A 32P-postlabeling method for the detection of adducts in the DNA of human fibroblasts exposed to sulfur mustard

Niu, Tianqi; Matijašević, Zdenka; Austin-Ritchie, Paula; Stering, Allen; Ludlum, David B.

doi:10.1016/s0009-2797(96)03690-3

Cited by 24 publications

(8 citation statements)

References 4 publications

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“…There are also numerous studies that examine the mustard-derived DNA adducts from treated cells in culture and animals, 46,48,50,54,56,59,64,65 including white blood cells of patients on chemotherapeutic regimens that included a nitrogen mustard. 53,57 A variety of methods have been used to identify these adducts, including 32 P-postlabeling, 47,58 cochromatography by HPLC with authentic standards, 8,9,24,48,57 and mass spectrometry. 18,20,[49][50][51][52]56,59,64 Recently, tandem mass spectrometry with stable isotope dilution has been employed to identify and quantitate some of these DNA adducts.…”

Section: ■ Discussionmentioning

confidence: 99%

“…The labile N7-G and N3-A nitrogen and sulfur mustard adducts and their cross-links are often thermally hydrolyzed f r o m D N A u n d e r n e u t r a l 8 − 1 0 , 5 1 , 5 7 , 6 1 o r acidic 9,10,24,46,48,50,52,53,59 conditions. Alternatively, enzymatic digestion to the nucleosides has also been employed 18,20,47,49,54,58,65 and is necessary for DNA modifications that do not increase base lability (e.g., O 6 -dG adducts). Fapy nucleosides can exist as a number of slowly interconverting isomeric constituents such as αand β-anomers, furnanose and hexanose forms of the deoxyribose, geometric isomers of the formamide group, and possible atroisomers.…”

Section: ■ Discussionmentioning

confidence: 99%

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Characterization of Nitrogen Mustard Formamidopyrimidine Adduct Formation of Bis(2-chloroethyl)ethylamine with Calf Thymus DNA and a Human Mammary Cancer Cell Line

Gruppi

Hejazi

Christov

et al. 2015

Chem. Res. Toxicol.

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A robust, quantitative ultraperformance liquid chromatography ion trap multistage scanning mass spectrometric (UPLC/MS3) method was established to characterize and measure five deoxyguanosine (dG) adducts formed by reaction of the chemotherapeutic nitrogen mustard (NM) bis-(2-chloroethyl)ethylamine with calf thymus (CT) DNA. In addition to the known N7-guanine (NM-G) adduct and its crosslink (G-NM-G), the ring-opened formamidopyrimidine (FapyG) mono-adduct (NM-FapyG) and cross-links in which one (FapyG-NM-G) or both (FapyG-NM-FapyG) guanines underwent ring-opening to FapyG units were identified. Authentic standards of all adducts were synthesized and characterized by NMR and mass spectrometry. These adducts were quantified in CT DNA treated with NM (1 μM) as their deglycosylated bases. A two-stage neutral thermal hydrolysis was developed to mitigate the artifactual formation of ring-opened FapyG adducts involving hydrolysis of the cationic adduct at 37 °C, followed by hydrolysis of the FapyG adducts at 95 °C. The limit of quantification values ranged between 0.3 and 1.6 adducts per 107 DNA bases, when the equivalent of 5 μg DNA hydrolysate was assayed on column. The principal adduct formed was the G-NM-G cross-link, followed by the NM-G mono-adduct; the FapyG-NM-FapyG adduct was at the limit of detection. The NM-FapyG adducts formed in CT DNA at a level of ~20% that of the NM-G adduct. NM-FapyG has not been previously quanitified and the FapyG-NM-G and FapyG-NM-FapyG adducts have not be previously characterized. Our validated analytical method was then applied to measure DNA adduct formation in the MDA-MB-231 mammary tumor cell line exposed to NM (100 μM) for 24 h. The major adduct formed was NM-G (970 adducts per 107 bases), followed by G-NM-G (240 adducts per 107 bases) and NM-FapyG (180 adducts per 107 bases), and lastly the FapyG-NM-G cross-link adduct (6.0 adducts per 107 bases). These lesions are expected to contribute to the NM-mediated toxicity and genotoxicity in vivo.

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Section: ■ Discussionmentioning

confidence: 99%

Section: ■ Discussionmentioning

confidence: 99%

Characterization of Nitrogen Mustard Formamidopyrimidine Adduct Formation of Bis(2-chloroethyl)ethylamine with Calf Thymus DNA and a Human Mammary Cancer Cell Line

Gruppi

Hejazi

Christov

et al. 2015

Chem. Res. Toxicol.

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“…Besides this theory, it can be also applied to chemically-injured veterans, whose meibomian glands injured and pave the way for chronic blepharitis. On the other hand, escalated levels of MMP-8, 9 in tear fluid of these veterans perhaps is as a result of mutagenic and destructive effects of SM respectively on DNA (Ludlum et al, 1994(Ludlum et al, , 1984(Ludlum et al, , 1986Niu et al, 1996;Lawley et al, 1969;Smith et al, 1993;Rosenthal et al, 1998) and epithelial cells of cornea which is followed by over-expression of genes responsible for MMP-8,9 or under-expression of their inhibitors (TIMPs) in corneal epithelial cells. So for further research, it is worth designing some experiments to assert this theory.…”

Section: Discussionmentioning

confidence: 99%

Levels of matrix metalloproteinases (MMPs) in tear fluid of mustard gas exposed patients with chronic dry-eye symptoms.

Hosseini¹

2012

Afr. J. Biochem. Res.

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The purpose of this study is to compare the levels of MMP-1, 2, 8, 9 in tear fluids of people exposed to mustard gas in the war between Iraq and Iran who had the chronic dry-eye symptoms compared to the normal group. In this study, 25 of the patients who were exposed to mustard gas and had chronic dry eye symptoms were compared to 25 patients as control group; consisting of 25 people who had common chronic dry eye symptoms with blepharitis and 25 healthy people as normal group. The level of MMP-1, 2, 8, 9 in tear fluid of people of these three groups was assessed by enzyme-linked immunosorbent assay (ELISA). The results for levels of MMP-1 (P=0.8) and MMP-2 (P=0.54) in tear fluid of patients in comparison with normal group do not indicate any significant difference. On the other hand, the calculated levels for MMP-8 (P=0.002) and MMP-9 (P=0.01) in tear fluid of patients in comparison with normal group shows a significant increase. The differences were considered statistically significant at p< 0.05. The effects of exposure to mustard gas on eyes of chemical-injured veterans destroy meibomian glands which paves the way for evaporative dry-eye. As a result, MMPs in tear fluid of these patients increase which eventually elevates the matrix metalloproteinases (MMPs) levels in tear fluid and results in later eye-impairments.

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“…In this kind of procedure, the synthesis of radio-labeled agent and special precautions for proper radiochemical handling are shortcomings. Immunochemical detection [12,13] and 32 P-postlabeling method [14] have also been used for HETEG quantitation for the high sensitivity. However, the onerous antibody and sample handling requirements in immunochemical detection systems and the false positives in 32 P-postlabeling method are disadvantages for the further application.…”

Section: Introductionmentioning

confidence: 99%

A sensitive high performance liquid chromatography–positive electrospray tandem mass spectrometry method for N7-[2-[(2-hydroxyethyl)thio]-ethyl]guanine determination

Yuxia¹,

Yue²,

Q³

et al. 2011

Journal of Chromatography B

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A 32P-postlabeling method for the detection of adducts in the DNA of human fibroblasts exposed to sulfur mustard

Cited by 24 publications

References 4 publications

Characterization of Nitrogen Mustard Formamidopyrimidine Adduct Formation of Bis(2-chloroethyl)ethylamine with Calf Thymus DNA and a Human Mammary Cancer Cell Line

Characterization of Nitrogen Mustard Formamidopyrimidine Adduct Formation of Bis(2-chloroethyl)ethylamine with Calf Thymus DNA and a Human Mammary Cancer Cell Line

Levels of matrix metalloproteinases (MMPs) in tear fluid of mustard gas exposed patients with chronic dry-eye symptoms.

A sensitive high performance liquid chromatography–positive electrospray tandem mass spectrometry method for N7-[2-[(2-hydroxyethyl)thio]-ethyl]guanine determination

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