. T., and Wong, J. (2002). Randomized controlled trial of transarterial lipiodol chemoembolization for unresectable hepatocellular carcinoma. Hepatology 35, 1164-1171.
Lockshin, R. A., and Zakeri, Z. (2004). Caspase-independent cell death? Oncogene 23, 2766-2773.
Lorenz, H. M., Herrmann, M., Winkler, T., Gaipl, U., and Kalden, J. R. (2000). Role of apoptosis in autoimmunity. Apoptosis 5, 443-449.
Lorenzo, H. K., Susin, S. A., Penninger, J., and Kroemer, G. (1999). Apoptosis inducing factor (AIF): a phylogenetically old, caspase-independent effector of cell death. Cell Death Differ 6, 516-524.
Luo, X., Budihardjo, I., Zou, H., Slaughter, C., and Wang, X. (1998). Bid, a Bcl2 interacting protein, mediates cytochrome c release from mitochondria in response to activation of cell surface death receptors. Cell 94, 481-490.
Lynch, D. H., Campbell, K. A., Miller, R. E., Badley, A. D., and Paya, C. V. (1996). FasL/Fas and TNF/TNFR interactions in the regulation of immune responses and disease. Behring Inst Mitt, 175-184.
Masuda, Y., Kawazoe, N., Nakajo, S., Yoshida, T., Kuroiwa, Y., and Nakaya, K. (1995). Bufalin induces apoptosis and influences the expression of apoptosis-related genes in human leukemia cells. Leuk Res 19, 549-556.
Mathur, A., Hong, Y., Kemp, B. K., Barrientos, A. A., and Erusalimsky, J. D. (2000). Evaluation of fluorescent dyes for the detection of mitochondrial membrane potential changes in cultured cardiomyocytes. Cardiovasc Res 46, 126-138.
McCarty, J. S., Toh, S. Y., and Li, P. (1999). Study of DFF45 in its role of chaperone and inhibitor: two independent inhibitory domains of DFF40 nuclease activity. Biochem Biophys Res Commun 264, 176-180.
McDonnell, M. A., Wang, D., Khan, S. M., Vander Heiden, M. G., and Kelekar, A. (2003). Caspase-9 is activated in a cytochrome c-independent manner early during TNFalpha-induced apoptosis in murine cells. Cell Death Differ 10, 1005-1015.
Mevorach, D. (1999). The immune response to apoptotic cells. Ann N Y Acad Sci 887, 191-198.
Molinete, M., Vermeulen, W., Burkle, A., Menissier-de Murcia, J., Kupper, J. H., Hoeijmakers, J. H., and de Murcia, G. (1993). Overproduction of the poly(ADP-ribose) polymerase DNA-binding domain blocks alkylation-induced DNA repair synthesis in mammalian cells. Embo J 12, 2109-2117.
Mosmann, T. (1983). Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. J Immunol Methods 65, 55-63.
Nechushtan, A., Smith, C. L., Hsu, Y. T., and Youle, R. J. (1999). Conformation of the Bax C-terminus regulates subcellular location and cell death. Embo J 18, 2330-2341.
Nechushtan, A., Smith, C. L., Lamensdorf, I., Yoon, S. H., and Youle, R. J. (2001). Bax and Bak coalesce into novel mitochondria-associated clusters during apoptosis. J Cell Biol 153, 1265-1276.
Nicholson, D. W., Ali, A., Thornberry, N. A., Vaillancourt, J. P., Ding, C. K., Gallant, M., Gareau, Y., Griffin, P. R., Labelle, M., Lazebnik, Y. A., and et al. (1995). Identification and inhibition of the ICE/CED-3 protease necessary for mammalian apoptosis. Nature 376, 37-43.
O'Reilly, L. A., Ekert, P., Harvey, N., Marsden, V., Cullen, L., Vaux, D. L., Hacker, G., Magnusson, C., Pakusch, M., Cecconi, F., et al. (2002). Caspase-2 is not required for thymocyte or neuronal apoptosis even though cleavage of caspase-2 is dependent on both Apaf-1 and caspase-9. Cell Death Differ 9, 832-841.
Partik, G., Kahl-Rainer, P., Sedivy, R., Ellinger, A., Bursch, W., and Marian, B. (1998). Apoptosis in human colorectal tumours: ultrastructure and quantitative studies on tissue localization and association with bak expression. Virchows Arch 432, 415-426.
Petros, A. M., Medek, A., Nettesheim, D. G., Kim, D. H., Yoon, H. S., Swift, K., Matayoshi, E. D., Oltersdorf, T., and Fesik, S. W. (2001). Solution structure of the antiapoptotic protein bcl-2. Proc Natl Acad Sci U S A 98, 3012-3017.
Rao, L., Modha, D., and White, E. (1997). The E1B 19K protein associates with lamins in vivo and its proper localization is required for inhibition of apoptosis. Oncogene 15, 1587-1597.
Rathmell, J. C., and Thompson, C. B. (2002). Pathways of apoptosis in lymphocyte development, homeostasis, and disease. Cell 109 Suppl, S97-107.
Reed, C. J. (2000a). Apoptosis and cancer: strategies for integrating programmed cell death. Semin Hematol 37, 9-16.
Reed, J. C. (1995). Regulation of apoptosis by bcl-2 family proteins and its role in cancer and chemoresistance. Curr Opin Oncol 7, 541-546.
Reed, J. C. (2000b). Mechanisms of apoptosis. Am J Pathol 157, 1415-1430.
Reed, J. C., Haldar, S., Cuddy, M. P., Croce, C., and Makover, D. (1989). Deregulated BCL2 expression enhances growth of a human B cell line. Oncogene 4, 1123-1127.
Rodriguez, J., and Lazebnik, Y. (1999). Caspase-9 and APAF-1 form an active holoenzyme. Genes Dev 13, 3179-3184.
Ruffolo, S. C., Breckenridge, D. G., Nguyen, M., Goping, I. S., Gross, A., Korsmeyer, S. J., Li, H., Yuan, J., and Shore, G. C. (2000). BID-dependent and BID-independent pathways for BAX insertion into mitochondria. Cell Death Differ 7, 1101-1108.
Sakahira, H., Enari, M., and Nagata, S. (1998). Cleavage of CAD inhibitor in CAD activation and DNA degradation during apoptosis. Nature 391, 96-99.
Sattler, M., Liang, H., Nettesheim, D., Meadows, R. P., Harlan, J. E., Eberstadt, M., Yoon, H. S., Shuker, S. B., Chang, B. S., Minn, A. J., et al. (1997). Structure of Bcl-xL-Bak peptide complex: recognition between regulators of apoptosis. Science 275, 983-986.
Sawada, M., Sun, W., Hayes, P., Leskov, K., Boothman, D. A., and Matsuyama, S. (2003). Ku70 suppresses the apoptotic translocation of Bax to mitochondria. Nat Cell Biol 5, 320-329.
Scaffidi, C., Fulda, S., Srinivasan, A., Friesen, C., Li, F., Tomaselli, K. J., Debatin, K. M., Krammer, P. H., and Peter, M. E. (1998). Two CD95 (APO-1/Fas) signaling pathways. Embo J 17, 1675-1687.
Selzner, M., Morse, M. A., Vredenburgh, J. J., Meyers, W. C., and Clavien, P. A. (2000). Liver metastases from breast cancer: long-term survival after curative resection. Surgery 127, 383-389.
Sharpe, J. C., Arnoult, D., and Youle, R. J. (2004). Control of mitochondrial permeability by Bcl-2 family members. Biochim Biophys Acta 1644, 107-113.
Shi, Y. (2002). Mechanisms of caspase activation and inhibition during apoptosis. Mol Cell 9, 459-470.
Shimizu, S., Narita, M., and Tsujimoto, Y. (1999). Bcl-2 family proteins regulate the release of apoptogenic cytochrome c by the mitochondrial channel VDAC. Nature 399, 483-487.
Soldani, C., Lazze, M. C., Bottone, M. G., Tognon, G., Biggiogera, M., Pellicciari, C. E., and Scovassi, A. I. (2001). Poly(ADP-ribose) polymerase cleavage during apoptosis: when and where? Exp Cell Res 269, 193-201.
Sporny, S., and Musial, J. (2004). [Pathogenesis and morphology of hepatocellular carcinoma]. Pol Merkuriusz Lek 16, 293-297.
Sun, Z., Lu, P., Gail, M. H., Pee, D., Zhang, Q., Ming, L., Wang, J., Wu, Y., Liu, G., and Zhu, Y. (1999). Increased risk of hepatocellular carcinoma in male hepatitis B surface antigen carriers with chronic hepatitis who have detectable urinary aflatoxin metabolite M1. Hepatology 30, 379-383.
Sureda, F. X., Gabriel, C., Comas, J., Pallas, M., Escubedo, E., Camarasa, J., and Camins, A. (1999). Evaluation of free radical production, mitochondrial membrane potential and cytoplasmic calcium in mammalian neurons by flow cytometry. Brain Res Brain Res Protoc 4, 280-287.
Susin, S. A., Lorenzo, H. K., Zamzami, N., Marzo, I., Snow, B. E., Brothers, G. M., Mangion, J., Jacotot, E., Costantini, P., Loeffler, M., et al. (1999). Molecular characterization of mitochondrial apoptosis-inducing factor. Nature 397, 441-446.
Suzuki, Y., Imai, Y., Nakayama, H., Takahashi, K., Takio, K., and Takahashi, R. (2001). A serine protease, HtrA2, is released from the mitochondria and interacts with XIAP, inducing cell death. Mol Cell 8, 613-621.
Thomas, A. L., O'Byrne, K., and Steward, W. P. (2000). Chemotherapy for upper gastrointestinal tumours. Postgrad Med J 76, 321-327.
Thorgeirsson, S. S., and Grisham, J. W. (2002). Molecular pathogenesis of human hepatocellular carcinoma. Nat Genet 31, 339-346.
Thornberry, N. A., and Lazebnik, Y. (1998). Caspases: enemies within. Science 281, 1312-1316.
Tounekti, O., Belehradek, J., Jr., and Mir, L. M. (1995). Relationships between DNA fragmentation, chromatin condensation, and changes in flow cytometry profiles detected during apoptosis. Exp Cell Res 217, 506-516.
Tsujimoto, Y. (2003). Cell death regulation by the Bcl-2 protein family in the mitochondria. J Cell Physiol 195, 158-167.
Tsukuma, H., Hiyama, T., Tanaka, S., Nakao, M., Yabuuchi, T., Kitamura, T., Nakanishi, K., Fujimoto, I., Inoue, A., Yamazaki, H., and et al. (1993). Risk factors for hepatocellular carcinoma among patients with chronic liver disease. N Engl J Med 328, 1797-1801.
van Gurp, M., Festjens, N., van Loo, G., Saelens, X., and Vandenabeele, P. (2003). Mitochondrial intermembrane proteins in cell death. Biochem Biophys Res Commun 304, 487-497.
Vaux, D. L., Haecker, G., and Strasser, A. (1994). An evolutionary perspective on apoptosis. Cell 76, 777-779.
Vaux, D. L., and Korsmeyer, S. J. (1999). Cell death in development. Cell 96, 245-254.
Verhagen, A. M., Ekert, P. G., Pakusch, M., Silke, J., Connolly, L. M., Reid, G. E., Moritz, R. L., Simpson, R. J., and Vaux, D. L. (2000). Identification of DIABLO, a mammalian protein that promotes apoptosis by binding to and antagonizing IAP proteins. Cell 102, 43-53.
Verrier, F., Mignotte, B., Jan, G., and Brenner, C. (2003). Study of PTPC composition during apoptosis for identification of viral protein target. Ann N Y Acad Sci 1010, 126-142.
Villani, G., Greco, M., Papa, S., and Attardi, G. (1998). Low reserve of cytochrome c oxidase capacity in vivo in the respiratory chain of a variety of human cell types. J Biol Chem 273, 31829-31836.
Wang, K., Yin, X. M., Copeland, N. G., Gilbert, D. J., Jenkins, N. A., Keck, C. L., Zimonjic, D. B., Popescu, N. C., and Korsmeyer, S. J. (1998). BID, a proapoptotic BCL-2 family member, is localized to mouse chromosome 6 and human chromosome 22q11. Genomics 53, 235-238.
Wang, X. (2001). The expanding role of mitochondria in apoptosis. Genes Dev 15, 2922-2933.
Wei, M. C., Lindsten, T., Mootha, V. K., Weiler, S., Gross, A., Ashiya, M., Thompson, C. B., and Korsmeyer, S. J. (2000). tBID, a membrane-targeted death ligand, oligomerizes BAK to release cytochrome c. Genes Dev 14, 2060-2071.
Williams, G. T., and Smith, C. A. (1993). Molecular regulation of apoptosis: genetic controls on cell death. Cell 74, 777-779.
Wolf, B. B., Schuler, M., Echeverri, F., and Green, D. R. (1999). Caspase-3 is the primary activator of apoptotic DNA fragmentation via DNA fragmentation factor-45/inhibitor of caspase-activated DNase inactivation. J Biol Chem 274, 30651-30656.
Wolter, K. G., Hsu, Y. T., Smith, C. L., Nechushtan, A., Xi, X. G., and Youle, R. J. (1997). Movement of Bax from the cytosol to mitochondria during apoptosis. J Cell Biol 139, 1281-1292.
Wu, G., Chai, J., Suber, T. L., Wu, J. W., Du, C., Wang, X., and Shi, Y. (2000). Structural basis of IAP recognition by Smac/DIABLO. Nature 408, 1008-1012.
Wyllie, A. H., Kerr, J. F., and Currie, A. R. (1980). Cell death: the significance of apoptosis. Int Rev Cytol 68, 251-306.
Xu, C. X., and He, J. H. (1999). Anticancer effect of Howiinol A and its mechanism of action. J Asian Nat Prod Res 2, 1-19.
Yang, E., Zha, J., Jockel, J., Boise, L. H., Thompson, C. B., and Korsmeyer, S. J. (1995). Bad, a heterodimeric partner for Bcl-XL and Bcl-2, displaces Bax and promotes cell death. Cell 80, 285-291.
Yang, H. I., Lu, S. N., Liaw, Y. F., You, S. L., Sun, C. A., Wang, L. Y., Hsiao, C. K., Chen, P. J., Chen, D. S., and Chen, C. J. (2002). Hepatitis B e antigen and the risk of hepatocellular carcinoma. N Engl J Med 347, 168-174.
Yang, J., Liu, X., Bhalla, K., Kim, C. N., Ibrado, A. M., Cai, J., Peng, T. I., Jones, D. P., and Wang, X. (1997). Prevention of apoptosis by Bcl-2: release of cytochrome c from mitochondria blocked. Science 275, 1129-1132.
Yang, Y., Fang, S., Jensen, J. P., Weissman, A. M., and Ashwell, J. D. (2000). Ubiquitin protein ligase activity of IAPs and their degradation in proteasomes in response to apoptotic stimuli. Science 288, 874-877.
Yang, Y. L., and Li, X. M. (2000). The IAP family: endogenous caspase inhibitors with multiple biological activities. Cell Res 10, 169-177.
Yao, F. Y., Ferrell, L., Bass, N. M., Watson, J. J., Bacchetti, P., Venook, A., Ascher, N. L., and Roberts, J. P. (2001). Liver transplantation for hepatocellular carcinoma: expansion of the tumor size limits does not adversely impact survival. Hepatology 33, 1394-1403.
Yoon, Y. S., Kim, J. W., Kang, K. W., Kim, Y. S., Choi, K. H., and Joe, C. O. (1996). Poly(ADP-ribosyl)ation of histone H1 correlates with internucleosomal DNA fragmentation during apoptosis. J Biol Chem 271, 9129-9134.
Yu, S. W., Wang, H., Poitras, M. F., Coombs, C., Bowers, W. J., Federoff, H. J., Poirier, G. G., Dawson, T. M., and Dawson, V. L. (2002). Mediation of poly(ADP-ribose) polymerase-1-dependent cell death by apoptosis-inducing factor. Science 297, 259-263.
Yuan, J., Murrell, G. A., Trickett, A., and Wang, M. X. (2003). Involvement of cytochrome c release and caspase-3 activation in the oxidative stress-induced apoptosis in human tendon fibroblasts. Biochim Biophys Acta 1641, 35-41.
Zamzami, N., and Kroemer, G. (2001). The mitochondrion in apoptosis: how Pandora's box opens. Nat Rev Mol Cell Biol 2, 67-71.
Zhang, D. L., Liu, S. G., Yan, L. F., Li, L. J., Huang, G. S., Fang, F. D., Xia, G. T., He, X. Y., Gao, B. X., Bai, X. H., et al. (2001a). Carcinogenesis or tumourigenicity testing of animal cell lines for vaccine preparation by colony formation on soft agar and by agglutination under plant lectins. Cell Biol Int 25, 997-1002.
Zhang, X. D., Zhang, X. Y., Gray, C. P., Nguyen, T., and Hersey, P. (2001b). Tumor necrosis factor-related apoptosis-inducing ligand-induced apoptosis of human melanoma is regulated by smac/DIABLO release from mitochondria. Cancer Res 61, 7339-7348.
Zimmermann, K. C., Bonzon, C., and Green, D. R. (2001). The machinery of programmed cell death. Pharmacol Ther 92, 57-70.
Zou, H., Henzel, W. J., Liu, X., Lutschg, A., and Wang, X. (1997). Apaf-1, a human protein homologous to C. elegans CED-4, participates in cytochrome c-dependent activation of caspase-3. Cell 90, 405-413.
Zou, H., Li, Y., Liu, X., and Wang, X. (1999). An APAF-1.cytochrome c multimeric complex is a functional apoptosome that activates procaspase-9. J Biol Chem 274, 11549-11556.