JOURNAL OF PHARMACEUTICAL AND BIOMEDICAL ANALYSIS
·
2019
·
IF 2.983
Yu Tang,Zhe Wang,Menglin Li,Ruiping Zhang,Jinlan Zhang
A rapid, sensitive and wide coverage ultra-high-performance liquid chromatography with tandem mass spectrometry (UHPLC-MS/MS) method has been developed and validated for the simultaneous quantitation of 14 alkylation DNA adducts in cell genomic DNA, RNA and cell contents isolated from the in vitro cultured human kidney cell line 293 T and the human liver cell line L02 exposed to 3 genotoxic reagents: N -methyl -N- nitrosourea (MNU), methyl methanesulfonate (MMS) and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK). After exposure, DNA was isolated and directly hydrolysed under acid conditions or digested by enzymes to obtain the hydrolysates containing DNA alkylation adducts followed by optimization of the pretreatment method and chromatographic separation conditions. Quantification was performed on a Waters ACQUITY UPLC BEH Amide column (1.7 μm, 2.1 × 150 mm) using an electrospray ionization (ESI) source in positive mode by selective reaction monitoring (SRM) at the precursor to product ion transitions of 14 analytes. The method showed selectivity, good linearity (r＞0.9950), accuracy (82.1%–115%), and intra-day (RSD%＜14%) and inter-day (RSD%＜15%) precision for 14 analytes. The recoveries of two pretreatment methods were all more than 50.5%, and no relative matrix effects were observed. Additionally, the samples were stable after short-term storage at 20 ℃ for 2 h, at 4 ℃ for 48 h or one cycle of freeze-thaw at −80 ℃. The established UHPLC-MS/MS method was used to evaluate the changes in alkylation DNA adducts and epigenetic modification-related methylcytosine after exposure to genotoxic reagents. For the first time, the results demonstrated that 3 genotoxic reagents induced different total amounts of adducts in the following sequence: MMS ＞ NNK ＞ MNU, and showed significant differences in the ratios of 7MeG to 1MeA and 1MeG to 1MeA in the 293 T cell model. Meanwhile, 293 T and L02 cells revealed significantly different DNA adduct formation characteristics in the contents of 1MeG and 1MeA. The DNA adduct formation relationships between DNA, RNA, and cell contents were probed to predict cancer risk and potential genotoxic exposure. This approach could be used to investigate the DNA adducts, their formation and the relationship to the mutagenicity or carcinogenicity of genotoxic reagents in future studies.
研究领域:
毒理学
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分析化学
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分子生物学
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DOI
PubMed
高影响因子
MUTAGENESIS
·
2025
·
IF 4.3
Hu Yushi,Lei Yuyang,Gao Zhenna,Cao Yiyi,Xi Jing,Ma Yuning,Gao Qinwen,Fu Jian,Zhang Xinyu,Luan Yang
Aristolochic acid I (AAI) is a carcinogen associated with various human cancers. However, its causal relationship with hepatocellular carcinoma remains controversial, and inconsistent results from rodent studies have suggested species-specific differences. Here we evaluated AAI genotoxicity using functional human-induced hepatocyte-like cells (hiHep cells), a model that closely mimics primary human hepatocytes in gene expression and function, thereby shedding light on its potential hepatocarcinogenic risk in humans. First, we assessed AAI genotoxicity by evaluating AAI-DNA adducts and micronucleus frequency. In hiHep cells, AAI (0.7–2.5 µM) induced up to 105 adducts per 108 nucleotides, indicating high metabolic activation of AAI. A concentration-dependent increase in micronucleus frequency indicated a significant increase in chromosomal aberrations in hiHep cells. Considering the evidence of AAI inducing oxidative stress, we assessed 8-hydroxy-2′-deoxyguanosine and reactive oxygen species levels to evaluate DNA oxidative damage. For both indicators, significantly elevated levels were observed. A mechanism involving oxidative damage was further supported by observations of mitochondrial dysfunction, including changes in mitochondrial membrane potential and mitochondrial complex activity. Ascorbate treatment decreased AAI-induced oxidative DNA damage and DNA adduct formation, providing direct cellular evidence for free radical intermediates in AAI metabolic activation—a mechanism previously hypothesized but not experimentally validated in a human-relevant hepatocyte model. Our study findings revealed the genotoxic effects of AAI on hiHep cells and implicated oxidative stress as the key mechanism. These findings strengthen the association between AAI exposure and liver disease and highlight the potential role of antioxidant therapies in mitigating AAI-associated carcinogenesis.
研究领域:
毒理学
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肝脏生物学
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癌症研究
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产品关联
DOI
PubMed
高影响因子
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
·
2018
·
IF 3.412
Yukun Feng,Saide Wang,Hui Wang,Ying Peng,Jiang Zheng
Methyleugenol (ME), a natural ingredient of several herbs and spices used in the human diet, is hepatocarcinogenic in rodents. Following metabolic activation to the reactive carbocation intermediate, ME can bind covalently to DNA, which is directly associated with its carcinogenicity. In this work, a non-invasive approach to determine ME exposure was established by monitoring the urinary N6-(methylisoeugenol-3′-yl)-2′-deoxyadenosine (ME-dA) adduct. The developed method entails liquid–liquid extraction enrichment of urinary ME-dA, incorporation of deuterated ME-dA as an internal standard, and analysis by liquid chromatography coupled tandem mass spectrometry. Male rats (10–12 weeks, 180–200 g) were treated (p.o.) with ME, and ME-dA was excreted in urine in a dose- and time-dependent manner. The non-invasive approach enabled us to successfully determine exposure to ME-containing herbs and spices. These results suggest that ME-dA can potentially serve as an effective biomarker of ME exposure in rats. It is expected that the developed approach of detecting urinary ME-dA will facilitate the investigation of ME carcinogenesis.
研究领域:
毒理学
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分析化学
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癌症研究
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产品关联
DOI
PubMed
高影响因子
JOURNAL OF CHROMATOGRAPHY A
·
2016
·
IF 3.926
Ning Zhang,Yuanyuan Song,Danni Wu,Tian Xu,Meiling Lu,Weibing Zhang,Hailin Wang
Crotonaldehyde (Cro) is one of widespread and genotoxic α,β-unsaturated aldehydes and can react with the exocyclic amino group of 2′-deoxyguanosine (dG) in genomic DNA to form 1, N 2 -propano-2′-deoxyguanosine (ProdG) adducts. In this study, two diastereomers of high purity were prepared, including non-isotope and stable isotope labeled ProdG adducts, and exploited stable isotope dilution-based calibration method. By taking advantage of synthesized ProdG standards, we developed a sensitive ultrahigh performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UHPLC-ESI–MS/MS) method for accurate quantification of two diastereomers of ProdG adducts. In addition to optimization of the UHPLC separation, ammonium bicarbonate (NH 4 HCO 3 ) was used as additive in the mobile phase for enhancing the ionization efficiency to ProdG adducts and facilitating MS detection. The limits of detection (LODs, S/N = 3) and the limits of quantification (LOQs, S/N = 10) are estimated about 50 amol and 150 amol, respectively. By the use of the developed method, both diastereomers of ProdG adducts can be detected in untreated human MRC5 cells with a frequency of 2.4–3.5 adducts per 10 8 nucleotides. Crotonaldehyde treatment dramatically increases the levels of ProdG adducts in human MRC5 in a concentration-dependent manner.
研究领域:
生物化学
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毒理学
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分析化学
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DOI
PubMed
高影响因子
ARCHIVES OF TOXICOLOGY
·
2021
·Vol. 95
·期 8
·2839-2850
·
IF 5.153
Wan Jingjing,Chen Ruixue,Yang Zhou,Xi Jing,Cao Yiyi,Chen Yu,Zhang Xinyu,Luan Yang
Aristolochic acids (AAs) are a family of natural compounds with AA I and AA II being known carcinogens, whose bioactivation causes DNA adducts formation. However, other congeners have rarely been investigated. This study aimed to investigate genotoxicity of AA IVa, which differs from AA I by a hydroxyl group, abundant in Aristolochiaceae plants. AA IVa reacted with 2'-deoxyadenosine (dA) and 2'-deoxyguanosine (dG) to form three dA and five dG adducts as identified by high-resolution mass spectrometry, among which two dA and three dG adducts were detected in reactions of AA IVa with calf thymus DNA (CT DNA). However, no DNA adducts were detected in the kidney, liver, and forestomach of orally dosed mice at 40 mg/kg/day for 2 days, and bone marrow micronucleus assay also yielded negative results. Pharmacokinetic analyses of metabolites in plasma indicated that AA IVa was mainly O -demethylated to produce a metabolite with two hydroxyl groups, probably facilitating its excretion. Meanwhile, no reduced metabolites were detected. The competitive reaction of AA I and AA IVa with CT DNA, with adducts levels varying with pH of reaction revealed that AA IVa was significantly less reactive than AA I, probably by hydroxyl deprotonation of AA IVa, which was explained by theoretical calculations for reaction barriers, energy levels of the molecular orbits, and charges at the reaction sites. In brief, although it could form DNA adducts in vitro, AA IVa was non-genotoxic in vivo, which was attributed to its low reactivity and biotransformation into an easily excreted metabolite rather than bioactivation.
研究领域:
药理学
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毒理学
·
分子生物学
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DOI
PubMed
高影响因子
JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES
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2017
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IF 2.603
Xiangyu Li,Lujuan Liu,Hongjuan Wang,Jian Chen,Beibei Zhu,Huan Chen,Hongwei Hou,Qingyuan Hu
A stable method, using isotope dilution liquid chromatography-tandem mass spectrometry (LC–MS/MS), to simultaneously determine six aldehyde-DNA adducts was developed and applied to the analysis of human salivary DNA samples. The detection limit of these six DNA adducts was in the range of 0.006–0.014 ng/mL and that of the quantification limit was 0.017–0.026 ng/mL. The intra-day and inter-day precision of all aldehyde-DNA adducts was <10%. The analysis was completed within 25 min. Additionally, a noninvasive technique was used to collect the DNA samples from human saliva. The new method was successfully applied for the analysis of salivary DNA of nonsmokers and smokers. Five aldehyde-DNA adducts were detected in both smoker and nonsmoker salivary DNA, while α-Acr-dG was not detected in all the samples. Among these detected DNA adducts, no significant differences were found between smoker and nonsmoker (p > 0.05). This may due to the individual detoxifying differences or environmental and endogenous exposure. Our study provides a rapid and selective method to simultaneously detect six aldehyde-DNA adducts and to assess potential DNA damage induced by aldehydes,
研究领域:
分析化学
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分子生物学
·
毒理学
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DOI
高影响因子
JOURNAL OF CHROMATOGRAPHY A
·
2024
·
IF 3.8
Ying Shi,Yi-Wen Wu,Qian-Xue Shen,Jun Cao
Aristolochic acids are one of the major compounds in aristolochia plants, which are nephrotoxic and carcinogenic. A method was established for the detection and identification of aristolochic acids and their DNA adducts in four different herbs using ultra-high performance liquid chromatography-ion mobility quadrupole time-of flight mass spectrometry. Solid phase extraction conditions were optimized to improve the sensitivity of the experiment by using 40 mg of C 18 as adsorbent and 100 μL ethanol as elution solvent. At a collision energy of 10-40 eV, these compounds and cleavage patterns were precisely identified and analyzed by secondary fragmentation and collision cross section values. The obtained mass spectrometry data were then analyzed by targeted metabolomics, including principal component analysis, partial least squares-discriminant analysis and hierarchical clustering analysis, and importing the samples in the established model, the confidence values can reach 0.61 and 0.76. All in all, this method can provide a useful tool for the detection of aristolochic acids and deoxyribonucleic acid adducts. In conclusion, this method was successfully used for the detection and identification of aristolochic acids and their DNA adducts.
研究领域:
药理学
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毒理学
·
分析化学
✓ 已验证
DOI
PubMed
高影响因子
Bioactive Materials
·
2022
·
IF 16.874
Yi Xiao,Haomin Yi,Jingzhi Zhu,Suhua Chen,Guofang Wang,Yilong Liao,Yuanyuan Lei,Liyin Chen,Xingcai Zhang,Fangfu Ye
Toxicity assessment is a major problem in pharmaceutical candidates and industry chemicals development. However, due to the lack of practical analytical methods for DNA adduct analysis, the safety evaluation of drug and industry chemicals was severely limited. Here, we develop a DNAzyme-based method to detect DNA adduct damage for toxicity assessment of drugs and chemicals. Among 18 structural variants of G4 DNAzyme, EA2 DNAzyme exhibits an obvious DNA damaging effect of styrene oxide (SO) due to its unstable structure. The covalent binding of SO to DNAzyme disrupts the Hoogsteen hydrogen bonding sites of G-plane guanines and affects the formation of the G4 quadruplex. DNA damage chemicals reduce the peroxidase activity of the G4 DNAzyme to monitor the DNA adduct damage by disrupting the structural integrity of the G4 DNAzyme. Our method for genotoxic assessment of pharmaceutical candidates and industrial chemicals can elucidate the complex chemical pathways leading to toxicity, predict toxic effects of chemicals, and evaluate possible risks to human health.
研究领域:
分子生物学
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毒理学
·
药学
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化学工程
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产品关联
DOI
PubMed
高影响因子
JOURNAL OF NANOBIOTECHNOLOGY
·
2022
·Vol. 20
·期 1
·1-17
·
IF 9.429
Xu Lihua,Zhang Kaixiang,Ma Xing,Li Yingying,Jin Yajie,Liang Chenglin,Wang Yong,Duan Wendi,Zhang Hongling,Zhang Zhenzhong,Shi Jinjin,Liu Junjie,Wang Yunlong,Li Wentao
Despite many nano-based strategies devoted to delivering cisplatin for tumor therapy, its clinical benefits are compromised by poor tissue penetration and limited DNA adducts formation of the drug. Herein, a cisplatin loading nanomotor based janus structured Ag-polymer is developed for cisplatin delivery of deeper tissue and increased DNA adducts formation. The nanomotor displayed a self‐propelled tumor penetration fueled by hydrogen peroxide (H2O2) in tumor tissues, which is catalytically decomposed into a large amount of oxygen bubbles by Ag nanoparticles (NPs). Notably, cisplatin could elevate the intracellular H2O2 level through cascade reactions, further promote the degradation of Ag NPs accompanied with the Ag+ release, which could downregulate intracellular Cl− through the formation of AgCl precipitate, thereby enhancing cisplatin dechlorination and Pt–DNA formation. Moreover, polymer can also inhibit the activity of ALKBH2 (a Fe2+-dependent DNA repair enzyme) by chelating intracellular Fe2+ to increase the proportion of irreparable Pt–DNA cross-links. It is found that deep tissue penetration, as well as the increased formation and maintenance of Pt–DNA adducts induced by the nanomotor afford 80% of tumor growth inhibition with negligible toxicity. This work provides an important perspective of resolving chemotherapeutic barriers for boosting cisplatin therapy.Graphical Abstract
研究领域:
癌症治疗
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纳米技术
·
化疗
·
药物递送系统
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产品关联
DOI
PubMed
高影响因子
SCIENCE OF THE TOTAL ENVIRONMENT
·
2023
·
IF 9.8
Hongmei Zou,Mingxuan Zhang,Jin Chen,Stanley Aniagu,Yan Jiang,Tao Chen
Benzo[ a ]pyrene (BaP) is a representative polycyclic aromatic hydrocarbon widely present in the environment. We previously reported that the aryl hydrocarbon receptor (AHR) mediates BaP-induced apoptosis and cardiac malformations in zebrafish embryos, but the underlying molecular mechanisms were unclear. Since BaP is a mutagenetic compound, we hypothesize that BaP induces apoptosis and heart defects via AHR-mediated DNA damage. In this study, zebrafish embryos were exposed to BaP at a concentration of 0.1 μM from 2 to 72 h post fertilization, either with or without inhibitors/agonists. AHR activity and levels of reactive oxygen species (ROS) were examined under a fluorescence microscope. mRNA expression levels were quantified by qPCR. DNA damage and apoptosis were detected by immunofluorescence. Our findings revealed that BaP exposure significantly increased BPDE-DNA adducts, mitochondrial damage, apoptosis and heart defects in zebrafish embryos. These effects were counteracted by inhibiting AHR/cyp1a1 using pharmaceutical inhibitors or genetic knockdown. Furthermore, we observed that spironolactone, an antagonist of nucleotide excision repair (NER), significantly enhanced BaP-induced BPDE-DNA adducts, mitochondrial damage, apoptosis and heart malformation rates. Conversely, SRT1720, a SIRT1 agonist, reduced the adverse effects of BaP. Supplementation with spironolactone also enhanced γ-H2AX signals in the heart of zebrafish embryos exposed to BaP. Additional experiments demonstrated that BaP suppressed the expression of SIRT1. We further established that AHR, when activated by BaP, directly inhibited SIRT1 transcription, leading to downregulation of XPC and XPA, which are essential NER genes involved in the recognition and verification steps of the NER process. Taken together, our results indicate that AHR mediates BaP-induced DNA damage in the heart of zebrafish embryos by inducing BPDE-DNA adduct formation via the AHR/Cyp1a1 signalling pathway, as well as suppressing NER via AHR-mediated inhibition of SIRT1.
研究领域:
毒理学
·
发育生物学
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分子生物学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
International Journal of Nanomedicine
·
2024
·
IF 6.6
Weitang Sun,Jingbin Zhong,Buyun Gao,Jieling Feng,Zijie Ye,Yueling Lin,Kelan Zhang,Wenqi Su,Shibo Zhu,Yinghua Li,Wei Jia
Purpose Hydroxyapatite-based nanoparticles have found diverse applications in drug delivery, gene carriers, diagnostics, bioimaging and tissue engineering, owing to their ability to easily enter the bloodstream and target specific sites. However, there is limited understanding of the potential adverse effects and molecular mechanisms of these nanoparticles with varying geometries upon their entry into the bloodstream. Here, we used two commercially available hydroxyapatite nanoparticles (HANPs) with different geometries (less than 100 nm in size each) to investigate this issue.Methods First, the particle size, Zeta potential, and surface morphology of nano-hydroxyapatite were characterized. Subsequently, the effects of 2~2000 μM nano-hydroxyapatite on the proliferation, migration, cell cycle distribution, and apoptosis levels of umbilical vein endothelial cells were evaluated. Additionally, the impact of nanoparticles of various shapes on the differential expression of genes was investigated using transcriptome sequencing. Additionally, we investigated the in vivo biocompatibility of HANPs through gavage administration of nanohydroxyapatite in mice.Results Our results demonstrate that while rod-shaped HANPs promote proliferation in Human Umbilical Vein Endothelial Cell (HUVEC) monolayers at 200 μM, sphere-shaped HANPs exhibit significant toxicity to these monolayers at the same concentration, inducing apoptosis/necrosis and S-phase cell cycle arrest through inflammation. Additionally, sphere-shaped HANPs enhance SULT1A3 levels relative to rod-shaped HANPs, facilitating chemical carcinogenesis-DNA adduct signaling pathways in HUVEC monolayers. In vivo experiments have shown that while HANPs can influence the number of blood cells and comprehensive metabolic indicators in blood, they do not exhibit significant toxicity.Conclusion In conclusion, this study has demonstrated that the geometry and surface area of HANPs significantly affect VEC survival status and proliferation. These findings hold significant implications for the optimization of biomaterials in cell engineering applications.
研究领域:
纳米技术
·
生物材料
·
细胞生物学
·
毒理学
·
药学
·
基因组学
✓ 已验证
DOI
PubMed
高影响因子
ANALYTICAL CHEMISTRY
·
2013
·
IF 5.695
Ruichuan Yin,Shengquan Liu,Chao Zhao,Meiling Lu,Moon-shong Tang,Hailin Wang
Acrolein (Acr), a ubiquitous environmental pollutant, can react directly with genomic DNA to form mutagenic adducts without undergoing metabolic activation. To sensitively and accurately quantify Acr–DNA adducts (including structural isomers and stereoisomers) in human leukocytes, we developed an enhanced stable isotope dilution ultrahigh performance liquid chromatography (UHPLC)–tandem mass spectrometry (MS/MS) method using ammonium bicarbonate (NH4HCO3), which is thermally unstable and degrades readily to carbon dioxide and ammonia in heated gas phase. Interestingly, ammonium bicarbonate (as an additive to the mobile phase) not only improves the protonation of AcrdG adducts but also suppresses the formation of MS signal-deteriorating metal–AcrdG complexes during electrospray ionization, leading to the enhancement of their MS detection by 2.3–8.7 times. In contrast, routinely used ammonium salts (ammonium acetate and ammonium formate) and formic acid do not show similar enhancement. The developed method is potentially useful for enhancing ESI-MS detection of other modified 2′-deoxyribonucleosides that have difficulty in protonation and may form excess metal complexes during electrospray ionization. The limits of detection (LODs, S/N = 3) are estimated to be about 40–80 amol. By the use of the developed method, we found that the Acr adducts of three nucleotides (dG, dA, and dC) can be detected in human leukocytes. In addition to the known γ-AcrdG, α-AcrdA is also identified as an Acr-adduct of high abundance (2.5–20 adducts per108 nts).
研究领域:
分析化学
·
环境科学
·
分子生物学
·
毒理学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）