Caspase-3蛋白在重离子辐照肺癌细胞中的功能研究
|
摘要
原发性肺癌是死亡率较高的恶性肿瘤之一,且发病率、死亡率有逐年升高趋势。放疗已成其治疗的主要手段,约85%的肺癌患者需要接受放射治疗。研究表明,电离辐照可通过多种信号传导途径诱导细胞凋亡,达到治疗目的。Caspase-3是Caspase家族中最重要的蛋白酶,是多种凋亡途径的共同下游效应部分,是细胞凋亡蛋白酶级联反应的必经之路。本实验通过研究12C6+射线辐照人肺癌细胞株H1299,观察12C6+于不同剂量、不同时间辐照细胞周期和细胞凋亡的动态变化及检测Caspase-3、Caspase-9、Bcl-2、Bax蛋白等的表达,探讨重离子辐照治疗肺癌的理论依据。实验结果如下：
1重离子辐照可抑制H1299细胞的增殖
流式细胞仪检测了细胞周期的分布情况：随着12C6+辐照剂量的加大,以G2/M期细胞阻滞为主,阻滞程度增加呈剂量依赖性,当12C6+辐照剂量超过6GyE时,G]前期细胞明显增加,出现细胞死亡。重离子束辐照能够有效地抑制肿瘤细胞复制,并促使肿瘤细胞死亡。
采用噻唑蓝比色法观察重离子束辐照H1299细胞生长的增殖能力,结果显示随着重离子束剂量增加及培养时间的延长其细胞增殖率逐渐下降,其生长抑制曲线呈剂量-时间相关性。
2重离子辐照对H1299细胞凋亡的影响
荧光显微镜下观察细胞形态显示,经12C6+辐照后培养24h,可见多数细胞形态变的不规则,细胞核出现碎裂,核固缩及凋亡小体。随着辐照剂量增加,细胞凋亡率增加。
流式细胞技术检测也显示了H1299细胞在重离子辐照下,随着剂量的增加细胞凋亡率也明显增加,揭示了重离子可体外诱导H1299细胞凋亡。
3重离子辐照与相关凋亡基因的表达
Caspase-3活性分析试剂盒检测显示,H1299细胞在重离子辐照后,Caspase-3蛋白酶活性随辐照剂量的增加而增加,揭示重离子辐照可提高Caspase-3蛋白酶活性,且与细胞凋亡率变化呈正相关。
Western-blotting法和RT-PCR检测实验结果显示,Caspase-3的基因条带随着剂量加大逐渐加深,表明12C6+辐照可匕调Caspase-3表达,并呈一定剂量依赖；免疫组化检测结果显示了在重离子辐照下,随着辐照剂量的增加,Caspase-3、Caspase-9和Bax蛋白表达均有增加,而Bcl-2蛋白表达呈下降趋势,Bcl-2/Bax蛋白比值下降；表明重离子辐照可激活Caspase-3的活性。而Bax的表达上调在一定程度上抑制了Bcl-2蛋白的功能,Caspase-3、Caspase-9蛋白共同参与了细胞凋亡,提示重离子辐照诱导H1299细胞凋亡可能是通过Caspase-3途径调控的。
4Caspase-3活性抑制实验：
当H1299细胞经Caspase-3抑制剂预处理后,观察培养24h的各组细胞活性和细胞凋亡率显示：Caspase-3抑制剂(Z-DEVD-fmk)能特异性阻断Caspase-3活性,同时降低细胞凋亡发生。
本研究从以上实验中得到下列结论：重离子辐照能够体外抑制H1299细胞增殖。可能是通过激活肺癌细胞H1299内Caspase-3而诱导P53非依赖性细胞凋亡,Caspase-3蛋白表达与辐照敏感性成正比。Caspase-3、Caspase-9蛋白共同参与了细胞凋亡。因此,重离子可能是通过激活H1299细胞内Caspase-3蛋白酶活性并降低Bcl-2/Bax比值而诱导细胞凋亡,Caspase-3蛋白参与了调控肺癌细胞H1299的细胞周期,提示重离子辐照诱导H1299细胞凋亡可能是通过Caspase-3途径调控的。通过调控Caspase-3基因的表达可能改变肿瘤细胞的放射敏感性,这有望为提高肿瘤放射治疗提供新思路与新方法。
The lung cancer is one of the highest mortality of malignant tumors, its morbidity and mortality tend to increase year by year. Radiotherapy has become the principal means of its treatment. About85%of lung cancer patients need to receive radiation therapy at some stage in their course of the disease. Studies have shown that ionizing radiation can induce apoptosis through a variety of signal transduction pathway for therapeutic purposes. Caspase-3is the most important protease in Caspase family. It is the common downriver effective parts of multi-apoptosis pathways and the only way to apoptotic enzyme reaction. We have studied12C6+-ray irradiation of human lung cell line H1299. Dynamic changes of the cell cycle and apoptosis were observed, and the expression caspase-3, Caspase-9, Bcl-2and Bax proteins of the cells irradiated by12C6+at different doses and times was detected. This study is to explore the theoretical mechanism and apoptosis pathway of heavy ion radiation therapy of lung cancer.
Several important experimental results were acquired as summarized as below:
1Heavy ion beam could depress the cancerous cell replication
FCM was used to check the cell cycle distributions of H1299cells after heavy ion beam irradiations. As the dose of the heavy ion radiation increased, the G2/M cells were increased obviously; the level of retardarce was dose-dependent. As the dose was over6GyE, the G1cells were increased and the cells become to die. That shows the heavy ion beam could depress the cancerous cell replication and induce cell death.
Assay OD value of the hole with MTT. The result shows that the inhibition is time-and dose-dependent. In the same time, as the dose of12C6+increased, the cell survival rate decreases
2Heavy ion beam could induce H1299cells undergone apoptosis
The cellular form through the fluorescence microscope after radiated for24h was observed. It clearly shown cell nuclear morphology changes such as membrane blebbing and formation of apoptotic body which are characteristic of apoptosis. Apoptosis induction increased is time and dose dependent manner.
FCM showed that the rate of apoptosis of H1299radiated by heavy ion beam was increased as the dose increased. That indicates the heavy ion can induce the apoptotic of H1299in vitro.
3Heavy ion irradiation and related apoptosis gene expression
Through the Caspase-3activeness analytical reagent box examination, with the dose of irradiation increasing, Caspase-3activity was significantly increased, and the dose-dependen. It clued on that enzyme activity of Caspase-3of H1299would be higher after being irradiated by12C6+and the change of apoptosis rate was positive correlation.
The expression of protein Caspase-3was detected with Western Blotting and RT-PCR in irradiated cell H1299. The results shown that:irradiation of12C6+can increase protein expression of Caspase-3, and there is dose-depended.
The immunohistochemistry test result demonstrates that the expression of gene Caspase-3,Caspase-9and Bax was increased with the increase of radial dose, but Bcl-2's was decreased, Bcl-2/Bax decreased as well. That indicates:the heavy ion radiation can activate Caspase-3. The Bax massive expressions have suppressed Bcl-2function to a certain extent. It suggests heavy ion radiation induces the H1299cell apoptosis possibly is weakly through the caspase-3way regulation.
4Caspase-3activity inhibition assay
When H1299cells were pretreated with Caspase-3inhibitor and cells were observed activity and apoptosis rate displayed after24h:Caspase-3inhibitor (Z-DEVD-fmk) that specifically blocks the activation of Caspase-3, while reducing cell apoptosis.
This research tests from above may obtain the following conclusion:The heavy ion radiation can suppress H1299cell multiplication in vitro. Radiation-inducing can lead H1299to apoptosis by the non-P53-dependent pathway. The expression of protein Caspase-3and the radio sensitivity are proportional. The heavy ion possibly is through activates Caspase-3in the H1299and reduces the Bcl-2/Bax ratio to induce the apoptosis. The protein Caspase-3participates in the regulation mitotic cycle, gives play to the anti-tumor. By regulating the expression of Caspase-3gene may change the radio sensitivity of tumor cells, which is expected to increase tumor radiation therapy to provide new ideas and new methods.
1重离子辐照可抑制H1299细胞的增殖
流式细胞仪检测了细胞周期的分布情况：随着12C6+辐照剂量的加大,以G2/M期细胞阻滞为主,阻滞程度增加呈剂量依赖性,当12C6+辐照剂量超过6GyE时,G]前期细胞明显增加,出现细胞死亡。重离子束辐照能够有效地抑制肿瘤细胞复制,并促使肿瘤细胞死亡。
采用噻唑蓝比色法观察重离子束辐照H1299细胞生长的增殖能力,结果显示随着重离子束剂量增加及培养时间的延长其细胞增殖率逐渐下降,其生长抑制曲线呈剂量-时间相关性。
2重离子辐照对H1299细胞凋亡的影响
荧光显微镜下观察细胞形态显示,经12C6+辐照后培养24h,可见多数细胞形态变的不规则,细胞核出现碎裂,核固缩及凋亡小体。随着辐照剂量增加,细胞凋亡率增加。
流式细胞技术检测也显示了H1299细胞在重离子辐照下,随着剂量的增加细胞凋亡率也明显增加,揭示了重离子可体外诱导H1299细胞凋亡。
3重离子辐照与相关凋亡基因的表达
Caspase-3活性分析试剂盒检测显示,H1299细胞在重离子辐照后,Caspase-3蛋白酶活性随辐照剂量的增加而增加,揭示重离子辐照可提高Caspase-3蛋白酶活性,且与细胞凋亡率变化呈正相关。
Western-blotting法和RT-PCR检测实验结果显示,Caspase-3的基因条带随着剂量加大逐渐加深,表明12C6+辐照可匕调Caspase-3表达,并呈一定剂量依赖；免疫组化检测结果显示了在重离子辐照下,随着辐照剂量的增加,Caspase-3、Caspase-9和Bax蛋白表达均有增加,而Bcl-2蛋白表达呈下降趋势,Bcl-2/Bax蛋白比值下降；表明重离子辐照可激活Caspase-3的活性。而Bax的表达上调在一定程度上抑制了Bcl-2蛋白的功能,Caspase-3、Caspase-9蛋白共同参与了细胞凋亡,提示重离子辐照诱导H1299细胞凋亡可能是通过Caspase-3途径调控的。
4Caspase-3活性抑制实验：
当H1299细胞经Caspase-3抑制剂预处理后,观察培养24h的各组细胞活性和细胞凋亡率显示：Caspase-3抑制剂(Z-DEVD-fmk)能特异性阻断Caspase-3活性,同时降低细胞凋亡发生。
本研究从以上实验中得到下列结论：重离子辐照能够体外抑制H1299细胞增殖。可能是通过激活肺癌细胞H1299内Caspase-3而诱导P53非依赖性细胞凋亡,Caspase-3蛋白表达与辐照敏感性成正比。Caspase-3、Caspase-9蛋白共同参与了细胞凋亡。因此,重离子可能是通过激活H1299细胞内Caspase-3蛋白酶活性并降低Bcl-2/Bax比值而诱导细胞凋亡,Caspase-3蛋白参与了调控肺癌细胞H1299的细胞周期,提示重离子辐照诱导H1299细胞凋亡可能是通过Caspase-3途径调控的。通过调控Caspase-3基因的表达可能改变肿瘤细胞的放射敏感性,这有望为提高肿瘤放射治疗提供新思路与新方法。
The lung cancer is one of the highest mortality of malignant tumors, its morbidity and mortality tend to increase year by year. Radiotherapy has become the principal means of its treatment. About85%of lung cancer patients need to receive radiation therapy at some stage in their course of the disease. Studies have shown that ionizing radiation can induce apoptosis through a variety of signal transduction pathway for therapeutic purposes. Caspase-3is the most important protease in Caspase family. It is the common downriver effective parts of multi-apoptosis pathways and the only way to apoptotic enzyme reaction. We have studied12C6+-ray irradiation of human lung cell line H1299. Dynamic changes of the cell cycle and apoptosis were observed, and the expression caspase-3, Caspase-9, Bcl-2and Bax proteins of the cells irradiated by12C6+at different doses and times was detected. This study is to explore the theoretical mechanism and apoptosis pathway of heavy ion radiation therapy of lung cancer.
Several important experimental results were acquired as summarized as below:
1Heavy ion beam could depress the cancerous cell replication
FCM was used to check the cell cycle distributions of H1299cells after heavy ion beam irradiations. As the dose of the heavy ion radiation increased, the G2/M cells were increased obviously; the level of retardarce was dose-dependent. As the dose was over6GyE, the G1cells were increased and the cells become to die. That shows the heavy ion beam could depress the cancerous cell replication and induce cell death.
Assay OD value of the hole with MTT. The result shows that the inhibition is time-and dose-dependent. In the same time, as the dose of12C6+increased, the cell survival rate decreases
2Heavy ion beam could induce H1299cells undergone apoptosis
The cellular form through the fluorescence microscope after radiated for24h was observed. It clearly shown cell nuclear morphology changes such as membrane blebbing and formation of apoptotic body which are characteristic of apoptosis. Apoptosis induction increased is time and dose dependent manner.
FCM showed that the rate of apoptosis of H1299radiated by heavy ion beam was increased as the dose increased. That indicates the heavy ion can induce the apoptotic of H1299in vitro.
3Heavy ion irradiation and related apoptosis gene expression
Through the Caspase-3activeness analytical reagent box examination, with the dose of irradiation increasing, Caspase-3activity was significantly increased, and the dose-dependen. It clued on that enzyme activity of Caspase-3of H1299would be higher after being irradiated by12C6+and the change of apoptosis rate was positive correlation.
The expression of protein Caspase-3was detected with Western Blotting and RT-PCR in irradiated cell H1299. The results shown that:irradiation of12C6+can increase protein expression of Caspase-3, and there is dose-depended.
The immunohistochemistry test result demonstrates that the expression of gene Caspase-3,Caspase-9and Bax was increased with the increase of radial dose, but Bcl-2's was decreased, Bcl-2/Bax decreased as well. That indicates:the heavy ion radiation can activate Caspase-3. The Bax massive expressions have suppressed Bcl-2function to a certain extent. It suggests heavy ion radiation induces the H1299cell apoptosis possibly is weakly through the caspase-3way regulation.
4Caspase-3activity inhibition assay
When H1299cells were pretreated with Caspase-3inhibitor and cells were observed activity and apoptosis rate displayed after24h:Caspase-3inhibitor (Z-DEVD-fmk) that specifically blocks the activation of Caspase-3, while reducing cell apoptosis.
This research tests from above may obtain the following conclusion:The heavy ion radiation can suppress H1299cell multiplication in vitro. Radiation-inducing can lead H1299to apoptosis by the non-P53-dependent pathway. The expression of protein Caspase-3and the radio sensitivity are proportional. The heavy ion possibly is through activates Caspase-3in the H1299and reduces the Bcl-2/Bax ratio to induce the apoptosis. The protein Caspase-3participates in the regulation mitotic cycle, gives play to the anti-tumor. By regulating the expression of Caspase-3gene may change the radio sensitivity of tumor cells, which is expected to increase tumor radiation therapy to provide new ideas and new methods.
引文
Ahamed M, Akhtar MJ, Raja M, Ahmad I, Siddiqui MK, AlSalhi MS, Alrokayan SA (2011) ZnO nanorod-induced apoptosis in human alveolar adenocarcinoma cells via p53, survivin and bax/bcl-2 pathways:role of oxidative stress[J]. Nanomedicine 7:904-913.
Alimonti JB, Ball TB, Fowke KR (2003) Mechanisms of CD4+T lymphocyte cell death in human immunodeficiency virus infection and AIDS[J]. J Gen Virol 84:1649-1661.
Avdeeva Zh I, Akol'zina SE, Alpatova NA, Medunitsyn NV (2005) [Effects of cytokines on the immunogenic properties of hepatitis A vaccine][J]. Vopr Virusol 50:23-27.
Bajt ML, Cover C, Lemasters JJ, Jaeschke H (2006) Nuclear translocation of endonuclease G and apoptosis-inducing factor during acetaminophen-induced liver cell injury[J]. Toxicol Sci 94:217-225.
Bassler N, Jakel O, Sondergaard CS, Petersen JB (2010) Dose-and LET-painting with particle therapy[J]. Acta Oncol 49:1170-1176.
Bechmann LP, Marquitan G, Jochum C, Saner F, Gerken G, Canbay A (2008) Apoptosis versus necrosis rate as a predictor in acute liver failure following acetaminophen intoxication compared with acute-on-chronic liver failure[J]. Liver Int 28:713-716.
Borner C (2003) The Bcl-2 protein family:sensors and checkpoints for life-or-death decisions[J]. Mol Immunol 39:615-647.
Budihardjo I, Oliver H, Lutter M, Luo X, Wang X (1999) Biochemical pathways of caspase activation during apoptosis[J]. Annu Rev Cell Dev Biol 15:269-290.
Canbay A, Higuchi H, Bronk SF, Taniai M, Sebo TJ, Gores GJ (2002) Fas enhances fibrogenesis in the bile duct ligated mouse:a link between apoptosis and fibrosis[J]. Gastroenterology 123:1323-1330.
Canman CE, Kastan MB (1997) Role of p53 in apoptosis[J]. Adv Pharmacol 41:429-460.
Castro MA, Schwartsmann G, Bernard EA, Moreira JC (1999) Phenotype modulation of cellular UV-sensitivity[J]. Cancer Lett 145:65-72.
Chen GG, Liang NC, Lee JF, Chan UP, Wang SH, Leung BC, Leung KL (2004) Over-expression of Bcl-2 against Pteris semipinnata L-induced apoptosis of human colon cancer cells via a NF-kappa B-related pathway[J]. Apoptosis 9:619-627.
Chen YC, Shen SC, Tsai SH (2005) Prostaglandin D(2) and J(2) induce apoptosis in human leukemia cells via activation of the caspase 3 cascade and production of reactive oxygen species[J]. Biochim Biophys Acta 1743:291-304.
Chen YH, Zhang YH, Zhang HJ, Liu DZ, Gu M, Li JY, Wu F, Zhu XZ, Li J, Nan FJ (2006) Design, synthesis, and biological evaluation of isoquinoline-1,3,4-trione derivatives as potent caspase-3 inhibitors[J]. J Med Chem 49:1613-1623.
Chintharlapalli SR, Jasti M, Malladi S, Parsa KV, Ballestero RP, Gonzalez-Garcia M (2005) BMRP is a Bcl-2 binding protein that induces apoptosis[J]. J Cell Biochem 94:611-626.
Choi KM, Zhong Y, Hoit BD, Grupp IL, Hahn H, Dilly KW, Guatimosim S, Lederer WJ, Matlib MA (2002) Defective intracellular Ca(2+) signaling contributes to cardiomyopathy in Type 1 diabetic rats[J]. Am J Physiol Heart Circ Physiol 283:H1398-1408.
Choy JC, Granville DJ, Hunt DW, McManus BM (2001) Endothelial cell apoptosis:biochemical characteristics and potential implications for atherosclerosis[J]. J Mol Cell Cardiol 33:1673-1690.
Chugh R, Dunn R, Zalupski MM, Biermann JS, Sondak VK, Mace JR, Leu KM, Chandler WF, Baker LH (2005) Phase Ⅱ study of 9-nitro-camptothecin in patients with advanced chordoma or soft tissue sarcoma[J]. J Clin Oncol 23:3597-3604.
Combs SE, Ellerbrock M, Haberer T, Habermehl D, Hoess A, Jakel O, Jensen A, Klemm S, Munter M, Naumann J, Nikoghosyan A, Oertel S, Parodi K, Rieken S, Debus J (2010) Heidelberg Ion Therapy Center (HIT):Initial clinical experience in the first 80 patients[J]. Acta Oncol 49:1132-1140.
Coultas L, Strasser A (2003) The role of the Bcl-2 protein family in cancer[J]. Semin Cancer Biol 13:115-123.
Dubray B, Breton C, Delic J, Klijanienko J, Maciorowski Z, Vielh P, Fourquet A, Dumont J, Magdelenat H, Cosset JM (1997) In vitro radiation-induced apoptosis and tumour response to radiotherapy:a prospective study in patients with non-Hodgkin lymphomas treated by low-dose irradiation[J]. Int J Radiat Biol 72:759-760.
Ferri CP, Prince M, Brayne C, Brodaty H, Fratiglioni L, Ganguli M, Hall K, Hasegawa K, Hendrie H, Huang Y, Jorm A, Mathers C, Menezes PR, Rimmer E, Scazufca M (2005) Global prevalence of dementia:a Delphi consensus study[J]. Lancet 366:2112-2117.
Fukumura A, Tsujii H, Kamada T, Baba M, Tsuji H, Kato H, Kato S, Yamada S, Yasuda S, Yanagi T, Hara R, Yamamoto N, Mizoe J, Akahane K, Fukuda S, Furusawa Y, Iwata Y, Kanai T, Kanematsu N, Kitagawa A, Matsufuji N, Minohara S, Miyahara N, Mizuno H, Murakami T, Nishizawa K, Noda K, Takada E, Yonai S (2009) Carbon-ion radiotherapy:clinical aspects and related dosimetry[J]. Radiat Prot Dosimetry 137:149-155.
Furusawa Y, Aoki M, Durante M (2002) Simultaneous exposure of mammalian cells to heavy ions and X-rays[J]. Adv Space Res 30:877-884.
Fushimi K, Uzawa K, Ishigami T, Yamamoto N, Kawata T, Shibahara T, Ito H, Mizoe JE, Tsujii H, Tanzawa H (2008) Susceptible genes and molecular pathways related to heavy ion irradiation in oral squamous cell carcinoma cells[J]. Radiother Oncol 89:237-244.
Giannopoulou I, Nakopoulou L, Zervas A, Lazaris AC, Stravodimos C, Giannopoulos A, Davaris PS (2002) Immunohistochemical study of pro-apoptotic factors Bax, Fas and CPP32 in urinary bladder cancer:prognostic implications[J]. Urol Res 30:342-345.
Gross A, McDonnell JM, Korsmeyer SJ (1999) BCL-2 family members and the mitochondria in apoptosis[J]. Genes Dev 13:1899-1911.
Gueven N, Becherel OJ, Birrell G, Chen P, DelSal G, Carney JP, Grattan-Smith P, Lavin MF (2006) Defective p53 response and apoptosis associated with an ataxia-telangiectasia-like phenotype[J]. Cancer Res 66:2907-2912.
Haddad JJ (2004) On the antioxidant mechanisms of Bcl-2:a retrospective of NF-kappaB signaling and oxidative stress[J]. Biochem Biophys Res Commun 322:355-363.
Hengartner MO (2001) Apoptosis:corralling the corpses[J]. Cell 104:325-328.
Hsia JY, Chen CY, Chen JT, Hsu CP, Shai SE, Yang SS, Chuang CY, Wang PY, Miaw J (2003) Prognostic significance of caspase-3 expression in primary resected esophageal squamous cell carcinoma[J]. Eur J Surg Oncol 29:44-48.
Imai R, Kamada T, Tsuji H, Sugawara S, Serizawa I, Tsujii H, Tatezaki S (2010) Effect of carbon ion radiotherapy for sacral chordoma:results of Phase Ⅰ-Ⅱ and Phase Ⅱ clinical trials[J]. Int J Radiat Oncol Biol Phys 77:1470-1476.
Imai R, Kamada T, Tsuji H, Yanagi T, Baba M, Miyamoto T, Kato S, Kandatsu S, Mizoe JE, Tsujii H, Tatezaki S (2004) Carbon ion radiotherapy for unresectable sacral chordomas[J]. Clin Cancer Res 10:5741-5746.
Isobe N, Onodera H, Mori A, Shimada Y, Yang W, Yasuda S, Fujimoto A, Ooe H, Arii S, Kitaichi M, Imamura M (2004) Caspase-3 expression in human gastric carcinoma and its clinical significance[J]. Oncology 66:201-209.
Johnson R, Staiano-Coico L, Austin L, Cardinale I, Nabeya-Tsukifuji R, Krueger JG (1996) PUVA treatment selectively induces a cell cycle block and subsequent apoptosis in human T-lymphocytes[J]. Photochem Photobiol 63:566-571.
Jonges LE, Nagelkerke JF, Ensink NG, van der Velde EA, Tollenaar RA, Fleuren GJ, van de Velde CJ, Morreau H, Kuppen PJ (2001) Caspase-3 activity as a prognostic factor in colorectal carcinoma[J]. Lab Invest 81:681-688.
Kanai T, Endo M, Minohara S, Miyahara N, Koyama-ito H, Tomura H, Matsufuji N, Futami Y, Fukumura A, Hiraoka T, Furusawa Y, Ando K, Suzuki M, Soga F, Kawachi K (1999) Biophysical characteristics of HIMAC clinical irradiation system for heavy-ion radiation therapy[J]. Int J Radiat Oncol Biol Phys 44:201-210.
Kanai T, Furusawa Y, Fukutsu K, Itsukaichi H, Eguchi-Kasai K, Ohara H (1997) Irradiation of mixed beam and design of spread-out Bragg peak for heavy-ion radiotherapy[J]. Radiat Res 147:78-85.
Kanai T, Matsufuji N, Miyamoto T, Mizoe J, Kamada T, Tsuji H, Kato H, Baba M, Tsujii H (2006) Examination of GyE system for HIMAC carbon therapy [J]. Int J Radiat Oncol Biol Phys 64:650-656.
Kane DJ, Sarafian TA, Anton R, Hahn H, Gralla EB, Valentine JS, Ord T, Bredesen DE (1993) Bcl-2 inhibition of neural death:decreased generation of reactive oxygen species[J]. Science 262:1274-1277.
Katsuda K, Kataoka M, Uno F, Murakami T, Kondo T, Roth JA, Tanaka N, Fujiwara T (2002) Activation of caspase-3 and cleavage of Rb are associated with p16-mediated apoptosis in human non-small cell lung cancer cells[J]. Oncogene 21:2108-2113.
Kim S, Ko H, Park JE, Jung S, Lee SK, Chun YJ (2002) Design, synthesis, and discovery of novel trans-stilbene analogues as potent and selective human cytochrome P450 1B1 inhibitors[J]. J Med Chem 45:160-164.
Kolenko V, Bloom T, Rayman P, Bukowski R, Hsi E, Finke J (1999) Inhibition of NF-kappa B activity in human T lymphocytes induces caspase-dependent apoptosis without detectable activation of caspase-1 and -3[J]. J Immunol 163:590-598.
Kondo S, Barna BP, Kondo Y, Tanaka Y, Casey G, Liu J, Morimura T, Kaakaji R, Peterson JW, Werbel B, Barnett GH (1996) WAF1/CIP1 increases the susceptibility of p53 non-functional malignant glioma cells to cisplatin-induced apoptosis[J]. Oncogene 13:1279-1285.
Korsmeyer SJ (1992) Bcl-2 initiates a new category of oncogenes:regulators of cell death[J]. Blood 80:879-886.
Kraft G (1990) The radiobiological and physical basis for radiotherapy with protons and heavier ions[J]. Strahlenther Onkol 166:10-13.
Kroemer G (1997) The proto-oncogene Bcl-2 and its role in regulating apoptosis[J]. Nat Med 3:614-620.
Kubota N, Suzuki M, Furusawa Y, Ando K, Koike S, Kanai T, Yatagai F, Ohmura M, Tatsuzaki H, Matsubara S, et al. (1995) A comparison of biological effects of modulated carbon-ions and fast neutrons in human osteosarcoma cells[J]. Int J Radiat Oncol Biol Phys 33:135-141.
Li P, Nijhawan D, Budihardjo I, Srinivasula SM, Ahmad M, Alnemri ES, Wang X (1997) Cytochrome c and dATP-dependent formation of Apaf-1/caspase-9 complex initiates an apoptotic protease cascade[J]. Cell 91:479-489.
Li X, Marani M, Yu J, Nan B, Roth JA, Kagawa S, Fang B, Denner L, Marcelli M (2001) Adenovirus-mediated Bax overexpression for the induction of therapeutic apoptosis in prostate cancer[J]. Cancer Res 61:186-191.
Loop T, Dovi-Akue D, Frick M, Roesslein M, Egger L, Humar M, Hoetzel A, Schmidt R, Borner C, Pahl HL, Geiger KK, Pannen BH (2005) Volatile anesthetics induce caspase-dependent, mitochondria-mediated apoptosis in human T lymphocytes in vitro[J]. Anesthesiology 102:1147-1157.
Lunkes A, Trottier Y, Mandel JL (1998) Pathological mechanisms in Huntington's disease and other polyglutamine expansion diseases[J]. Essays Biochem 33:149-163.
Mahieux R, Pise-Masison C, Gessain A, Brady JN, Olivier R, Perret E, Misteli T, Nicot C (2001) Arsenic trioxide induces apoptosis in human T-cell leukemia virus type 1- and type 2-infected cells by a caspase-3-dependent mechanism involving Bcl-2 cleavage[J]. Blood 98:3762-3769.
Marcelli M, Cunningham GR, Walkup M, He Z, Sturgis L, Kagan C, Mannucci R, Nicoletti I, Teng B, Denner L (1999) Signaling pathway activated during apoptosis of the prostate cancer cell line LNCaP:overexpression of caspase-7 as a new gene therapy strategy for prostate cancer[J]. Cancer Res 59:382-390.
Masunaga S, Ando K, Uzawa A, Hirayama R, Furusawa Y, Koike S, Sakurai Y, Nagata K, Suzuki M, Kashino G, Kinashi Y, Tanaka H, Maruhashi A, Ono K (2008) Radiobiologic significance of response of intratumor quiescent cells in vivo to accelerated carbon ion beams compared with gamma-rays and reactor neutron beams[J].Int J Radiat Oncol Biol Phys 70:221-228.
Miyazaki H, Kuwano K, Yoshida K, Maeyama T, Yoshimi M, Fujita M, Hagimoto N, Yoshida R, Nakanishi Y (2004) The perforin mediated apoptotic pathway in lung injury and fibrosis[J]. J Clin Pathol 57:1292-1298.
Mizoe JE, Tsujii H, Hasegawa A, Yanagi T, Takagi R, Kamada T, Tsuji H, Takakura K (2007) Phase I/II clinical trial of carbon ion radiotherapy for malignant gliomas:combined X-ray radiotherapy, chemotherapy, and carbon ion radiotherapy[J]. Int J Radiat Oncol Biol Phys 69:390-396.
Moll UM, Zaika A (2001) Nuclear and mitochondrial apoptotic pathways of p53[J]. FEBS Lett 493:65-69.
Mori S, Yanagi T, Hara R, Sharp GC, Asakura H, Kumagai M, Kishimoto R, Yamada S, Kato H, Kandatsu S, Kamada T (2010) Comparison of respiratory-gated and respiratory-ungated planning in scattered carbon ion beam treatment of the pancreas using four-dimensional computed tomography [J]. Int J Radiat Oncol Biol Phys 76:303-312.
Muchmore SW, Sattler M, Liang H, Meadows RP, Harlan JE, Yoon HS, Nettesheim D, Chang BS, Thompson CB, Wong SL, Ng SL, Fesik SW (1996) X-ray and NMR structure of human Bcl-xL, an inhibitor of programmed cell death[J]. Nature 381:335-341.
Naka T, Boitze C, Kuester D, Schulz TO, Schneider-Stock R, Kellner A, Samii A, Herold C, Ostertag H, Roessner A (2005) Alterations of Gl-S checkpoint in chordoma:the prognostic impact of p53 overexpression[J]. Cancer 104:1255-1263.
Neumar RW, Xu YA, Gada H, Guttmann RP, Siman R (2003) Cross-talk between calpain and caspase proteolytic systems during neuronal apoptosis[J]. J Biol Chem 278:14162-14167.
Norberg E, Gogvadze V, Ott M, Horn M, Uhlen P, Orrenius S, Zhivotovsky B (2008) An increase in intracellular Ca2+ is required for the activation of mitochondrial calpain to release AIF during cell death[J]. Cell Death Differ 15:1857-1864.
Nordsmark M, Bentzen SM, Rudat V, Brizel D, Lartigau E, Stadler P, Becker A, Adam M, Molls M, Dunst J, Terris DJ, Overgaard J (2005) Prognostic value of tumor oxygenation in 397 head and neck tumors after primary radiation therapy. An international multi-center study[J]. Radiother Oncol 77:18-24.
Ogata M, Inanami O, Nakajima M, Nakajima T, Hiraoka W, Kuwabara M (2003) Ca(2+)-dependent and caspase-3-independent apoptosis caused by damage in Golgi apparatus due to 2,4,5,7-tetrabromorhodamine 123 bromide-induced photodynamic effects[J]. Photochem Photobiol 78:241-247.
Olive PL, Durand RE (1997) Apoptosis:an indicator of radiosensitivity in vitro?[J]. Int J Radiat Biol 71:695-707.
Pan G, Li J, Zhou Y, Zheng H, Pei D (2006) A negative feedback loop of transcription factors that controls stem cell pluripotency and self-renewal[J]. FASEB J 20:1730-1732.
Penning LC, Rasch MH, Ben-Hur E, Dubbelman TM, Havelaar AC, Van der Zee J, Van Steveninck J (1992) A role for the transient increase of cytoplasmic free calcium in cell rescue after photodynamic treatment[J]. Biochim Biophys Acta 1107:255-260.
Puolakkainen PA, Brekken RA, Muneer S, Sage EH (2004) Enhanced growth of pancreatic tumors in SPARC-null mice is associated with decreased deposition of extracellular matrix and reduced tumor cell apoptosis[J]. Mol Cancer Res 2:215-224.
Putt KS, Chen GW, Pearson JM, Sandhorst JS, Hoagland MS, Kwon JT, Hwang SK, Jin H, Churchwell MI, Cho MH, Doerge DR, Helferich WG, Hergenrother PJ (2006) Small-molecule activation of procaspase-3 to caspase-3 as a personalized anticancer strategy[J]. Nat Chem Biol 2:543-550.
Reed JC (1998) Bcl-2 family proteins[J]. Oncogene 17:3225-3236.
Reya T, Morrison SJ, Clarke MF, Weissman IL (2001) Stem cells, cancer, and cancer stem cells[J]. Nature 414:105-111.
Saikumar P, Dong Z, Mikhailov V, Denton M, Weinberg JM, Venkatachalam MA (1999) Apoptosis: definition, mechanisms, and relevance to disease[J]. Am J Med 107:489-506.
Samis JA, Back DW, Graham EJ, DeLuca CI, Elce JS (1991) Constitutive expression of calpain Ⅱ in the rat uterus during pregnancy and involution[J]. Biochem J 276 (Pt 2):293-299.
Scheil S, Bruderlein S, Liehr T, Starke H, Herms J, Schulte M, Moller P (2001) Genome-wide analysis of sixteen chordomas by comparative genomic hybridization and cytogenetics of the first human chordoma cell line, U-CH1 [J]. Genes Chromosomes Cancer 32:203-211.
Shinoura N, Muramatsu Y, Yoshida Y, Asai A, Kirino T, Hamada H (2000) Adenovirus-mediated transfer of caspase-3 with Fas ligand induces drastic apoptosis in U-373MG glioma cellsfJ]. Exp Cell Res 256:423-433.
Skarsgard LD (1998) Radiobiology with heavy charged particles:a historical review[J]. Phys Med 14 Suppl 1:1-19.
Sliskovic I, Mutus B (2006) Reversible inhibition of caspase-3 activity by iron(Ⅲ):potential role in physiological control of apoptosis[J]. FEBS Lett 580:2233-2237.
Spira A, Ettinger DS (2004) Multidisciplinary management of lung cancer[J]. N Engl J Med 350:379-392.
Stone HB, Dewey WC, Wallace SS, Coleman CN (1998) Molecular Biology to Radiation Oncology:A Model for Translational Research? Opportunities in basic and translational research. From a workshop sponsored by the National Cancer Institute, Radiation Research Program, January 26-28,1997, Bethesda, Maryland[J]. Radiat Res 150:134-147.
Suzuki A, Ito T, Kawano H, Hayashida M, Hayasaki Y, Tsutomi Y, Akahane K, Nakano T, Miura M, Shiraki K (2000) Survivin initiates procaspase 3/p21 complex formation as a result of interaction with Cdk4 to resist Fas-mediated cell death[J]. Oncogene 19:1346-1353.
Suzuki Y, Oka K, Ohno T, Kato S, Tsujii H, Nakano T (2009) Prognostic impact of mitotic index of proliferating cell populations in cervical cancer patients treated with carbon ion beam[J]. Cancer 115:1875-1882.
Tang MJ, Tai IT (2007) A novel interaction between procaspase 8 and SPARC enhances apoptosis and potentiates chemotherapy sensitivity in colorectal cancers[J]. J Biol Chem 282:34457-34467.
Tatsukawa H, Sano T, Fukaya Y, Ishibashi N, Watanabe M, Okuno M, Moriwaki H, Kojima S (2011) Dual induction of caspase 3- and transglutaminase-dependent apoptosis by acyclic retinoid in hepatocellular carcinoma cells[J]. Mol Cancer 10:4.
Thibaut S, Bourre L, Hernot D, Rousset N, Lajat Y, Patrice T (2002) Effects of BAPTA-AM, Forskolin, DSF and Z.VAD.fmk on PDT-induced apoptosis and m-THPC phototoxicity on B16 cells[J]. Apoptosis 7:99-106.
Thompson CB (1995) Apoptosis in the pathogenesis and treatment of disease[J]. Science 267:1456-1462.
Tsujii H, Kamada T (2012) A review of update clinical results of carbon ion radiotherapy[J]. Jpn J Clin Oncol 42:670-685.
Vittar NB, Awruch J, Azizuddin K, Rivarola V (2010) Caspase-independent apoptosis, in human MCF-7c3 breast cancer cells, following photodynamic therapy, with a novel water-soluble phthalocyanine[J]. Int J Biochem Cell Biol 42:1123-1131.
Vogelstein B, Lane D, Levine AJ (2000) Surfing the p53 network[J]. Nature 408:307-310.
Wakatsuki M, Tsuji H, Ishikawa H, Yanagi T, Kamada T, Nakano T, Suzuki H, Akakura K, Shimazaki J, Tsujii H (2008) Quality of life in men treated with carbon ion therapy for prostate cancer[J]. Int J Radiat Oncol Biol Phys 72:1010-1015.
Wang Y, Huang JC, Zhou ZL, Yang W, Guastella J, Drewe J, Cai SX (2004) Dipeptidyl aspartyl fluoromethylketones as potent caspase-3 inhibitors:SAR of the P2 amino acid[J]. Bioorg Med Chem Lett 14:1269-1272.
Weissman IL (2000) Stem cells:units of development, units of regeneration, and units in evolution[J]. Cell 100:157-168.
Willis SN, Fletcher JI, Kaufmann T, van Delft MF, Chen L, Czabotar PE, lerino H, Lee EF, Fairlie WD, Bouillet P, Strasser A, Kluck RM, Adams JM, Huang DC (2007) Apoptosis initiated when BH3 ligands engage multiple Bcl-2 homologs, not Bax or Bak[J]. Science 315:856-859.
Winter RN, Kramer A, Borkowski A, Kyprianou N (2001) Loss of caspase-1 and caspase-3 protein expression in human prostate cancer[J]. Cancer Res 61:1227-1232.
Wu CH, Jeng JH, Wang YJ, Tseng CJ, Liang YC, Chen CH, Lee HM, Lin JK, Lin CH, Lin SY, Li CP, Ho YS (2002) Antitumor effects of miconazole on human colon carcinoma xenografts in nude mice through induction of apoptosis and G0/G1 cell cycle arrest[J]. Toxicol Appl Pharmacol 180:22-35.
Yamabe K, Shimizu S, Ito T, Yoshioka Y, Nomura M, Narita M, Saito I, Kanegae Y, Matsuda H (1999) Cancer gene therapy using a pro-apoptotic gene, caspase-3[J]. Gene Ther 6:1952-1959.
Yamakawa N, Takahashi A, Mori E, Imai Y, Furusawa Y, Ohnishi K, Kirita T, Ohnishi T (2008) High LET radiation enhances apoptosis in mutated p53 cancer cells through Caspase-9 activation[J]. Cancer Sci 99:1455-1460.
Yang E, Zha J, Jockel J, Boise LH, Thompson CB, Korsmeyer SJ (1995) Bad, a heterodimeric partner for Bcl-XL and Bcl-2, displaces Bax and promotes cell death[J]. Cell 80:285-291.
Yonai S, Matsufuji N, Kanai T (2009) Monte Carlo study on secondary neutrons in passive carbon-ion radiotherapy:identification of the main source and reduction in the secondary neutron dose[J]. Med Phys 36:4830-4839.
Zacharias T, Dorr W, Enghardt W, Haberer T, Kramer M, Kumpf R, Rothig H, Scholz M, Weber U, Kraft G, Herrmann T (1997) Acute response of pig skin to irradiation with 12C-ions or 200 kV X-rays[J]. Acta Oncol 36:637-642.
Zapata JM, Krajewska M, Krajewski S, Huang RP, Takayama S, Wang HG, Adamson E, Reed JC (1998) Expression of multiple apoptosis-regulatory genes in human breast cancer cell lines and primary tumors[J]. Breast Cancer Res Treat 47:129-140.
Zhan Q, Chen IT, Antinore MJ, Fornace AJ, Jr. (1998) Tumor suppressor p53 can participate in transcriptional induction of the GADD45 promoter in the absence of direct DNA binding[J]. Mol Cell Biol 18:2768-2778.
丁小凡,李文建,李德杏(2005a)重离子对小鼠移植性Lewis肺癌分次照射的生物效应[J].中华放射医学与防护杂志25：45-47.
丁小凡,陈龙华,李文建(2005b)碳离子局部照射对荷瘤小鼠存活时间和血像的影响[J].第一军医大学学报25：32-34.
王艳明,卫韦华(2011.11.10)我国重离子治疗深层肿瘤技术和临床试验获重要突破[J].兰州晨报.
李文建,周光明,王菊芳(2003)重离子辐射哺乳动物细胞敏感性的分子机理[J].原子核物理评论20：42-43.
李文建,周光明,卫增泉(2002)碳离子辐照细胞的相对生物学效率[J].高能物理与核物理26：742-746.
李连弟,鲁风珠,张思维(1997)中国恶性肿瘤死亡率20年变化趋势和近期预测分析[J].中华肿瘤杂志19：3-9.
周光明,李文建,高清祥(2003)DNA双链断裂产额的新算法[J1.原子核物理评论20：52-53.
段纪俊,陈万青,张思维(2009)中国恶性肿瘤死亡率的国际比较[J].中国社会医学杂志26：377-378.
刘哲林,姜涛(2004)Caspase-3抑制剂研究进展[J].广东药学院学报20：669-670.
卫生部卫生统计信息中心(2003)中国恶性肿瘤危险因素研究[M].北京：中国协和医科大学出版社66-70.
吴运生,程清洲(2003)大肠癌及腺瘤中Caspase-3和Bcl-2的表达及临床意义[J].数理医药学杂志16：300-302.
张百红,姜宁雨(2002)肝癌组织Caspase-3基因表达与细胞凋亡的关系[J].第四军医大学学报23：35-38.
Alimonti JB, Ball TB, Fowke KR (2003) Mechanisms of CD4+T lymphocyte cell death in human immunodeficiency virus infection and AIDS[J]. J Gen Virol 84:1649-1661.
Avdeeva Zh I, Akol'zina SE, Alpatova NA, Medunitsyn NV (2005) [Effects of cytokines on the immunogenic properties of hepatitis A vaccine][J]. Vopr Virusol 50:23-27.
Bajt ML, Cover C, Lemasters JJ, Jaeschke H (2006) Nuclear translocation of endonuclease G and apoptosis-inducing factor during acetaminophen-induced liver cell injury[J]. Toxicol Sci 94:217-225.
Bassler N, Jakel O, Sondergaard CS, Petersen JB (2010) Dose-and LET-painting with particle therapy[J]. Acta Oncol 49:1170-1176.
Bechmann LP, Marquitan G, Jochum C, Saner F, Gerken G, Canbay A (2008) Apoptosis versus necrosis rate as a predictor in acute liver failure following acetaminophen intoxication compared with acute-on-chronic liver failure[J]. Liver Int 28:713-716.
Borner C (2003) The Bcl-2 protein family:sensors and checkpoints for life-or-death decisions[J]. Mol Immunol 39:615-647.
Budihardjo I, Oliver H, Lutter M, Luo X, Wang X (1999) Biochemical pathways of caspase activation during apoptosis[J]. Annu Rev Cell Dev Biol 15:269-290.
Canbay A, Higuchi H, Bronk SF, Taniai M, Sebo TJ, Gores GJ (2002) Fas enhances fibrogenesis in the bile duct ligated mouse:a link between apoptosis and fibrosis[J]. Gastroenterology 123:1323-1330.
Canman CE, Kastan MB (1997) Role of p53 in apoptosis[J]. Adv Pharmacol 41:429-460.
Castro MA, Schwartsmann G, Bernard EA, Moreira JC (1999) Phenotype modulation of cellular UV-sensitivity[J]. Cancer Lett 145:65-72.
Chen GG, Liang NC, Lee JF, Chan UP, Wang SH, Leung BC, Leung KL (2004) Over-expression of Bcl-2 against Pteris semipinnata L-induced apoptosis of human colon cancer cells via a NF-kappa B-related pathway[J]. Apoptosis 9:619-627.
Chen YC, Shen SC, Tsai SH (2005) Prostaglandin D(2) and J(2) induce apoptosis in human leukemia cells via activation of the caspase 3 cascade and production of reactive oxygen species[J]. Biochim Biophys Acta 1743:291-304.
Chen YH, Zhang YH, Zhang HJ, Liu DZ, Gu M, Li JY, Wu F, Zhu XZ, Li J, Nan FJ (2006) Design, synthesis, and biological evaluation of isoquinoline-1,3,4-trione derivatives as potent caspase-3 inhibitors[J]. J Med Chem 49:1613-1623.
Chintharlapalli SR, Jasti M, Malladi S, Parsa KV, Ballestero RP, Gonzalez-Garcia M (2005) BMRP is a Bcl-2 binding protein that induces apoptosis[J]. J Cell Biochem 94:611-626.
Choi KM, Zhong Y, Hoit BD, Grupp IL, Hahn H, Dilly KW, Guatimosim S, Lederer WJ, Matlib MA (2002) Defective intracellular Ca(2+) signaling contributes to cardiomyopathy in Type 1 diabetic rats[J]. Am J Physiol Heart Circ Physiol 283:H1398-1408.
Choy JC, Granville DJ, Hunt DW, McManus BM (2001) Endothelial cell apoptosis:biochemical characteristics and potential implications for atherosclerosis[J]. J Mol Cell Cardiol 33:1673-1690.
Chugh R, Dunn R, Zalupski MM, Biermann JS, Sondak VK, Mace JR, Leu KM, Chandler WF, Baker LH (2005) Phase Ⅱ study of 9-nitro-camptothecin in patients with advanced chordoma or soft tissue sarcoma[J]. J Clin Oncol 23:3597-3604.
Combs SE, Ellerbrock M, Haberer T, Habermehl D, Hoess A, Jakel O, Jensen A, Klemm S, Munter M, Naumann J, Nikoghosyan A, Oertel S, Parodi K, Rieken S, Debus J (2010) Heidelberg Ion Therapy Center (HIT):Initial clinical experience in the first 80 patients[J]. Acta Oncol 49:1132-1140.
Coultas L, Strasser A (2003) The role of the Bcl-2 protein family in cancer[J]. Semin Cancer Biol 13:115-123.
Dubray B, Breton C, Delic J, Klijanienko J, Maciorowski Z, Vielh P, Fourquet A, Dumont J, Magdelenat H, Cosset JM (1997) In vitro radiation-induced apoptosis and tumour response to radiotherapy:a prospective study in patients with non-Hodgkin lymphomas treated by low-dose irradiation[J]. Int J Radiat Biol 72:759-760.
Ferri CP, Prince M, Brayne C, Brodaty H, Fratiglioni L, Ganguli M, Hall K, Hasegawa K, Hendrie H, Huang Y, Jorm A, Mathers C, Menezes PR, Rimmer E, Scazufca M (2005) Global prevalence of dementia:a Delphi consensus study[J]. Lancet 366:2112-2117.
Fukumura A, Tsujii H, Kamada T, Baba M, Tsuji H, Kato H, Kato S, Yamada S, Yasuda S, Yanagi T, Hara R, Yamamoto N, Mizoe J, Akahane K, Fukuda S, Furusawa Y, Iwata Y, Kanai T, Kanematsu N, Kitagawa A, Matsufuji N, Minohara S, Miyahara N, Mizuno H, Murakami T, Nishizawa K, Noda K, Takada E, Yonai S (2009) Carbon-ion radiotherapy:clinical aspects and related dosimetry[J]. Radiat Prot Dosimetry 137:149-155.
Furusawa Y, Aoki M, Durante M (2002) Simultaneous exposure of mammalian cells to heavy ions and X-rays[J]. Adv Space Res 30:877-884.
Fushimi K, Uzawa K, Ishigami T, Yamamoto N, Kawata T, Shibahara T, Ito H, Mizoe JE, Tsujii H, Tanzawa H (2008) Susceptible genes and molecular pathways related to heavy ion irradiation in oral squamous cell carcinoma cells[J]. Radiother Oncol 89:237-244.
Giannopoulou I, Nakopoulou L, Zervas A, Lazaris AC, Stravodimos C, Giannopoulos A, Davaris PS (2002) Immunohistochemical study of pro-apoptotic factors Bax, Fas and CPP32 in urinary bladder cancer:prognostic implications[J]. Urol Res 30:342-345.
Gross A, McDonnell JM, Korsmeyer SJ (1999) BCL-2 family members and the mitochondria in apoptosis[J]. Genes Dev 13:1899-1911.
Gueven N, Becherel OJ, Birrell G, Chen P, DelSal G, Carney JP, Grattan-Smith P, Lavin MF (2006) Defective p53 response and apoptosis associated with an ataxia-telangiectasia-like phenotype[J]. Cancer Res 66:2907-2912.
Haddad JJ (2004) On the antioxidant mechanisms of Bcl-2:a retrospective of NF-kappaB signaling and oxidative stress[J]. Biochem Biophys Res Commun 322:355-363.
Hengartner MO (2001) Apoptosis:corralling the corpses[J]. Cell 104:325-328.
Hsia JY, Chen CY, Chen JT, Hsu CP, Shai SE, Yang SS, Chuang CY, Wang PY, Miaw J (2003) Prognostic significance of caspase-3 expression in primary resected esophageal squamous cell carcinoma[J]. Eur J Surg Oncol 29:44-48.
Imai R, Kamada T, Tsuji H, Sugawara S, Serizawa I, Tsujii H, Tatezaki S (2010) Effect of carbon ion radiotherapy for sacral chordoma:results of Phase Ⅰ-Ⅱ and Phase Ⅱ clinical trials[J]. Int J Radiat Oncol Biol Phys 77:1470-1476.
Imai R, Kamada T, Tsuji H, Yanagi T, Baba M, Miyamoto T, Kato S, Kandatsu S, Mizoe JE, Tsujii H, Tatezaki S (2004) Carbon ion radiotherapy for unresectable sacral chordomas[J]. Clin Cancer Res 10:5741-5746.
Isobe N, Onodera H, Mori A, Shimada Y, Yang W, Yasuda S, Fujimoto A, Ooe H, Arii S, Kitaichi M, Imamura M (2004) Caspase-3 expression in human gastric carcinoma and its clinical significance[J]. Oncology 66:201-209.
Johnson R, Staiano-Coico L, Austin L, Cardinale I, Nabeya-Tsukifuji R, Krueger JG (1996) PUVA treatment selectively induces a cell cycle block and subsequent apoptosis in human T-lymphocytes[J]. Photochem Photobiol 63:566-571.
Jonges LE, Nagelkerke JF, Ensink NG, van der Velde EA, Tollenaar RA, Fleuren GJ, van de Velde CJ, Morreau H, Kuppen PJ (2001) Caspase-3 activity as a prognostic factor in colorectal carcinoma[J]. Lab Invest 81:681-688.
Kanai T, Endo M, Minohara S, Miyahara N, Koyama-ito H, Tomura H, Matsufuji N, Futami Y, Fukumura A, Hiraoka T, Furusawa Y, Ando K, Suzuki M, Soga F, Kawachi K (1999) Biophysical characteristics of HIMAC clinical irradiation system for heavy-ion radiation therapy[J]. Int J Radiat Oncol Biol Phys 44:201-210.
Kanai T, Furusawa Y, Fukutsu K, Itsukaichi H, Eguchi-Kasai K, Ohara H (1997) Irradiation of mixed beam and design of spread-out Bragg peak for heavy-ion radiotherapy[J]. Radiat Res 147:78-85.
Kanai T, Matsufuji N, Miyamoto T, Mizoe J, Kamada T, Tsuji H, Kato H, Baba M, Tsujii H (2006) Examination of GyE system for HIMAC carbon therapy [J]. Int J Radiat Oncol Biol Phys 64:650-656.
Kane DJ, Sarafian TA, Anton R, Hahn H, Gralla EB, Valentine JS, Ord T, Bredesen DE (1993) Bcl-2 inhibition of neural death:decreased generation of reactive oxygen species[J]. Science 262:1274-1277.
Katsuda K, Kataoka M, Uno F, Murakami T, Kondo T, Roth JA, Tanaka N, Fujiwara T (2002) Activation of caspase-3 and cleavage of Rb are associated with p16-mediated apoptosis in human non-small cell lung cancer cells[J]. Oncogene 21:2108-2113.
Kim S, Ko H, Park JE, Jung S, Lee SK, Chun YJ (2002) Design, synthesis, and discovery of novel trans-stilbene analogues as potent and selective human cytochrome P450 1B1 inhibitors[J]. J Med Chem 45:160-164.
Kolenko V, Bloom T, Rayman P, Bukowski R, Hsi E, Finke J (1999) Inhibition of NF-kappa B activity in human T lymphocytes induces caspase-dependent apoptosis without detectable activation of caspase-1 and -3[J]. J Immunol 163:590-598.
Kondo S, Barna BP, Kondo Y, Tanaka Y, Casey G, Liu J, Morimura T, Kaakaji R, Peterson JW, Werbel B, Barnett GH (1996) WAF1/CIP1 increases the susceptibility of p53 non-functional malignant glioma cells to cisplatin-induced apoptosis[J]. Oncogene 13:1279-1285.
Korsmeyer SJ (1992) Bcl-2 initiates a new category of oncogenes:regulators of cell death[J]. Blood 80:879-886.
Kraft G (1990) The radiobiological and physical basis for radiotherapy with protons and heavier ions[J]. Strahlenther Onkol 166:10-13.
Kroemer G (1997) The proto-oncogene Bcl-2 and its role in regulating apoptosis[J]. Nat Med 3:614-620.
Kubota N, Suzuki M, Furusawa Y, Ando K, Koike S, Kanai T, Yatagai F, Ohmura M, Tatsuzaki H, Matsubara S, et al. (1995) A comparison of biological effects of modulated carbon-ions and fast neutrons in human osteosarcoma cells[J]. Int J Radiat Oncol Biol Phys 33:135-141.
Li P, Nijhawan D, Budihardjo I, Srinivasula SM, Ahmad M, Alnemri ES, Wang X (1997) Cytochrome c and dATP-dependent formation of Apaf-1/caspase-9 complex initiates an apoptotic protease cascade[J]. Cell 91:479-489.
Li X, Marani M, Yu J, Nan B, Roth JA, Kagawa S, Fang B, Denner L, Marcelli M (2001) Adenovirus-mediated Bax overexpression for the induction of therapeutic apoptosis in prostate cancer[J]. Cancer Res 61:186-191.
Loop T, Dovi-Akue D, Frick M, Roesslein M, Egger L, Humar M, Hoetzel A, Schmidt R, Borner C, Pahl HL, Geiger KK, Pannen BH (2005) Volatile anesthetics induce caspase-dependent, mitochondria-mediated apoptosis in human T lymphocytes in vitro[J]. Anesthesiology 102:1147-1157.
Lunkes A, Trottier Y, Mandel JL (1998) Pathological mechanisms in Huntington's disease and other polyglutamine expansion diseases[J]. Essays Biochem 33:149-163.
Mahieux R, Pise-Masison C, Gessain A, Brady JN, Olivier R, Perret E, Misteli T, Nicot C (2001) Arsenic trioxide induces apoptosis in human T-cell leukemia virus type 1- and type 2-infected cells by a caspase-3-dependent mechanism involving Bcl-2 cleavage[J]. Blood 98:3762-3769.
Marcelli M, Cunningham GR, Walkup M, He Z, Sturgis L, Kagan C, Mannucci R, Nicoletti I, Teng B, Denner L (1999) Signaling pathway activated during apoptosis of the prostate cancer cell line LNCaP:overexpression of caspase-7 as a new gene therapy strategy for prostate cancer[J]. Cancer Res 59:382-390.
Masunaga S, Ando K, Uzawa A, Hirayama R, Furusawa Y, Koike S, Sakurai Y, Nagata K, Suzuki M, Kashino G, Kinashi Y, Tanaka H, Maruhashi A, Ono K (2008) Radiobiologic significance of response of intratumor quiescent cells in vivo to accelerated carbon ion beams compared with gamma-rays and reactor neutron beams[J].Int J Radiat Oncol Biol Phys 70:221-228.
Miyazaki H, Kuwano K, Yoshida K, Maeyama T, Yoshimi M, Fujita M, Hagimoto N, Yoshida R, Nakanishi Y (2004) The perforin mediated apoptotic pathway in lung injury and fibrosis[J]. J Clin Pathol 57:1292-1298.
Mizoe JE, Tsujii H, Hasegawa A, Yanagi T, Takagi R, Kamada T, Tsuji H, Takakura K (2007) Phase I/II clinical trial of carbon ion radiotherapy for malignant gliomas:combined X-ray radiotherapy, chemotherapy, and carbon ion radiotherapy[J]. Int J Radiat Oncol Biol Phys 69:390-396.
Moll UM, Zaika A (2001) Nuclear and mitochondrial apoptotic pathways of p53[J]. FEBS Lett 493:65-69.
Mori S, Yanagi T, Hara R, Sharp GC, Asakura H, Kumagai M, Kishimoto R, Yamada S, Kato H, Kandatsu S, Kamada T (2010) Comparison of respiratory-gated and respiratory-ungated planning in scattered carbon ion beam treatment of the pancreas using four-dimensional computed tomography [J]. Int J Radiat Oncol Biol Phys 76:303-312.
Muchmore SW, Sattler M, Liang H, Meadows RP, Harlan JE, Yoon HS, Nettesheim D, Chang BS, Thompson CB, Wong SL, Ng SL, Fesik SW (1996) X-ray and NMR structure of human Bcl-xL, an inhibitor of programmed cell death[J]. Nature 381:335-341.
Naka T, Boitze C, Kuester D, Schulz TO, Schneider-Stock R, Kellner A, Samii A, Herold C, Ostertag H, Roessner A (2005) Alterations of Gl-S checkpoint in chordoma:the prognostic impact of p53 overexpression[J]. Cancer 104:1255-1263.
Neumar RW, Xu YA, Gada H, Guttmann RP, Siman R (2003) Cross-talk between calpain and caspase proteolytic systems during neuronal apoptosis[J]. J Biol Chem 278:14162-14167.
Norberg E, Gogvadze V, Ott M, Horn M, Uhlen P, Orrenius S, Zhivotovsky B (2008) An increase in intracellular Ca2+ is required for the activation of mitochondrial calpain to release AIF during cell death[J]. Cell Death Differ 15:1857-1864.
Nordsmark M, Bentzen SM, Rudat V, Brizel D, Lartigau E, Stadler P, Becker A, Adam M, Molls M, Dunst J, Terris DJ, Overgaard J (2005) Prognostic value of tumor oxygenation in 397 head and neck tumors after primary radiation therapy. An international multi-center study[J]. Radiother Oncol 77:18-24.
Ogata M, Inanami O, Nakajima M, Nakajima T, Hiraoka W, Kuwabara M (2003) Ca(2+)-dependent and caspase-3-independent apoptosis caused by damage in Golgi apparatus due to 2,4,5,7-tetrabromorhodamine 123 bromide-induced photodynamic effects[J]. Photochem Photobiol 78:241-247.
Olive PL, Durand RE (1997) Apoptosis:an indicator of radiosensitivity in vitro?[J]. Int J Radiat Biol 71:695-707.
Pan G, Li J, Zhou Y, Zheng H, Pei D (2006) A negative feedback loop of transcription factors that controls stem cell pluripotency and self-renewal[J]. FASEB J 20:1730-1732.
Penning LC, Rasch MH, Ben-Hur E, Dubbelman TM, Havelaar AC, Van der Zee J, Van Steveninck J (1992) A role for the transient increase of cytoplasmic free calcium in cell rescue after photodynamic treatment[J]. Biochim Biophys Acta 1107:255-260.
Puolakkainen PA, Brekken RA, Muneer S, Sage EH (2004) Enhanced growth of pancreatic tumors in SPARC-null mice is associated with decreased deposition of extracellular matrix and reduced tumor cell apoptosis[J]. Mol Cancer Res 2:215-224.
Putt KS, Chen GW, Pearson JM, Sandhorst JS, Hoagland MS, Kwon JT, Hwang SK, Jin H, Churchwell MI, Cho MH, Doerge DR, Helferich WG, Hergenrother PJ (2006) Small-molecule activation of procaspase-3 to caspase-3 as a personalized anticancer strategy[J]. Nat Chem Biol 2:543-550.
Reed JC (1998) Bcl-2 family proteins[J]. Oncogene 17:3225-3236.
Reya T, Morrison SJ, Clarke MF, Weissman IL (2001) Stem cells, cancer, and cancer stem cells[J]. Nature 414:105-111.
Saikumar P, Dong Z, Mikhailov V, Denton M, Weinberg JM, Venkatachalam MA (1999) Apoptosis: definition, mechanisms, and relevance to disease[J]. Am J Med 107:489-506.
Samis JA, Back DW, Graham EJ, DeLuca CI, Elce JS (1991) Constitutive expression of calpain Ⅱ in the rat uterus during pregnancy and involution[J]. Biochem J 276 (Pt 2):293-299.
Scheil S, Bruderlein S, Liehr T, Starke H, Herms J, Schulte M, Moller P (2001) Genome-wide analysis of sixteen chordomas by comparative genomic hybridization and cytogenetics of the first human chordoma cell line, U-CH1 [J]. Genes Chromosomes Cancer 32:203-211.
Shinoura N, Muramatsu Y, Yoshida Y, Asai A, Kirino T, Hamada H (2000) Adenovirus-mediated transfer of caspase-3 with Fas ligand induces drastic apoptosis in U-373MG glioma cellsfJ]. Exp Cell Res 256:423-433.
Skarsgard LD (1998) Radiobiology with heavy charged particles:a historical review[J]. Phys Med 14 Suppl 1:1-19.
Sliskovic I, Mutus B (2006) Reversible inhibition of caspase-3 activity by iron(Ⅲ):potential role in physiological control of apoptosis[J]. FEBS Lett 580:2233-2237.
Spira A, Ettinger DS (2004) Multidisciplinary management of lung cancer[J]. N Engl J Med 350:379-392.
Stone HB, Dewey WC, Wallace SS, Coleman CN (1998) Molecular Biology to Radiation Oncology:A Model for Translational Research? Opportunities in basic and translational research. From a workshop sponsored by the National Cancer Institute, Radiation Research Program, January 26-28,1997, Bethesda, Maryland[J]. Radiat Res 150:134-147.
Suzuki A, Ito T, Kawano H, Hayashida M, Hayasaki Y, Tsutomi Y, Akahane K, Nakano T, Miura M, Shiraki K (2000) Survivin initiates procaspase 3/p21 complex formation as a result of interaction with Cdk4 to resist Fas-mediated cell death[J]. Oncogene 19:1346-1353.
Suzuki Y, Oka K, Ohno T, Kato S, Tsujii H, Nakano T (2009) Prognostic impact of mitotic index of proliferating cell populations in cervical cancer patients treated with carbon ion beam[J]. Cancer 115:1875-1882.
Tang MJ, Tai IT (2007) A novel interaction between procaspase 8 and SPARC enhances apoptosis and potentiates chemotherapy sensitivity in colorectal cancers[J]. J Biol Chem 282:34457-34467.
Tatsukawa H, Sano T, Fukaya Y, Ishibashi N, Watanabe M, Okuno M, Moriwaki H, Kojima S (2011) Dual induction of caspase 3- and transglutaminase-dependent apoptosis by acyclic retinoid in hepatocellular carcinoma cells[J]. Mol Cancer 10:4.
Thibaut S, Bourre L, Hernot D, Rousset N, Lajat Y, Patrice T (2002) Effects of BAPTA-AM, Forskolin, DSF and Z.VAD.fmk on PDT-induced apoptosis and m-THPC phototoxicity on B16 cells[J]. Apoptosis 7:99-106.
Thompson CB (1995) Apoptosis in the pathogenesis and treatment of disease[J]. Science 267:1456-1462.
Tsujii H, Kamada T (2012) A review of update clinical results of carbon ion radiotherapy[J]. Jpn J Clin Oncol 42:670-685.
Vittar NB, Awruch J, Azizuddin K, Rivarola V (2010) Caspase-independent apoptosis, in human MCF-7c3 breast cancer cells, following photodynamic therapy, with a novel water-soluble phthalocyanine[J]. Int J Biochem Cell Biol 42:1123-1131.
Vogelstein B, Lane D, Levine AJ (2000) Surfing the p53 network[J]. Nature 408:307-310.
Wakatsuki M, Tsuji H, Ishikawa H, Yanagi T, Kamada T, Nakano T, Suzuki H, Akakura K, Shimazaki J, Tsujii H (2008) Quality of life in men treated with carbon ion therapy for prostate cancer[J]. Int J Radiat Oncol Biol Phys 72:1010-1015.
Wang Y, Huang JC, Zhou ZL, Yang W, Guastella J, Drewe J, Cai SX (2004) Dipeptidyl aspartyl fluoromethylketones as potent caspase-3 inhibitors:SAR of the P2 amino acid[J]. Bioorg Med Chem Lett 14:1269-1272.
Weissman IL (2000) Stem cells:units of development, units of regeneration, and units in evolution[J]. Cell 100:157-168.
Willis SN, Fletcher JI, Kaufmann T, van Delft MF, Chen L, Czabotar PE, lerino H, Lee EF, Fairlie WD, Bouillet P, Strasser A, Kluck RM, Adams JM, Huang DC (2007) Apoptosis initiated when BH3 ligands engage multiple Bcl-2 homologs, not Bax or Bak[J]. Science 315:856-859.
Winter RN, Kramer A, Borkowski A, Kyprianou N (2001) Loss of caspase-1 and caspase-3 protein expression in human prostate cancer[J]. Cancer Res 61:1227-1232.
Wu CH, Jeng JH, Wang YJ, Tseng CJ, Liang YC, Chen CH, Lee HM, Lin JK, Lin CH, Lin SY, Li CP, Ho YS (2002) Antitumor effects of miconazole on human colon carcinoma xenografts in nude mice through induction of apoptosis and G0/G1 cell cycle arrest[J]. Toxicol Appl Pharmacol 180:22-35.
Yamabe K, Shimizu S, Ito T, Yoshioka Y, Nomura M, Narita M, Saito I, Kanegae Y, Matsuda H (1999) Cancer gene therapy using a pro-apoptotic gene, caspase-3[J]. Gene Ther 6:1952-1959.
Yamakawa N, Takahashi A, Mori E, Imai Y, Furusawa Y, Ohnishi K, Kirita T, Ohnishi T (2008) High LET radiation enhances apoptosis in mutated p53 cancer cells through Caspase-9 activation[J]. Cancer Sci 99:1455-1460.
Yang E, Zha J, Jockel J, Boise LH, Thompson CB, Korsmeyer SJ (1995) Bad, a heterodimeric partner for Bcl-XL and Bcl-2, displaces Bax and promotes cell death[J]. Cell 80:285-291.
Yonai S, Matsufuji N, Kanai T (2009) Monte Carlo study on secondary neutrons in passive carbon-ion radiotherapy:identification of the main source and reduction in the secondary neutron dose[J]. Med Phys 36:4830-4839.
Zacharias T, Dorr W, Enghardt W, Haberer T, Kramer M, Kumpf R, Rothig H, Scholz M, Weber U, Kraft G, Herrmann T (1997) Acute response of pig skin to irradiation with 12C-ions or 200 kV X-rays[J]. Acta Oncol 36:637-642.
Zapata JM, Krajewska M, Krajewski S, Huang RP, Takayama S, Wang HG, Adamson E, Reed JC (1998) Expression of multiple apoptosis-regulatory genes in human breast cancer cell lines and primary tumors[J]. Breast Cancer Res Treat 47:129-140.
Zhan Q, Chen IT, Antinore MJ, Fornace AJ, Jr. (1998) Tumor suppressor p53 can participate in transcriptional induction of the GADD45 promoter in the absence of direct DNA binding[J]. Mol Cell Biol 18:2768-2778.
丁小凡,李文建,李德杏(2005a)重离子对小鼠移植性Lewis肺癌分次照射的生物效应[J].中华放射医学与防护杂志25：45-47.
丁小凡,陈龙华,李文建(2005b)碳离子局部照射对荷瘤小鼠存活时间和血像的影响[J].第一军医大学学报25：32-34.
王艳明,卫韦华(2011.11.10)我国重离子治疗深层肿瘤技术和临床试验获重要突破[J].兰州晨报.
李文建,周光明,王菊芳(2003)重离子辐射哺乳动物细胞敏感性的分子机理[J].原子核物理评论20：42-43.
李文建,周光明,卫增泉(2002)碳离子辐照细胞的相对生物学效率[J].高能物理与核物理26：742-746.
李连弟,鲁风珠,张思维(1997)中国恶性肿瘤死亡率20年变化趋势和近期预测分析[J].中华肿瘤杂志19：3-9.
周光明,李文建,高清祥(2003)DNA双链断裂产额的新算法[J1.原子核物理评论20：52-53.
段纪俊,陈万青,张思维(2009)中国恶性肿瘤死亡率的国际比较[J].中国社会医学杂志26：377-378.
刘哲林,姜涛(2004)Caspase-3抑制剂研究进展[J].广东药学院学报20：669-670.
卫生部卫生统计信息中心(2003)中国恶性肿瘤危险因素研究[M].北京：中国协和医科大学出版社66-70.
吴运生,程清洲(2003)大肠癌及腺瘤中Caspase-3和Bcl-2的表达及临床意义[J].数理医药学杂志16：300-302.
张百红,姜宁雨(2002)肝癌组织Caspase-3基因表达与细胞凋亡的关系[J].第四军医大学学报23：35-38.