菲啰啉对不同氧化剂和多柔比星所致CHL细胞DNA链断裂的影响

目的　为了研究菲口罗啉对 2种氧化剂和抗癌药多柔比星诱发细胞DNA损伤的影响 ,并初步探讨其损伤机制。方法　用不同浓度菲口罗啉预处理CHL细胞 30min ,再分别加入 3种不同染毒受试物 ,共同培养一定时间 (0 .3mmol·L- 1重铬酸钾 :10 5min ;0 .5μmol·L- 1多柔比星 :75min ;0 .4mmol·L- 1过氧化氢(H2 O2 ) :2 5min)后 ,用碱性单细胞凝胶电泳方法 (AS CGE)测定DNA链断裂情况 ,并同时以菲口罗啉与二甲亚砜 (DMSO ,0 .33mol·L- 1)比较对H2 O2 致DNA损伤中·OH的产生和清除。结果　 3种染毒受试物均可明显引起CHL细胞DNA链断裂 ;而当 3μmol·L- 1菲口罗啉预处理后 ,可使重铬酸钾、H2 O2 所致DNA迁移长度和细胞拖尾率明显降低 ,并超过DMSO降低H2 O2 的损伤作用 ,当菲口罗林浓度升至 12 μmol·L- 1时 ,可完全消除这两种因素所致的DNA链断裂损伤 ;10 μmol·L- 1菲口罗啉可抑制多柔比星所致DNA损伤 ,但浓度直至 60 μmol·L- 1仍不能完全消除多柔比星的损伤作用。结论　菲口罗啉对 2种氧化剂和多柔比星所致DNA损伤均有不同程度的防护作用 ,同时提示重铬酸钾和H2 O2 所致的DNA损伤主要与需过渡金属离子参与的·OH产生有关 ,而多柔比星所致损伤仅部分与此有关     

菲啰啉对氧化剂所致CHL细胞DNA链断裂的影响

目的　了解金属螯合剂菲啉对不同氧化相关因素所致ＣＨＬ细胞ＤＮＡ损伤的影响 ,初步探讨损伤机制。方法　ＣＨＬ细胞用不同浓度菲啉处理 30ｍｉｎ后加入氧化相关因素培养一定时间 (过氧化氢 :2 5ｍｉｎ ;重铬酸钾 :1ｈ 45ｍｉｎ ;阿霉素 :1ｈ 15ｍｉｎ) ,然后用ＳＣＧＥ(单细胞凝胶电泳 )测定ＤＮＡ链断裂情况 ,分析菲啉对ＤＮＡ氧化损伤的影响。结果　0 4ｍｍｏｌ ＬＨ2 Ｏ2 、0 .3ｍｍｏｌ ＬＫ2 Ｃｒ2 Ｏ7、0 5 μｍｏｌ Ｌ阿霉素均可明显引起ＣＨＬ细胞ＤＮＡ链断裂。当菲啉浓度为 3μｍｏｌ Ｌ时 ,对Ｈ2 Ｏ2 、Ｋ2 Ｃｒ2 Ｏ…     

1-17 菲啰啉对氧化剂所致CHL细胞DNA链断裂的影响

目的了解金属螯合剂菲啰啉对不同氧化相关因素所致CHL细胞DNA损伤的影响,初步探讨损伤机制.方法CHL细胞用不同浓度菲啰啉处理30min后加入氧化相关因素培养一定时间(过氧化氢25min;重铬酸钾1h45min;阿霉素1h15min),然后用SCGE(单细胞凝胶电泳)测定DNA链断裂情况,分析菲啰啉对DNA氧化损伤的影响.结果0.4mmol/LH2O2、0.3mmol/LK2Cr2O2、0.5μmol/L阿霉素均可明显引起CHL细胞DNA链断裂.当菲啰啉浓度为3μmol/L时,对H2O2、K2Cr2O2所致的损伤即有明显的保护作用;当浓度升至12μmol/L时,则可完全消除这两种因素所致的DNA链断裂损伤.当菲啰啉浓度为10、30μmol/L时,对阿霉素所致的损伤有明显保护作用;但浓度直至60μmol/L,仍不能完全消除阿霉素的损伤作用.结论菲啰啉对三者所致DNA损伤有不同程度的防保作用,提示H2O2和K2Cr2O2所致DNA损伤主要与过渡金属离子参与的@OH产生有关,而阿霉素所致损伤只有部分与此有关.     

共振瑞利散射法测定多柔比星脂质体中游离多柔比星的含量

目的:建立共振瑞利散射法(RRS)测定多柔比星(DOX)脂质体中游离 DOX 含量。方法:在 pH 2.6的 Britton-Robinson(BR)缓冲液中,刚果红(CR)与 DOX 通过分子间作用力形成离子缔合物,在λ_(ex)=λ_(em)=380 nm 波长时能使 RRS 信号显著增强。结果:RRS 法在1～12 μg·mL~(-1)范围内呈线性关系,检出限为0.05~(-1)g·mL~(-1)。对6个不同批号的 DOX 脂质体混悬液中游离 DOX 的含量测定,结果与紫外可见分光光度法相符,RSD(n=6)为1.9%～3.2%,平均回收率为94.3%。结论:此方法灵敏度较高,可以用于测定 DOX 脂质体中游离 DOX 的含量。     

多柔比星的毒性研究进展

多柔比星是一种具有细胞周期非特异性杀伤作用的蒽环类抗肿瘤抗生素,因其具有抗肿瘤谱广、活性强等特性,广泛用于治疗各种肿瘤。多柔比星主要用于急性白血病的治疗,对乳腺癌、肺癌、膀胱癌等多种肿瘤也有一定的疗效,但其强烈的细胞毒性作用对机体可产生广泛的生物化学反应,随着药物累积剂量的增加,其不良反应的发生率也相应增高。f     

褪黑素对多柔比星所致肾病的保护作用

本研究旨在观察褪黑素 ( MT)对大鼠“阿霉素肾病”的保护作用 .将所有受试动物分为 5组 ,即正常组 ,MT( d0 - d7,5mg·kg-1·d-1,ig)组 ,多柔比星模型组 ( d 1 ,Dox,5mg· kg-1,iv)组和MT( d 0 - d 7,0 .5,5mg· kg-1· d-1) + Dox( d 1 ,5mg· kg-1,iv)组 .检测大鼠 d7,d1 4,d2 1和 d2 8时尿蛋白 ,尿丙二醛排泄量和 d 2 8时血浆生化指标 .结果显示 ,Dox组大鼠呈典型的肾病综合征 ,MT+ Dox组大鼠尿蛋白减少 ,血浆蛋白明显回升 ,血脂降低 ;同时 ,Dox组动物尿丙二醛显著增加 ,MT+ Dox组丙二醛降低 .这些结果表明 ,MT可减轻“阿霉素肾病”大鼠肾损害 .     

表柔比星联用右丙亚胺和多柔比星脂质体对化疗所致心脏毒性的临床研究

目的观察表柔比星联用右丙亚胺和多柔比星脂质体对化疗所致心脏毒性的保护作用。方法将96例乳腺癌术后患者分为右丙亚胺组、多柔比星脂质体组和对照组,三组患者均接受以蒽环类药物为基础的术后辅助化疗方案4周期,通过监测各时期心电图异常率、心脏彩超左心室射血分数（LVEF）、短轴缩短率（SF）来评估心脏功能。结果对照组化疗后心电图异常率最高,右丙亚胺组、多柔比星脂质体组均能明显降低心电图异常率,且从化疗后1周期就显示出心脏保护作用;而LVEF及SF的变化在三组患者化疗前后差异无统计学意义（P〉0.05）。结论表柔比星化疗时加用DEX或者使用多柔比星脂质体对减轻蒽环类药物心脏毒性是有效且安全的。     

盐酸多柔比星的高效毛细管电泳法分析

用高效毛细管电泳法测定盐酸多柔比星的含量。考察不同试验条件下盐酸多柔比星的胶束动电毛细管电泳行为。选择５０ｍｍｏｌ／Ｌ十二烷基硫酸钠，５０ｍｍｏｌ／Ｌ硼 溶液（ｐＨ８．７）含１０％乙腈为运行缓冲液，以β－萘磺酸钠为内标，于２３１ｎｍ波长处测定。在０．１７～０．８５ｇ／Ｌ的范围内呈良好的线性关系，ｒ＝０．９９９４，重现性实验ＲＳＤ为２．５％（ｎ＝５）。     

药物对多柔比星诱导肝损伤保护作用的研究进展

多柔比星具有很好的抗肿瘤作用,但因其心、肝、肾等毒副作用明显而限制了其应用.近年来的研究表明,多柔比星诱导的肝损伤发生机制以脂质过氧化和自由基损伤为主,因此应用具有抗氧化作用的药物预防和治疗多柔比星诱导的肝损伤成为研究热点.目前已知多种抗氧化药物,如降脂类药物、维生素类药物、含硫醇类药物等都具有很好的肝保护作用,此外一些植物提取物也可以作为肝保护剂缓解多柔比星诱导的肝毒性.     

Zinc chromate induces chromosome instability and DNA double strand breaks in human lung cells

Hexavalent chromium Cr(VI) is a respiratory toxicant and carcinogen, with solubility playing an important role in its carcinogenic potential. Zinc chromate, a water insoluble or ‘particulate’ Cr(VI) compound, has been shown to be carcinogenic in epidemiology studies and to induce tumors in experimental animals, but its genotoxicity is poorly understood. Our study shows that zinc chromate induced concentration-dependent increases in cytotoxicity, chromosome damage and DNA double strand breaks in human lung cells. In response to zinc chromate-induced breaks, MRE11 expression was increased and ATM and ATR were phosphorylated, indicating that the DNA double strand break repair system was initiated in the cells. In addition, our data show that zinc chromate-induced double strand breaks were only observed in the G2/M phase population, with no significant amount of double strand breaks observed in G1 and S phase cells. These data will aid in understanding the mechanisms of zinc chromate toxicity and carcinogenesis.     

Production of DNA strand breaks in vitro and reactive oxygen species in vitro and in HL-60 cells by PCB metabolites.

PCBs are industrial chemicals that continue to contaminate our environment. They cause various toxic effects in animals and in exposed human populations. The mechanisms of toxicity, however, are not completely understood. PCBs are metabolized by cytochromes P450 to mono- and dihydroxylated compounds. Dihydroxy-PCBs can potentially be oxidized to the corresponding quinones. We hypothesized that reactive oxygen species (ROS) are produced by redox reactions of PCB metabolites. We tested several synthetic dihydroxy- and quinoid-PCBs with 1-3 chlorines for their potential to produce ROS in vitro and in HL-60 human leukemia cells, and DNA strand breaks in vitro. All dihydroxy-PCBs tested produced superoxide. The quinones generated superoxide only in the presence of GSH, probably during the autoxidation of the glutathione conjugates. We observed increased superoxide production with decreasing halogenation. Incubation of dihydroxy-PCBs or PCB quinones + GSH with plasmid DNA resulted in DNA strand break induction in the presence of Cu(II). Tests with various ROS scavengers indicated that hydroxyl radicals and singlet oxygen are likely involved in this strand break induction. Finally, dihydroxy- and quinoid PCBs also produced ROS in HL-60 cells in a dose- and time-dependent manner. We conclude that dihydroxylated PCBs, and PCB quinones after reaction with GSH, produce superoxide and other ROS both in vitro and in HL-60 cells, and oxidative DNA damage in the form of DNA strand breaks in vitro. The reactions seen in vitro and in cells may well be a predictor of the toxicity of PCBs in animals.     

DNA strand breaks and apoptosis induced by oxaliplatin in cancer cells

Platinum anticancer drugs, such as cisplatin, are thought to exert their activity by DNA damage. Oxaliplatin, a clinically active diaminocyclohexane platinum compound, however, requires fewer DNA-Pt adducts than cisplatin to achieve cell growth inhibition. Here we investigated whether secondary DNA damage and apoptotic responses to oxaliplatin compensate for the reduced formation of DNA adducts. Oxaliplatin treatment of leukemic CEM and ovarian A2780 cancer cells resulted in early (4 hr) induction of DNA single-strand breaks measured by nucleoid sedimentation. These infrequent early lesions progress with time into massive double-stranded DNA fragmentation (fragments >50k bp) paralleled by characteristic apoptotic changes revealed by cell morphology and multivariate flow cytometry. Profound oxaliplatin-induced apoptotic DNA fragmentation was detectable following a 24 hr treatment of A2780 and CEM cells with 2 and 10 microM oxaliplatin, respectively. This DNA fragmentation was inhibited completely by the broad-spectrum caspase inhibitor Z-VAD-fmk. Cisplatin, which forms markedly more DNA-Pt adducts in CEM and A2780 cells than equimolar oxaliplatin, was similarly potent as oxaliplatin in terms of early strand breaks and later apoptotic responses. Oxaliplatin was also profoundly apoptotic in several other tumor cell lines of prostate origin but had only a marginal effect in normal prostate PrEC cells. Collectively, the results demonstrate that, relative to the magnitude of the primary DNA-Pt lesions, oxaliplatin is disproportionately more potent than cisplatin in the induction of apoptosis. Apoptosis induction, possibly enhanced by a contribution of targets other than DNA, seems to be an important factor in the mechanism of action of oxaliplatin.     

甲基丙烯酸环氧丙酯体外诱导的人胚肺成纤维细胞DNA链断裂

甲基丙烯酸环氧丙酯 (GMA)导致人胚肺成纤维细胞 (HELFs)恶性转化及其潜在致癌机理尚未阐明 .本研究用 0 .5- 5.0 mg· L-1的 GMA给体外培养的 HELFs染毒 2 h和 1 2 h,或用 5.0 mg· L-1的 GMA染毒 1 5min- 2 4 h,应用单细胞凝胶电泳技术 (彗星试验 )对 GMA引起的 HELFs DNA断裂作用进行了初步探讨 .琼脂糖凝胶电泳及流式细胞仪检测等方法观察了 GMA对细胞凋亡的诱导作用 .结果表明 ,GMA可导致染毒细胞 DNA发生剂量和时间依赖性链断裂 .用 5.0 mg· L-1的 GMA染毒后仅 1 h断裂作用即已显著 ,并随染毒时间延长而递增 ,至 2 4 h时损伤最为严重 .但在同样条件下未观察到 GMA对细胞凋亡的诱导作用 .结果提示 GMA导致的非凋亡性 DNA链断裂可能是其诱导 HELFs恶性转化早期重要的遗传事件之一     

甲基丙烯酸环氧丙酯体外诱导的人胚肺成纤维细胞DNA链断裂

甲基丙烯酸环氧丙酯（GMA）导致人胚肺成纤维细胞（HELFs）恶性转化及其潜在致癌机理尚未阐明. 本研究用0.5-5.0 mg·L^-1的GMA给体外培养的HELFs染毒2 h和12 h,或用5.0 mg·L^-1的GMA染毒15 min-24 h, 应用单细胞凝胶电泳技术（彗星试验）对GMA引起的HELFs DNA断裂作用进行了初步探讨. 琼脂糖凝胶电泳及流式细胞仪检测等方法观察了GMA对细胞凋亡的诱导作用. 结果表明, GMA可导致染毒细胞DNA发生剂量和时间依赖性链断裂. 用5.0 mg·L^-1的GMA染毒后仅1 h断裂作用即已显著, 并随染毒时间延长而递增, 至24 h时损伤最为严重. 但在同样条件下未观察到GMA对细胞凋亡的诱导作用. 结果提示GMA导致的非凋亡性DNA链断裂可能是其诱导HELFs恶性转化早期重要的遗传事件之一.     

Carcinogen metabolism and carcinogen-induced strand breaks in DNA of isolated liver cells

Viable rat hepatocytes were isolated and incubated for various times in suspension or monolayer. RNA and DNA synthesis and the contents of cytochromes P₄₅₀ and b₅, as well as microsomal functions, were studied throughout the incubation. Incubation with dimethylnitrosamine (DMN) resulted in protein and RNA alkylation that were linear with time over a period of 4 hours. Correspondingly the appearance of single strand breaks in DNA could be demonstrated following in vivo or in vitro administration of the carcinogen.     

Low concentration of arsenite exacerbates UVR-induced DNA strand breaks by inhibiting PARP-1 activity

Epidemiological studies have associated arsenic exposure with many types of human cancers. Arsenic has also been shown to act as a co-carcinogen even at low concentrations. However, the precise mechanism of its co-carcinogenic action is unknown. Recent studies indicate that arsenic can interfere with DNA-repair processes. Poly(ADP-ribose) polymerase (PARP)-1 is a zinc-finger DNA-repair protein, which can promptly sense DNA strand breaks and initiate DNA-repair pathways. In the present study, we tested the hypothesis that low concentrations of arsenic could inhibit PAPR-1 activity and so exacerbate levels of ultraviolet radiation (UVR)-induced DNA strand breaks. HaCat cells were treated with arsenite and/or UVR, and then DNA strand breaks were assessed by comet assay. Low concentrations of arsenite (≤ 2 µM) alone did not induce significant DNA strand breaks, but greatly enhanced the DNA strand breaks induced by UVR. Further studies showed that 2 µM arsenite effectively inhibited PARP-1 activity. Zinc supplementation of arsenite-treated cells restored PARP-1 activity and significantly diminished the exacerbating effect of arsenite on UVR-induced DNA strand breaks. Importantly, neither arsenite treatment, nor zinc supplementation changed UVR-triggered reactive oxygen species (ROS) formation, suggesting that their effects upon UVR-induced DNA strand breaks are not through a direct free radical mechanism. Combination treatments of arsenite with PARP-1 inhibitor 3-aminobenzamide or PARP-1 siRNA demonstrate that PARP-1 is the target of arsenite. Together, these findings show that arsenite at low concentration exacerbates UVR-induced DNA strand breaks by inhibiting PARP-1 activity, which may represent an important mechanism underlying the co-carcinogenicity of arsenic.     

Effects of Brussels sprouts extracts on hydrogen peroxide-induced DNA strand breaks in human lymphocytes

Aqueous Brussels sprouts extracts inhibit oxidation of isolated DNA in vitro, possibly through scavenging oxygen radicals. We have studied the effect of preincubating human lymphocytes with aqueous extracts of raw, cooked and autolysed Brussels sprouts and the glucosinolate, sinigrin, on hydrogen peroxide-induced DNA damage, strand breaks and base oxidation, in vitro by means of the Comet assay. DNA repair enzymes endonuclease III (EndoIII) and formamidopyrimidine–DNA glycosylase (FPG) were used to examine the levels of oxidised pyrimidines and purines in DNA, respectively. Aqueous extracts of cooked and autolysed Brussels sprouts and sinigrin decreased DNA strand breaks in human lymphocytes exposed to 100 μ H₂O₂ for 5 min on ice, although the level of EndoIII and FPG sensitive sites was not reduced. The maximum inhibition was by 38 and 39% at concentrations of cooked and autolysed extracts of 10 μg/ml and 5 μg/ml, respectively, whereas the inhibitory effect decreased with increasing concentrations up to 100 μg/ml. The maximum inhibition by sinigrin was by 54% at 2 μg/ml. Extracts of raw Brussels sprouts or green beans had no DNA-protective effect. The results indicate that compounds, including sinigrin, in cooked and autolysed Brussels sprouts can enhance lymphocyte resistance towards H₂O₂-induced DNA strand breaks in vitro.     

Synergistic cytotoxicity and DNA strand breaks in cells and plasmid DNA exposed to uranyl acetate and ultraviolet radiation

Depleted uranium (DU) has a chemical toxicity that is independent of its radioactivity. The purpose of this study was to explore the photoactivation of uranyl ion by ultraviolet (UV) radiation as a chemical mechanism of uranium genotoxicity. The ability of UVB (302 nm) and UVA (368 nm) radiation to photoactivate uranyl ion to produce single strand breaks was measured in pBR322 plasmid DNA, and the presence of adducts and apurinic/apyrimidinic sites that could be converted to single strand breaks by heat and piperidine was analyzed. Results showed that DNA lesions in plasmid DNA exposed to UVB‐ or UVA‐activated DU were only slightly heat reactive, but were piperidine sensitive. The cytotoxicity of UVB‐activated uranyl ion was measured in repair‐proficient and repair‐deficient Chinese hamster ovary cells and human keratinocyte HaCaT cells. The cytotoxicity of co‐exposures of uranyl ion and UVB radiation was dependent on the order of exposure and was greater than co‐exposures of arsenite and UVB radiation. Uranyl ion and UVB radiation were synergistically cytotoxic in cells, and cells exposed to photoactivated DU required different DNA repair pathways than cells exposed to non‐photoactivated DU. This study contributes to our understanding of the DNA lesions formed by DU, as well as their repair. Results suggest that excitation of uranyl ion by UV radiation can provide a pathway for uranyl ion to be chemically genotoxic in populations with dermal exposures to uranium and UV radiation, which would make skin an overlooked target organ for uranium exposures. Copyright © 2014 John Wiley & Sons, Ltd.