The Inhibitory Effect of Extracts From Fructus lycii and Rhizoma polygonati on in vitro DNA Breakage by Alternariol

Alternariol caused DNA single-strand breakage. Conversion of the closed circular double-stranded supercoiled DNA (pBR 322) to the nicked circular form and linear form was used to investigate the effect of extracts of some Chinese medical herbs on DNA nicking induced by alternariol. Some substances in the extracts of Rhizoma polygonati (RP) and Fructus lycii (FL) were shown to protect DNA from the attack by alternariol.Some substance in the RP may bind to plasmid DNA, and this binding reduces the electrophoretic mobility of DNA. These results indicate that substances from FL and RP may be used as DNA protectors. It is possible that they play an important role in preventing cancer.

Inhibition of Rare Earth Chlorinate on Ni_(2)O_(3)-Induced DNA Strand Breakage and Effect on Oxy-Radicals

Inhibition of RECl3 on Ni2O3-induced DNA breakage of human embryo lung cell (HEL) and reduced content of negative superoxidative ion (O2) in guinea alveolar macrophage (AM ) was observed by means of single cell gel electrophoresis assay (comet assay) and cytochrome C assay respectively. Incubated with 2×10 cell/ml human embryo lung cell for 1 h at 37℃, 20μg. ml-1 of Ni2O3 could obviously induce DNA strand breakage compared with the control (P< 0.01). Add 10 μg.ml-1 CeCl3 or 10μg RECl3 with 20 μg·ml-1 Ni2O3 simultaneously in to HEL culture, the DNA strand breakage caused by Ni2O3 reduction significantly. Culture with 4×10 cell/ ml AM for 1 h at 37℃, 10, 20 μg· ml-1 of Ni2O3 could distinctly increase·O2 content in AM compared with the control (P< 0.05). In the similar way, RECl3, CeCl3 or LaCl3 could evidently decrease·O2 content induced by Ni2O3 in AM (P<0.01), but both kinds of RE compounds can not suppress·O2 content in AM.

DNA Double-Strand Breaks,Potential Targets for HBV Integration

Hepatitis B virus(HBV)-induced hepatocellular carcinoma(HCC) is one of the most fre-quently occurring cancers.Hepadnaviral DNA integrations are considered to be essential agents which can promote the process of the hepatocarcinogenesis.More and more researches were designed to find the relationship of the two.In this study,we investigated whether HBV DNA integration occurred at sites of DNA double-strand breaks(DSBs),one of the most detrimental DNA damage.An 18-bp I-SceI homing endonuclease recognition site was introduced into the DNA of HepG2 cell line by stable DNA transfection,then cells were incubated in patients’ serum with high HBV DNA copies and at the same time,DSBs were induced by transient expression of I-SceI after transfection of an I-SceI expression vector.By using nest PCR,the viral DNA was detected at the sites of the break.It appeared that integra-tion occurred between part of HBV x gene and the I-SceI induced breaks.The results suggested that DSBs,as the DNA damages,may serve as potential targets for hepadnaviral DNA insertion and the integrants would lead to widespread host genome changes necessarily.It provided a new site to investi-gate the integration.

Codon evolution in double-stranded organelle DNA: strong regulation of homonucleotides and their analog alternations

In our previous study, complete single DNA strands which were obtained from nuclei, chloroplasts and plant mitochondria obeyed Chargaff’s second parity rule, although those which were obtained from animal mitochondria deviated from the rule. On the other hand, plant mitochondria obeyed another different rule after their classification. Complete single DNA strand sequences obtained from chloroplasts, plant mitochondria, and animal mitochondria, were divided into the coding and non-coding regions. The non-coding region, which was the complementary coding region on the reverse strand, was incorporated as a coding region in the forward strand. When the nucleotide contents of the coding region or non-coding regions were plotted against the composition of the four nucleotides in the complete single DNA strand, it was determined that chloroplast and plant mitochondrial DNA obeyed Chargaff’s second parity rule in both the coding and non-coding regions. However, animal mitochondrial DNA deviated from this rule. In chloroplast and plant mitochondrial DNA, which obey Chargaff’s second parity rule, the lines of regression for G (purine) and C (pyrimidine) intersected with regression lines for A (purine) and T (pyrimidines), respectively, at around 0.250 in all cases. On the other hand, in animal mitochondrial DNA, which deviates from Chargaff’s second parity rule, only regression lines due to the content of homonucleotides or their analogs in the coding or non-coding region against those in the complete single DNA strand intersected at around 0.250 at the horizontal axis. Conversely, the intersection of the two lines of regression (G and A or C and T) against the contents of heteronucleotides or their analogs shifted from 0.25 in both coding and non-coding regions. Nucleotide alternations in chloroplasts and plant mitochondria are strictly regulated, not only by the proportion of homonucleotides and their analogs, but also by the heteronucleotides and their analogs. They are strictly regulated in animal mitochondria only by the content of homonucleotides and their analogs.

Maternal gene Ooep may participate in homologous recombination-mediated DNA double-strand break repair in mouse oocytes

DNA damage in oocytes can cause infertility and birth defects. DNA double-strand breaks （DSBs） are highly deleterious and can substantially impair genome integrity. Homologous recombination （HR）-mediated DNA DSB repair plays dominant roles in safeguarding oocyte quantity and quality. However, little is known regarding the key players of the HR repair pathway in oocytes. Here, we identified oocyte-specific gene Ooep as a novel key component of the HR repair pathway in mouse oocytes. OOEP was required for efficient ataxia telangiectasia mutated （ATM） kinase activation and Rad51 recombinase （RAD51） focal accumulation at DNA DSBs. Ooep null oocytes were defective in DNA DSB repair and prone to apoptosis upon exogenous DNA damage insults. Moreover, Ooep null oocytes exhibited delayed meiotic maturation. Therefore, OOEP played roles in preserving oocyte quantity and quality by maintaining genome stability. Ooep expression decreased with the advance of maternal age, suggesting its involvement in maternal aging.

Double-stranded DNA breaks and gene functions in recombination and meiosis

Meiotic prophase I is a long and complex phase. Homologous recombination is an important process that occurs between homologous chromosomes during meiotic prophase I. Formation of chiasmata, which hold homologous chromosomes together until the metaphase I to anaphase I transition, is critical for proper chromosome segregation. Recent studies have suggested that the SPO 11 proteins have conserved functions in a number of organisms in generating sites of double-stranded DNA breaks （DSBs） that are thought to be the starting points of homologous recombination. Processing of these sites of DSBs requires the function of RecA homologs, such as RAD5 1, DMC 1, and others, as suggested by mutant studies; thus the failure to repair these meiotic DSBs results in abnormal chromosomal alternations, leading to disrupted meiosis. Recent discoveries on the functions of these RecA homologs have improved the understanding of the mechanisms underlying meiotic homologous recombination.

Mechanical properties of double-stranded DNA biofilm with Gaussian distribution

In microcantilever-based label-free biodetection technologies, deflection changes induced by adsorptions of double-stranded DNA （dsDNA） molecules on Au-layer surface are greatly affected by the mechanical, thermal and electrical properties of DNA biofilm. In this paper, the elastic properties of dsDNA biofilm are studied. First, the Parsegian＇s empirical potential based on a mesoscopic liq- uid crystal theory is employed to describe the interaction energy among coarse-grained DNA cylinders. Then, con- sidering a Gaussian distribution of DNA interaxial distance, the thought experiment method is used to derive an analyti- cal expression for Young＇s modulus of DNA biofilm with a stochastic packing pattern for the first time. Results show that Young＇s modulus of DNA biofilm is on the order of 10 MPa. These findings could provide a simple and effective method to evaluate the mechanical properties of soft biofilm on snbstrate.

Adsorption dynamics of double-stranded DNA on a graphene oxide surface with both large unoxidized and oxidized regions

The adsorption dynamics of double-stranded DNA(dsDNA)molecules on a graphene oxide(GO)surface are important for applications of DNA/GO functional structures in biosensors,biomedicine and materials science.In this work,molecular dynamics simulations were used to examine the adsorption of different length dsDNA molecules(from 4 bp to24 bp)on the GO surface.The dsDNA molecules could be adsorbed on the GO surface through the terminal bases and stand on the GO surface.For short dsDNA(4 bp)molecules,the double-helix structure was partially or totally broken and the adsorption dynamics was affected by the structural fluctuation of short dsDNA and the distribution of the oxidized groups on the GO surface.For long dsDNA molecules(from 8 bp to 24 bp)adsorption is stable.By nonlinear fitting of the contact angle between the axis of the dsDNA molecule and the GO surface,we found that a dsDNA molecule adsorbed on a GO surface has the chance of orienting parallel to the GO surface if the length of the dsDNA molecule is longer than 54 bp.We attributed this behavior to the flexibility of dsDNA molecules.With increasing length,the flexibility of dsDNA molecules also increases,and this increasing flexibility gives an adsorbed dsDNA molecule more chance of reaching the GO surface with the free terminal.This work provides a whole picture of adsorption of dsDNA molecules on the GO surface and should be of benefit for the design of DNA/GO based biosensors.

Exposure to Long Magnetic Resonance Imaging Thermometry Does Not Cause Significant DNA Double-Strand Breaks on CF-1 Mice

The purpose of the study was to investigate if the high gradient strength and slew rate used for long MRI-thermometry monitoring could cause DNA double-stranded breaks (DSBs). To this end, an enzyme-linked immunosorbent assay (ELISA) was used to quantify γH2AX, a molecular marker for DSBs, in the blood of mice after a 6-hour exposure to magnetic resonance imaging (MRI). Fourteen CF-1 female mice were separated into 4 experimental groups: Untreated negative control, MRI-treated, MRI-Control, and exposed to ionizing radiation positive control. Untreated negative control was used as a baseline for ELISA to quantify γH2AX. MRI-treated consisted of a 6-hour continuous magnetic resonance imaging (MRI) echo planar imaging (EPI) sequence with a slew rate of 192 mT/m/s constituting a significantly longer imaging time than routine clinical imaging. MRI-control mice were maintained under the same conditions outside the MRI scanner for 6-hours. Mice in the irradiation group served as a positive control of DSBs and were exposed to either 2 Gy, 5 Gy or 10 Gy of ionizing radiation. DSBs in the blood lymphocytes from the treatment groups were analyzed using the γH2AX ELISA and compared. Total protein concentration in lysates was determined for each blood sample and averaged 1 ± 0.35 mg/mL. Irradiated positive controls were used to test radiation dose-dependency of the γH2AX ELISA assay where a linear dependency on radiation exposure was observed (r2 = 0.93) between untreated and irradiated samples. Mean and standard error mean of γH2AX formation were calculated and compared between each treatment group. Repeated measures 1-way ANOVA showed statistically significant differences between the means of irradiated controls and both the MRI-control and MRI-treated groups. There was no statistically significant difference between the MRI-treated samples and the MRI-control groups. Our results show that long MRI exposure at a high slew rate did not cause increased levels of γH2AX when compared to control mice, suggesting that no increase in DSBs was caused by the long MR thermometry imaging session. The novelty of this work contradicts other studies that have suggested MRI may cause DSBs;this work suggests an alternative cause of DNA damage.

螺旋藻提取物对DNA拓扑异构酶活性的抑制及对DNA的直接影响

探讨螺旋藻提取物对DNA拓扑异构酶活性的影响以及对DNA的直接作用。方法 :TOPOI介导的负超螺旋 pBR322解旋反应检测对TOPOI酶活力的影响 ;TOPOII介导的kDNA去连环作用检测对TOPOII活力的影响 ;采用负超螺旋pBR322和kDNA检测对DNA的直接作用。结果 :螺旋藻提取物的水溶性成分和DMSO可溶性成分在浓度分别为3 2μg/ml和80μg/ml时,能完全抑制TOPOI介导的负超螺旋 pBR322解旋反应;两者对TOPOII介导的kDNA去连环作用完全抑制的浓度分别为16μg/ml和2000μg/ml。此外 ,螺旋藻提取物水溶性成分在高浓度时可直接引起DNA的双链断裂。结论 :TOPO酶 ,主要是TOPOI 。

不同浓度甲醛致小鼠肾细胞DNA损伤效应研究

为了探讨甲醛导致生物机体内的DNA损伤效应及其剂量-效应关系,通过不同浓度的液态甲醛对小鼠肾细胞进行染毒,并分别运用了单细胞凝胶电泳实验、荧光检测法实验和KCl-SDS沉淀法实验进行研究.结果显示,甲醛在低浓度(5μmo·lL-1)时,具有致DNA断裂的作用;在中等浓度(25μmo·lL-1)时,对DNA的损伤作用以DNA-DNA交联为主;在高浓度(125、625μmo·lL-1)时,对DNA的损伤作用以DNA-蛋白质交联为主.

DNA断裂检测方法──单细胞凝胶电泳法

单细胞凝胶电泳（ｓｉｎｇｌｅｃｅｌｌｇｅｌｅｌｅｃｔｒｏｐｈｏｒｅｓｉｓａｓｓａｙ，ＳＣＧＥ）也叫彗星试验（ｃｏｍｅｔａｓｓａｙ），是一种快速、敏感、简便、廉价的检测单个哺乳动物细胞ＤＮＡ断裂的技术，目前已用于检测氧化、紫外线和电离辐射引起的损伤，以及三氯乙烷、丙烯酰胺等化学物及老化、吸烟所致损害的研究．文章介绍ＳＣＧＥ的发展、检测分析方法、原理及其在ＤＮＡ损伤与修复、生物监测、遗传毒理研究、肿瘤治疗方案优化和疗效研究方面的应用前景．

金属离子介导活性氧引起DNA氧化损伤及机制研究

应用高效液相色谱 电化学检测技术 ,以及单细胞凝胶电泳技术 ,研究了 11种不同价态金属离子体外诱导核酸 (DNA)氧化损伤产生 8 羟基脱氧鸟苷和DNA链断裂的化学机制 .结果表明 ,能够氧化为高价态的金属离子如Fe2 + 、Cu2 + 、Cr3 + 等能够介导活性氧造成DNA的氧化损伤 ,介导能力呈现Fe2 + Cu2 + >Cr3 + 的顺序 .另外一些金属离子 ,如V3 + 、Ni2 + 、Cd2 + 、Zn2 + 、Co2 + 、Fe3 + 等 ,不能介导核酸氧化损伤形成 8 OH dG加合物及引发DNA断裂 ,但可能诱导核酸链交联的发生 .

钬元素对小鼠肝脏细胞DNA损伤的影响

背景与目的: 通过研究钬离子溶液对小鼠肝脏细胞DNA的损伤 ,探讨钬元素对诱导动物细胞凋亡的影响。材料与方法:处理组1:小鼠定时灌胃氯化钬溶液50mg/(kg·d),连续5d;组2～3 :小鼠分别定时腹腔注射氯化钬溶液60mg/(kg·d)和120mg/(kg·d) ,连续2d ;组4:小鼠一次性腹腔注射氯化钬溶液320mg/kg;每次染毒相间24h,组1～4:小鼠均在末次染毒24h后处死 ,提取肝脏DNA。组5:小鼠一次性腹腔注射等体积生理盐水 ;组6～9 :小鼠一次性腹腔注射氯化钬溶液80mg/kg,分别于注射后12、24、48和96h处死小鼠提取肝脏DNA。通过琼脂糖凝胶电泳研究各组DNA带型变化。 结果 :处理组7DNA电泳中出现连续的弥散带型 ,其它各组均未观察到明显的拖尾现象。也未观察到“DNAladder"。 结论 :钬离子对小鼠肝脏细胞DNA的断裂作用可能与其剂量大小、处理时间及DNA修复作用有关 ,而且无特异性。本实验结果表明 。

DNA链断裂作为醛类污染物接触标志物的研究

应用单细胞凝胶电泳技术,通过对小鼠脾淋巴细胞染毒试验,测定了甲醛、乙醛、丙烯醛单独及联合作用下对细胞DNA分子链的断裂损伤.结果表明,3种醛均能引起DNA分子发生链断裂,对于乙醛和丙烯醛存在明确的剂量(反应关系,3种化合物还存在着一定的协同作用.上述结果提示醛类污染物潜在致癌性的一个可能机制以及DNA断裂作为醛类化合物接触标志物的可能性.

白血病细胞与正常外周血淋巴细胞的PARP活性及本底DNA链断裂的比较研究

聚腺苷二磷酸核糖聚合酶[poly(ADP-ribose)polymerase,PARP]是广泛存在于有核细胞中的多功能酶。当细胞DNA发生损伤时,PARP可作为细胞内的分子感受器,识别、结合到DNA断裂处,并被激活。激活的PARP可催化组蛋白H1、拓扑异构酶Ⅰ及Ⅱ、DNA聚合酶、RNA聚合酶、DNA连接酶、Ca^(2+)/Mg^(2+)依赖的核酸内切酶及PARP自身等多种核内受体蛋白的聚ADP-核糖基化作用,传递损伤信息,引发损伤所致级联反应,最终决定细胞的命运:修复损伤或凋亡。因此,PARP对维持细胞的正常功能具有极为重要的作用,其异常可能是导致细胞死亡失控的因素之一,癌变正是这种失控的一种后果。为了探讨PARP与细胞恶性变之间的关系,采集了14例健康供血员血样和4例初诊白血病病人血样,比较研究了白血病细胞与正常人外周血淋巴细胞的PARP活性。同时,应用彗星法(comet assay)研究了细胞内本底DNA链断裂及染色质结构。研究发现,与正常人外周血淋巴细胞相比,外周血白血病细胞中含有异常高的PAPP活性,在急性淋巴细胞白血病、慢性淋巴细胞白血病、慢性粒细胞白血病细胞中检出大量的本底DNA链断裂。这一结果表明:①外周血白血病细胞中异常高的PARP活性和大量存在的本底DNA链断裂可能为其临床诊断提供有效的辅助指标。

彗星试验检测DNA交联的研究

为了寻求一种简便、快速的DNA交联检测方法 ,本文以过氧化氢 (H2 O2 )为标准断裂剂 ,用彗星试验检测了甲醛 (formaldehyde,FA)诱导的TK6细胞的DNA交联作用 ,并对两种实验方案进行了比较。结果显示 :随着甲醛浓度的增加 ,由过氧化氢造成的DNA迁移距离减小 ;当甲醛浓度为 10 0 0 μmol L时 ,无一个细胞拖尾 ,提示甲醛完全抑制了过氧化氢诱导的DNA迁移。此外 ,两种实验方案均可检出甲醛的DNA交联作用 ,但以先加交联剂、后加断裂剂的方案优于先加断裂剂、后加交联剂的方案 ,因为前一方案操作更简便 ,不受DNA修复的影响 ,所测DNA交联的剂量—反应关系更好。因此 ,彗星试验确为一种灵敏、简便和快速的DNA交联检测方法。

气态甲醛对小鼠外周血白细胞的DNA损伤作用

为研究气态甲醛对外周血白细胞DNA的损伤作用,选用SPF级昆明种纯系雄性小鼠作为研究对象,用浓度为0.5 mg/m2、1.0 mg/m2、3.0 mg/m3气态甲醛对小鼠进行连续动态染毒72 h,染毒结束后,观察外周血白细胞的DNA断裂和DNA-蛋白质交联的状况.实验表明:气态甲醛能诱导小鼠外周血白细胞的DNA断裂,在各染毒浓度组与对照组均有及显著差异,在0.5 mg/m3浓度组DNA断裂效应比1.0 mg/m2,3.0 mg/m3浓度组高;1.0 mg/m2,3.0 mg/m3浓度组出现明显的DNA-蛋白质交联作用.研究结果表明:气态甲醛暴露对小鼠外周血白细胞的DNA有明显的损伤作用.

稀土对氧化镍致细胞DNA损伤及对氧自由基作用的影响

利用单细胞凝胶电泳技术 ,细胞色素C等技术分别观察CeCl3和RECl3对Ni2 O3所致人胚肺细胞DNA链损伤和对肺巨噬细胞 (AM)氧自由基 ,如超氧阴离子 (·O- 2 ) ,羟基自由基 (·OH)的影响。RECl3,CeCl3,LaCl3可明显降低Ni2 O3对人胚细胞DNA链的损伤 ,抑制Ni2 O3诱导AM产生过多的·O- 2 。

高LET的^7Li离子致DNA损伤的直接和间接作用研究

在HI-13串列加速器加速的具有高LET值的7Li离子辐照不同浓度的pUC19质粒DNA水溶液、加自由基清除剂(甘露醇)的DNA水溶液以及干状DNA样品,利用高分辨的原子力显微镜技术,研究7Li致DNA损伤的直接作用和间接作用。结果显示,在相同剂量下,7Li离子比低LET辐射能诱发更多的双链断裂,形成更多的集团损伤,使DSB的分布更局部和更密集。对于水溶液DNA,7Li离子的水辐解产生的自由基的间接作用在DNA分子链断裂的产生方面发挥着重要作用,而且自由基清除剂甘露醇能有效地保护DNA分子。