32P-post-labelling analysis of DNA adducts formed in the upper gastrointestinal tissue of mice fed bracken extract or bracken spores.

2.50
Hdl Handle:
http://hdl.handle.net/10541/95883
Title:
32P-post-labelling analysis of DNA adducts formed in the upper gastrointestinal tissue of mice fed bracken extract or bracken spores.
Authors:
Povey, Andrew C; Potter, D; O'Connor, Peter J
Abstract:
Bracken toxicity to both domestic and laboratory animals is well established and tumours are formed when rodents are treated with either bracken extracts or bracken spores. In this study we have administered bracken spores and extract to mice in order to investigate whether such exposure leads to the formation of DNA adducts. DNA, isolated from the upper gastrointestinal tract and liver, was digested to 3'-nucleotides. Adducts were extracted with butanol, 32P-post-labelled, separated by thin layer chromatography (TLC) and visualised and quantified using storage-phosphor technology. A cluster of adducts was clearly seen in the DNA of the upper gastrointestinal tract, but not liver, 5 and 24 h after treatment with bracken extract or bracken spores. These adducts were not observed in DNA extracted from vehicle-treated animals. Whereas, after 5 h adduct levels in extract and spore-treated animals were similar, after 24 h adduct levels in the extract-treated animals had diminished by > 75%, but levels in spore-treated animals remained similar to those found after 5 h. This suggests that the DNA-reactive compounds were being released slowly from the spores, even though the spores had been sonicated before administration. Adducts were also quantified after the addition of an internal standard (deoxyinosine 3'-monophosphate) by comparing the amount of label incorporated into the adducts with that found in a known amount of the internal standard. Adduct levels using this internal standard approach were similar to those found by direct measurement of radioactivity incorporated into the adduct, indicating that the labelling of adducts was quantitative. We have tried, unsuccessfully, to synthesise ptaquiloside, the principal carcinogenic component present within bracken. However, similar patterns of adducts were observed when two other compounds, (1-(4-chlorophenyl sulphonyl)-l-cyclopropane carbonitrile and 3-cyclopropylindeno [1,2-c] pyrazol-4-(O-methyl)oxime), which both contain a cyclopropyl ring, were administered to mice. The adducts detected in bracken-treated animals may, thus, have arisen from ptaquiloside but, whether these adducts arise directly from the compounds and bracken spores/extract themselves or via an indirect mechanism, remains to be determined. As bracken-induced DNA adducts are detectable in rodent tissues by a 32P-post-labelling procedure commonly employed to investigate DNA damage in human populations, it may prove possible to apply such approaches to determine human exposure.
Affiliation:
Department of Carcinogenesis, Paterson Institute for Cancer Research, Manchester, UK.
Citation:
32P-post-labelling analysis of DNA adducts formed in the upper gastrointestinal tissue of mice fed bracken extract or bracken spores. 1996, 74 (9):1342-8 Br. J. Cancer
Journal:
British Journal of Cancer
Issue Date:
Nov-1996
URI:
http://hdl.handle.net/10541/95883
PubMed ID:
8912527
Type:
Article
Language:
en
ISSN:
0007-0920
Appears in Collections:
All Paterson Institute for Cancer Research

Full metadata record

DC FieldValue Language
dc.contributor.authorPovey, Andrew Cen
dc.contributor.authorPotter, Den
dc.contributor.authorO'Connor, Peter Jen
dc.date.accessioned2010-04-07T13:44:56Z-
dc.date.available2010-04-07T13:44:56Z-
dc.date.issued1996-11-
dc.identifier.citation32P-post-labelling analysis of DNA adducts formed in the upper gastrointestinal tissue of mice fed bracken extract or bracken spores. 1996, 74 (9):1342-8 Br. J. Canceren
dc.identifier.issn0007-0920-
dc.identifier.pmid8912527-
dc.identifier.urihttp://hdl.handle.net/10541/95883-
dc.description.abstractBracken toxicity to both domestic and laboratory animals is well established and tumours are formed when rodents are treated with either bracken extracts or bracken spores. In this study we have administered bracken spores and extract to mice in order to investigate whether such exposure leads to the formation of DNA adducts. DNA, isolated from the upper gastrointestinal tract and liver, was digested to 3'-nucleotides. Adducts were extracted with butanol, 32P-post-labelled, separated by thin layer chromatography (TLC) and visualised and quantified using storage-phosphor technology. A cluster of adducts was clearly seen in the DNA of the upper gastrointestinal tract, but not liver, 5 and 24 h after treatment with bracken extract or bracken spores. These adducts were not observed in DNA extracted from vehicle-treated animals. Whereas, after 5 h adduct levels in extract and spore-treated animals were similar, after 24 h adduct levels in the extract-treated animals had diminished by > 75%, but levels in spore-treated animals remained similar to those found after 5 h. This suggests that the DNA-reactive compounds were being released slowly from the spores, even though the spores had been sonicated before administration. Adducts were also quantified after the addition of an internal standard (deoxyinosine 3'-monophosphate) by comparing the amount of label incorporated into the adducts with that found in a known amount of the internal standard. Adduct levels using this internal standard approach were similar to those found by direct measurement of radioactivity incorporated into the adduct, indicating that the labelling of adducts was quantitative. We have tried, unsuccessfully, to synthesise ptaquiloside, the principal carcinogenic component present within bracken. However, similar patterns of adducts were observed when two other compounds, (1-(4-chlorophenyl sulphonyl)-l-cyclopropane carbonitrile and 3-cyclopropylindeno [1,2-c] pyrazol-4-(O-methyl)oxime), which both contain a cyclopropyl ring, were administered to mice. The adducts detected in bracken-treated animals may, thus, have arisen from ptaquiloside but, whether these adducts arise directly from the compounds and bracken spores/extract themselves or via an indirect mechanism, remains to be determined. As bracken-induced DNA adducts are detectable in rodent tissues by a 32P-post-labelling procedure commonly employed to investigate DNA damage in human populations, it may prove possible to apply such approaches to determine human exposure.en
dc.language.isoenen
dc.subject.meshAnimals-
dc.subject.meshDNA Adducts-
dc.subject.meshDigestive System-
dc.subject.meshFemale-
dc.subject.meshMice-
dc.subject.meshMice, Inbred C57BL-
dc.subject.meshMice, Inbred DBA-
dc.subject.meshPlant Extracts-
dc.subject.meshPlants, Toxic-
dc.subject.meshSpores-
dc.title32P-post-labelling analysis of DNA adducts formed in the upper gastrointestinal tissue of mice fed bracken extract or bracken spores.en
dc.typeArticleen
dc.contributor.departmentDepartment of Carcinogenesis, Paterson Institute for Cancer Research, Manchester, UK.en
dc.identifier.journalBritish Journal of Canceren
All Items in Christie are protected by copyright, with all rights reserved, unless otherwise indicated.