Ku蛋白表达与鼻咽癌细胞放射敏感性 关系初探

目的探讨不同放射敏感性鼻咽癌细胞株CNE-1（鼻咽高分化鳞癌细胞株）、CNE-2（鼻咽低分化鳞癌细胞株）中DNA依赖蛋白激酶（DNA—PK）的调节亚基Ku70、Ku80基因的表达与鼻咽癌细胞放射敏感性的关系。方法通过克隆形成实验测定CNE-1、CNE-2不同剂量的存活分数，并用线性二次模型拟合剂量存活曲线求出放射生物学参数α、β、SF2、MID值，以及四氮唑蓝比色分析法（MTT法）检测^60Co γ射线4Gy照射后的细胞群体生长情况，以评价两株细胞的放射敏感性。逆转录实时荧光定量PCR技术（RT-FQ-PCR）检测照射前、后不同时间及不同剂量CNE-1、CNE-2细胞mRNA水平Ku70、Ku80基因的定量表达。结果CNE-1在各个剂量点的存活分数均比CNE-2高，MID值分别为2．78、1．61，SF2值分别为0．627、0．341；4Gy照射后的存活率分别为88．2％、72．3％；RT-FQ-PCR显示两株细胞中均有Ku70、Ku80基因的表达，其相对表达量之比分别为1．76±0．54（P=0．07）、13．82±3．78（P=0．004），Ku80基因在CNE-2细胞中存在剂量依赖关系。结论实验验证了CNE-2比CNE-1对射线更敏感，Ku80基因的表达与鼻咽癌细胞的放射敏感性有关。     

Ku80表达抑制细胞模型的建立及其功能检测

目的 建立利用RNAi技术抑制Ku80表达的HeLa细胞模型，探讨Ku80在放射生物学方面的功能。方法构建靶向抑制Ku80的siRNA表达质粒，转染HeLa细胞，筛选稳定表达的转化克隆；Westernblotting检测Ku80表达变化；6MVX线照射6Gy后，流式细胞术检测细胞凋亡率和细胞周期；克隆形成实验检测细胞SF2、D0等值。结果构建的质粒转染HeLa细胞获得3个稳定转染克隆，Westernblotting分析表明2个阳性克隆Ku80蛋白抑制率分别达到89．3％和96．4％；Ku80表达抑制细胞受x线照射后48及72h的凋亡率高于对照细胞（P〈0．05），但4株细胞的细胞周期变化无统计学意义（P〉0．05）；2个阳性克隆细胞株的D0和SF2值明显降低，在D10剂量时的增敏比分别为1．315和1．365。结论运用RNAi技术建立的Ku80表达抑制克隆细胞可以成为简单实用的细胞模型；Ku80-siRNA可以抑制Ku80的表达，从而促进HeLa细胞对X线的敏感性。     

辐射联合外源性野生型p53基因对不同p53状态的黑色素瘤细胞系的生物学效应

目的研究辐射结合腺病毒（Ad CMV）载体介导的p53基因转导对不同p53状态的人黑色素瘤细胞系基因转移效率、凋亡和辐射敏感性的影响。方法用复制缺陷的重组腺病毒载体（AdCMV-p53）介导入p53基因转导1Gy X射线预照射的黑色素瘤细胞系A375（wt p53）和WM983a（mu p53），RT-PCR检测mRNA水平，流式细胞仪测定细胞周期阻滞及外源性P53蛋白表达情况，Tunel法检测细胞凋亡，克隆形成率测定辐射后细胞存活率。用携带报道基因的复制缺陷重组腺病毒载体AdCMV-GFP作为对照。结果1Gy X射线照射可较高地增加AdCMV-p53对A375和WM983a细胞系的基因转导效率，转导的外源性野生型p53可在两种细胞中高效表达，并诱导细胞周期G1期阻滞；单纯转导p53对A375（wt p53）细胞无明显诱导凋亡和生长抑制效应，但可部分诱导WM983a（mu p53）细胞凋亡；而转导p53基因48h后给予X射线辐射，两种细胞的克隆存活率较其对照组均明显减低，外源性p53基因对WM983a（mu p53）细胞的辐射增敏作用较A375（wt p53）细胞明显。结论外源性野生型p53基因过表达可增加黑色素瘤细胞系A375和WM983a的辐射敏感性，但对WM983a细胞系的辐射增敏作用高于A375细胞系。表明p53是基因治疗黑色素瘤较好的候选基因。     

p21在电离辐射诱导EL-4细胞G_1期阻滞中的作用

目的　探讨 p2 1在电离辐射诱导EL 4细胞G1期阻滞中的作用。方法　采用North ernblot检测 p2 1WAF1mRNA水平的变化 ;采用流式细胞术检测 p2 1蛋白表达及细胞周期的变化。结果　 4 0GyX射线照射后 12h、2 4h、48hG1期EL 4细胞百分数明显高于假照射组 ;p2 1WAF1mRNA水平从照射后 1h开始升高 ,4h达峰值 ,持续至照后 12h ;p2 1蛋白表达在照射后 2～ 48h明显增高。结论　 4 0GyX射线照射可诱导EL 4细胞G1期阻滞 ,p2 1在电离辐射诱导EL 4细胞G1期阻滞中起重要作用。     

60Coγ射线对大鼠星形胶质瘤细胞增殖抑制作用的机理研究

目的 研究^60Coγ射线对大鼠星形胶质瘤细胞的增殖抑制作用的机制。方法 ^60Coγ射线单次照射，分为对照组、4、16和64Gy组。四甲基偶氮唑蓝（MTT）法测定细胞生长曲线。照射后48h采用流式细胞仪检测细胞凋亡和细胞周期分布，免疫组化法检测P53及P21蛋白的表达。结果 16和64G主y组C6细胞出现明显的增殖抑制。随着吸收剂量的增大，凋亡比例显著增加（P〈0．01），G0／G1期的细胞比例增加，S期的细胞比例降低。野生型p53与p21随着照射剂量增加，表达增强。P53与P21蛋白表达强度呈正相关，相关系数斜率为0．889。结论 γ射线对大鼠星形胶质瘤细胞系C6细胞的增殖抑制通过诱导细胞凋亡和G1期阻滞实现，G1期阻滞的分子调控通路可能为p53-p21通路。     

西妥昔单抗联合照射对结直肠癌CL187细胞的抑制作用及机制探讨

目的 研究西妥昔单抗(C225)联合外照射对结直肠癌CL187细胞生物学效应的影响,并探讨相关分子机制。方法 结直肠癌CL187细胞分单纯照射组和C225处理的联合照射组,受6 MV X 射线照射0、4和8 Gy后24和48 h,用四甲基偶氮唑盐(MTT)比色法测吸光度(A)值,比较两组细胞死亡率的差异。利用克隆形成实验比较两组细胞增殖能力的差异。采用流式细胞仪,检测细胞周期和细胞凋亡。蛋白免疫印迹法分析两组细胞DNA-PKcs、Ku70和Ku80蛋白表达量的变化。结果 联合照射组较单纯照射组死亡细胞比例增加(t=-6.14、-6.53,P<0.05),细胞克隆形成能力下降。C225增强了射线对细胞的杀伤作用,放射增敏比SER为1.38。联合照射组G₀/G₁期细胞阻滞增加(t=-4.64, P<0.05),细胞凋亡比例增加(t=-9.16, P<0.05),DNA修复相关蛋白DNA-PKcs、Ku70和Ku80蛋白表达量减少。结论 西妥昔单抗增强照射对结直肠癌CL187细胞的杀伤作用,可能是通过影响细胞周期、细胞凋亡和DNA损伤修复基因实现的。     

二氢青蒿素对人宫颈癌细胞株照射后周期的影响及相关机制

目的 观察二氢青蒿素及X射线对肿瘤细胞周期的影响,并研究其具体作用机制。方法 选用已知p53突变的人宫颈癌HeLa细胞,并以p53功能正常的人宫颈癌SiHa细胞作为对照。采用流式细胞术分析X射线(6 Gy)、二氢青蒿素(20及100 μmol/L)对两种细胞的细胞周期的影响;应用蛋白印迹法(Western blot)检测细胞周期相关蛋白表达量的变化。结果 X射线照射明显导致HeLa细胞G₂期阻滞,照射后G₂期细胞比例由14.45%上升至73.58%,在二氢青蒿素联合照射作用后,HeLa细胞G₂期细胞比例由单纯照射组的73.58%降至48.31%;而对照组SiHa细胞G₂期变化不明显。在单纯照射组,随着细胞G₂期阻滞的增加,HeLa细胞中Wee1蛋白表达量增加,Cyclin B1蛋白表达量降低,而在二氢青蒿素联合照射作用后,细胞内Wee1蛋白表达量较单纯照射组减少,Cyclin B1蛋白表达量较单纯照射组增高,与该药能去除电离辐射导致细胞G₂期阻滞过程相一致。结论 对于p53突变的人宫颈癌HeLa细胞,二氢青蒿素能抑制辐射所引起的细胞G₂期阻滞,其机理可能与细胞周期调控蛋白Wee1、Cyclin B1表达变化有关;对p53功能正常的SiHa细胞,辐射主要引起细胞G₁期阻滞,故二氢青蒿素对其周期的影响作用不明显。     

电离辐射诱导的细胞G2期阻滞

哺乳动物受X射线照射后,可以使细胞周期延迟或阻滞,包括G1期阻滞、S期延迟和G2期阻滞。G1期阻滞仅在野生型p53基因存在时出现,在清除DNA损伤的细胞中具有重要作用;而G2期阻滞更有利于损伤后DNA的修复和细胞存活,并且与p53基因存在状态无关。因此,对电离辐射诱导细胞G2期阻滞机制的探讨成为近年来国内外放射生物学领域的研究热点。     

腺病毒介导p53基因转染增加人胃癌细胞的放射敏感性

目的评价腺病毒介导p53基因(Adp53)转染对人胃癌细胞的凋亡效应和放射增敏作用。方法以重组腺病毒介导p53基因感染4种不同p53状况的人胃癌细胞，用免疫组织化学法和Western blot法检测P53蛋白在胃癌细胞中的表达；用细胞集落形成法检测细胞存活率；用TUNEL法检测细胞凋亡。胃癌细胞感染Adp53后照射4Gy，用流式细胞仪检测细胞周期分布和凋亡；胃癌细胞种植肿瘤内注射Adp53后照射6Gy，以肿瘤相对体积增长曲线观察肿瘤抑制情况。结果1：100效靶比(MOI)Adp53产生细胞高转染率，以及p53基因在4种胃癌细胞中均高表达，并产生G2/M期阻滞、凋亡增加和细胞增殖抑制。如果以凋亡评价放射效应，Adp53转染对4Gy照射4种细胞的凋亡率比值为：W细胞3．0，M细胞3．6，neo细胞2．2，823细胞2．5。体内实验结果显示，Adp53对w细胞肿瘤6Gy照射的抑瘤率比值为1．41，而对M细胞肿瘤为1．91。结论腺病毒介导p53基因转染产生细胞凋亡并提高人胃癌细胞的放射敏感性，这种作用不依赖于细胞内在的p53状况。     

X射线全身照射诱导小鼠派伊氏板细胞凋亡及其机理的研究

目的 研究不同剂量X射线全身照射后小鼠派伊氏板细胞凋亡相关基因蛋白表达的变化规律。初步阐明凋亡调控的部分分子机理。方法 应用光、电镜技术观察小鼠派伊氏板细胞凋亡的形态改变,并用流式细胞术检测小鼠派伊氏板细胞Bcl-xL及Fas-L蛋白表达的变化。结果 2GyX射线全身照射后派伊氏板细胞凋亡增加,且Bcl-xL蛋白表达下降,Fas-L蛋白表达升高;75mGyX射线全身照射后,派伊氏板细胞凋亡减少,且Bcl-xL蛋白表达升高,Fas-L蛋白表达下降。结论 凋亡相关基因Bcl-xL、Fas-L在电离辐射诱导的派伊氏板细胞凋亡方面起重要的调控作用。     

Threshold effect for teratogenic risk of radiation depends on dose-rate and p53-dependent apoptosis

PURPOSE: To obtain evidence that the p53 gene is indispensable for reduction of high teratogenic risk of radiation at a high dose-rate to zero risk by lowering the dose-rate. MATERIALS AND METHODS: Wild-type p53(+/+), heterozygous p53(+/-) and null p53(-/-) mice were exposed to gamma-rays at high or low dose-rates during days 9.5-10.5 of gestation. The incidence of malformations and prenatal deaths was studied. Frequencies of cells dying by apoptosis were measured during or after protracted irradiation. RESULTS: After irradiation with 2 Gy, the frequency of apoptotic cells increased to 20% for p53(+/+) mice and did not increase at all for p53(-/-) mice. For p53(+/+) mice, 2 Gy y-rays induced 70% malformations when given at 1.06 Gy/min, but no malformations above the control when given at 1.2 mGy/min. In contrast, after irradiation of p53(-/-) foetuses with 2 Gy at 1.2mGy/min, the incidence of malformations increased 12% above control levels. CONCLUSION: Foetal irradiation with 2 Gy at 1.2 mGy/min was not teratogenic for p53(+/+) mice but teratogenic for p53(-/-) mice. This indicates that the p53 gene is indispensable for a threshold effect in the risk of radiation at low doses or dose-rates.     

X射线诱导非小细胞肺癌A549细胞凋亡的适应性反应及相关miRNA筛选的研究

目的 研究X射线辐射诱导非小细胞肺癌（NSCLC）A549细胞凋亡的适应性反应,并筛选适应性反应相关的微RNA（miRNA）。 方法 将NSCLC A549细胞分为6组,包括 50 mGy+20 Gy、200 mGy+20 Gy、20 Gy、50 mGy、200 mGy照射组及对照组（0 Gy）,前2组细胞分别用50、200 mGy初始剂量进行照射,培养6 h后用20 Gy的效应剂量进行照射,20 Gy、50 mGy、200 mGy照射组同时进行照射。培养24 h后使用流式细胞仪检测细胞凋亡情况。利用小RNA测序技术筛选差异表达miRNA,并对其靶基因进行基因本体（GO）及京都基因与基因组百科全书（KEGG）通路的功能富集分析。采用实时荧光定量PCR（qRT-PCR）对部分差异表达miRNA进行验证。2组间数据的比较采用Welch t检验。 结果 50 mGy+20 Gy照射组和200 mGy+20 Gy照射组的A549细胞早期凋亡率分别为（1.81±0.11）%和（2.17±0.19）%,低于20 Gy照射组的（4.54±0.23）%,且差异均有统计学意义（t=10.680、8.006,均P<0.01）。与20 Gy照射组相比,50 mGy+20 Gy照射组和200 mGy+20 Gy照射组共同差异表达趋势miRNA有1个上调（miR-3662）、15个下调（miR-185-3p、miR-1908-5p、miR-1307-5p、miR-182-3p、miR-92a-3p、miR-582-5p、miR-501-3p、miR-138-5P、miR-1260b、miR-484、miR-378d、miR-193b-3P、miR-127-3p、miR-1303及miR-654-5p）。GO富集分析结果显示,差异表达miRNA调控靶基因功能显著富集于细胞通讯调节、代谢过程的正向调节、代谢信号的调节、酶结合及催化活性等过程。KEGG富集分析结果显示,靶基因相关信号通路显著富集于溶酶体、丝裂原活化蛋白激酶、Ras和内吞作用等信号通路。qRT-PCR检测结果显示,miRNA表达情况与基因芯片结果趋势一致（10个miRNA表达水平得到验证）。 结论 X射线50、200 mGy照射剂量均能诱导NSCLC A549细胞凋亡的适应性反应,并筛选到一组共同差异表达的miRNA,可能在X射线辐射诱导细胞凋亡的适应性反应中发挥了重要作用,有可能成为调节电离辐射生物效应的潜在靶点。     

Lentivirus-mediated RNA interference of Ku70 to enhance radiosensitivity of human mammary epithelial cells

Purpose:?To investigate the radiosensitising effect of Ku autoantigen 70 (Ku70) and Ku autoantigen 80 (Ku80) knockdown by lentivirus-mediated RNA interference (RNAi) in the MCF10A immortalised human mammary epithelial cell line.

Materials and methods:?MCF10A cells were infected with lentiviral vectors for RNAi of Ku70. The Ku70-knockdown cell line (Ku70i) and a mock-infected control cell line (LVTHM) were used to perform radiation experiments. For the in?vitro Micronucleus (MN) assay, both cell lines were irradiated with doses of 2 and 4 Gy ⁶⁰Co γ-rays. For cell survival experiments, doses ranging between 0 and 8 Gy were used.

Results:?Western blot analysis showed that the Ku70 lentiviral vector was effective in silencing the expression of both Ku70 and Ku80. A significantly higher radiation-induced MN yield was obtained in the Ku70i cell line compared to the control LVTHM cell line. RNAi of Ku70 also resulted in a lower survival yield after irradiation compared to the control cell line. Analysis of cell death mechanisms showed that MCF10A cells (Ku70i and LVTHM) do not undergo apoptosis, but undergo post-irradiation cellular senescence.

Conclusion:?RNAi of Ku70 resulted in increased chromosomal and cellular radiosensitivity in the MCF10A human mammary cell line after irradiation with ⁶⁰Co γ-rays. These results further strengthen the role of the Ku protein in correct DNA double strand break (DSB) repair.     

High sensitivity of rat foetal germ cells to low dose-rate irradiation

Purpose : To investigate the response of germ and Sertoli cells to γ-irradiation at two distinct periods of testicular development in rat foetuses. Materials and methods : Pregnant rats were exposed to 60 Co γ-rays at days 15, 19 or 21 post-coitum (p.c.), at doses ranging from 0.1 to 1.5Gy, and at different dose-rates. Testicular weight, seminiferous tubule condition and the number of germ and Sertoli cells were measured at early and late times after irradiation. Apoptosis was studied by the ISEL method and p53 expression was studied by immunohistochemistry Results : At high dose-rates (≥3.3 Gy min -1) , 1.5Gy radiation at day 15 p.c. had a short-term effect on germ cell survival. A large proportion of these cells rapidly underwent p53-independent apoptosis. Apoptotic cells were strongly clustered. The remaining germ cells divided and differentiated normally leading to a majority of normal tubules in the adult testis. However, at low dose-rate (0.6 mGy min -1) , much greater depopulation of the seminiferous tubules occurred. When irradiation was given at day 19 p.c., the same dose had a delayed effect on germ cells, leading to sterility. Sertoli cells had a normal survival for irradiation at day 15 p.c. Their proliferation became higher in prepubescent testis compared with controls, when irradiation occurred at day 19 p.c. Conclusion : The position of gonocytes in the cell cycle at the time of irradiation seems to be a determining parameter for inducing gonocyte apoptosis. The strong effect of irradiation on germ cells at very low dose-rate and the appearance of clusters of apoptotic gonocytes may be a consequence of the syncitial organization of germ cells, favouring their cell synchronisation or the transmission of death signalling when they are in a radiosensitive period.     

High sensitivity of rat foetal germ cells to low dose-rate irradiation

PURPOSE: To investigate the response of germ and Sertoli cells to gamma-irradiation at two distinct periods of testicular development in rat foetuses. MATERIALS AND METHODS: Pregnant rats were exposed to 60Co gamma-rays at days 15, 19 or 21 post-coitum (p.c.), at doses ranging from 0.1 to 1.5Gy, and at different dose-rates. Testicular weight, seminiferous tubule condition and the number of germ and Sertoli cells were measured at early and late times after irradiation. Apoptosis was studied by the ISEL method and p53 expression was studied by immunohistochemistry. RESULTS: At high dose-rates (> or = 3.3 Gy min(-1)), 1.5 Gy radiation at day 15 p.c. had a short-term effect on germ cell survival. A large proportion of these cells rapidly underwent p53-independent apoptosis. Apoptotic cells were strongly clustered. The remaining germ cells divided and differentiated normally leading to a majority of normal tubules in the adult testis. However, at low dose-rate (0.6mGy min(-1)), much greater depopulation of the seminiferous tubules occurred. When irradiation was given at day 19 p.c., the same dose had a delayed effect on germ cells, leading to sterility. Sertoli cells had a normal survival for irradiation at day 15 p.c. Their proliferation became higher in prepubescent testis compared with controls, when irradiation occurred at day 19 p.c. CONCLUSION: The position of gonocytes in the cell cycle at the time of irradiation seems to be a determining parameter for inducing gonocyte apoptosis. The strong effect of irradiation on germ cells at very low dose-rate and the appearance of clusters of apoptotic gonocytes may be a consequence of the syncitial organization of germ cells, favouring their cell synchronisation or the transmission of death signalling when they are in a radiosensitive period.     

Radioresistance in a tumour cell line correlates with radiation inducible Ku 70/80 end-binding activity

Purpose: The aims of the present study were to better understand the role of Ku 80, which is involved in double-strand break repair in mammalian cells in the mechanism of radiation resistance and to verify the possibility of increasing cell radiosensitivity by targeted inhibition of Ku autoantigen 80 (Ku 80).

Materials and methods: Western blot and electrophoretic mobility shift assay (EMSA) were performed on the human bladder carcinoma cell line RT112 (radioresistant) and on the human colorectal carcinoma cell line SW48 (radiosensitive) to assess the expression levels of DNA-dependent protein kinase (DNA-PK) components and the DNA-binding activity of the Ku 70/80 heterodimer after exposure to radiation, respectively. Ku 80 silencing was carried out with the use of small interfering RNA (siRNA).

Results: Greater differences in the DNA-binding activity of Ku 70/80 and Ku 80 phosphorylation level were observed in RT112 as compared to SW48 after X-ray treatment. There is no correlation between Ku expression and DNA-binding activity at lower doses. A significant increase in nuclear Ku 80 expression was observed one hour after the exposure, only at the higher doses, while the DNA-PK catalytic subunits (DNA-PKcs) and Ku 70 levels did not change significantly. Inhibition of Ku 80 expression by siRNA induced radiosensitivity in the RT112 cell line.

Conclusions: Our data demonstrate that in a bladder tumour cell line up-regulation of Ku end-binding activity without any marked change in Ku expression underlie radiation resistance.     

不同剂量X射线对同步化HeLaS3细胞周期的影响

目的　研究不同剂量Ｘ射线对同步化ＨｅＬａＳ３ 细胞周期的影响,为进一步探讨其分子调控机理和临床肿瘤放疗提供基础资料。方法　采用胸苷（ＴｄＲ） 双阻断法和流式细胞术（ＦＣＭ） 检测了ＨｅＬａＳ３ 细胞同步化后分别于其细胞周期各时相进行７５ ｍ Ｇｙ 和２－０ Ｇｙ Ｘ 射线照射以及于Ｇ２＋ Ｍ 期进行０－０２５～２－０ Ｇｙ 照射后其细胞周期进程的变化。结果　２－０ Ｇｙ 照射时细胞无论处于Ｇ０／Ｇ１ 、Ｓ期还是Ｇ２ ＋ Ｍ 期,从释放点后９～１５ 小时内均发生明显的Ｓ期延迟和Ｇ２ 阻滞,但其中以Ｇ２ 期照射者阻滞最显著;于Ｇ０／Ｇ１ 和Ｇ２ ＋ Ｍ 期进行７５ ｍ Ｇｙ 照射,分别在９ 和１２ ｈ 时发生一过性的明显的Ｇ２ 阻滞,至１１ 和１５ｈ 时完全从这一阻滞中脱离,甚至促进了其细胞周期的进程;而在Ｓ期进行７５ ｍ Ｇｙ 照射时,并没有发生这种阻滞,反而加速了其细胞周期由Ｇ２ ＋ Ｍ→Ｇ０／Ｇ１ 的移行,其中尤以９ｈ 时最为显著。剂量效应关系研究表明,于Ｇ２ ＋ Ｍ 期进行０－０２５ ～２－０ Ｇｙ 照射后,在释放点后１１ ｈ 时均发生Ｇ０／Ｇ１ 期细胞数减少,Ｇ２ 阻滞,且有剂量依赖性,而Ｓ期细胞数的变化在０－０２５ ～０－１ Ｇｙ 和０－５ ～２－０ Ｇｙ     

Radioresistance in a tumour cell line correlates with radiation inducible Ku 70/80 end-binding activity

PURPOSE: The aims of the present study were to better understand the role of Ku 80, which is involved in double-strand break repair in mammalian cells in the mechanism of radiation resistance and to verify the possibility of increasing cell radiosensitivity by targeted inhibition of Ku autoantigen 80 (Ku 80). MATERIALS AND METHODS: Western blot and electrophoretic mobility shift assay (EMSA) were performed on the human bladder carcinoma cell line RT112 (radioresistant) and on the human colorectal carcinoma cell line SW48 (radiosensitive) to assess the expression levels of DNA-dependent protein kinase (DNA-PK) components and the DNA-binding activity of the Ku 70/80 heterodimer after exposure to radiation, respectively. Ku 80 silencing was carried out with the use of small interfering RNA (siRNA). RESULTS: Greater differences in the DNA-binding activity of Ku 70/80 and Ku 80 phosphorylation level were observed in RT112 as compared to SW48 after X-ray treatment. There is no correlation between Ku expression and DNA-binding activity at lower doses. A significant increase in nuclear Ku 80 expression was observed one hour after the exposure, only at the higher doses, while the DNA-PK catalytic subunits (DNA-PKcs) and Ku 70 levels did not change significantly. Inhibition of Ku 80 expression by siRNA induced radiosensitivity in the RT112 cell line. CONCLUSIONS: Our data demonstrate that in a bladder tumour cell line up-regulation of Ku end-binding activity without any marked change in Ku expression underlie radiation resistance.     

Induction of cytogenetic adaptive response of somatic and germ cells in vivo and in vitro by low-dose X-irradiation

The cytogenetic adaptive response induced by low-level radiation was studied using human and rabbit lymphocytes in vitro and bone marrow cells and germ cells in vivo. The inductive dose of X-rays was 10 mGy for the in vitro studies at a dose rate of 10 mGy/min, and 2, 10, 50, 75 and 100 mGy for the in vivo studies at a dose rate of 50 mGy/min. The challenging dose was 1.5 Gy X-rays for the in vitro experiments and 0.65 or 0.75 Gy for the in vivo experiments at a dose rate of 0.44 Gy/min. The results reported here, in addition to those that have appeared in the literature, show the following characteristics documented for the first time: (1) 10 mGy could induce the adaptive response in human as well as rabbit lymphocytes irradiated not only in G1, S and G2 phases, but also in the Go state; (2) although the induced adaptive response could only last three cell cycles, it could be revived when the inductive dose was repeated after the third cell cycle; (3) the adaptive response could be induced by low-dose X-rays in somatic cells, both in vitro (lymphocytes) and in vivo (bone marrow cells), and also in germ cells (spermatocytes); (4) the magnitude of the adaptive response induced by whole-body irradiation was found to be dose-dependent--the lower the inductive dose the more the reduction of the frequency of chromatid aberrations following the challenging dose.     

低剂量辐射对人骨髓间充质干细胞影响的研究

目的 探讨低剂量辐射(LDR)对人骨髓间充质干细胞(hBM-MSC)生物学特性的影响。方法 应用体外培养传代的第四代hBM-MSC,采用X射线照射,照射剂量分别为50 mGy、75mGy、100 mGv,剂量率为12.5 mGy/min,分别观察LDR后hBM-MSC生长曲线、细胞周期与凋亡的变化及细胞因子、干细胞因子(SCF)、白细胞介素6(IL-6)、巨噬细胞集落刺激因子(M-CSF)表达量的变化。结果 LDR后,hBM-MSC从72 h起生长明显加快;LDR照射hBM-MSC后,在G0-G1期百分率逐渐减少,S期百分率在照射后48 h、72 h逐渐明显增多,以75 mGy照射后72 h的S期百分率增多最明显,为68.88%,而细胞凋亡的变化结果是LDR后在24 h、48 h有增多趋势,照射后72 h,凋亡细胞百分率有减少趋势;LDR照射hBM。MSC后24 h、48 h SCF分泌量均有升高趋势;50mGv、75mgy、100mGyX射线照射hBM-MSC后在培养的24 h、48 h,IL-6分泌量均有升高趋势;LDR照射hBM-MSC后,除50mGy照射后72h外,M-CSF分泌量在不同剂量照射后24 h、48 h、72 h均持续升高。结论 LDR后hBM-MSC从生长曲线、细胞周期与凋亡的变化及细胞因子表达量的变化均表现出兴奋性效应。