Direct human DNA protection by Coriolus versicolor (Yunzhi) extract

Abstract

Context and objective: Scientific evidence has shown Coriolus versicolor (L. ex Fr.) Quel (also known as Yunzhi) has the role of immunomodulator in therapeutic effect. The aim of this in vitro study was to investigate the antioxidative effect of Yunzhi and to explore the mechanisms behind its DNA protection.

Materials and methods: Commercial Yunzhi extract was dissolved in water and diluted in five concentrations (10¹–10⁵?μg/L) with appropriate buffers. Lymphocytes harvested from three healthy subjects were incubated with Yunzhi extract for 30?min. Cells were then subjected to 5?min oxidant challenge by 45?μM hydrogen peroxide. The standard alkaline comet (SAC) assay and lysed cell comet (LCC) assay were performed in parallel. DNA damage of each treatment was scored under a fluorescence microscope and compared with the cells without Yunzhi pretreatment.

Results: U-shaped dose–response was seen in both versions of the comet assay. Yunzhi at 10⁴?μg/L demonstrated a genoprotective effect against oxidative damage in the SAC assay (25% decrease in comet score). In the LCC assay, a trend of protection in lymphocytes was observed but it did not reach statistical significance.

Conclusion: A direct antioxidant effect of Yunzhi against oxidant challenge on the DNA of lymphocytes was evidenced. The active component in Yunzhi was likely to be membrane permeable.     

Genotoxic and chemopreventive assessment of Cynara scolymus L. aqueous extract in a human-derived liver cell line

Cynara scolymus L., popularly known as artichoke, is consumed as food and used as tea infusions for pharmacological purposes to treat liver dysfunctions and other conditions. Scientific data on the safety and protective effect of artichoke in human-derived liver cells is missing. This study investigated the genotoxic and modulatory effect of a liophilized extract suspended in water of C. scolymus L. leaves. Four extract concentrations (0.62, 1.25, 2.5 and 5.0?mg/mL) were evaluated using the comet assay on human hepatocyte cultures, HepG2 cells. Genotoxicity was assessed after two treatment periods, 1 and 24?h. Antigenotoxicity was evaluated against oxidative lesions induced by hydrogen peroxide in pre-, simultaneous and post-treatment protocols. Artichoke leaves aqueous extract induced genotoxic effects in HepG2 cells after 1- and 24-h treatments. In turn, extract concentrations of 0.62, 1.25 and 2.5?mg/mL, exhibited a protective effect in pretreatment, compared to hydrogen peroxide alone. However, in simultaneous and post-treatment protocols, only the lowest concentration reduced the frequency of DNA damage induced by hydrogen peroxide. In addition, in the simultaneous treatment protocol, the highest artichoke extract concentration increased hydrogen peroxide genotoxicity. It can be concluded that artichoke is genotoxic, in vitro, to HepG2 cells, but can also modulate hydrogen peroxide DNA damage.     

Protective effect of amifostine on busulfan induced DNA damage in human hepatoma cells

Busulfan is one of the most effective chemotherapeutic agents used for the treatment of chronic myeloid leukemia. However, as a bifunctional alkylating agent, during clinical use several side effects may occur. In addition, several in vivo and in vitro studies of busulfan have shown a range of genotoxic effects including DNA strand break and inhibition of DNA synthesis. Amifostine, an organic thiophosphate compound, has been shown to exert an important cyto-protective effect in many tissues. The aim of this study was to explore whether amifostine protects against busulfan-induced genotoxicity in HepG2 cell line. Our results showed that amifostine reduced the genotoxic effects of busulfan significantly in both type of experiment conditions, as measured via comet assay. Furthermore, amifostine decreased the intracellular ROS generation induced by busulfan and also increased the intracellular GSH levels in HepG2 cells. Altogether, our results suggest a protective action of amifostine against busulfan cytotoxicity and genotoxicity via various pathways. The most protective effect was observed with amifostine when it was administrated 24?h before busulfan treatment.     

Protective effect of basil (Ocimum basilicum L.) against oxidative DNA damage and mutagenesis.

Mutagenic and antimutagenic properties of essential oil (EO) of basil and its major constituent Linalool, reported to possess antioxidative properties, were examined in microbial tests. In Salmonella/microsome and Escherichia. coli WP2 reversion assays both derivatives (0.25–2.0 μl/plate) showed no mutagenic effect. Salmonella. typhimurium TA98, TA100 and TA102 strains displayed similar sensitivity to both basil derivatives as non-permeable E. coli WP2 strains IC185 and IC202 oxyR. Moreover, the toxicity of basil derivatives to WP2 strains did not depend on OxyR function. The reduction of t-BOOH-induced mutagenesis by EO and Linalool (30–60%) was obtained in repair proficient strains of the E. coli K12 assay (Nikoli?, B., Stanojevi?, J., Miti?, D., Vukovi?-Ga?i?, B., Kne?evi?-Vuk?evi?, J., Simi?, D., 2004. Comparative study of the antimutagenic potential of vitamin E in different E. coli strains. Mutat. Res. 564, 31–38), as well as in E. coli WP2 IC202 strain. EO and Linalool reduced spontaneous mutagenesis in mismatch repair deficient E. coli K12 strains (27–44%). In all tests, antimutagenic effect of basil derivatives was comparable with that obtained with model antioxidant vitamin E. Linalool and vitamin E induced DNA strand breaks in Comet assay on S. cerevisiae 3A cells, but at non-genotoxic concentrations (0.075 and 0.025 μg/ml, respectively) they reduced the number of H₂O₂-induced comets (45–70% Linalool and 80–93% vitamin E). Obtained results indicate that antigenotoxic potential of basil derivatives could be attributed to their antioxidative properties.     

Protective effects of isothiocyanates towards N-nitrosamine-induced DNA damage in the single-cell gel electrophoresis (SCGE)/HepG2 assay

The aim of this study was to investigate the protective effect of isothiocyanates towards N-nitrosamine-induced DNA damage in the single-cell gel electrophoresis (SCGE)/HepG2 assay. None of the isothiocyanates (ITCs) concentrations tested in the presence or absence of formamidopyrimidine-DNA glycosylase (Fpg) caused DNA damage per se. Combined treatments of HepG2 cells with phenethyl isothiocyanate (PEITC), allyl isothiocyanate (AITC) or indol-3-carbinol (I3C) and N-nitrosopyrrolidine (NPYR) or N-nitrosodimethylamine (NDMA) reduced the genotoxic effects of the N-nitrosamines in a dose-dependent manner. The protective effect of the three ITCs tested was higher towards NPYR-induced oxidative DNA damage than against NDMA. The greatest protective effect towards NPYR-induced oxidative DNA damage was shown by I3C (1 microM, 79%) and by PEITC (1 microM, 67%) and I3C (1 microM, 61%) towards NDMA (in presence of Fpg enzyme). However, in absence of Fpg enzyme, AITC (1 microM, 72%) exerted the most drastic reduction towards NPYR-induced oxidative DNA damage, and PEITC (1 microM, 55%) towards NDMA. Our results indicate that ITCs protect human-derived cells against the DNA damaging effect of NPYR and NDMA, two carcinogenic compounds that occur in the environment.     

阿魏酸乙酯对过氧化氢损伤人血管内皮细胞的保护作用

目的 :观察阿魏酸乙酯对人血管内皮细胞的保护作用。方法 :实验分对照组、模型组、阿魏酸 4个剂量组、阿魏酸乙酯 4个剂量组。过氧化氢诱导血管内皮细胞损伤 ,用四甲基偶氮唑蓝比色法检测细胞存活数量 ,用比色法测细胞培养液中过氧化物丙二醛含量及乳酸脱氢酶的活性。结果 :阿魏酸乙酯及阿魏酸不同剂量 (2 ,4 ,8,16nmol·L-1)对过氧化氢损伤的人血管内皮细胞具有保护作用 ,可抑制过氧化氢损伤的血管内皮细胞释放乳酸脱氢酶和丙二醛。相同剂量组的阿魏酸乙酯的作用强于阿魏酸。结论 :阿魏酸乙酯具有减轻过氧化氢损伤 ,保护血管内皮细胞的作用 ,并且其作用强于阿魏酸     

Protective effects of kahweol and cafestol against hydrogen peroxide-induced oxidative stress and DNA damage

There is an increasing evidence that oxidative stress is implicated in the processes of inflammation and carcinogenesis. It has been shown that kahweol and cafestol, coffee-specific diterpenes, exhibit chemoprotective effects. This study investigated the effects of kahweol and cafestol, coffee-specific diterpenes, on the hydrogen peroxide (H(2)O(2))-induced oxidative stress and DNA damage in NIH3T3 cells. When the cells were treated with kahweol or cafestol, cytotoxicity, lipid peroxidation, and reactive oxygen species production induced by H(2)O(2) were markedly reduced in a dose-dependent manner. Moreover, kahweol and cafestol were shown to be highly protected against H(2)O(2)-induced oxidative DNA damage as determined by the Comet (single cell gel electrophoresis) assay and the measurement of 8-oxoguanine content in NIH3T3 cells. Kahweol and cafestol also protected hydroxyl radical-induced 2-deoxy-d-ribose degradation by ferric ion-nitrilotriacetic acid and H(2)O(2). In addition, kahweol and cafestol efficiently removed the superoxide anion generated from the xanthine/xanthine oxidase system. These results suggest that kahweol and cafestol are effective in protecting against H(2)O(2)-induced oxidative stress and DNA damage, probably via scavenging free oxygen radicals, and that kahweol and cafestol act as antioxidants.     

In vivo protective efficacy of astaxanthin against ionizing radiation-induced DNA damage

Astaxanthin, a carotenoid pigment, is believed to be effective in the repair of DNA damage. Our study evaluates the effect of astaxanthin on DNA damage in rats exposed to whole-body radiotherapy using the comet assay. Thirty-two male rats were randomly divided into four groups (control, ionizing radiation, astaxanthin, and radiation+astaxanthin). The radiation and radiation+astaxanthin groups were exposed to X-rays at a dose of 8 gray (0.62 gray/min). Astaxanthin was administered at 4 mg/kg by gavage for 7 days starting from irradiation. The %TailDNA parameter was chosen as an indicator of DNA damage and the results were compared using one-way ANOVA. %TailDNA was 3.24 ± 3.12 in the control group, 2.85 ± 2.73 in the astaxanthin group, 4.11 ± 7.90 in the radiation group, and 3.59 ± 4.05 in the radiation+astaxanthin group. There was a significant increase in DNA damage in the radiation group, compared with the control and astaxanthin groups (p < .001). DNA damage was reduced in the radiation+astaxanthin group compared with the radiation group (p < .05). Although this decrease did not reduce damage to the level of the control group, it was significant. The decrease in radiation-induced DNA damage by astaxanthin administration in our study supports the hypothesis that astaxanthin is a promising agent for against/reducing DNA damage.     

Ebselen拮抗H_2O_2对HL-60细胞的毒性及DNA损伤作用(英文)

目的:以H_2O_2造成人白血病HL-60细胞的氧化损伤,研究含硒化合物Ebselen(Ebs)对氧化性损伤的影响。方法:MTT比色法测定细胞增殖,TBA法检测膜脂过氧化水平,单细胞电泳法确定DNA损伤程度,并用荧光探针DCFH-DA检测细胞内活性氧自由基(ROS)水平的变化。结果:H_2O_2(100μmol·L~(-1)~)可显著抑制HL-60细胞增殖,引起膜脂过氧化水平升高,Ebs对此表现出浓度依赖性的抑制效应,Ebs20μmol·L~(-1)对H_2O_2 100μmol·L~(-1)引起的膜脂过氧化水平抑制率达56.4％;同样,Ebs对H_2O_2 100μmol·L~(-1)造成的DNA损伤和细胞内活性氧水平的升高均有拮抗效应,也呈明显量效关系。结论:含硒化合物Ebs对由活性氧诱发的细胞毒性和DNA损伤有强的防护效应。     

Protective effects of enalapril in streptozotocin-induced diabetic rat: studies of DNA damage, apoptosis and expression of CCN2 in the heart, kidney and liver

Diabetes mellitus is characterized by hyperglycemia, which induces oxidative stress and perturbs a number of pathways, leading to tissue injury. One of the pathological responses to tissue injury is the development of fibrosis and cell death. Enalapril is a non-thiol angiotensin-converting enzyme inhibitor that is commonly used in the treatment of diabetes-associated hypertension. The present study examines the possible beneficial effects of enalapril on the development of diabetes associated fibrosis and DNA damage in rats. Sprague-Dawley rats (250 ± 10 g) were used in the study. Enalapril (10 mg kg(-1) per oral) was administered for four consecutive weeks to the streptozotocin (STZ)-induced diabetic rats. After 4 weeks, all the animals were sacrificed and comet assay (normal and modified) was performed to detect the normal as well as oxidative DNA damage. Expression of profibrotic marker CCN2 and fibrosis was examined in the heart, kidney and liver of diabetic rats. Enalapril treatment significantly restored the malondialdehyde and glutathione content as well as the DNA damage in the heart, kidney and liver of diabetic rat. Significant decrease in the expression of CCN2 was observed in the heart, kidney and liver of diabetic rat receiving enalapril treatment as compared with the diabetic group. Further, the enalapril treatment led to significant decrease in the fibrosis and CCN2 expression in the diabetic group as compared with control. The results of the present study clearly demonstrate that enalapril ameliorates the DNA damage, cell death and expression of CCN2 in the heart, kidney and liver of the STZ-induced diabetic rat.     

DNA damage in brain cells and behavioral deficits in mice after treatment with high doses of amantadine

Amantadine (AMA) is an uncompetitive antagonist of the N‐methyl‐d ‐aspartate receptor, with clinical application, acting on treatment of influenza A virus and Parkinson's disease. It has been proposed that AMA can indirectly modulate dopaminergic transmission. In high doses, the central nervous system is its primary site of toxicity. To examine deleterious effects on CNS induced by AMA, this study evaluated possible neurobehavioral alterations induced by AMA such as stereotyped behavior, the effects on locomotion and memory and its possible genotoxic/mutagenic activities. Adult male CF‐1 mice were treated with a systemic injection of AMA (15, 30 or 60 mg kg^?1) 20 min before behavioral tasks on open field and inhibitory avoidance. Higher AMA doses increased the latency to step‐down inhibitory avoidance test in the training session in the inhibitory avoidance task. At 60 mg kg^?1 AMA induced impairing effects on locomotion and exploration and hence impaired habituation to a novel environment. Stereotyped behavior after each administration in a 3‐day trial was observed, suggesting effects on dopaminergic system. Amantadine was not able to induce chromosomal mutagenesis or toxicity on bone marrow, as evaluated by the micronucleus assay. At the lowest dose tested, AMA did not induce DNA damage and it was unable to impair memory, locomotion, exploration or motivation in mice. However, higher AMA doses increased DNA damage in brain tissue, produced locomotor disturbances severe enough to preclude testing for learning and memory effects, and induced stereotypy, suggesting neurotoxicity.     

纳米银对HL-7702肝细胞DNA的损伤作用

目的探讨纳米银对HL-7702肝细胞DNA的损伤效应。方法设对照组、微米银组和纳米银组,微米银和纳米银10,25,50,100,250mg·L-1对HL-7702肝细胞进行染毒,24h后用单细胞凝胶电泳技术检测HL-7702细胞DNA的损伤程度,用CASP软件分析彗星图像,并计算尾部DNA百分率和尾矩。结果与对照组和微米银组相比,纳米银染毒后HL-7702肝细胞DNA损伤效应明显增强;随纳米银染毒浓度的升高,HL-7702肝细胞尾部DNA百分率和尾矩均呈逐渐升高趋势;与对照组相比,染毒剂量≥25mg·L-1时,尾部DNA百分率及尾矩则显著升高(P<0.05,P<0.01)。结论纳米银可导致HL-7702肝细胞DNA损伤,微米银无明显的损伤作用。     

Protective effects of Ginkgo biloba extract against lysophosphatidylcholine-induced vascular endothelial cell damage

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Protective effects of Ginkgo biloba extract against lysophosphatidylcholine-induced vascular endothelial cell damage.

AIM: To study the protective effects of Ginkgo biloba extract (GbE) against endothelial cell damage induced by lysophosphatidylcholine (LPC). METHODS: The vasorelaxation response to acetylcholine (ACh) were investigated in the isolated rabbit thoracic aorta. Lipid peroxidation products were determined by measuring thiobarbituric acid reactive substance. RESULTS: GbE attenuated the inhibition of vasorelaxation response to ACh and prevented the LPC-induced increase of malondialdehyde (MDA) content both in thoracic aortae. GbE prevented the leakage of LDH and the increase of MDA content in cultured endothelial cells in a concentration-dependent manner. GbE also markedly increased epoprostenol level in cultured endothelial cells treated with LPC. CONCLUSION: GbE protected endothelial cells against LPC-induced damage due to reduction in lipid peroxidation and facilitation of synthesis and/or release of epoprostenol.     

In vitro evaluation of the cytotoxic and genotoxic effects of artemether,an antimalarial drug,in a gastric cancer cell line (PG100)

Artemisinin is a sesquiterpene lactone endoperoxide, obtained from Artemisia annua, and extensively used as an antimalarial drug. Many studies have reported the genotoxic and cytotoxic effects of artemisinins; however, there are no studies that compare such effects between cancer cell lines and normal human cells after treatment with artemether, an artemisinin derivative. Gastric cancer is the fourth most frequent type of cancer and the second highest cause of cancer mortality worldwide. Thus, the aim of this study was to evaluate the in vitro genotoxic and cytotoxic effects induced by artemether in gastric cancer cell line (PG100) and compare them with the results obtained in human lymphocytes exposed to the same conditions. We used MTT (3‐(4,5‐methylthiazol‐2‐yl)‐2, 5‐diphenyl‐tetrazolium bromide) assay, comet assay and ethidium bromide/acridine orange viability staining to evaluate the cytotoxic and genotoxic effects of artemether in PG100. MTT assay showed a decrease in the survival percentages for both cell types treated with different concentrations of artemether (P < 0.05). PG100 also showed a significant dose‐dependent increase in DNA damage index at concentrations of 119.4 and 238.8 µg ml^?1 (P < 0.05). Our results showed that artemether induced necrosis in PG100 at concentrations of 238.8 and 477.6 µg ml^?1, for all the tested harvest times (P < 0.05). In lymphocytes, artemether induced both apoptosis and necrosis at concentrations of 238.8 and 477.6 µg ml^?1, for all the tested harvest times (P < 0.05). In conclusion, human lymphocytes were more sensitive to the cytotoxic effects of the antimalarial drug than the gastric cancer cell line PG100. Copyright © 2011 John Wiley & Sons, Ltd.     

Comet‐FISH studies for evaluation of genetic damage of citalopram in somatic cells of the mouse

Damage to DNA can lead to many different acute and chronic pathophysiological conditions, ranging from cancer to endothelial damage. The present study was designed to evaluate the DNA damage of an antidepressant drug, citalopram, at the recommended human doses in somatic cells of mice in vivo. Mice exposed to citalopram at varying oral doses of 12 or 24 mg kg^–1 for 7 days exhibited a significant increase in the level of DNA‐strand breaking and micronuclei formation as detected by a bone marrow comet assay and micronucleus test, respectively. Furthermore, using fluorescence in situ hybridization (FISH) analysis with the centromeric mouse‐satellite DNA‐probe for erythrocyte micronuclei it could be shown that citalopram is aneugen as well as clastogen in somatic cells in vivo. Colchicine (COL) and mitomycin C (MMC) were used as positive controls and these compounds produced the expected responses. Both the clastogenic and the aneugenic potential of citalopram can give rise to the development of secondary tumours and abnormal reproductive outcomes. Overall, the results suggest that citalopram at the recommended human doses induces some genetic alterations, which can adversely affect the normal cellular functioning in mice. The mechanism(s) by which citalopram cause this adverse effect appear related, in part, to primary DNA strand breakage as detected by the comet assay as well as clastogenic and aneugenic events as detected by the FISH assay. Therefore, the clinical use of citalopram must be weighed against the risks of genetic damages in citalopram users. Copyright © 2013 John Wiley & Sons, Ltd.     

白藜芦醇苷体外对过氧化氢导致小鼠肝细胞损伤的保护作用

目的　观察中药虎杖的活性成分白藜芦醇苷对过氧化氢 (H2 O2 )所致肝细胞损伤的影响。方法　用邻苯三酚自氧化法测过氧化物歧化酶 ,用硫代巴比妥酸 (TBA)法测丙二醛 (MDA)含量 ,用改良Hafeman方法测还原谷胱甘肽含量 ,用 5 ,5’ 二巯基 2 ,2’ 二硝基苯甲酸 (DTNB)法测谷胱甘肽过氧化物酶活性。谷丙转氨酶 (ALT)、一氧化氮 (NO)和一氧化氮合酶 (NOS)采用测试药盒测定。结果　白藜芦醇苷系列浓度 (0 0 5 ,0 1,0 5 ,1,2 ,4mmol·L-1)作用肝细胞后 ,能显著降低H2 O2 引起的NO和MDA水平升高 ,抑制NOS活性 ,升高SOD和GSH px活性 ,减少GSH消耗 ,明显减少了H2 O2 导致的肝细胞悬液中ALT浓度增高。结论　白藜芦醇苷在一定浓度范围内对H2 O2 所致的小鼠肝细胞氧化损伤具有保护作用。     

Absence of genotoxic effects of the coumarin derivative 4‐methylesculetin in vivo and its potential chemoprevention against doxorubicin‐induced DNA damage

4‐Methylesculetin (4‐ME) is a synthetic derivative of coumarin that displays a potent reactive oxygen species (ROS) scavenger and metal chelating agent and therefore has been produced to help reduce the risk of human disease. The main objective of this study was to investigate the in vivo genotoxicity of 4‐ME and initially to verify its potential antigenotoxicity on doxorubicin (DXR)‐induced DNA damage. Different doses of 4‐ME (500, 1000 and 2000 mg kg^–1 body weight) were administered by gavage only or with a simultaneous intraperitoneal (i.p.) injection of DXR (80 mg kg^–1). The following endpoints were analyzed: DNA damage in peripheral blood, liver, bone marrow, brain and testicle cells according to an alkaline (pH > 13) comet assay and micronucleus induction in bone marrow cells. Cytotoxicity was assessed by scoring polychromatic (PCE) and normochromatic (NCE) erythrocytes (PCE/NCE ratio). No differences were observed between the negative control and the groups treated with a 4‐ME dose for any of the endpoints analyzed, indicating that it lacks genotoxic and cytotoxic effects. Moreover, 4‐ME demonstrated protective effects against DXR‐induced DNA damage at all tested doses and in all analyzed cell types, which ranged from 34.1% to 93.3% in the comet assay and 54.4% to 65.9% in the micronucleus test. Copyright © 2012 John Wiley & Sons, Ltd.     

Protective effects of the aqueous extract of Scutellaria baicalensis against acrolein-induced oxidative stress in cultured human umbilical vein endothelial cells

Context:?Scutellaria baicalensis Georgi (Labiatae) (SbG), one of the fifty fundamental herbs of Chinese herbology, has been reported to have anti-asthmatic, antifungal, antioxidative, and anti-inflammatory activities.

Objective:?This study was designed to determine the protective effects of the extract of SbG against the acrolein-induced oxidative stress in cultured human umbilical vein endothelial cells (HUVEC).

Materials and methods:?The MTT reduction assay was employed to determine cell viability. The total cellular glutathione (GSH) level was detected using a colorimetric GSH assay kit. Cellular GSH production was conducted by detecting the mRNA expression levels of γ-glutamylcysteine ligase catalytic subunit and modifier subunit.

Results:?Concentration-dependent cytotoxic effects of acrolein were observed while SbG could effectively protect the acrolein-induced oxidative damage. The protective mechanism was investigated, showing that the increased GSH content in the SbG-incubated HUVE cells was associated with the protective effects of SbG-treated cells. Further RT-PCR data confirmed the elevated mRNA expressions of GSH synthesis enzymes.

Discussion and conclusion:?The current study strongly indicated that SbG could be a potential antioxidant against oxidative stress in treating cardiovascular diseases.