枸杞对DNA损伤后修复作用的研究

寻找天然抗致癌的物质对人类肿瘤的预防作用早已引起人们的重视。枸杞是我国传统中药。既是食品，又是药品，久服可坚筋骨，轻身不老，而耐寒暑。现代医学研究发现，枸杞具抗衰延寿，提高免疫功能，抑制癌细胞生长等作用。以小鼠骨髓细胞染色体畸变率为指标，进一步探讨枸...     

二氨胺基酸对V79细胞DNA损伤效应的研究

我们采用单细胞凝胶电泳新技术和ＳＣＥ试验研究了８３－１除草剂的主要代谢产物２，４－二氯－６－胺基酚（ＤＣＡＰ）对Ｖ７９细胞的ＤＮＡ损伤作用。结果表明，ＤＣＡＰ浓度≥５０μｇ／ｍｌ时，细胞出现明显的ＤＮＡ迁移和姐妹染色单体交换，说明ＤＣＡＰ是ＤＮＡ损伤剂，结合前期研究结果推断，６位硝基还原成胺基是该除草剂在体内活化致癌的基本途径，结果还表明，单细胞凝胶电泳经ＳＣＥ试验更灵敏，可能是研究环境低剂量暴露     

螺旋藻多糖对紫外线诱发的人胚肺二倍体细胞DNA损伤修复的影响

目的：观察螺旋藻多糖（polysaaccharide from Spirulina platensis,PSP)对紫外线诱发的人胚肺二倍体细胞DNA损伤修复的影响。方法：使用单细胞凝胶电泳技术（single cell gel clectrophoresis assay SCGE),检测紫外线（UV）照射30min及照射后30,60,90min各组细胞DNA的损伤情况,以DNA迁移距离（migrate length of DNA,MLD)作为损伤指标。结果PSP能预防UV诱发的DNA损伤,并能促进UV诱发损伤后的DNA修复合成。结论：PSP有增强核酸内切酶和连接酶活性的作用,并且这种作用呈剂量依赖性。     

异佛尔酮二异氰酸酯诱发DNA损伤的研究

对异佛尔酮二异氰酸醋(Isophorone Diisocyanate,IPDI)诱导大肠杆菌SOS反应、人淋巴细胞程序外DNA合成(UDS)和小鼠骨髓细胞DNA合成的影响进行了初步研究。IPDI对大肠杆菌PQ_(35)和PQ_(37)两个菌株均无明显的诱导SOS反应的能力,但IPDI可明显诱发人淋巴细胞UDS,对培养小鼠骨髓细胞DNA的合成具有明显的抑制作用,而且呈剂量-效应关系。IPDI对哺乳动物细胞DNA具有损伤作用。     

线粒体DNA氧化损伤热点的形成及其修复速率的比较

本研究旨在探讨线粒体DNA氧化损伤热点的形成是否与该点DNA修复速率慢有关 .BRL 3A细胞暴露于 5 0mmol·L- 1过氧化氢 (H2 O2 ) 30min后 ,分别于 0 ,1,4及 2 4h收获细胞 ,应用连接介导聚合酶链反应分析线粒体DNA氧化损伤热点的修复百分率并与线粒体DNA氧化损伤的总修复百分率比较 ,通过狭缝印迹法 ,比较了各时间点细胞内p5 3/线粒体DNA比率来验证其修复的可靠性 .结果表明 :在 4及2 4h ,热点的修复速率分别为 5 .2 %和 4 2 .1% .而线粒体DNA的总修复百分率分别为 76 .9%和 84 .1% ,后者与前者相比 ,其修复速率快 1～ 14倍左右 .应用狭缝印迹发现各时间点细胞内p5 3/线粒体DNA比率无明显变化 .因此 ,线粒体DNA氧化损伤热点的形成与其修复速率较慢有关 .狭缝印迹结果表明所观测的线粒体DNA的修复是可信的     

非同源末端连接修复相关因子对DNA损伤修复调控及肿瘤治疗作用的研究进展

细胞在内源性或外源性因子的胁迫作用下会产生各种损伤,包括遗传物质DNA的双链断裂(DSB)。非同源末端连接(NHEJ)是哺乳动物细胞中DSB损伤修复的一种主要机制。NHEJ过程中一些主要因子如DNA依赖性蛋白激酶、DNA交联修复蛋白1C、X射线修复交叉互补蛋白4/DNA连接酶Ⅳ和X射线修复交叉互补蛋白4类似因子对DNA损伤修复(DDR)具有重要的调控作用,其中任何一种因子的改变都会影响DDR的效率。此外,NHEJ相关因子与肿瘤发生息息相关。本文针对NHEJ相关因子调控DSB修复方面的研究作一简要综述,并对NHEJ修复相关因子在肿瘤治疗中的研究进行总结。     

脉冲场凝胶电泳检测电离辐射诱发DNA损伤及其修复

目的：评价脉冲场凝胶电泳（PFGE）法检测电离辐射诱发小鼠脾细胞DNA损伤及其修复的可行性.方法：采用PFGE法测定不同剂量γ射线诱发小鼠脾细胞DNA单、双链断裂及4 Gyγ射线照射后不同时间脾细胞DNA单、双链断裂及其修复情况,并与未照射组（对照组）进行比较.结果：γ射线照射小鼠后,脾细胞DNA单、双链断裂数目随照射剂量的增加均呈增加趋势,单链断裂数目多于双链,1 Gy所致DNA双链断裂及2 Gy所致DNA单链断裂显著高于对照组（t=2.668,P＜0.05;t=5.117,P＜0.01）.以4 Gy照射后经过不同时间修复,单、双链断裂均呈下降趋势,起初DNA链断裂的修复为快速修复,1 h后大多数损伤已得到修复.单链断裂的修复速度高于双链,当修复时间超过2 h后,单链断裂又呈现上升趋势.结论：本实验方法有可能成为一种快速、敏感地检测活体动物细胞DNA损伤及其修复的方法.     

Cytogenetic evaluation and DNA interaction studies of the food colorants amaranth,erythrosine and tartrazine

Food coloring agents, amaranth, erythrosine and tartrazine have been tested at 0.02–8 mM in human peripheral blood cells in vitro, in order to investigate their genotoxic, cytotoxic and cytostatic potential. Amaranth at the highest concentration (8 mM) demonstrates high genotoxicity, cytostaticity and cytotoxicity. The frequency of SCEs/cell was increased 1.7 times over the control level. Additionally, erythrosine at 8, 4 and 2 mM shows a high cytotoxicity and cytostaticity. Finally, tartrazine seems to be toxic at 8 and 4 mM. No signs of genotoxicity were observed. Reversely, tartrazine showed cytotoxicity at 1 and 2 mM. Furthermore, spectroscopic titration studies for the interaction of these food additives with DNA showed that these dyes bind to calf thymus DNA and distinct isosbestic points are observed clearly suggesting binding of the dyes to DNA. Additionally DNA electrophoretic mobility experiments showed that these colorants are obviously capable for strong binding to linear dsDNA causing its degradation. PCR amplification of all DNA fragments (which previously were pre-treated with three different concentrations of the colorants, extracted from agarose gel after separation and then purified), seems to be attenuated with a manner dye concentration-dependent reflecting in a delayed electrophoretic mobility due to the possible binding of some molecules of the dyes. Evaluation of the data and curves were obtained after quantitative and qualitative analysis of the lanes of the gel by an analyzer computer program. Our results indicate that these food colorants had a toxic potential to human lymphocytes in vitro and it seems that they bind directly to DNA.     

两种萘胺对DNA损伤及致细胞转化的研究

用碱洗脱荧光分析法检测两种萘胺对DNA交联性损伤的研究结果表明,浓度在7.1～14.3μg/ml时1-萘胺未造成DNA的横向交联性损伤,而2-萘胺却引起这种类型的损伤,并呈现剂量效应关系。在细胞转化试验中,浓度在1.8～14.3μg/m时,1-萘胺为阴性结果,2-萘胺为阳性结果。     

Synthesis and studies on three-compartment flavone-containing combi-molecules designed to target EGFR, DNA, and MEK

In order to induce a tandem targeting of EGFR, DNA, and MEK, we built complex combi-molecules containing an EGFR targeting quinazoline and an aminoethyltriazene moiety linking the entire molecule to PD98059. Two complex molecules were synthesized: one with a short aminoethyl spacer, AL232, and the other AL414 with a longer aminoethylaminoethyl spacer. AL414 was a more potent inhibitor of EGFR tyrosine kinase than AL232. Both combi-molecules blocked EGFR phosphorylation in whole cells and downregulated extracellular signaling-regulated kinases (ERK1,2). However, only AL414 was capable of inducing DNA damage. Thus, it was taken in vivo for metabolic analysis. The results showed that 3 h after injection, AL414 was hydrolyzed to an EGFR inhibitor FD105, which was further acetylated to FD105Ac, a more potent inhibitor of EGFR. The detected flavone derivative was PD98059 linked to the hydroxyalkyl moiety resulting from the decomposition of the alkyldiazonium species. Independent synthesis of the latter metabolite and further in vitro analysis showed that it was deprived of antiproliferative activity. The results in toto suggest that while AL414 is a three-compartment combi-molecule, only the EGFR and DNA targeting species can be released and the cleavage to the intact MEK inhibitor PD98059 was mitigated by the stability of the carbamate.     

紫外线致DNA突变的快速测定研究(英文)

毛细管电泳、变性色谱、免疫吸附、DNA测序方法等已广泛用于DNA碱基错配、氧化缺失、链断裂等变化的检测,但有时如进行药物筛选时,只需定性地检测DNA是否变化或变化程度。本文采用褶合光谱法定性地检测紫外线致DNA变化程度,将突变后DNA的褶合光谱与未变异前DNA自身标准比较,并以差谱值δ量化地表示突变程度。当褶合光谱δ高达11.48%时,才能从二阶导数光谱上发现差异,表明方法的灵敏度远远高于前者。加入DMSO后,溶液在254 nm照射时,δ升高,表现为DNA变异诱导剂;溶液在365 nm照射时,δ降低,表现为DNA变异保护剂。褶合光谱法快速、简便、灵敏、经济,可以作为检测DNA突变、筛选抗突变药物的一种新型方法。     

A Fast Determination of DNA Mutation Induced by Ultraviolet Radiation

Electrophoresis, chromatography, immunoassay, sequencing and other time consuming ap-proaches have been developed to determine DNA base mismatching, oxidative lesion or strand breaks. Sometimes,however, only qualitative information is enough to decide whether mutation has happened to DNA and its extent.Convolution spectrometry (CS), a new technique to discover ultrafme difference on ultraviolet (UV) absorption ofdifferent substances, is originally employed to find out any subtle mutation of DNA induced by UV radiation. Muta-tive DNA is compared with ego criteria based on the spectra of the former DNA, any difference is quantitatively ex-pressed by dispersion (5). Visible changes cannot be observed on second -derivative spectra until the mutation gets 5up to 11.48%. Dimethyl sulfoxide is an intensifier of UV 254 nm induced DNA mutation and protector at 365 nm,which is simply confirmed by increasing and decreasing 5. Every convolution procedure takes less than 1 min. Convolution spectrometry provides a fast, simple, sensitive and inexpensive alternative to determine DNA mutation, andto screen anti-mutational medicines.     

2种盐酸非索非那定制剂的人体生物等效性研究

目的:研究盐酸非索非那定口腔崩解片与普通片在正常人体内的生物等效性。方法:采用随机双周期交叉试验设计,20名男性健康志愿者分别单剂量口服盐酸非索非那定口腔崩解片(受试制剂)与普通片(参比制剂)60mg,采用高效液相色谱-电喷雾串联质谱(LC-MS/MS)法测定非索非那定血药浓度,用DASver2.1软件计算二者药动学参数以考察生物等效性。结果:受试者单剂量口服受试制剂与参比制剂后的Cmax分别为(240.01±77.66)、(237.76±89.97)ng·mL-1,tmax分别为(1.85±1.02)、(2.18±0.81)h,AUC0～72分别为(1309.9±467.7)、(1236.3±433.1)ng·h·mL-1,AUC0～∞分别为(1319.2±468.8)、(1245.9±436.2)ng·h·mL-1。受试制剂相对于参比制剂的生物利用度为(112±36)%。结论:盐酸非索非那定口腔崩解片与普通片具有生物等效性。     

Influence of PCB153 on Oxidative DNA Damage and DNA Repair-Related Gene Expression Induced by PBDE-47 in Human Neuroblastoma Cells In Vitro

We studied the relationship between 2,2,4,4-tetrabromodiphenylether (PBDE-47) and oxidative DNA damage as well as the modeof interaction between PBDE-47 and 2,2,4,4,5,5-hexachlorobiphenyl(PCB153) by incubating SH-SY5Y cells in four doses of PBDE-47(0, 1, 5, 10µM) and/or 5µM PCB153 and 100µMNAC (N-acetylcysteine) for 24 h. Results showed that reactiveoxygen species (ROS) production in the 5µM PBDE-47 + PCB153and 10µM PBDE-47 + PCB153 groups were significantly higherthan that of the control group (p < 0.05). DNA strand breakageand 8-hydroxy-2'-deoxyguanosine (8-OHdG) levels were significantlyincreased in the 10µM PBDE-47, 5µM PBDE-47 + PCB153,and 10µM PBDE-47 + PCB153 groups compared with the control(p < 0.05). Furthermore, ROS formation and DNA strand breakagewere dramatically increased in the 5µM PBDE-47 + PCB153and 10µM PBDE-47 + PCB153 groups compared with the correspondingPBDE-47 only group and the PCB153 group (p< 0.05). The levelof 8-OHdG was significantly increased in the 10µM PBDE-47+ PCB153 group compared with the corresponding PBDE-47 onlygroup and the PCB153 group (p < 0.05). The PBDE-47 groupcoincubated with NAC decreased the ROS level and amelioratedPBDE-47–mediated DNA damage. The mRNA expression levelsof X-ray repair cross-complementing gene 1 (Xrcc1) were significantlydecreased in the 10µM PBDE-47, 5µM PBDE-47 + PCB153,and 10µM PBDE-47 + PCB153 groups, whereas X-ray repaircross-complementing gene 3 (Xrcc3) were significantly increasedin the 10µM PBDE-47 and 10µM PBDE-47 + PCB153 groupscompared with the control (p < 0.05). The PBDE-47 groupscoincubated with NAC, however, considerably increased Xrcc1while decreasing Xrcc3 mRNA expression (p < 0.05). Theseresults indicate that PBDE-47 induced oxidative DNA damage andthat PBDE-47 combined with PCB153 may increase such effectsin SH-SY5Y cells in vitro. Furthermore, our results suggestthat oxidative stress is responsible for DNA damage inducedby PBDE-47.     

4种抗菌药物诱导肺炎克雷伯菌释放内毒素特性的研究

目的 :探讨4种抗菌药物不同浓度、不同作用时间诱导细菌释放内毒素能力的大小 ,为临床合理用药提供参考。方法 :选择4种敏感抗菌药物单独作用于肺炎克雷伯菌 ,测定1倍、10倍、100倍最低抑菌浓度 (MIC)作用于细菌1、2、4、8h释放内毒素的量。结果 :头孢曲松钠和美罗培南诱导细菌释放内毒素量最大 ,环丙沙星中等 ,庆大霉素较低 ;细菌释放内毒素量1倍MIC>10倍MIC>100倍MIC ;抗菌药物作用时间越长 ,内毒素释放的量越大。结论 :临床医师选用抗菌药物应参考其诱导细菌释放内毒素的特性。     

细菌DNA免疫增强作用的研究

探讨DNA佐剂对蛋白和多糖抗原的免疫增强作用。从大肠杆菌和黄杆菌中提取纯化DNA，采用超声波进行物理降解。制备不同分子量、不同剂量的DNA佐剂，混合抗原后对小鼠进行免疫。实验结果表明，DNA佐剂能诱导Th1型体液免疫反应；血清中IgG滴度和IgG2a／IgG1l的比率随DNA用量增加，DNA片断的增大而提高。