Inhibitory effects of Cassia tora L. on benzo[a]pyrene-mediated DNA damage toward HepG2 cells

The effects of water extracts from Cassia tora L. (WECT) treated with different degrees of roasting on benzo[a]pyrene (B[a]P)-induced DNA damage in human hepatoma cell line HepG2 were investigated via the comet assay without exogenous activation mixtures, such as S9 mix. WECT alone, at concentrations of 0.1-2 mg/mL, showed neither cytotoxic nor genotoxic effect toward HepG2 cells. B[a]P-induced DNA damage in HepG2 cells could be reduced by WECT in a dose-dependent manner (P < 0.05). At a concentration of 1 mg/mL, the inhibitory effects of WECT on DNA damage were in the order unroasted (72%) > roasted at 150 degrees C (60%) > roasted at 250 degrees C (23%). Ethoxyresorufin-O-dealkylase activity of HepG2 cells was effectively inhibited by WECT, and a similar trend of inhibition was observed in the order unroasted (64%) > roasted at 150 degrees C (42%) > roasted at 250 degrees C (18%). The activity of NADPH cytochrome P-450 reductase was also decreased by unroasted and 150 degrees C-roasted samples (50% and 38%, respectively). Furthermore, glutathione S-transferase activity was increased by treatment with unroasted (1.26-fold) and 150 degrees C-roasted (1.35-fold) samples at 1 mg/mL. In addition, the contents of anthraquinones (AQs) in WECT, including chrysophanol, emodin, and rhein, were decreased with increasing roasting temperature. Each of these AQs also demonstrated significant antigenotoxic activity in the comet assay. The inhibitory effects of chrysophanol, emodin, and rhein on B[a]P-mediated DNA damage in HepG2 cells were 78, 86, and 71%, respectively, at 100 microM. These findings suggested that the decreased antigenotoxicity of the roasted samples might be due to a reduction in their AQs content.     

莫能菌素对蚯蚓（Eisenia fetida）活体DNA的损伤研究

通过单细胞凝胶电泳实验研究了不同浓度莫能菌素暴露对赤子爱胜蚓（Eisenia fetida）体腔细胞DNA的损伤,结果显示,50mg.kg-1莫能菌素处理组尾部DNA含量值最大、尾长值最大、Olive尾矩值最大,分别为34.539%、107.736μm和29.354;随着莫能菌素暴露剂量的增加,尾部DNA含量、Olive尾矩和尾长损伤频率增加;尾部DNA含量对莫能菌素暴露最为敏感,对照组和各处理组的尾部DNA含量之间均存在显著性差异（P〈0.05）;对照组与15、25、50 mg.kg-1处理组的Olive尾矩和尾长损伤频率之间均存在显著性差异（P〈0.05）;暴露浓度与尾部DNA含量、Olive尾矩和尾长具有良好的剂量-效应关系（P〈0.05）。实验结果表明,蚯蚓体腔细胞DNA损伤可作为指示莫能菌素影响的生物标志物,彗星试验是检测莫能菌素暴露对赤子爱胜蚓活体基因损伤的有效手段。     

Zinc ion enhances GABA tea-mediated oxidative DNA damage

GABA tea is a tea product that contains a high level of γ-aminobutyric acid (GABA). Previous study has demonstrated a synergistic effect of GABA tea and copper ions on DNA breakage. This study further explored whether zinc (Zn), a nonredox metal, modulated DNA cleavage induced by GABA tea extract. In a cell-free system, Zn(2+) significantly enhanced GABA tea extract and (-)-epigallocatechin-3-gallate (EGCG)- or H(2)O(2)-induced DNA damage at 24 h of incubation. Additionally, low dosages of GABA tea extract (1-10 μg/mL) possessed pro-oxidant activity to increase H(2)O(2)/Zn(2+)-induced DNA cleavage in a dose-dependent profile. By use of various reactive oxygen scavengers, it was observed that glutathione, catalase, and potassium iodide effectively inhibited DNA degradation caused by the GABA tea extract/H(2)O(2)/Zn(2+) system. Moreover, the data showed that the GABA tea extract itself (0.5-5 mg/mL) could induce DNA cleavage in a long-term exposure (48 h). EGCG, but not the GABA tea extract, enhanced H(2)O(2)-induced DNA cleavage. In contrast, GABA decreased H(2)O(2)- and EGCG-induced DNA cleavage, suggesting that GABA might contribute the major effect on the antioxidant activity of GABA tea extract. Furthermore, a comet assay revealed that GABA tea extract (0.25 mg/mL) and GABA had antioxidant activity on H(2)O(2)-induced DNA breakage in human peripheral lymphocytes. Taken together, these findings indicate that GABA tea has the potential of both pro-oxidant and antioxidant. It is proposed that a balance between EGCG-induced pro-oxidation and GABA-mediated antioxidation may occur in a complex mixture of GABA tea extract.     

Hydrogen peroxide induced oxidative stress damage and antioxidant enzyme response in Caco-2 human colon cells

Studies were conducted to evaluate the cell damage caused by exposing human colon carcinoma cells, Caco-2, to hydrogen peroxide at concentrations varying from 0 to 250 microM for 30 min. Evaluation of cell viability, as measured by trypan blue dye exclusion test, showed that the loss of viability was < 5% at concentrations up to 250 microM hydrogen peroxide. Cell membrane damage and DNA damage as measured by the leakage of lactate dehydrogenase and the comet assay, respectively, were significantly high at concentrations >100 microM hydrogen peroxide compared to those of the control. Antioxidant mechanisms in Caco-2 cells were evaluated by measuring catalase, superoxide dismutase, and glutathione peroxidase activities. Catalase activities remained constant in cells treated with 50-250 microM hydrogen peroxide. Superoxide dismutase activity decreased, whereas glutathione peroxidase activity increased in cells treated with H(2)O(2) concentrations of >50 microM. This study showed that with increasing hydrogen peroxide concentration, cell membrane leakage and DNA damage increased, whereas the three antioxidant enzymes responded differently, as shown by mathematical models.     

Modulating effects of thyme and its major ingredients on oxidative DNA damage in human lymphocytes

The present study was carried out to investigate the modulating effects of thyme and its major components against the oxidative DNA damage induced by H(2)O(2). The human lymphocytes with thymol, carvacrol, and gamma-terpinene incubated with or without 0.1 mM H(2)O(2) for 30 min at 37 degrees C and the DNA damage were evaluated by singe cell gel electropheresis (comet assay). Concentrations above 0.1 mM thymol and gamma-terpinene and 0.05 mM carvacrol significantly induced DNA damage in human lymphocytes, but at the smaller concentrations no additional DNA strand breakage has been observed. At the all concentrations studied, gamma-terpinene did not show any protective effect against H(2)O(2) induced oxidative DNA damage, but the phenolic compounds thymol and carvacrol at concentrations below 0.2 and 0.1 mM, respectively, significantly reduced the oxidative DNA damage (p < 0.001). The n-hexane and ethyl acetate fractions prepared from the methanolic extracts of Thymus spicata also were found to inhibit DNA damage.     

纳米氧化锌对人胚肾HEK293细胞的遗传毒性

纳米氧化锌在人体内积蓄产生生物毒性,引起了人们对其安全性的重视.为了评价纳米氧化锌的遗传毒性,设置10、25、50、75和100 mg/L 5个剂量组的纳米氧化锌培养液与人胚肾细胞(HEK293细胞)分别接触培养12、24和48 h后,采用单细胞凝胶电泳(SCGE)试验分析纳米氧化锌对细胞DNA的损伤情况,体外微核试验检测细胞微核率.结果显示,随着培养基中纳米氧化锌浓度的增加,与对照组相比,染毒细胞头部DNA含量显著降低,尾部DNA含量、尾矩、Olive尾矩数值显著升高(P＜0.05);微核试验发现染毒组的微核率显著升高.研究结果提示,高浓度的纳米氧化锌可引起HEK293胚肾细胞DNA和染色体水平损伤,表现出遗传毒性效应,为纳米氧化锌的安全性提供了可靠的理论依据.     

Protective effects of fermented filtrate from Antrodia camphorata in submerged culture against CCl4-induced hepatic toxicity in rats

The protective effects and the possible mechanisms of dry matter of fermented filtrate (DMF) from Antrodia camphorata in submerged culture (ACSC) on H(2)O(2)-induced cytotoxicity in HepG2 and carbon tetrachloride (CCl(4))-induced hepatotoxicity in Sprague-Dawley rats were investigated. The results showed that the inhibitory effect of DMF and its crude triterpenoids on lipid peroxidation occurred in a dose-response manner in an AAPH/linoleic acid system. When HepG2 cells were pretreated with DMF at the concentration of 0.10 mg/mL for 4 h and then induced by 1 h of treatment with H(2)O(2) (100 microM), lipid peroxidation was significantly (p < 0.05) decreased, as measured by the formation of malondialdehyde. The oral pretreatment with DMF [0.25 and 0.50 mg/kg of body weight (bw)] for 5 consecutive days prior to the administration of a single dose of 40% CCl(4) (0.10 mL/100 g of bw, ip) significantly prevented the increase in serum levels of hepatic enzyme markers (alanine and aspartate aminotransferase) and liver lipid peroxidation (p < 0.05). Histopathological evaluation of the rat liver revealed that DMF reduced the incidence of liver lesions, including neutrophil infiltration, hydropic swelling, and necrosis induced by CCl(4) in rats. Moreover, reduced glutathione (GSH)-dependent enzymes (glutathione peroxidase, glutathione reductase, and glutathione S-transferase) and the GSH/GSSG ratio were significantly improved in the oral pretreatment DMF of rats (p < 0.01). The results suggest that DMF may play a role in preventing oxidative damage in living systems by up-regulating hepatic GSH-dependent enzymes to preserve the normal GSH/GSSH ratio and scavenging free radicals formed during CCl(4) metabolism.     

Effects of chlorophyll-related compounds on hydrogen peroxide induced DNA damage within human lymphocytes

Chlorophylls (Chl's) are the most abundant natural plant pigments. Four chlorophyll-related compounds (CRCs), including chlorophyllide a and b (Chlide a and b) and pheophorbide a and b (Pho a and b), were investigated for their antioxidative capacities to protect human lymphocyte DNA from hydrogen peroxide (H(2)O(2)) induced strand breaks and oxidative damage ex vivo. Lymphocytes exposed to H(2)O(2) at concentrations of 10 and 50 microM revealed an increased frequency of DNA single-strand breaks (ssb's; as measured by the comet assay) and also an increased level of oxidized nucleoside (as measured by 8-hydroxydeoxyguanosine, 8-OHdG). All Chl's reduced the level of DNA ssb's and 8-OHdG within human lymphocytes following exposure to 10 microM H(2)O(2). Only Pho a and b were able to decrease DNA ssb's and 8-OHdG following treatment of lymphocytes with 50 microM H(2)O(2), in a concentration-dependent fashion. It was demonstrated herein that Pho a and b were more antioxidative than others. We applied DPPH free-radical scavenge assays in vitro, and got similar results. Pho a and b had higher ability in scavenging capacities than others. We conclude that water-extract Chl's are able to enhance the ability of human lymphocytes to resist H(2)O(2)-induced oxidative damage, especially for Pho a and b.     

外源抗坏血酸和谷胱甘肽对荔枝保鲜效果的影响

为了探讨外源抗坏血酸（AsA）和谷胱甘肽（GSH）对荔枝保鲜效果的影响。分别在多菌灵500 mg/L+施保克 3 mL/L+ 柠檬酸15.0 mmol/L溶液中分别加入抗坏血酸50.0 mmol/L（AsA处理）和谷胱甘肽50.0 mmol/L（GSH处理）浸果5 min,以多菌灵500 mg/L+施保克3 mL/L+柠檬酸15.0 mmol/L浸果5 min为对照处理,分别置于常温和6℃低温贮藏、相对湿度为80%。结果表明：AsA和GSH处理均能降低荔枝果皮多酚氧化酶（PPO）活性和相对电导率;提高果肉的超氧化物岐化酶（SOD）和过氧化氢酶（CAT）活性;降低果肉过氧化氢（H2O2）和丙二醛（MDA）的含量,维持果肉中较高的维生素C和GSH含量。低温下,AsA和GSH处理均能提高荔枝保鲜效果,降低果实的腐烂率,但AsA处理的保鲜效果要优于GSH处理的效果,而常温下GSH处理并不能提高荔枝的保鲜效果。     

(-)-Anonaine induces DNA damage and inhibits growth and migration of human lung carcinoma h1299 cells

The anticancer effects of (-)-anonaine were investigated in this current study. (-)-Anonaine at concentration ranges of 50-200 μM exhibited significant inhibition to cell growth and migration activities on human lung cancer H1299 cells at 24 h, albeit cell cycle analyses showed that (-)-anonaine at the above concentration ranges did not cause any significant changes in cell-cycle distributions. Significant nuclear damages of H1299 cells were observed with 10-200 μM (-)-anonaine treatment in a comet assay, whereas higher concentrations (6 and 30 mM) of (-)-anonaine concentrations were required to cause DNA damages in an in vitro plasmid cleavage assay. In summary, our results demonstrated that (-)-anonaine exhibited dose-dependent antiproliferatory, antimigratory, and DNA-damaging effects on H1299 cells. We inferred that (-)-anonaine can cause cell-cycle arrest and DNA damage to hamper the physiological behavior of cancer cells at 72 h, and therefore, it can be useful as one of the potential herbal supplements for chemoprevention of human lung cancer.     

Influence of dietary quercetin on glutathione redox status in mice

Flavonoids such as quercetin have been shown to serve as a protective defense against oxidative damage in vivo. However, the bioavailability of quercetin depends on the food source and type of glycosidic moiety linked to the molecule. In this study, mice were fed 1 mg/day quercetin in the form of quercetin aglycone, rutin, apple, or onion, and reduced glutathione (GSH), oxidized glutathione (GSSG), and protein-GSH mixed disulfides were determined to investigate the influence of dietary quercetin on the GSH redox status in metabolically active tissues, mitochondria, and plasma of mice. All quercetin treatment groups produced increases in the GSH:GSSG ratio and decreases in mixed disulfide levels in hepatic tissue. Cardiac tissue did not change in response to dietary quercetin; however, cardiac mitochondria demonstrated a reduction in the GSH:GSSG ratio and an increase in protein mixed disulfide levels. No significant changes were observed in the plasma GSH:GSSG ratio, but mixed disulfide levels were decreased for all of the diets. The changes in plasma redox status did not parallel the changes in the tissues. Onion fed mice demonstrated the greatest increases in GSH:GSSG ratios and the greatest decreases in protein mixed disulfide levels of all diets compared. For all treatment groups, increases in the GSH:GSSG ratios corresponded with decreases in protein mixed disulfide levels. The results of this study indicate that quercetin influences GSH:GSSG ratios and protein thiolation in a tissue-specific manner and that these effects are dependent on food source and bioavailability.     

铀胁迫对大豆幼苗细胞DNA损伤的彗星试验研究

通过室内培养方法,用八氧化三铀配制浓度为100、200、400、600、800、1000μmol·L^-1的6种铀溶液和对照组培养大豆幼苗,采用彗星试验研究了铀胁迫对大豆幼苗细胞DNA的损伤情况。结果表明,铀溶液能对大豆幼苗细胞DNA造成损伤;配制的6种浓度的铀溶液均对大豆幼苗根部细胞DNA造成损伤,其中浓度为800和600μmol·L^-1的铀溶液对大豆幼苗根部细胞DNA的损伤最严重;铀浓度为1000μmol·L^-1的铀溶液对大豆幼苗茎部细胞DNA的损伤最严重;配制的6种浓度的铀溶液对大豆幼苗叶部细胞DNA的损伤不明显。     

Use of experimental design methodology to prepare Maillard reaction products from glucose and cysteine inhibitors of polyphenol oxidase from eggplant (Solanum melongena)

Polyphenol oxidase (PPO) from eggplant was extracted and partially purified by a two-step fractionation-precipitation using ammonium sulfate and phenylsepharose hydrophobic interaction chromatography. The eggplant PPO extract was characterized concerning its kinetic properties. Optimal conditions to obtain Maillard reaction products (MRPs) with a maximal inhibitory potency (IP) toward PPO activity were determined using the surface response methodology and a four-factor and five-level experimental design. The MRPs were prepared from cysteine (0.25 M) and glucose (0-1 M), at several initial pH values (2-6) and at differing heating times (3-19 h) and temperatures (95-115 degrees C). The maximal IP was obtained after heating a model system of glucose/cysteine (1/0.25 M) at pH 2 for 3 h 20 min at 115 degrees C. The soluble part of this MRP, called MRP(IPmax), was a noncompetitive inhibitor toward eggplant PPO. The IP of MRP(IPmax) on PPO activity was very potent as compared to that displayed by benzoic, p-coumaric, and t-cinnamic acids, as well as sorbic acid and 4-hexylresorcinol. The activity of preincubated PPO at 0 degrees C with MRP(IPmax) was only slightly restored after dialysis or gel filtration.     

Brassica oleracea L. Var. costata DC and Pieris brassicae L. aqueous extracts reduce methyl methanesulfonate-induced DNA damage in V79 hamster lung fibroblasts

Brassica oleracea L. var. costata DC leaves and Pieris brassicae L. larvae aqueous extracts were assayed for their potential to prevent/induce DNA damage. None of them was mutagenic at the tested concentrations in the Ames test reversion assay using Salmonella His(+) TA98 strains, with and without metabolic activation. In the hypoxanthine-guanine phosphoribosyltransferase mutation assay using mammalian V79 fibroblast cell line, extracts at 500 μg/mL neither induced mutations nor protected against the mutagenicity caused by methyl methanesulfonate (MMS). In the comet assay, none of the extracts revealed to be genotoxic by itself, and both afforded protection, more pronounced for larvae extracts, against MMS-induced genotoxicity. As genotoxic/antigenotoxic effects of Brassica vegetables are commonly attributed to isothiocyanates, the extracts were screened for these compounds by headspace-solid-phase microextraction/gas chromatography-mass spectrometry. No sulfur compound was detected. These findings demonstrate that both extracts could be useful against damage caused by genotoxic compounds, the larvae extract being the most promising.     

Modulation of hepatic drug metabolizing enzymes and oxidative status by rooibos (Aspalathus linearis) and Honeybush (Cyclopia intermedia), green and black (Camellia sinensis) teas in rats

Rooibos and honeybush teas significantly (P < 0.05) enhanced the activity of cytosolic glutathione S-transferase alpha. A significant (P < 0.05) to marginal (P < 0.1) increase in the activity of the microsomal UDP-glucuronosyl transferase was obtained with unprocessed rooibos and honeybush teas, respectively. Oxidized glutathione (GSSG) levels were significantly (P < 0.05) reduced in the liver of all tea treated rats while reduced glutathione (GSH) was markedly increased in the liver of the herbal tea treated rats. These changes resulted in a significant (P < 0.05) increase in the GSH/GSSG ratio by the unprocessed, processed rooibos and unprocessed honeybush teas. Green and black teas markedly to significantly decreased the oxygen radical absorbance capacity in liver homogenates, respectively. Modulation of phase II drug metabolizing enzymes and oxidative status in the liver may be important events in the protection against adverse effects related to mutagenesis and oxidative damage.     

Genotoxicity in the Offspring of Rats Exposed to Contaminated and Acidified Experimentally Soils

The aim of this study was to evaluate the genotoxic and mutagenic potential of contaminated soil diluted in acidic solutions and not acidic, in the offspring of rats exposed during pregnancy and neonatal periods. To this end, a comet assay and micronucleus test were performed. Soil samples were solubilized in the following three solvents: distilled water (control group), acid solvent at pH 5.2, and acid solvent at pH 3.6. Soil and solvent were mixed in a rate of 1:2 in g/mL, and hydrofluoric acid was used in the acid solvents. In the comet assay, the tail length, percentage of DNA within the tail and tail moment was analyzed in the whole blood of the pups that were studied. The number of micronuclei found in the bone marrow cells was counted in the micronucleus test. In all parameters evaluated in the comet assay, the group exposed to the lowest pH value when associated with contaminated soil (p < 0.05) had the most damage. However, the micronucleus test showed differences between all exposed groups and the control group (p < 0.05). These results reaffirm the health risks related to the exposure to soil contaminants.     

壬基酚对中华大蟾蜍蝌蚪的毒性效应

为评价水体中低浓度壬基酚（NP）对中华大蟾蜍（Bufo bufogargarizans）蝌蚪的毒性效应,采用室内饲养、测定方法,研究了NP在不同暴露时间（8、16、24、32d）,不同浓度（0.002、0.005、0.010mg·L-1）条件下,对蝌蚪生长发育、过氧化氢酶（CAT）活性、丙二醛（MDA）含量和DNA损伤的影响。结果表明：暴露于0.010mg·L-1NP的蝌蚪,其生长发育被极显著抑制（P〈0.01）;CAT活性在两个较低浓度组（0.002、0.005mg·L-1）表现为先诱导后恢复,在0.010mg·L-1浓度组表现为诱导-恢复-诱导（P〈0.05）;MDA含量在两个较低浓度组（0.002、0.005mg·L-1）持续升高（P〈0.05）,呈明显的时间-效应关系;各处理组DNA损伤水平均显著高于对照组（P〈0.05）,但随暴露时间的延长而有所下降,具有一定的剂量-效应关系。总之,即使水体中的NP符合灌溉标准,也可能抑制蝌蚪的生长发育,并对其造成氧化损伤和遗传损伤。     

微囊藻毒素MC-LR对罗非鱼（Oreochromis niloticus）肝脏谷胱甘肽含量及其相关酶活性的影响

采用腹腔注射的方式研究了微囊藻毒素MC-LR（注射剂量为500μg.kg-1 BW）胁迫下罗非鱼（Oreochromis niloticus）肝脏谷胱甘肽（GSH）含量及其相关酶γ-谷氨酰半胱氨酸合成酶（γ-GCS）、谷胱甘肽硫转移酶（GST）、谷胱甘肽还原酶（GR）和谷胱甘肽过氧化物酶（GPx）活性的动态变化。结果表明,MC-LR能够对罗非鱼肝脏GSH含量及其相关酶活性产生明显影响。在MC-LR胁迫下,与对照组相比,GSH含量呈现先下降后上升趋势,总体上被诱导;罗非鱼肝脏γ-GCS和GST在试验过程中出现两次显著升高现象,在GST作用下,GSH与MC-LR结合会造成GSH的消耗,γ-GCS和GR的活性增强能够使GSH含量升高,从而使罗非鱼肝脏GSH能够维持一定水平;GR和GPx的活性均表现为先上升后下降,它们能有效调节罗非鱼肝脏GSH-GSSG缓冲系统,从而在减轻或消除由MC-LR侵入而造成的肝细胞氧化胁迫中发挥重要作用。