利用超分次技术进行肿瘤治疗的探讨

以人类肝癌细胞SMMC-7721和正常肝细胞L02为研究对象,以这两种细胞0.3 Gy时超敏感性的存活数据为基础,从理论上探讨了γ射线照射时,用超分次技术治疗肝癌的可能性。经过计算发现:如果目标肿瘤和周围的正常组织超敏感性的存活差异提高到3%,即可利用超分次技术对肿瘤进行治疗。应用超分次进行分次照射时,照射的结果与分次的间隔时间有关。对这一现象的机理进行了一定的探讨,发现时间间隔与细胞G2期的长短可能存在一定的相关性。

辐射诱导p53腺病毒重组体转染对结直肠癌细胞周期的影响

以复制缺陷型重组腺病毒载体(AdCMV-GFP)为对照,用复制缺陷型p53重组腺病毒载体 (AdCMV-p53)转染经0．5、1．0、2．0Gyγ射线照射的HT-29细胞,克隆形成法检测对细胞的抑制作用,流式细胞分析法检测细胞周期和细胞凋亡,探讨辐射诱导对AdCMV-p53转染p53突变型结直肠癌细胞(HT-29细胞系)细胞周期的影响。结果显示,0．5-1．0Gy辐射诱导明显增强40 MOI AdCMV-p53转染对HT-29细胞的抑制。与AdCMV-p53转染对照相比,1 d后,辐射诱导转染组G0／G1期细胞减少5%-15％,S期细胞增加 2%-19％,2．0Gy辐射诱导80 MOI AdCMV-p53转染组G2/M期细胞增加12％;3d后,0．5、1．0Gy辐射诱导40 MOI AdCMV-p53转染组G2／M期细胞分别增加10%-13％。辐射诱导AdCMV-p53转染组细胞凋亡与辐射诱导剂量和AdCMV-p53转染剂量相关。以上结果表明,辐射诱导加速AdCMV-p53转染细胞由G0／G1期到S期的进程,促进S期阻滞和G2/M期阻滞发生。

80MeV/u~(12)C~(6+)离子辐照小鼠头部对脾脏细胞周期分布的影响

研究重离子辐照小鼠头部对脾脏细胞周期分布的影响,为重离子放射治疗癌症和太空防护提供基础数据。80MeV/u能量的12C6+对BALB/c小鼠头部给以0、0.5、1、2、4、10Gy的照射,用流式细胞仪测脾脏细胞周期分布。重离子辐照后36h,小鼠脾脏细胞S期细胞随着辐照剂量的增加显著减少(p<0.05);0.5Gy组、4Gy组和10Gy组出现G1/G0期阻滞明显阻滞(p<0.05),1Gy组和2Gy组无显著变化(p﹥0.05);0.5Gy组G2/M期细胞显著减少(p<0.01),其它剂量组明显阻滞(p<0.05)。重离子辐照小鼠头部对小鼠脾脏细胞周期分布有明显影响。

3Gyγ射线对不同肿瘤细胞细胞周期进程的影响

本实验旨在研究电离辐射对不同肿瘤细胞细胞周期进程的影响,为肿瘤的放射治疗提供科学依据。以3Gyγ射线照射指数生长期的肿瘤细胞,用流式细胞术测定细胞DNA含量并确定细胞周期各时相的细胞百分比。实验结果显示,γ射线照射后,SMMC-7721,hepG2和HO-8910细胞的G2/M期发生明显阻滞,且都在辐射后12h累积达到最大值,但S期只发生短暂的延迟,即辐射后6h明显累积,之后下降至接近对照水平或低于对照;Hela细胞的G2/M期和S期均发生显著阻滞。实验结果说明,电离辐射后,不同肿瘤细胞的G2/M期和S期检查点的激活和维持是有差异的。

利用PCC技术预测γ射线对肝癌细胞的辐射效应

利用早熟染色体凝集技术预测研究了γ射线对肝癌细胞SMMC7721的辐射效应。结果表明,G1和G2期细胞内的染色单体和等点染色单体断裂数与照射剂量之间存在线性相关性,染色单体断裂总数与细胞存活率之间存在良好的线性相关性。说明辐射诱导的染色单体断裂可以作为预测SMMC7721细胞内在辐射敏感性的指标,也可为临床诊断和治疗肝癌提供依据。

电离辐射对不同肿瘤细胞细胞周期进程的影响

Human tumor cell lines of SMMC-7721 (liver cancer), hepG2 (liver cancer), HO-8910 (ovary cancer), and Hela (cervix cancer) were irradiated to 3Gy by Co γ rays and different cell cycle responses were found. The re- 60 sults showed that the SMMC-7721, hepG2 and HO-8910 cells displayed G2 / M phase arrest and S phase tempo- rally delayed after the irradiation. The Hela cells had an increased number of cells in both G2 / M and S phase, which indicated that the G2 / M and S checkpoints were both activated.

低剂量电离辐射预处理对高剂量辐射诱导的肝癌细胞hepG2细胞周期阻滞的影响

以人肝癌细胞 hepG2 为研究对象,用流式细胞术测定并分析了低剂量 γ 射线(5cGy)预照射对高剂量照射(3Gy)诱导的细胞周期阻滞的影响。结果显示:(1)低剂量照射可引起 hepG2 细胞在 G2/M 期短暂累积(至照射后 4h);(2)低剂量照射促进 hepG2 细胞的生长;(3)高剂量照射后,hepG2 细胞的 G2期发生阻滞,而 S 期只发生短暂延迟;(4)与单纯高剂量照射相比,低剂量照射预处理后 4h 给予高剂量照射可进一步促进hepG2 细胞在 G2/M 期累积,但预处理后 8h 给予高剂量可促进细胞通过 G2/M 期。实验结果表明,低剂量照射预处理对高剂量照射诱导的细胞周期阻滞的影响,取决于两次照射的时间间隔。

低剂量电离辐射预处理对不同肿瘤细胞周期的影响

实验研究了低剂量γ射线预照射对人肝癌细胞系hepG2和人宫颈癌细胞系HeLa的细胞周期进程的影响.结果显示(1)低剂量(5cGy)辐射后,两种细胞的G2/M期细胞短暂累积;(2)低剂量辐射促进肿瘤细胞的生长;(3)高剂量(3Gy)辐射后,hepG2细胞发生G2期阻滞,HeLa细胞发生S期和G2期阻滞;(4)与单纯高剂量照射相比,低剂量辐射预处理后4h,再给予高剂量辐射,可进一步促进hepG2细胞在G2/M期累积,但是预照射对HeLa细胞的周期进程没有明显影响.因此,低剂量辐射预处理对高剂量诱导的细胞周期阻滞的影响依赖于肿瘤细胞的类型.

Prematurely condensed chromosome fragments in human lymphocytes induced by high doses of high-linear-energy-transfer irradiation

This study provides a useful biodosimetry protocol for radiation accidents that involve high doses of heavy particle radiation. Human peripheral blood lymphocytes (PBLs) were irradiated in vitro with high doses (5–50 Gy) of charged heavy-ion particles (carbon ions, at an effective linear-energy-transfer (LET) of 34.6 keV/ m), and were then stimulated to obtain dividing cells. PBLs were treated with 100nMcalyculin A to force chromosomes to condense prematurely, and chromosome spreads were obtained and stained with Giemsa. The G2 prematurely condensed chromosome (G2-PCC) index and the number of G2-PCC including fragments (G2-PCC-Fs) per cell for each radiation dose point were scored. Dose-effect relationships were obtained by plotting the G2-PCC indices or G2-PCC-Fs numbers against radiation doses. The G2-PCC index was greater than 5% up to doses of 15 Gy; even after a 30Gy radiation dose, the index was 1 to 2%. At doses higher than 30 Gy, however, the G2-PCC indices were close to zero. The number of G2-PCC-Fs increased steeply for radiation doses up to 30 Gy at a rate of 1.07 Gy−1. At doses higher than 30 Gy, the numbers of G2-PCC-Fs could not be accurately indexed because of the limited numbers of cells for analysis. Therefore, the number of G2-PCC-Fs could be used to estimate radiation doses up to 30 Gy. In addition, a G2-PCC index close to zero could be used as an indicator for radiation doses greater than 40 Gy.

Effect of 80.55 MeV/u(12)C(6+) ions on radiosensitivity and cell cycle of human hepatoma cell lines

In this paper, the relationship between radiosensitivity, cell cycle alteration and the change of apoptosis in different human hepatoma cell lines irradiated by heavy ions were studied with the aim of building up the base data for clinical therapy. Exponentially growing hepatoma cell lines were irradiated by 80.55 MeV/u(12)C(6+) ions at a dose of 0 Gy, 0.5 Gy, 1 Gy, 2 Gy, 4 Gy and 8 Gy. The radiosensitivity was assessed by means of the colony-forming assay. The DNA content, the percentage of each cell-cycle phase and the apoptosis rate were obtained with flow cytometry methods. After the irradiation, the SF2 (survival fraction at 2 gray) of SMMC-7721 cells were evidently lower than that of HepG2 cells. The S phase arrest, G2/M phase arrest delay and the apoptosis in the two hepatoma cell lines varied with the increase of the dose and repair time. The heavy ions could obviously kill the human hepatoma cell lines. Compared to HepG2 cells, SMMC-7721 cells were more radiosensitive to C-12(6+) ions.

Biodosimetry estimate for high-LET irradiation

The purpose of this paper is to prepare for an easy and reliable biodosimeter protocol for radiation accidents involving high-linear energy transfer (LET) exposure. Human peripheral blood lymphocytes were irradiated using carbon ions (LET: 34.6 keV mu m(-1)), and the chromosome aberrations induced were analyzed using both a conventional colcemid block method and a calyculin A induced premature chromosome condensation (PCC) method. At a lower dose range (0-4 Gy), the measured dicentric (dics) and centric ring chromosomes (cRings) provided reasonable dose information. At higher doses (8 Gy), however, the frequency of dics and cRings was not suitable for dose estimation. Instead, we found that the number of Giemsa-stained drug-induced G2 prematurely condensed chromosomes (G2-PCC) can be used for dose estimation, since the total chromosome number (including fragments) was linearly correlated with radiation dose (r = 0.99). The ratio of the longest and the shortest chromosome length of the drug-induced G2-PCCs increased with radiation dose in a linear-quadratic manner (r = 0.96), which indicates that this ratio can also be used to estimate radiation doses. Obviously, it is easier to establish the dose response curve using the PCC technique than using the conventional metaphase chromosome method. It is assumed that combining the ratio of the longest and the shortest chromosome length with analysis of the total chromosome number might be a valuable tool for rapid and precise dose estimation for victims of radiation accidents.

Comparison of clonogenic assay with premature chromosome condensation assay in prediction of human cell radiosensitivity

Aim: To determine whether the number of non-rejoining G2-chromatid breaks can predict the radiosensitivity of human cell lines. Methods: Cell lines of human ovary carcinoma cells (HO8910), human hepatoma cells (HepG2) and liver cells (L02) were irradiated with a range of doses and assessed both of cell survival and non-rejoining G2-chromatid breaks at 24 h after irradiation. Cell survival was documented by a colony assay. Non-rejoining G2-chromatid breaks were measured by counting the number of non-rejoining G2 chromatid breaks at 24 h after irradiation, detected by the prematurely chromosome condensed (PCC) technique. Results: A linear-quadratic survival curve was observed in three cell lines, and HepG2 was the most sensitive to gamma-radiation. A dose-dependent linear increase was observed in radiation-induced non-rejoining G2-PCC breaks measured at 24 h after irradiation in all cell lines, and HepG2 was the most susceptible to induction of non-rejoining G2-PCC breaks. A close correlation was found between the clonogenic radiosensitivity and the radiation-induced non-rejoining G2-PCC breaks (r=0.923). Furthermore, survival-aberration correlations for two or more than two doses lever were also significant. Conclusion: The number of non-rejoining G2 PCC breaks holds considerable promise for predicting the radiosensitivity of normal and tumor cells when two or more than two doses lever is tested.

Protective effects of melatonin against low- and high-LET irradiation

To investigate the protective effects of melatonin against high-LET ionizing radiation, V79 Chinese hamster cells were irradiated with 100 keV/mu m carbon beam. Parallel experiments were performed with 200 kV X-rays. To avoid the impact from extra solvents, melatonin was dissolved directly in culture medium. Cells were cultured in melatonin medium for 1 hr before irradiation. Cell inactivation was measured with conventional colony forming assay, medium containing 6-thioguanine was used for the selection of mutants at hprt locus, and the cell cycle was monitored by flow cytometry. Both carbon beam and X-rays induced cell inactivation, hprt gene mutation and cell cycle G2 block dose-dependently. But carbon beam showed stronger effects as indicated by all three endpoints and the relative biological effectiveness (RBE) was 3.5 for cell killing (at 10% survival level) and 2.9 for mutation induction (at 5 x 10(-5) mutants/ cell level). Melatonin showed protective effects against ionizing radiation in a dose-dependent manner. In terms of cell killing, melatonin only increased the survival level of those samples exposed to 8Gy or larger of X-rays or 6 Gy or larger of carbon beam. In the induction of hprt mutation and G2 block, melatonin reduced such effects induced by carbon beam but not by X-rays. The results suggest that melatonin reduces the direct interaction of particles with cells rather than an indirect interaction. Further studies are required to disclose the underlying mechanisms.

Potential role of DNA-dependent protein kinase in cellular resistance to ionizing radiation

In this paper, we study the ability of DNA-PK-deficient (M059J) and -proficient (M059K) cells to undergo the rate of cellular proliferation, cell cycle distribution and apoptosis after 10 Gy X-ray irradiation, and the role of DNA-PK in radiosensitivity. The results showed that M059J cells exhibited hyper-radiosensitivity compared with M059K cells. A strong G2 phase arrest was observed in M059J cells post irradiation. Significant accumulation in the G2 phase in M059J cells was accompanied by apoptosis at 12 h. Altogether, the data suggested that DNA-PK may have two roles in mammalian cells after DNA damage, a role in DNA DSB repair and a second role in DNA-damaged cells to traverse a G2 checkpoint, by which DNA-PK may affect cellular sensitivity to ionizing radiation. 地址: [Li Ning; Zhang Hong; Wang Yanling; Hao Jifang] Chinese Acad Sci, Inst Modern Phys, Lanzhou 730000, Peoples R China; [Li Ning; Zhang Hong; Wang Yanling; Hao Jifang] Key Lab Heavy Ion Radiat Med Gansu Prov, Lanzhou 730000, Peoples R China; [Li Ning; Wang Yanling] Chinese Acad Sci, Grad Sch, Beijing 100039, Peoples R China; [Wang Xiaohu] Gansu Tumor Hosp, Dept Radiotherapy, Lanzhou 730050, Peoples R China

抑制survivin表达增强人肝癌HepG2细胞对高线性能量转移射线的辐射敏感性；Inhibiting Survivin Expression Increases The Radiosensitivity of Human Hepatoma HepG2 Cells to High-LET Radiation

探讨了肿瘤细胞中survivin的表达对高线性能量转移(LET)射线辐射敏感性的影响.根据Gen Bank提供的survivin序列,合成特异性survivin-siRNA寡核苷酸,转染人肝癌HepG2细胞,抑制survivin的表达.发现siRNA转染后诱导了HepG2细胞G2/M期阻滞,增加了自发性和辐射诱导的细胞凋亡.在高线性能量转移(LET)碳离子辐照后,siRNA转染细胞的克隆存活率明显下降.这些结果表明survivin表达是HepG2细胞产生对高LET射线辐射抗性的关键因素.