Quercetin-induced lipid peroxidation and DNA damage in isolated rat-liver nuclei

The extent of lipid peroxidation and DNA damage induced by quercetin were studied under aerobic conditions in isolated rat-liver nuclei. The effects of iron and copper ions on these two toxic oxidative processes were also investigated. Quercetin induced significant (P less than 0.05) concentration-dependent nuclear lipid peroxidation concurrent with DNA degradation; these effects were enhanced by iron and copper ions. The results suggest that the reactive oxygen species generated by quercetin autoxidation, catalyzed by iron and copper ions, are responsible for the concurrent lipid peroxidation and DNA damage in isolated rat-liver nuclei.     

Oxidative DNA damage, lipid peroxidation and nephrotoxicity induced in the rat kidney after ferric nitrilotriacetate administration.

8-Hydroxydeoxyguanosine (8-OH-dG) was examined in the kidneys of rats after single i.p. administration of ferric nitrilotriacetate (Fe-NTA) for variable periods of time at various doses along with the measurement of lipid peroxidation and serum biochemical parameters and histopathological examination. Though lipid peroxide level increased rapidly and decreased sharply after reaching a much higher peak 1 h after treatment, significant higher levels of 8-OH-dG were observed at 1, 6 and 24 h after injection. On the other hand, the increase of 8-OH-dG formation was observed in a similar dose-dependent manner to the appearance of nephrotoxic responses in terms of serum biochemical and histopathological changes.     

缺血修饰白蛋白检测对多柔比星心肌毒性诊断价值的探讨

目的:探讨缺血修饰白蛋白(IMA)对多柔比星导致急性心肌毒性反应早期诊断的临床意义,对多柔比星导致急性心肌毒性反应患者作出早期诊断及预测.方法:根据临床表现、心电图、患者IMA和肌钙蛋白T(cTnT)血清结果,将106例用多柔比星治疗的患者血清分为发病人群发病时血清(A组)、发病人群给药24 h后血清(B组)、高危人群给药24 h后血清(C组)及未发病人群给药24 h后血清(D组)4组血清,对4组中的血液指标检验结果升高例数进行组内及组间x2检验,比较其中差异;并对106例多柔比星诱发的急性心肌毒性患者的515份心电图与515份血清中的IMA指标升高例数进行相关性分析.结果:多柔比星急性心肌毒性反应患者发病时血清与正常人群血清IMA升高例数差异有统计学意义,x2 =30.94,P=0.00.多柔比星心肌毒性反应患者在用药24 h后与正常人群血清中IMA及cTnT升高例数差异有统计学意义(x2=36.43,P=0.00;x2=128.23,P=0.00).106例患者的515份异常心电图与515份血清中的IMA值异常例数进行相关性分析,结果提示发生急性心肌毒性反应患者中IMA的升高与心电图之间存在着相关性,r=0.783,P＜0.05.结论:多柔比星急性心肌毒性反应患者的IMA血液检验指标的异常提示,IMA检测对诊断多柔比星心肌毒性反应存在一定价值.     

Effects of antioxidants on quercetin-induced nuclear DNA damage and lipid peroxidation

The effects of catalase, superoxide dismutase, mannitol, glutathione, and diallyl sulfide on quercetin-induced DNA damage and lipid peroxidation were investigated in a model system of isolated rat-liver nuclei under aerobic conditions and in the presence of equimolar iron or copper. Mannitol produced a small but significant inhibition of the concurrent nuclear DNA damage and lipid peroxidation induced by quercetin in the presence of iron or copper. Catalase significantly decreased quercetin-induced nuclear DNA damage only in the presence of iron and had no significant effect on lipid peroxidation. Superoxide dismutase showed no significant effect on nuclear DNA damage, but stimulated the quercetin-induced lipid peroxidation only in the presence of copper. Glutathione significantly inhibited the nuclear lipid peroxidation but enhanced the DNA damage. Diallyl sulfide significantly enhanced the nuclear DNA damage but stimulated the lipid peroxidation only in the presence of iron. These results suggest that the reactive oxygen species, especially the hydroxyl radicals, are responsible for the concurrent lipid peroxidation and DNA damage induced by quercetin in the presence of iron or copper in isolated rat-liver nuclei.     

Enhanced adriamycin—induced delayed gastrointestinal radiosensitivity in tumor-bearing mice

Intestinal proliferative activity in BDF, mice bearing the Lewis lung tumor (LL_ca/BDF₁) was markedly depressed with increasing tumor burden. When compared with non-tumor-bearing mice (BDF₁), integrated cell production over 7 days was reduced to 56% in animals with small (400 mm³) tumors and to 30% in animals with large (2500 mm³) tumors. Gastrointestinal radiosensitivity was measured by proliferative compensatory response kinetics to a radiation dose of 600 rad. The presence of tumor (mean tumor volume = 859 ± 209 mm³) delayed the jejunal response to radiation by 24 hr and reduced the integrated cell production from 136% in BDF₁ mice to 119% in the LL_ca/BDF₁ mice. While the presence of tumor did not alter the temporal response of the colonic epithelium to radiation, the compensatory peak was reduced from 248% (BDF₁) to 200% (LL_ca/BDF₁). Adriamycin (Adr; 10 mg/kg) given 60 days prior to radiation failed to enhance the jejunal radiosensitivity in BDF₁ mice. However, when tumor-bearing LL_ca/BDF₁ mice were treated under an identical dose and time configuration, the jejunal response to 600 rad was significantly impaired: proliferative peaks were reduced from 182 to 115% ; integrated cell production was reduced from 119 to 72%. In the colon of tumor-bearing mice, pretreatment with AdR reduced the proliferative compensatory peak from the subsequent radiation dose to 120% of pretreatment levels.     

缬沙坦对阿霉素所致心肌细胞损伤保护作用的初步研究

目的 研究缬沙坦（valsartan,VAT）对阿霉素（doxorubicin,DOX）所致心肌细胞损伤的保护作用。方法 采用不同浓度（0.5 μmol/L、1 μmol/L、2 μmol/L、4 μmol/L）DOX处理H9C2心肌细胞建立DOX心肌细胞损伤模型。CCK-8法检测细胞存活率,流式细胞仪检测细胞凋亡率和细胞周期变化。结果 与对照组比较,不同浓度（0.5 μmol/L、1 μmol/L、2 μmol/L、4 μmol/L）DOX作用H9C2心肌细胞12 h和24 h,细胞存活率均随药物浓度增加而降低（P＜0.01）。其中1 μmol/L DOX作用24 h即可明显引起H9C2心肌细胞损伤,细胞存活率为（75.5±5.6）%,细胞凋亡率为（11.94±2.07）%,与对照组比较差异有统计学意义（P＜0.05）;10 μmol/L的 VAT预处理H9C2心肌细胞24 h后可明显抑制1 μmol/L DOX引起的心肌毒性作用,与单用 DOX组比较,细胞存活率升高［（74.2±3.0）% vs （89.3±4.0）%,P＜0.01］、细胞凋亡率下降［（13.15±6.07）% vs （11.01±3.17）%,P＜0.01］、G₀/G₁期细胞所占比例明显减少［（69.21±2.03）%  vs （50.28±1.21）%,P＜0.05］。结论 VAT能抑制DOX所致的心肌细胞损伤,保护作用可能与其调控细胞凋亡有关。     

右丙亚胺对表柔比星诱导大鼠心肌损伤干预机制研究

目的：比较不同剂量右丙亚胺对表柔比星诱导大鼠心肌损伤的干预作用及可能机制。方法：35只SD大鼠随机分5组,每组7只。对照组：于大鼠双侧腹腔分别注射生理盐水5mL/kg,间隔时间为30min,隔日1次,共4次（7d）;模型组（表柔比星＋生理盐水）：大鼠腹腔内注射表柔比星4.5mg/kg,隔日1次,共4次（7d）,注射表柔比星前30min于另一区域腹腔内注射生理盐水5mL/kg;表柔比星＋低、中、高剂量右丙亚胺组：大鼠腹腔内注射表柔比星4.5mg/kg,隔日1次,共4次（7d）,注射表柔比星前30min于另一区域腹腔内注射右丙亚胺45、67.5和90mg/kg,隔日1次,共4次（7d）。处死大鼠后检测各组大鼠心肌组织微量丙二醛（MDA）含量、总超氧化物歧化酶（T-SOD）活性、血浆乳酸脱氢酶（LDH）及肌钙蛋白I（cTnI）水平,观察心肌组织病理形态学改变及心肌细胞凋亡情况。结果：模型组较对照组SOD活性降低,分别为（75.10±5.14）和（101.81±13.21）U/mL,F=5.7,P=0.00;MDA含量升高,分别为（13.60±2.88）和（5.28±3.14）nmol/mg,F=7.31,P=0.00;血浆LDH升高,分别为（5.27±0.58）×103和（2.23±0.47）×103 U/L,F=23.7,P=0.00;cTnI升高,分别为（483.38±52.07）和（264.16±52.07）ρg/mL,F=20.13,P=0.00;心肌细胞病理评分升高,分别为2.70±0.20和0,F=8.65,P=0.00;凋亡指数明显升高,分别为（66.54±3.46）%和（1.55±0.74）%,F=126.86,P=0.00。而加用右丙亚胺各组均较模型组提高SOD活性,降低MDA、血浆LDH及cTnI含量,减少心肌病理评分及心肌细胞凋亡指数,P〈0.01或P〈0.05。结论：右丙亚胺对表柔比星诱导的大鼠心肌损伤有保护作用,其机制可能与减少氧自由基的产生、降低脂质过氧化物含量以及调节心肌细胞凋亡机制有关。     

Mammary tumors induced in rats by adriamycin and daunomycin

The two anthracycline antitumor antibiotics, Adriamycin and daunomycin (DM), induced a high incidence of mammary tumors, both fibroadenomas and adenocarcinomas, in female rats that received a single i.v. dose, thus confirming previous results. The incidence of DM-induced adenocarcinomas increased with the dose of the drug, whereas the incidence of Adriamycin-induced adenocarcinomas showed a plateau at 5 mg/kg and above. Adriamycin- and DM-induced fibroadenomas showed a peak at lower doses (about 5 to 6 mg/kg). With the highest DM dose (12.5 mg/kg) used, there was a slight prevalence of adenocarcinomas over fibroadenomas.     

Induction by estrogens of lipid peroxidation and lipid peroxidederived malonaldehyde-DNA adducts in male Syrian hamsters: role of lipid peroxidation in estrogen-induced kidney carcinogenesis

Estrogen-induced kidney carcinogenesis in male Syrian hamstershas previously been postulated to be mediated by free radicalsgenerated by redox cycling of catecholestrogen metabolites.As part of our examination of this hypothesis, we have studiedthe induction of lipid peroxidation and lipid peroxide-derivedmalondialdehyde (MDA)-DNA adducts in kidney and liver of hamsterstreated with single injections of diethylstilbestrol (DES) orwith estradiol (E implants for various lengths of time. Treatmentof hamsters with 54) and 100 mg/kg DES increased concentrationsof both lipid hydroperoxides and of MDA-DNA adducts. In hamsterstreated with E Implants for up to 50 days, lipid peroxide levelsin liver were double control values 3 h after hormone implantation,and then decreased to plateau values of 30% over controls. Thosein kidney rose to 2- to 3-fold above controls 3 days after hormoneimplantation and then decreased to plateau values of 51% abovecontrols. MBA-DNA adduct levels were two or three times higherthan those of controls in liver and kidney of hamsters treatedwith hormone implants for 3 and 7 days. Renal lipid peroxideconcentrations were raised by chronic treat ment with E₂, butnot by weakly carcinogenic estrogens ethinylestradiol or 2-fluoroestradiol.In contrast, MDA- DNA adduct levels were raised by all threesteroidal estrogens 3 days after estrogen implantation. Theincreases in lipid peroxides and in MDA-DNA adducts in estrogen-treated hamsters support a mechanism of carcinogenesis by freeradical generation via redox cycling of catcholestrogen metabolites.Lipid peroxides are postulated to play a dual role in estrogen-inducedcarcinogenesis, (i) as cofactors for cytochrome P450-mediatedformation of catecholestrogen metabolites and their redox cycling,and (ii) as precursors of MDA, a DNA adduct-forming endogenouselectrophile.     

Promotion of colon carcinogenesis through increasing lipid peroxidation induced in rats by a high cholesterol diet

To examine the influence of hypercholesteremia on 1,2-dimethylhydrazine (DMH)-induced rat colon cancer, Sprague-Dawley rats received dietary cholesterol (CH, 0–2%) and cholic acid (CA, 0.25%) with or without DMH (20 mg/kg, s.c. injection) for 18 weeks. The rats receiving dietary cholesterol and cholic acid all significantly increased total serum cholesterol and lipids but only a high cholesterol diet (2% CH plus 0.25% CA) decreased the activity of glutathione peroxidase (GSH-Px) and increased the formation of peroxides in the colon (P < 0.01). The rats that received the combination of DMH and high cholesterol diet enhanced these effects. At the end of the experiment, the diet group administered DMH and high cholesterol (2% CH plus 0.25% CA) developed colon adenoma at 50% of incidence in pathological examination, but no colon adenoma formed in the rats treated with high cholesterol alone. It is supposed that a non-carcinogenic agent like cholesterol may potentiate the carcinogenicity of DMH in rats via an increase of lipid peroxidation and decrease in the activity of peroxidase in the target organ.     

Trans-4-hydroxy-2-nonenal, an aldehydic lipid peroxidation product, lacks genotoxicity in lacI transgenic mice

In order to cast light on the significance of lipid peroxidation products for carcinogenesis, the lacI mutant frequency (MF), micronucleus induction and cell proliferation were analyzed in lacI transgenic mice treated with trans-4-hydroxy-2-nonenal (HNE), a typical example. Male mice were ip injected with HNE at doses of 0, 5 or 50 mg/kg bw and 48 h thereafter, peripheral blood was collected for analyzing micronucleus induction. After 14 days, the mice were sacrificed to allow tissue sampling for examination of lacI MF and cell proliferative activity. Sixty percent of the mice given 50 mg/kg HNE died within 5 days after the treatment, but no other mortalities were observed. Histopathologically, marked pulmonary hemorrhage was found in the 50 mg/kg HNE group mice that survived until day 14. Immunohistochemically, HNE-modified proteins were detected in their alveolar macrophages. The HNE treatment did not increase lacI MF in the liver, kidney and lung and no significant increase in micronucleus induction or cell proliferation in major organs was found in either treatment. Moreover, no tumors developed in the 5 mg/kg HNE-treated mice which survived until week 78. Our results thus indicate that HNE lacks in vivo genotoxicity in lacI transgenic mice even when lethal doses are applied.     

Inhibition of human erythrocyte inositol lipid metabolism by adriamycin

Incubation of human erythrocytes with Adriamycin prevented their morphological transition from discocytes to echinocytes when they were either depleted of ATP or loaded with calcium. This effect was dependent upon drug concentration and cell density. Adriamycin (10(-5) M) prevented, by greater than 90%, the echinocytosis of 10(7) cells/ml (S. B. Chahwala and J. A. Hickman, Cancer Res., 45: 4986-4989, 1985), and 5 X 10(-4) M prevented that of 10(9) cells/ml. There was a poor correlation between the effects of Adriamycin as a modulator of this morphological transition and its potency as an inhibitor of calmodulin. Using inside-out red blood cell vesicles, Adriamycin inhibited calmodulin dependent Ca2+ uptake with a 50% inhibitory concentration of 5 X 10(-4) M. Adriamycin thus differs from other amphipathic drugs, such as those of the phenothiazine class, where inhibition of calmodulin correlated well with effects on erythrocyte morphology (G. A. Nelson, M. L. Andrews, and M. J. Karnovsky, J. Cell Biol., 96: 730-735, 1983). After 12 h of ATP depletion, levels of phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2] extracted from 10(9) erythrocytes/ml fell by 57% and after 48 h they fell by 97%, changes which were coincident with a 100% transition of morphology to echinocytes. Adriamycin, 5 X 10(-4) M-1 X 10(-3) M, maintained 10(9) cells/ml in a discocyte morphology and maintained PtdIns(4,5)P2 levels at 60-70% of the time zero controls, independently of the size of the fall in PtdIns(4,5)P2 levels. The data suggested that Adriamycin inhibited a discrete pool of PtdIns(4,5)P2 which may be responsible for the maintenance of a discocyte morphology. Neomycin, 10(-3) M, had no effect on the ATP depletion-induced discocyte-echinocyte transition of 10(9) erythrocytes/ml or on the fall in PtdIns(4,5)P2 levels. Adriamycin, like neomycin, prevented the calcium-induced breakdown of erythrocyte membrane vesicle PtdIns(4,5)P2 to inositol trisphosphate (50% inhibitory concentration, 7 X 10(-4) M) but, unlike neomycin (50% inhibitory concentration, 4.25 X 10(-4) M) it was able to inhibit breakdown by 100% at higher concentrations.     

Microsomal lipid peroxidation induced by Adriamycin,epirubicin, daunorubicin and mitoxantrone: a comparative study

Summary Rat-liver microsomes and NADPH could reduce Adriamycin, epirubicin and daunorubicin to their free radical forms, which enhanced peroxidation of microsomal lipids less than 2-fold in air but 3- to 5-fold at a pO₂ of 4 mm Hg. Mitoxantrone was not reduced by microsomes and had no effect on microsomal peroxidation. Daunorubicin caused more lipid peroxidation than similar concentrations of either Adriamycin or epirubicin, which were equally efficient. In each case peroxidation was iron-dependent and could be catalysed by ferritin. The antioxidants -carotene and -tocopherol inhibited lipid peroxidation at low or high pO₂. The dose-for-dose difference in the cardiotoxicity of epirubicin compared with Adriamycin is not explained by its effect on microsomal lipid peroxidation. However, the lower incidence of cardiotoxicity with mitoxantrone may be a consequence of its inability to form free radical species and promote lipid peroxidation.     

Rapid lipid peroxidation in the nuclear fraction of rat liver induced by a diet deficient in choline and methionine

A diet deficient in choline and methionine, known to produce hepatocellular carcinoma in the absence of any added chemical carcinogen, induced lipid peroxidation in the nuclear fraction of the liver when fed to male Fischer 344 rats. This lipid peroxidation was detected within 1 day of feeding the diet by the appearance of diene conjugates and increased progressively up to 3 days. It was prevented completely by the addition of choline chloride to the diet. The close proximity of DNA may make it a possible target for attack by free radicals.     

Multiple tumoricidal effector mechanisms induced by adriamycin

The antitumor cytotoxic mechanisms of Adriamycin-elicited peritoneal exudate cells were investigated. Peritoneal exudate cells from mice collected 1 day after an i.p. injection of Adriamycin (10 mg/kg) displayed enhanced cytotoxicity against P815 (natural killer-insensitive, macrophage-sensitive) but not YAC-1 (natural killer-sensitive) tumor cell lines. These cells contained a sufficient concentration of the drug to be cytotoxic for P815 tumor cells in 18-hr chromium release assays. Freeze-thaw lysates of these peritoneal exudate cells were found to be as cytotoxic to P815 as their corresponding whole cells. The lytic activity of these lysates was removed by centrifugation at 100,000 X g, indicating the insolubility of the effector moiety. These cells were also shown to produce significant amounts of superoxide anion and H2O2 in response to phorbol myristate acetate. A catalase-inhibitable augmentation of the cytotoxicity of these cells against P815 was observed when phorbol myristate acetate was added to the assay. Neutrophils and not macrophages were likely responsible for this effect. Peritoneal lymphocytes from mice given injections of Adriamycin 5 to 7 days previously were cytotoxic to YAC-1 tumor cells in 4-hr assays. Finally, peritoneal macrophages harvested 5 to 7 days after Adriamycin administration were cytotoxic to P815 in the absence of detectable Adriamycin. The addition of phorbol myristate acetate inhibited the lysis of P815 by these cells.     

DNA damage and repair in tumour and non-tumour tissues of mice induced by nicotinamide.

In vivo DNA damage and repair was induced by nicotinamide (NAM) in adenotype 12 virus-induced mouse sarcoma A12B3 and sarcoma F inoculated into CBA mice. DNA damage, NAM and NAD concentrations were measured after in vivo exposure to NAM, in tumours and spleens by alkaline elution and by HPLC analysis. Our results indicate that NAM between 100-1000 mg kg-1 causes a high level of in vivo DNA strand breaks in tumours and normal tissues in mice bearing the immunogenic sarcoma A12B3 but not in the non-immunogenic sarcoma F. The repair process was also delayed by the NAM treatment probably owing to inhibition of the DNA repair enzyme, poly(ADP-ribose)polymerase, as evidenced by accumulation of NAM and NAD. These data are consistent with NAM having a mechanism of action as a radiosensitiser at least in part by DNA repair inhibition. In addition, it should also be considered that high doses of NAM might cause considerable complications to normal tissue in tumour-bearing individuals.     

Plasma xanthophyll carotenoids correlate inversely with indices of oxidative DNA damage and lipid peroxidation.

Post hoc analysis of data obtained from a study designed to modulate oxidative damage by dietary intervention revealed consistently strong inverse correlations between plasma xanthophyll carotenoids and oxidative damage indices. Thirty-seven women participated in a 14-day dietary intervention that increased mean vegetable and fruit (VF) consumption to approximately 12 servings/day. An additional 10 subjects participated in an intervention that limited VF consumption to less than four servings per day. 8-Hydroxy-2'-deoxyguanosine (8-OHdG) in DNA isolated from peripheral lymphocytes and 8-OHdG excreted in urine were measured as indices of oxidative DNA damage. Lipid peroxidation was assessed by measuring 8-epiprostaglandin F2alpha (8-EPG) in urine. Plasma levels of selected carotenoids were also determined, with the intention of using a-carotene as a biochemical index of VF consumption. Urinary 8-OHdG and 8-EPG were measured by ELISA, and plasma carotenoids were measured by high performance liquid chromatography. Lymphocyte 8-OHdG was measured by reverse phase high performance liquid chromatography with electrochemical detection. We observed that the structurally related xanthophyll carotenoids, lutein and beta-cryptoxanthin, which occur in dissimilar botanical families, were consistently inversely associated with these oxidative indices. Statistically significant inverse correlations were observed between plasma lutein and/or beta-cryptoxanthin levels and lymphocyte 8-OHdG and urinary 8-EPG. Moreover, an inverse correlation was observed between change in plasma xanthophylls and change in lymphocyte 8-OHdG concentration that occurred during the course of the study. These data lead us to hypothesize that lutein and beta-cryptoxanthin serve as markers for the antioxidant milieu provided by plants from which they are derived. Whether these carotenoids are directly responsible for the observed antioxidant phenomena merits further investigation.     

硼替佐米联合阿霉素诱导T细胞淋巴瘤细胞凋亡的实验研究

目的 探讨蛋白酶抑制剂硼替佐米对淋巴瘤细胞株Jurkat细胞的凋亡诱导作用及其可能机制,同时观察联合硼替佐米与阿霉素对Jurkat细胞有无协同作用.方法 采用四甲基偶氮唑蓝(MTT)比色法检测硼替佐米、阿霉素对Jurkat细胞体外生长的抑制作用,油镜下观察细胞形态变化.采用DNA的碘化丙啶(PI)染色及Annexin V-PI双标记法检测细胞凋亡率,Western blot法检测硼替佐米、阿霉素对Jurkat细胞caspase-3、caspase-8和聚(腺苷二磷酸-核糖)多聚酶(PARP)蛋白表达水平的影响.结果 10～320 ng/ml硼替佐米处理Jurkat细胞24、48和72 h,均能明显抑制细胞增殖,且生长抑制率与药物作用浓度呈正相关,硼替佐米作用24、48和72 h的相关系数分别为0.900、0.849和0.679(均P<0.01),呈浓度依赖性.以10～320 ng/ml硼替佐米单药或联合低剂量阿霉素(125ng/ml)处理Jurkat细胞24 h,硼替佐米的IC_(50)从(137.64±6.82)ng/ml降为(20.44±2.85)ng/ml.细胞凋亡率与硼替佐米的作用浓度呈正相关(P<0.01).硼替佐米作用Jurkat细胞后,caspase-3、caspase-8和PARP蛋白均出现明显的剪切带.结论 硼替佐米具有诱导Jurkat细胞凋亡的作用,通过外源性途径诱导凋亡是其机制之一.硼替佐米与阿霉素有一定的协同作用,二者联合能增强Jurkat细胞对硼替佐米的敏感性.