组蛋白修饰影响细胞辐射敏感性的研究进展

组蛋白修饰在细胞DNA损伤修复过程发挥重要作用。近年来多项研究表明,组蛋白修饰可以在参与招募DNA损伤修复因子、创建染色质开放结构和建立组蛋白抑制性标记等方面影响细胞对辐射的应答。同时,调控组蛋白修饰方式可以影响DNA损伤修复的过程,进而影响辐射敏感性。本文就组蛋白修饰影响DNA损伤修复过程与辐射敏感性的机制进行综述。     

染色质重构因子CHD蛋白家族的研究进展

染色质重构是DNA修复、基因表达调控过程中的一个重要环节。染色质重构使染色质组织结构发生一系列重要的变化,如染色质去凝集,核小体变成开放式的疏松结构,使转录因子等更易接近并结合核小体DNA,从而调控基因转录等。CHD蛋白是目前已知的染色质重构复合物之一。目前已鉴定了6个人类CHD蛋白成员,主要有3种功能结构域:N端的两个染色质调节域,位于中部的类SWI2/SNF2 ATP酶/解旋酶域,以及C端的DNA结合域。CHD基因突变或表达异常与人类某些疾病有关。     

MicroRNAs与结直肠癌辐射敏感性的关系及作用机制

结直肠癌目前是世界第三大肿瘤,手术治疗后约50%的患者会复发和转移,目前美国国立综合癌症网络（NCCN）肿瘤学临床实践指南推荐常规接受适型外照射治疗。由于放疗将显著增加不良反应,如何最大程度减小辐射剂量,提高辐射敏感性至关重要。近年来人们不仅发现了microRNAs参与了结直肠癌的发病和演进,而且越来越多的证据表明,microRNAs在结直肠癌的辐射敏感性中发挥了重要的作用。辐射引起的DNA损伤反应包括ATM的激活,组蛋白修饰和染色质重塑,细胞周期停滞,损伤修复和凋亡等系列过程,microRNAs可以通过作用于任何一个环节调节DNA损伤修复过程,从而调控肿瘤的辐射敏感性。本综述重点阐述microRNAs影响DNA损伤修复的作用机制,并展望了microRNAs通过影响肿瘤辐射敏感性在临床上的应用。     

基因表达调控与肿瘤转移

肿瘤转移是癌细胞从原发部位转移到远端部位形成肿瘤的过程。近年来的研究表明,细胞中基因表达的改变与肿瘤侵袭转移的发生、发展有着密切的联系。除DNA序列自身的改变外,DNA甲基化、微小RNA（microRNAs）的作用、组蛋白修饰以及染色质重塑等亦是造成基因表达改变的调控途径。本文主要就这些调控途径在肿瘤侵袭转移中的作用进行综述。     

组蛋白H2AX的磷酸化及其在放射生物学中的潜在应用

染色质核小体是由组蛋白H3、H4、H2A和H2B各2个分子组成八聚体核心，外围缠绕DNA链而成，每个组蛋白通过其球形结构域与DNA和其他组蛋白作用，但它们的NH2-端和C-端暴露在外，因此可以发生多种修饰，如磷酸化、乙酰化、甲基化和泛素化等，这些修饰可以改变染色质的结构和DNA的转录特性。H2AX是组蛋白H2A家族中的一种，依细胞系和组织的不同可占哺乳动物H2A的2％-25％，与其他H2A成员不一样的是，H2AX的C-端尾部是一个由4个氨基酸SQ（E，D）（I，L，F，Y）组成的保守序列，简称SQE基序（SQE motif）。近几年的研究表明，组蛋白H2AX在DNA损伤修复、细胞周期检查点调控、基因组稳定的维持和肿瘤抑制中起重要作用，而且在放射生物学中具有应用前景。     

三种细胞辐射敏感性相关酶的作用研究

拓朴异构酶Ⅰ和Ⅱ、聚腺苷二磷酸核糖聚合酶等是参与调节染色质结构的关键酶，在ＤＮＡ复制、转录及损伤修复等重要的生命事件中起着十分重要的作用。本实验采用正交设计方法分析它们在细胞染色质调控方面的功能以及对辐射所致ＤＮＡ损伤的影响。结果表明，经过相应的酶抑制剂作用后，染色质结构的松弛程度增加，受照后ＤＮＡ的初始损伤加重，且三种酶之间存在交互作用，提示出这些酶可能在使染色质结构紧密化及提高辐射抗性方面起有利作用；所观察到的交互作用可能反映了三种酶之间的联系。     

泛素连接酶CUL4A-DDB1复合体参与DNA损伤修复的研究进展

泛素化修饰在DNA损伤信号中发挥重要功能,包括细胞周期监控、DNA修复、细胞衰老和程序性死亡的调控。CUL4A-DDB1泛素连接酶通过DCAFs靶向调控特异性的底物,启动DNA切除修复机制对受损DNA进行修复。近期的研究表明CUL4A-DDB1泛素连接酶协助DNA修复因子与受损DNA的识别,来维持基因组的稳定性和正确性。     

环状RNA在DNA损伤修复中作用的研究进展

环状RNA（CircRNA）是一类由外显子、内含子或基因间区经反向剪接形成的非编码RNA,具有种类丰富、序列保守、结构稳定和细胞组织特异性等特点。CircRNA在多种恶性肿瘤中处于失调状态,其可通过调节放化疗后细胞DNA双链断裂的损伤修复功能,使肿瘤细胞发生增殖失控、远处转移和凋亡受阻等一系列不良反应,进而影响治疗效果和预后。笔者综述了CircRNA的分子生物学特性及其在DNA损伤修复（特别是DNA双链断裂损伤修复）中发挥的作用,并对CircRNA在肿瘤患者的治疗、预后和减轻放化疗产生的不良反应等方面可能发挥的作用进行展望。     

Smad4诱导大规模染色质伸展结构域定位

目的：了解Smad4能否调节大规模染色质结构改变并对其功能区域进行定位。方法：将Smad4及其缺失突变体与pRC-lac载体上lac的阻遏物融合并在AO3-1细胞中表达。该细胞株由CHO细胞衍变而来，整合有256个拷贝串联排列的lac操纵基因。在lac操纵子基因上游含有DHFR基因，可用甲氨蝶呤(MTX)进行基因共扩增。在细胞中lac阻遏物识别和结合lac操纵子基因。用抗lac阻遏物抗体进行免疫组化试验，荧光显微镜下观察染色质结构的改变。结果：野生型Smad4(1～552aa)有一定的诱导染色质伸展能力，且不依赖于TGF-β。N端区域(1～150aa)、中间连接区域(130～325aa)没有诱导大规模染色质伸展活性。C端区域(260～552aa)有一定的诱导染色质伸展能力，而C端缺失38个氨基酸残基的区域(260～514aa)能强烈地诱导大规模染色质伸展。结论：Smad4诱导大规模染色质伸展结构域位于260～514位氨基酸之间，C端38个氨基酸对Smad4诱导大规模染色质伸展有抑制作用。对Smad4诱导大规模染色质伸展活性特性的研究为揭示Smad4在肿瘤发生发展中的作用机制提供了新的途径。     

靶向PARP-1调控肿瘤细胞放射敏感性的研究进展

放疗是晚期肿瘤的主要治疗手段,然而,由于肿瘤耐受和抵抗的出现,其治疗效果不佳.因此,纠正肿瘤放疗抵抗或提高其放疗敏感性成为迫切需要解决的问题.聚腺苷二磷酸核糖聚合酶(PARP)1是一种功能丰富的核蛋白,鉴于其在染色质结构调节和DNA损伤修复等细胞过程中的重要作用,PARP-1被认为是最具有潜力的一种放射增敏靶点.笔者就...     

The influence of differently polarised microwave radiation on chromatin in human cells

Purpose: To determine the possible biological effects of differently polarised microwave radiation on the chromatin state in human cells.

Materials and methods: Isolated human buccal epithelium cells were irradiated by microwaves of frequency f = 35 GHz and surface power density E = 30 μW/cm². The state of chromatin in human cells was determined by methods of light and electron microscopy. The state of cell membranes was evaluated by the method of vital indigo carmine staining.

Results: The microwave-induced condensation of chromatin in human cells is revealed. Degree of microwave-induced condensation depends on the state of polarisation of electromagnetic wave: In some cases left circularly polarised waves induce less effect than linearly polarised radiation. The linearly polarised electromagnetic waves induce cell membrane damage revealed by increase of cell staining. The data obtained are discussed in connection with mechanisms of biological effects of electromagnetic fields.

Conclusion: The data obtained in this work demonstrate important biological effects of monochromatic microwave irradiation at 35 GHz. Low-level microwave irradiation induces chromatin condensation in human cells and damages of cell membranes.     

Protection of DNA from radiation-induced double-strand breaks: influence of replication and nuclear proteins

PURPOSE: To study the protective effect of histone and non-histone proteins on double-strand break (dsb) induction in replicating S-phase DNA as well as bulk DNA of plateau phase human tumour cells. MATERIALS AND METHODS: Induction of dsb was studied in two human adenocarcinoma cell lines: Colo320HSR and MCF-7. To assess the influence of chromatin structure on radiation-induced DNA dsb, different nuclear preparations of cells, either continuously labelled with 14C or pulse labelled with 3H, were assessed by pulsed-field gel electrophoresis (PFGE). RESULTS AND CONCLUSIONS: Stepwise removal of DNA-bound proteins from the chromatin increased the amount of radiation-induced dsb in both cell lines. However, the protective effect of DNA-associated proteins on dsb induction was significantly reduced in DNA of replicating S-phase cells compared with bulk DNA of plateau phase cells. These data show that proteins associated with the DNA have a different protective effect on radiation-induced dsb, rendering replicating DNA with open chromatin structure more sensitive to dsb induction by ionizing radiation.     

沉默BLAP75基因对电离辐射诱导DNA损伤的影响

目的 研究BLM结合蛋白75（BLAP75）在电离辐射诱导DNA损伤反应中的生物学效应。 方法 运用RNA干扰技术,在细胞中特异性沉默BLAP75基因,然后通过单细胞凝胶电泳技术来研究电离辐射诱导DNA损伤程度的变化,并且通过再次表达BLAP75来拯救沉默BLAP75所致的实验表型,以及应用Western blot方法研究DNA损伤反应中的磷酸化修饰。 结果 与未转染的293T对照组细胞相比,电离辐射在沉默了BLAP75基因的细胞中会诱导更多的DNA断裂,并且在此细胞中重新表达BLAP75则能降低DNA断裂数量至对照组细胞水平;γ射线照射后,细胞周期检查点激酶2（Chk2）的磷酸化程度比阴性对照组细胞增强。 结论 BLAP75能减少电离辐射诱导的DNA损伤,在电离辐射损伤修复中可能具有重要作用。     

Protection of DNA from radiation-induced double-strand breaks: influence of replication and nuclear proteins

Purpose : To study the protective effect of histone and non-histone proteins on double-strand break (dsb) induction in replicating S-phase DNA as well as bulk DNA of plateau phase human tumour cells. Materials and methods : Induction of dsb was studied in two human adenocarcinoma cell lines: Colo320HSR and MCF-7. To assess the influence of chromatin structure on radiation-induced DNA dsb, different nuclear preparations of cells, either continuously labelled with 14 C or pulse labelled with 3 H, were assessed by pulsed-field gel electrophoresis (PFGE). Results and conclusions : Stepwise removal of DNA-bound proteins from the chromatin increased the amount of radiation-induced dsb in both cell lines. However, the protective effect of DNA-associated proteins on dsb induction was significantly reduced in DNA of replicating S-phase cells compared with bulk DNA of plateau phase cells. These data show that proteins associated with the DNA have a different protective effect on radiation-induced dsb, rendering replicating DNA with open chromatin structure more sensitive to dsb induction by ionizing radiation.     

Regulation of DNA repair kinetics in proliferative and quiescent tumor cells

The radiosensitivity and kinetics of repair of radiation-induced DNA damage were determined for proliferative (P) and quiescent (Q) cells of the mouse mammary adenocarcinoma line 67. 67 Q cells are more radiosensitive than 67 P cells. Radiation induced the same amount of DNA damage in both 67 P and 67 Q cells. Both 67 P and 67 Q cells repaired their DNA damage with biphasic kinetics, but the half-times for the fast and slow phase were longer in 67 Q cells. Q cell DNA appeared to be in a more compact or condensed chromatin structure and was less accessible to enzymatic digestion than P cell DNA. These data suggest that 67 Q cells are more sensitive to ionizing radiation than 67 P cells because they repair their radiation-induced DNA damage more slowly, perhaps as a result of their more condensed chromatin structure.     

Mobile chest imaging of neonates in incubators: Optimising DR and CR acquisitions

IntroductionNeonates are a particularly vulnerable patient group with complex medical needs requiring frequent radiographic examinations. This study aims to compare computed radiography (CR) and direct digital radiography (DDR) portable imaging systems used to acquire chest x-rays for neonates within incubators.MethodsAn anthropomorphic neonatal chest phantom was imaged under controlled conditions using one portable machine but captured using both CR and DDR technology. Other variables explored were: image receptor position (direct and incubator tray), tube current and kV. All other parameters were kept consistent. Contrast-to-noise ratio (CNR) was measured using ImageJ software and dose-area-product (DAP) was recorded. Optimisation score was calculated by dividing CNR with the DAP for each image acquisition.ResultsThe images with the highest CNR were those acquired using DDR direct exposures and the images with lowest CNR were those acquired using CR with the image receptor placed within the incubator tray. This is also supported by the optimisation scores which demonstrated DDR direct produced the optimal combination with regards to CNR and radiation dose. The CNR had a mean increase of 50.3% when comparing DDR direct with CR direct respectively. This was also evident when comparing DDR and CR for in-tray acquisitions, with CNR increasing by a mean of 43.5%. A mean increase of 20.4% was seen in CNR when comparing DDR tray exposures to CR direct.ConclusionDDR direct produced images of highest CNR, with incubator tray reducing CNR for both CR and DDR. However, DDR tray still had better image quality compared to CR direct.Implications for practiceWhere possible, DDR should be the imaging system of choice for portable examinations on neonates owing to its superior image quality at lower radiation dose.     

The shape of DNA elution dose-response curves under non-denaturing conditions: the contribution of the degree of chromatin condensation.

We have re-examined the effect of detergent type, pH and temperature of lysis on the shape of the DNA elution dose response under non-denaturing conditions using plateau-phase CHO cells. Results practically identical to those previously published (Okayasu and Iliakis, 1989) were obtained, with a 1 h incubation at 60 degrees C during lysis with sodium-N-laurylsarcosine (NLS) resulting in almost linear dose-response curves. We also examined chromatin decondensation as a contributing factor in the linearization observed in the elution dose-response curve under the above conditions. When nuclei with condensed chromatin were prepared from irradiated cells, applied on the filter and lysed with NLS at room temperature, a shoulder-type elution dose-response curve was obtained only slightly higher than that of cells lysed under the same conditions. However, when nuclei prepared from irradiated cells were applied on the filter after relaxation of chromatin by incubation in low ionic strength buffer and lysed with NLS at room temperature, an almost linear dose-response curve was obtained similar to that of cells lysed with NLS at 60 degrees C. Lysis with NLS at 60 degrees C of nuclei with relaxed chromatin did not further modify the DNA elution dose-response curve. Based on these results we propose that the linearization of the DNA elution dose-response curve observed after chromatin decondensation reflects a reduction in the degree of chromatin compactness in the nuclear complexes that leads to a relatively uniform distribution of the DNA on the filter and reduces trapping of elutable material in the compact nuclear structures otherwise present. Since high radiation doses dissolve compact nuclear structures, trapping of elutable material is expected to be highest at low doses of radiation, leading to the observed reduction in the fraction of DNA eluted per Gy at low versus high radiation doses and thus to the observed shoulder. Furthermore, we propose that the linearization of the DNA elution dose-response curves observed in cells lysed in NLS at 60 degrees C may also be due to a decondensation of the nuclear complexes on the filter as a result of the combined action of detergent and high temperature. The notion of a correlation between DNA elution dose response and cell radiosensitivity was examined in two human (SQ20B, SCC61) and two Chinese hamster (V-79, irs-2) cell lines with widely different radiosensitivities.(ABSTRACT TRUNCATED AT 400 WORDS)     

Image acquisition in general radiography: The utilisation of DDR

ObjectiveThis article explores image acquisition with DDR. General radiographic technology continues to advance therefore it remains paramount to continually reflect on DDR hardware and software amongst radiographers in an imaging modality that constitutes approximately 90% of all radiological examinations.MethodThis article reports findings from a wider ethnographic study of two general radiography environments in the United Kingdom (UK). Participant observation and semi-structured interviews were the methods used to uncover original data.ResultsTwo key themes are discussed. Firstly, ‘the extent of DDR knowledge’ amongst radiographers is examined. The findings uncover that not all radiographers have an adequate knowledge base with DDR technology. Secondly, ‘pitfalls and near misses with DDR’ is discussed. This theme highlights the potential danger of radiographers ‘over-repeating’ X-ray examinations, coincided with the occurrence of radiological incidents whereby a patient is exposed to ionising radiation with no added benefit.ConclusionThis paper concludes by challenging the current ‘skill base’ to operate DDR equipment. In addition, new pitfalls and near misses are highlighted, which may help forestall radiation incidents in the future. Dose and image optimisation remain central tenets to the role of the radiographer.Advances in knowledgeFew studies have challenged image acquisition with DDR. This study adds to existing knowledge by uncovering original phenomena that may initiate discussions within the radiography community and continually enhance healthcare delivery.