无铅汽油新型添加剂甲基叔丁基醚的致突变性研究

目的；研究甲基叔丁基醚（MTBE）的致突变性及遗传毒性。方法：采用Ames试验检测MTBE的遗传毒性，并用单细胞凝胶电泳法检测国产MTBE大鼠经口亚慢性染毒后血液有核细胞的DNA的损伤状况。结果：Ames试验（TA98和TA100菌株）中，在加与不加S9的情况下，MTBE均未表现出明显的致突变性。染毒大鼠血液有核细胞DNA的断裂损伤也较对照组未见统计学差异。结论：本研究结果未发现MTBE对受试系统有致突变性。     

阿莫西林对人HepG2细胞DNA有短暂损伤作用

目的 为了探讨阿莫西林对人HepG2细胞DNA是否有损伤作用.方法 培养的人HepG2细胞经不同浓度(2、10、50和250μmol/L)阿莫西林处理1h或经50μmol/L阿莫西林处理不同时间(20、40、60、120和180min)后,运用单细胞凝胶电泳(SCGE)技术结合彗星图像软件(CASP)分析细胞尾部DNA百分率(tail DNA%)变化情况.结果 不同浓度阿莫西林处理后结果显示,HepG2细胞尾部DNA百分率明显升高,至50μmol/L阿莫西林时达到最高值,各浓度处理组与不处理对照组相比差异皆有显著性(P<0.01);而同一浓度(50μmol/L)阿莫西林处理不同时间后结果显示,HepG2细胞尾部DNA百分率逐渐升高,至1h处理时间点时达到最高值,其后随着处理时间延长HepG2细胞尾部DNA百分率逐渐降低.结论 阿莫西林对人HepG2细胞DNA有短暂损伤作用,阿莫西林诱发的HepG2细胞DNA损伤可能随时间延长逐渐被HepG2细胞本身修复除去.     

Protective effect of bixin on cisplatin-induced genotoxicity in PC12 cells

Bixin is the main carotenoid found in annatto seeds (Bixa orellana L.) and is responsible for their reddish-orange color. The antioxidant properties of this compound are associated with its ability to scavenge free radicals, which may reduce damage and protect tissues against toxicity caused by anticancer drugs such as cisplatin. In this study, the genotoxicity and antigenotoxicity of bixin on cisplatin-induced toxicity in PC12 cells was assessed. Cytotoxicity was evaluated using the MTT assay, mutagenicity, genotoxicity, and protective effect of bixin were evaluated using the micronucleus test and comet assay. PC12 cells were treated with bixin (0.05, 0.08, and 0.10μg/mL), cisplatin (0.1μg/mL) or a combination of both bixin and cisplatin. Bixin was neither cytotoxic nor genotoxic compared to the controls. In the combined treatment bixin significantly reduced the percentage of DNA in tail and the frequency of micronuclei induced by cisplatin. This result suggests that bixin can function as a protective agent, reducing cisplatin-induced DNA damage in PC12 cells, and it is possible that this protection could also extend to neuronal cells. Further studies are being conducted to better understand the mechanisms involved in the activity of this protective agent prior to using it therapeutically.     

Antioxidant potential of aqueous extract of Phyllanthus amarus in rats

Objective:

Increased levels of oxidative stress may be implicated in the etiology of many pathological conditions. Protective antioxidant action imparted by many plant extracts and plant products make them promising therapeutic drugs for free radical induced pathologies. In this study we assessed the antioxidant potential of Phyllanthus amarus (Euphorbiaceae).

Materials and Methods:

Experimental rats were divided into two groups: Control and Phyllanthus amarus (P. amarus) treated. Treated rats received P. amarus aqueous extract (PAAEt) at a dose of 200 mg/kg body wt/day for 8 weeks. After the treatment period of 8 weeks lipid peroxidation (LPO), vitamin C, uric acid and reduced glutathione (GSH) were estimated in plasma and antioxidant enzymes: Glutathione peroxidase (GPx), catalase (CAT) and superoxide dismutase (SOD) were also assayed. Genotoxicity of PAAEt was assessed by single cell gel electrophoresis (SCGE) of lymphocytes under both in vitro and in vivo conditions. The protective role of PAAEt against hydrogen peroxide (H₂O₂), streptozotocin (STZ) and nitric oxide generating system induced lymphocyte DNA damage was also assessed by SCGE.

Results:

PAAEt treated rats showed a significant decrease in plasma LPO and a significant increase in plasma vitamin C, uric acid, GSH levels and GPx, CAT and SOD activities. SCGE experiment reveals that PAAEt was devoid of genotoxicity and had a significant protective effect against H₂O₂, STZ and nitric oxide (NO) induced lymphocyte DNA damage.

Conclusion:

The results suggest the non-toxic nature of PAAEt and consumption of PAAEt can be linked to improved antioxidant status and reduction in the risk of oxidative stress.     

Validation of the comet-X assay as a pharmacodynamic assay for measuring DNA cross-linking produced by the novel anticancer agent RH1 during a phase I clinical trial

Purpose RH1 is a novel anticancer agent with potent DNA-cross linking activity. RH1 has the potential to be activated within tumors over expressing NQO1, giving maximal antitumour activity with reduced toxicity in normal tissues. RH1 has recently completed a Cancer Research UK sponsored phase I clinical trial at two different centers in the United Kingdom. The comet-X assay was a secondary endpoint in this trial and assay validation was necessary. We describe here this validation process. Whilst it is impossible to cover all variations/conditions of a pharmacodynamic assay, we have strived to evaluate and demonstrate that this assay conforms to the three R’s of validation, that is robustness, reliability and reproducibility. Methods K562 and peripheral blood mononuclear cells were treated with either radiation alone, or with a combination of radiation and drug. These samples were then embedded in low melting point agarose and subjected to a modified version of the alkaline single cell gel electrophoresis (Comet) assay, described here as the comet-X assay. Variations in the preparation, electrophoresis, storage and scoring of these samples was investigated. In addition radiation and drug dose response curves were constructed. Finally stability of QC standards was investigated over a 30-month period. Results We have demonstrated a linear radiation-dose response in cells up to 20 Gy and drug induced DNA cross-linking up to 50 nM. From the radiation dose response curves we were able to show that the relative inaccuracy measured against a global mean value was less than 25% and the relative (within day) imprecision was less than 30% over all doses. Between day runs produced an intra assay imprecision of 21.2%. Variables involved in the electrophoresis process showed the voltage across all slides in the tank ranged from 3.1 to −2.0 (mV) whilst the current ranged from 0.8–5.5 mA. QC standards were prepared from PBMCs of healthy donors and frozen at −80°C. The stability of these frozen QC standards was measured over a 30-month period. No significant deterioration in any of the control, irradiated or drug treated samples was observed. Conclusions The comet-X assay has been shown to be a robust, reliable and reproducible assay. It is ideally suited for the evaluation of the pharmacodynamic effects of DNA cross-linking agents undergoing early clinical trials. Furthermore, this assay may provide valuable data, in conjunction with pharmacokinetics, when measuring toxicity and efficacy as part of the RH1 phase I clinical trial.     

单细胞电泳观察重离子辐照人肝癌细胞所致DNA损伤

目的 研究重离子对肿瘤细胞DNA的损伤程度，为重离子治癌的临床应用积累必要的基础数据。方法 用80MeV／u氖离子射线诱发SMMC-7721细胞DNA辐射损伤，利用单细胞凝胶电泳(single cell gelelectrophoresis，SCGE)实验方法处理细胞，并对结果进行统计分析。结果 氖离子辐照以剂量依赖的方式引起SMMC-7721细胞DNA迁移长度的增加，且拖尾长度与剂量呈线性正相关；同时，拖尾的细胞数(即损伤的细胞数)在剂量大于2Gy时100％细胞损伤。结论 重离子射线对DNA有强致损效应。     

砷与5-氮胞苷对人淋巴细胞DNA损伤的联合作用

为探讨砷和5－氮胞苷对人淋巴细胞DNA损伤及修复的联合作用，应用单细胞凝胶电泳（SCGE）技术比较研究了5－氮胞苷与砷同时和前后作用于人类淋巴细胞产生的联合毒性，结果显示10μmol/L5-氮胞苷和10μmol/L砷单独处理人淋巴细胞2h引起明显的DNA泳动（彗星尾），但两试剂引起的DNA泳动（彗星尾）间无显差异，5－氮胞苷前处理与砷后处理2h引起的彗星尾与其单独处理组比较非常显，砷前处理与5－氮胞苷后处理引起的彗星尾与其单独处理组比较无显性差异，但较对照组差异显，10μmol/L5-氮胞苷和10μmol/L砷分别单独处理2h引起了人淋巴细胞显的DNA损伤（链断裂），5－氮胞苷与砷在对淋巴细胞DNA的损伤上表现为单纯相加作用。5－氮胞苷前处理显增加了细胞对砷的基因毒性的敏感性，或砷后处理显增加了5－氮胞苷引起的DNA损伤，5－氮胞苷后处理2h显抑制了细胞对砷所致DNA损伤的修复。     

单细胞凝胶电泳分析地黄提取物对仓鼠肺成纤维细胞DNA的损伤作用

目的 采用单细胞凝胶电泳（彗星实验）技术研究地黄提取物致仓鼠肺成纤维细胞（CHL细胞）DNA的损伤作用,为该药物临床前安全性评价提供参考。方法 采用0.2、1.0、5.0 mg/mL地黄提取物,用双氧水作为阳性对照药,处理细胞,24 h后收获细胞,进行彗星电泳实验。结果/b> 双氧水处理后造成CHL细胞产生明显的DNA损伤,呈现不同彗尾长度的彗星,与阴性对照组比较有显著差异。不同质量浓度的地黄提取物处理细胞后,彗星长、彗尾长度等各项指标与阴性对照组比较无显著差异。结论 单细胞凝胶电泳检测DNA损伤的敏感度较高,不同质量浓度的地黄提取物对CHL细胞未产生DNA损伤。     

Genotoxicity of monosodium glutamate

Monosodium glutamate (MSG) is one of the most widely used flavor enhancers throughout the world. The aim of this study is to investigate the genotoxic potential of MSG by using chromosome aberrations (CAs), sister-chromatid exchanges (SCEs), cytokinesis-blocked micronucleus (CBMN), and random amplified polymorphic DNA-polimerase chain reaction (RAPD-PCR) in cultured human lymphocytes and alkaline comet assays in isolated human lymphocytes, which were incubated with six concentrations (250, 500, 1000, 2000, 4000 and 8000 μg/mL) of MSG. The result of this study indicated that MSG significantly and dose dependently increased the frequencies of CAs, SCE and MN in all treatments and times, compared with control. However, the replication (RI) and nuclear division indices (NDI) were not affected. In this paper, in vitro genotoxic effects of the MSG was also investigated on human peripheral lymphocytes by analysing the RAPD-PCR with arbitrary 10-mer primers. The changes occurring in RAPD profiles after MSG treatment include increase or decrease in band intensity and gain or loss of bands. In the comet assay, this additive caused DNA damage at all concentrations in isolated human lymphocytes after 1-h in vitro exposure. Our results demonstrate that MSG is genotoxic to the human peripheral blood lymphocytes in vitro.     

用SCGE法检测燃煤污染型砷中毒患者血细胞DNA的损伤作用

目的 探讨砷中毒的致癌机理及寻找灵敏、实用的ＤＮＡ损伤监测指标,方法 应用单细胞凝胶电泳法（ＳＣＧＥ）对贵州省兴仁县燃煤型砷中毒重病区１７５名砷接触者的血细胞ＤＮＡ损伤进行了检测,并分析了相关因素的影响。结果 砷能引起人体血细胞ＤＮＡ单逻断裂,其损伤发生早于临床表现,并与尿砷,发砷呈正相关关系,与皮肤损害进展一致,吸烟和氟的影响可加重睦ＤＮＡ的损伤作用,结论 ＳＣＧＥ具有灵敏、快速、简便等优点,可