12C和X射线辐照人肺癌细胞H1299的生物学效应

用高传能线密度（LET）的12C离子束和低LET的X射线辐照体外培养的非小细胞肺癌H1299(p53基因缺失), 研究它们的辐照生物学效应的差异。 用克隆形成率法测定了细胞对射线的辐射敏感性; 用AnnexinV/PI试剂盒检测了细胞早期凋亡; 用流式细胞仪检测了细胞周期变化。 实验结果表明, 12C离子束辐照H1299细胞的存活率明显低于用X射线辐照的; 12C离子束引起H1299细胞的早期凋亡率明显高于X射线辐照引起的, 且持续时间更长; 12C离子束引起的H1299细胞G2/M期的抑制更明显。 说明H1299细胞对高LET的12C离子束的辐射敏感性高于对X射线的, 重离子对p53基因缺失型肿瘤的治疗可实施较低的照射剂量、 较少的照射次数和较长的时间间隔。     

和厚朴酚对非小细胞肺癌细胞的辐射增敏效应

研究了和厚朴酚（HNK）对非小细胞肺癌（NSCLC）细胞系A549和H1299对低线性能量转移（LET） X射线和高LET碳离子的辐射增敏效应。首先用CCK-8检测了HNK对A549和H1299细胞的生长抑制情况,发现20 μmol/L的HNK处理对细胞的生长抑制作用较弱。用该浓度HNK预处理细胞2 h后给予不同剂量X射线或碳离子的照射,克隆存活法检测细胞的辐射敏感性,Annexin-PI双染法检测细胞凋亡,γH2AX焦点法检测DNA的双链断裂（DSB）损伤。实验结果显示:与X射线相比,NSCLC细胞对碳离子更敏感,HNK预处理仅对碳离子照射有辐射增敏作用;与碳离子单独照射相比,HNK预处理联合碳离子照射诱导了更明显的细胞凋亡;在照射后24 h,HNK预处理联合碳离子照射引起的细胞γH2AX焦点阳性率维持在较高水平,而X射线照射没有这些效应。实验结果表明,HNK预处理抑制了NSCLC细胞DNA的DSB修复,诱导了细胞凋亡的发生,从而提高了细胞对碳离子的辐射敏感性。The radiosensitizing effect of Honokiol (HNK) on non-small cell lung carcinoma (NSCLC) cell lines A549 and H1299 to low-linear energy transfer (LET) X-rays and high-LET carbon ions was investigated in this study. First, the inhibitory effects of HNK on the growth of A549 and H1299 cells were detected by CCK-8 assay, and 20 μmol/L HNK treatment was found to induce a growth inhibitory effect slightly in these two cell lines. Cells were pre-treated with HNK and then irradiated with X-rays and carbon ions of different doses. Cellular radiosensitivity, apoptosis and DNA damage were analyzed by clonogenic survival, Annexin-PI staining and γH2AX foci, respectively. The results showed the cells were more sensitive to carbon ion irradiation compared to X-rays and the radiosensitization of HNK was only observed after carbon ion irradiation. Furthermore, the co-treatment led to higher apoptosis rate 48 h after irradiation and increased the positive rate of γH2AX foci 24 h after irradiation in A549 and H1299 cells compared with those in the groups treated with carbon ion irradiation alone. These phenomena were not observed after X-ray irradiation. Our data suggest that the pre-treatment with HNK inhibited DNA DSB repair, induced apoptosis and then enhanced the cellular radiosensitivity to carbon ions in NSCLC cells.     

灵芝酸A对人肝癌细胞的辐射增敏效应的初步研究

本研究旨在初步探讨灵芝酸A(GAA)对人肝癌细胞系HepG2在高LET中子和低LET的γ射线条件下的辐射敏感性的影响及差异。研究中,我们用CCK-8方法检测不同浓度GAA对HepG2增殖抑制作用。选取低浓度(5μmol/L)GAA预处理细胞24 h,分别给予不同剂量的中子辐照或γ射线辐照,分别检测克隆存活率、细胞凋亡和γH2AX蛋白的foci的形成。结果表明:在不加GAA的情况下,高LET中子辐射比低LET的γ射线对细胞产生的凋亡比例高;在添加了GAA后,与未加GAA对照组相比,诱导细胞凋亡的比例明显增加;另外,加GAA处理后,细胞增殖抑制率也随着辐照剂量的增加而增高。即GAA能增加HepG2细胞的辐射敏感性,而在同样GAA剂量下,HepG2细胞对高LET中子辐射比低LET的γ射线更敏感。由此,这项研究说明灵芝酸或可开发成为一种天然辐射增敏剂,从而为癌症特别是肝癌的放疗提供新的辅助治疗方法。     

羧甲基-β-1,3-葡聚糖对人肝癌HepG2细胞的放射增敏作用

本研究旨在探讨羧甲基-β-1,3葡聚糖（CMG）对人肝癌HepG2细胞X射线或12C6+离子束辐射敏感性的影响。首先用CCK-8法检测CMG对HepG2细胞的生长抑制情况,得到半数抑制浓度（IC50）为120.6μg/mL。用浓度为0.1×IC50的CMG预处理HepG2细胞24 h,再给予2 Gy X射线或12C6+离子束辐照（CMG+辐照组）;CMG未处理组直接接受2 Gy X射线或12C6+离子束辐照（辐照组）。对比分析辐照组和CMG+辐照组细胞的克隆存活、DNA损伤、凋亡与周期分布、细胞内活性氧（ROS）水平。发现:与X射线辐照组相比,相同剂量的12C6+离子辐照组克隆存活率更小,DNA损伤和周期阻滞更加严重,细胞凋亡率和细胞内ROS水平也更高。与单独X射线或12C6+离子束辐照组相比,CMG+辐照组克隆存活率明显降低,细胞凋亡率随辐照后CMG作用时间的延长而明显增加,CMG使辐照后细胞内ROS维持在一个较高的水平,同时CMG明显加重了单独辐照诱导的DNA损伤和周期阻滞。结果表明,与X射线相比,HepG2细胞对相同剂量的12C6+离子辐射更敏感;CMG可增加HepG2细胞对X射线或12C6+离子辐射的敏感性;CMG可能通过增加受照HepG2细胞内的ROS水平,加剧辐照诱导的DNA损伤,促进辐射诱导细胞凋亡而起到辐射增敏作用。This study aims to investigate the effect of carboxymethy-β-1, 3-glucan (CMG) on the sensitivity of human hepatoma HepG2 cells to X-rays or 12C6+ ions irradiation. First, the inhibitory effect of CMG on the growth of HepG2 cells was detected by CCK-8 assay, and the half maximal inhibitory concentration (IC50) was 120.6 μg/mL. HepG2 cells were pretreated with CMG at a concentration of 0.1×IC50 for 24 h and then irradiated with 2 Gy X-ray or 12C6+ ion beams (CMG + irradiation group). CMG untreated group was directly irradiated by 2 Gy X-rays or 12C6+ ions beam (irradiation group). The clone survival, DNA damage, cell apoptosis, cell cycle distribution, and intracellular reactive oxygen species (ROS) levels in irradiation group and CMG + irradiation group were comparatively analyzed. The results showed that the clone survival rate was lower, DNA damage and cycle arrest were more serious, and the rate of apoptosis and intracellular ROS levels were higher in 12C6+ ions irradiation group than those in the same dose of X-rays irradiation group. Compared with X-rays or 12C6+ ions irradiation group, the clone survival rate of CMG + irradiation group was significantly decreased, and the apoptosis rate significantly increased with the prolongation of CMG treatment post-irradiation; CMG maintained intracellular ROS at a higher level after irradiation, CMG also significantly aggravated radiation-induced DNA damage and cycle arrest. These results indicated that HepG2 cells were more sensitive to 12C6+ ions radiation than those at the same dose of X-rays. CMG increased the sensitivity of HepG2 cells to X-rays or 12C6+ ions irradiation by increasing intracellular ROS level, exacerbating radiation-induced DNA damage and promoting radiation-induced apoptosis in irradiated HepG2 cells.     

Survivin表达在高LET射线诱导细胞凋亡中作用机理的初步研究

先前的研究表明,肿瘤细胞中survivin的高表达与细胞对高传能线密度(LET)射线的辐射抗性相关。研究了survivin表达在高LET射线诱导的细胞凋亡中的作用,发现抑制survivin表达对高LETC离子辐射诱导的Bcl-2和Bax表达没有明显的影响。在高LET射线辐照中,survivin可能通过抑制caspase-3和-9活性的途径,抑制了细胞凋亡。     

辐照诱导人正常肝细胞系HL-7702 细胞远后的微卫星不稳定性

采用高传能线密度(LET) 的12C6+离子束和低LET 的X 射线辐照人正常肝细胞系HL-7702 细胞,利用微卫星不稳定性(MSI) 检测来分析直接受照射细胞和通过转移培养基方式旁细胞传代八代子细胞以MSI 表征的远后效应。实验结果表明,12C6+离子束诱导的远后效应较X射线的低;旁细胞的远后效应较直接受照射细胞的高;辐射引起的MSI 与杂合性丢失(LOH) 的发生率具有位点特异性。结果提示,重离子放射治疗较X 射线放射治疗对正常组织引发的辐射风险要小,可通过对MSI 高发位点的筛选来评估放疗后患者长期生存状况和二次癌症发生风险。Human normal liver cell line HL-7702 cells were irradiated with high linear energy transfer (LET) 12C6+ ions and low-LET X-rays, respectively. Delayed effect in terms of microsatellite instability (MSI) in progenies of the directly irradiated cells and bystander cells, obtained in the way of medium transfer at the 8th passage postirradiation,were examined. The delayed effect induced by the high-LET 12C6+ ions was different from that induced by the low-LET X-rays, and a higher incidence of MSI was observed in the progenies of the cells after exposure to the X-rays than to the 12C6+ ions. We also found that the delayed effect in the progenies of the bystander cells was much more severe than thoseof directly irradiated cells. Furthermore, the events of MSI and loss of heterozygosity (LOH) induced by the ionizing radiations were not randomly distributed throughout the genome and specific loci existed indeed. These results imply that the radiation risk to normal tissues is lower in heavy ion therapy than in conventional X-ray radiotherapy, and the analysis of microsatellite loci with MSI high frequency occurrence can be applied to access long-term survival condition and second cancer risk for the patients after radiotherapy.     

Survivin表达在高LET射线诱导细胞凋亡中

先前的研究表明, 肿瘤细胞中survivin的高表达与细胞对高传能线密度(LET)射线的辐射抗性相关。 研究了survivin表达在高LET射线诱导的细胞凋亡中的作用, 发现抑制survivin表达对高LET C离子辐射诱导的Bcl-2和Bax表达没有明显的影响。 在高LET射线辐照中, survivin可能通过抑制caspase-3和-9活性的途径, 抑制了细胞凋亡。It has been proven that over expression of survivin in cancerous cell lines is related to the radioresistance of cells to high LET radiation in previous work. In this study, action mechanisms of survivin gene in apoptosis induced by high LET radiation were investigated. We found that inhibiting survivin by siRNA had no notable influence on Bcl-2 and Bax expressions induced by carbon ions. Survivin depressed cell apoptosis through the inhibition of the activities of caspase-3 and -9 possibly in cell apoptosis induced by high LET radiation.     

重离子诱导细胞衰老研究

综述了细胞衰老诱导因素和衰老细胞的特征,提出细胞衰老在癌症发生发展过程中具有双重作用效应。同时,以X射线和12C6+离子束对黑色素瘤细胞系92-1或人正常成纤维细胞系MRC5进行辐照,观察分析DNA损伤应答效应和细胞衰老诱导。实验结果表明,相对于X射线,12C6+离子束能导致DNA团簇损伤,持续激活DNA损伤应答,更容易诱导细胞衰老;12C6+离子束能同时诱导癌细胞和正常细胞衰老。这些实验结果暗示开展12C6+离子束诱导细胞衰老研究具有紧迫性。     

辐照诱导的人正常肝细胞系HL-7702细胞延迟效应

利用X射线辐照人正常肝细胞系HL-7702细胞, 运用胞质分离阻滞微核法实验检测细胞微核率,AnnexinV FITC细胞凋亡检测试剂盒检测细胞凋亡率, 细胞微核率和凋亡率随着辐照剂量的增加而显著增加。X射线照射后细胞传代培养, 第7代时不同剂量辐照后子代细胞微核率和凋亡率同未辐照细胞相比已无明显区别。 对不同剂量辐照后传代7代的细胞再次照射2.5 Gy的相同剂量,发现它们细胞微核率和凋亡率存在明显差异,即初次受辐照剂量高的细胞, 再次以相同剂量辐照后的微核率和凋亡率也高. 这些结果表明,X射线辐照导致了HL-7702细胞基因组不稳定性这一辐射延迟效应,再次辐照使得辐射的延迟效应得以明显的表现。 Human normal liver cell line HL-7702 cells were irradiated with different doses of X rays. Micronucleus and apoptosis rates in the irradiated cells were measured with cytokinesis block micronucleus method and Annexin V FITC apoptosis detection kit, respectively. Experimental data showed that the micronucleus and apoptosis rates increased obviously with increasing irradiation dose. After seven population doublings, the micronucleus and apoptosis rates of the cells surviving exposure to the X rays reduced to the same levels as non irradiated control cells; the progenies of the cells were secondly exposed to X rays at the same dose of 2.5 Gy. We found that the progenies of the cells surviving the first irradiations of the various doses showed markedly differential micronucleus frequencies and apoptotic rates. Although the same dose of 2.5 Gy was applied in the second irradiations, the micronucleus frequencies and apoptotic rates of the progenies of the cells initially exposed at higher doses were significantly higher than the others. These results indicate that X rays lead to genomic instability in HL 7702 cells, which is an important manifestation of radiation induced delayed effect, and a second radiation stimulus makes the delayed effect in the progeny of the previously irradiated cells be expressed obviously.     

12C6+离子预辐射对小鼠胸腺脾脏细胞周期进程的影响

以0, 0.05, 0.1, 0.25, 0.5 Gy 12C6+ 离子全身预辐照昆明小鼠, 间隔4 h后再对小鼠进行4 Gy全身辐射。 辐照后12 h用流式细胞仪检测小鼠胸腺脾脏细胞在各细胞周期时相的百分率, 同时用单细胞电泳检测受辐射小鼠胸腺脾脏细胞DNA损伤程度。 结果显示, 相对于大剂量预照射组, 各低剂量预照射组胸腺细胞S期细胞百分率显著减少; 脾脏细胞G0/G1期细胞百分率明显减少; 同时胸腺脾脏细胞的拖尾率及拖尾长度明显减少, 以0.1 Gy预辐照效果最为明显。 这些结果表明, 低剂量预辐射处理可以减弱胸腺细胞的S期阻滞及脾脏细胞的G1期阻滞, 并明显减轻胸腺脾脏细胞的DNA损伤程度。     

GANRA-5对X射线和~(12)C~(6+)离子辐射防护作用研究

研究了一种新的恶唑酮类衍生物GANRA-5对于人胚肺细胞MRC-5的辐射防护作用。以MTT评价其对于细胞的毒性,以γH2AX foci形成法检测其对于辐照后细胞中双链断裂的影响,发现其对于受到X射线和12C6+离子照射的细胞具有较强的辐射防护作用,并进一步发现其能够显著清除辐照后细胞内的自由基。这些结果表明,GANRA-5具有较低的细胞毒性,并能够通过清除自由基发挥较强的针对X射线和12C6+离子的辐射防护作用,有望开发为高效的辐射防护药物。     

氧化钆纳米粒子对A549肺癌细胞的辐射增敏效应研究

采用氧化钆纳米粒子（GON）,研究钆基纳米粒子对X射线和碳离子束的辐射增敏效应。首先,通过透射电镜观察材料粒径,使用DLS检测材料的水合半径及Zeta电位,并用紫外吸收谱证实GON在培养基中稳定性较好;研究发现钆（Gd）浓度为10.0 μg/mL的GON对30 keV/μm碳离子束辐照水溶液产生的羟自由基的增强系数为1.13;GON对A549肺癌细胞和正常MRC-5肺细胞没有明显的毒性,且在人肺癌A549细胞中的摄取量随共培养浓度的增加而增加,在10.0 μg/mL共培养浓度下,细胞摄入Gd的量为0.73 pg/cell;进一步采用克隆存活实验证明,GON的加入对X射线和碳离子辐照A549细胞所产生的损伤具有明显的增强,在10%的细胞存活水平下,GON对A549细胞在X射线及碳离子辐照下的辐射增敏分别达15.5%和10.1%。鉴于钆材料常被用于磁共振成像（MRI）,所获得的GON有望作为X射线和碳离子的诊疗一体化材料。     

X射线致HeLa细胞损伤过程中NADPH氧化酶的作斥

以HeLa细胞为实验材料,探讨了NADPH氧化酶在X射线诱导细胞损伤过程中的作用。结果显示,12Cry X射线辐照后细胞内活性氧（ROS）明显增加,在用NADPH氧化酶抑制剂处理后再辐照,则细胞内ROS降低到未辐照水平;同时辐照后NADPH氧化酶细胞质亚基p47^phox在细胞质积聚并和细胞膜亚基gp91^phox结合;Western blotting检测结果显示,NADPH氧化酶的关键亚基gp91^phox的表达量明显增加。以上结果说明,NADPH氧化酶可以被X射线激活,由其介导产生的ROS在X射线诱导HeLa细胞损伤过程中扮演重要角色。     

两种保护剂对C离子诱导小鼠急性肝损伤的应答

比较了N-乙酰半胱氨酸(NAC)及乙酰左旋肉毒碱(ALCAR)对12C6+离子照射小鼠的损伤效应,并探讨了其可能的作用机制。利用4Gy剂量的12C6+离子束对预先给予NAC(100mg/kg)和ALCAR(100mg/kg)保护的昆明小鼠进行单次全身照射。随后检测肝组织中总抗氧化能力(TAC)、DNA单链断裂和细胞凋亡率。结果显示,与照射对照组相比,提前给予NAC和ALCAR均极显著地增强了肝组织的抗氧化能力(P0.001),减轻了12C6+离子导致的肝组织中DNA断裂(P0.001)和细胞凋亡(P0.001)。此外,还发现ALCAR组抗重离子辐照损伤的能力显著地高于NAC组(P0.05)。实验结果提示了NAC和ALCAR可通过抵御组织内的氧化胁迫,阻止DNA链的断裂和细胞的凋亡,实现对C离子辐照损伤的保护效应。而且ALCAR比NAC可能更适合成为有潜力、有希望的抗C重离子辐射药物。     

利用拉曼光谱技术研究低剂量X射线辐射对人神经母细胞瘤细胞的影响

低剂量电离辐射引发的生物效应复杂而多样,其研究往往又受到辐射标志物和检测技术手段的限制。将拉曼光谱技术应用于低剂量辐射生物效应研究,利用10 mW,532 nm共聚焦拉曼光谱对经过100,200和500 mGy三种辐射剂量的X射线辐照之后的人神经母细胞瘤细胞进行检测,发现细胞嘌呤核苷酸（722～728和1 572～1 581 cm-1等等）、嘧啶核苷酸（770～785 cm-1等等）等DNA相关的拉曼特征峰受到电离辐射影响而发生变化,说明低剂量X射线辐照造成细胞DNA水平改变。采用流式细胞术对同样条件辐照后培养6 h的人神经母细胞瘤细胞进行细胞周期分析发现,三种剂量的X射线电离辐射均造成细胞在G2期阻滞,同样提示电离辐射引起DNA水平升高。通过划痕实验分析辐照后20 h的细胞迁移能力,结果显示,相较于未接受X射线照射的对照细胞,受到三种剂量电离辐射的人神经母细胞瘤细胞均出现迁移水平下降。研究结果表明,通过拉曼光谱分析发现低剂量X射线电离辐射引起人神经母细胞瘤细胞DNA水平变化,其结果与细胞周期分析和迁移分析的结果相一致,但检测时间大大提前,利用拉曼光谱技术可以实现低剂量辐射损伤等细胞生物学效应的早期发现与监测。     

人肝癌SMMC-7721细胞对低剂量γ射线辐射超敏感性的研究

研究了人肝癌细胞SMMC-7721在低剂量γ射线照射下超敏感性和增强的辐射抗性响应。选用对数生长期细胞接受0—6 Gy不同剂量的^60Coγ射线的照射。利用流式细胞仪对细胞进行分选计数,并用克隆形成法检测细胞存活率。发现SMMC-7721细胞存在低剂量辐射超敏感性和增强的辐射抗性响应,即在0—0．3Gy之间细胞表现出单位剂量杀伤增强现象,在0．3—1Gy细胞表现一定的辐射抗性,在1Gy以上,细胞的存活符合线性平方模型。     

5—200范围激光等离子体X射线辐射特性研究

利用带有针孔的透射式光栅光谱仪研究了激光等离子体X射线辐射的原子序数依赖性和激光功率密度对辐射的影响。得到了波长为1.06μm,平均功率密度为5×10~(14)W/cm~2的激光辐照条件下Z=6(C)到Z=79(Au)的不同原子序数激光等离子体X射线发射光谱。点聚焦和线聚焦激光照射方式下Al,Au等离子体X射线发射的对照实验结果表明,激光功率密度对低Z等离子体X射线发射的影响比对高Z的影响更明显。     

~(12)C~(6+)离子束辐照紫苏干种子当代效应

利用兰州重离子研究装置(HIRFL)提供的12C6+离子束辐照紫苏干种子(辐照剂量为40,80和120Gy,剂量率4Gy/min),探讨了重离子束辐照对紫苏M1代的生物学效应。结果发现,经不同剂量的12C6+离子束辐照后,紫苏种子的发芽率、发芽势、存活率、株高、分枝数、单株产量和千粒重等生物学性状均发生了变化,其中发芽势、单株产量和千粒重随辐照剂量的提高而降低,且有明显的剂量效应关系,但发芽率、大田成活率、株高和分枝数却随辐照剂量的增大,呈现出明显的"抛物线"趋势;紫苏幼苗根尖细胞的微核率和染色体畸变率随辐照剂量增加呈线性增加关系。这表明:12C6+重离子束辐照紫苏种子,具有明显的当代损伤效应,在本试验剂量范围内,低剂量辐照对发芽率和成活率有促进作用。     

重离子辐照玉米种子M1代诱变效应研究

用重离子^7Li和^12C以及^60Co的γ射线分别辐照玉米种子胚，研究重离子束辐照玉米种子M1代的生物学效应．结果表明:^60Co的丫射线使种子的发芽势和发芽率降低，重离子^7Li和^12C对发芽势和发芽率的抑制作用不明显；重离子辐照后，单株之间光合速率差异增大；M1代植株在雄性育性、叶片颜色及株高等方面发生了多种变化，并产生了一株双胚苗．应用随机多态性DNA分析技术，发现双胚苗的大、小苗之间及其与亲本自交系478在随机多态性DNA指纹上存在的差异，从分子水平上初步证实了双胚苗为自交系478的变异类型．In order to study M_(1) biological effects of heavy ion irradiation on maize seeds, the embryos of dry maize seeds are irradiated with~(7)Li ions and~(12)C ions as well as ~( 60)Co gamma rays. The results are as follows: ~( 60)Co gamma rays decrease germination impetus and germination rate of maize seeds, while the inhibitory effects of germination impetus and germination rate induced by the heavy ions~(7)Li and~(12)C are not significant. The difference of photosynthetic rates among the plants irradiate...     

5—200?范围激光等离子体X射线辐射特性研究

利用带有针孔的透射式光栅光谱仪研究了激光等离子体X射线辐射的原子序数依赖性和激光功率密度对辐射的影响。得到了波长为1.06μm,平均功率密度为5×10¹⁴W/cm²的激光辐照条件下Z=6(C)到Z=79(Au)的不同原子序数激光等离子体X射线发射光谱。点聚焦和线聚焦激光照射方式下Al,Au等离子体X射线发射的对照实验结果表明,激光功率密度对低Z等离子体X射线发射的影响比对高Z的影响更明显。 关键词：