DNA Strand breaks in testicular cells from humans and rats following in vitro exposure to 1,2-dibromo-3-chloropropane (DBCP)

Bjørge, Christine; Wiger, Richard; Holme, Jørn A.; Brunborg, Gunnar; Scholz, Tim; Dyeing, Erik; Søderlund, Erik J.

doi:10.1016/0890-6238(95)02018-7

“…Human testicular biopsies were obtained from organ donors, and single-cell suspensions of testicular cells were prepared as described ( Bjorge et al, 1996a ). Cells (unfrozen) were processed for comet assay analysis, stained with ethidium bromide, and analyzed using the Fenestra Comet image analysis system (Kinetic Imaging LTD, Liverpool, UK; Bjorge et al, 1996b ). Comet Tail Moments (TM; these experiments were partly done in the early days of the comet assay, when TM was often used) and total fluorescence intensity (TFI) were recorded for each cell; these parameters are already integrated in Fenestra.…”

Section: Methodsmentioning

confidence: 99%

“…We applied alkaline filter elution to measure DNA damage in testicular cell populations partly purified by means of centrifugal elutriation. We subsequently found that different testicular cell types could be identified when using the comet assay, which – unlike alkaline elution – allows measurement of DNA damage in individual cells ( Bjorge et al, 1996b ). This was possible since the fluorescent signal in the comet assay is related to the amount of DNA, which depends on cell ploidy.…”

Section: Introductionmentioning

confidence: 99%

Reference cells and ploidy in the comet assay

Brunborg

¹

,

Collins

²

,

Graupner

³

et al. 2015

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In the comet assay single cells are analyzed with respect to their level of DNA damage. Discrimination of the individual cell or cell type based on DNA content, with concomitant scoring of the DNA damage, is useful since this may allow analysis of mixtures of cells. Different cells can then be characterized based on their ploidy, cell cycle stage, or genome size. We here describe two applications of such a cell type-specific comet assay: (i) Testicular cell suspensions, analyzed on the basis of their ploidy during spermatogenesis; and (ii) reference cells in the form of fish erythrocytes which can be included as internal standards to correct for inter-assay variations. With standard fluorochromes used in the comet assay, the total staining signal from each cell – whether damaged or undamaged – was found to be associated with the cell’s DNA content. Analysis of the fluorescence intensity of single cells is straightforward since these data are available in scoring systems based on image analysis. The analysis of testicular cell suspensions provides information on cell type specific composition, susceptibility to genotoxicants, and DNA repair. Internal reference cells, either untreated or carrying defined numbers of lesions induced by ionizing radiation, are useful for investigation of experimental factors that can cause variation in comet assay results, and for routine inclusion in experiments to facilitate standardization of methods, and comparison of comet assay data obtained in different experiments or in different laboratories. They can also be used – in combination with a reference curve – to quantify the DNA lesions induced by a certain treatment. Fish cells of a range of genome sizes, both greater and smaller than human, are suitable for this purpose, and they are inexpensive.

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“…Metabolism of DBCP occurs largely in the liver via the microsomal cytochrome p450 system [8], where liver enzymes catalyze the biotransformation of DBCP to metabolites that are excreted in the bile and urine. DBCP is converted by glutathione S-transferases to a reactive, cytotoxic episulfonium ion in the testicular seminiferous tubules [9,10], and this metabolite can bind covalently to DNA, producing single-strand breaks [11]. This pathophysiologic activity most likely accounts for DBCP's specific toxicity during the spermatogenic cycle.…”

Section: Scientific Review Of Basic Mechanismsmentioning

confidence: 99%

“…Investigators have clearly demonstrated that DBCP is biotransformed to a greater extent to metabolites that covalently bind to the cells' DNA in rat testicular cells as compared to human testicular cells in vitro [11]. Investigators found a DBCP concentration-dependent increase in single-strand DNA breaks in rat DNA but no significant damage in human DNA was found at any concentration tested [11].…”

Section: Investigations Of Dbcp's Gonadotoxic Potency Have Been Carrimentioning

confidence: 99%

“…Investigators have clearly demonstrated that DBCP is biotransformed to a greater extent to metabolites that covalently bind to the cells' DNA in rat testicular cells as compared to human testicular cells in vitro [11]. Investigators found a DBCP concentration-dependent increase in single-strand DNA breaks in rat DNA but no significant damage in human DNA was found at any concentration tested [11]. This evidence suggests that testicular epithelium in humans receives a lower DBCP dose, metabolically activates less, detoxifies more, and experiences significantly less DNA damage and cytotoxicity than that in rats in response to equivalent DBCP inhalation exposures.…”

Section: Investigations Of Dbcp's Gonadotoxic Potency Have Been Carrimentioning

confidence: 99%

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Gonadotoxic Effects of DBCP: A Historical Review and Current Concepts

Hwang¹,

Walters²,

Lipshultz³

2013

TOUNJ

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Dibromo-chloro-propane (DBCP), a persistent liphophilic brominated organochlorine, has been produced for agricultural purposes as a nematocide since the 1950s. Widespread use due to its effectiveness as a pesticide continued until the late 1970s when early reports of its toxicity emerged from the laboratories, particularly its impact on spermatogenesis and other adverse reproductive health effects. Since then innumerable cases and studies have surfaced with clear impact after exposure to DBCP, however, the sustained effect of this exposure has yet to be completely understood. As a result of these studies, environmental agencies banned almost all agricultural uses of DBCP in the United States in the late 1970s. This review will try to balance the known toxicity of DBCP with a scientific assessment of published data and a summary of the legal issues that have resulted.

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1,2‐Dibrom‐3‐chlorpropan [MAK Value Documentation in German language, 2002]

2012

The MAK‐Collection for Occupational Health and Safety

0

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Veröffentlicht in der Reihe Gesundheitsschädliche Arbeitsstoffe , 34. Lieferung, Ausgabe 2002 Der Artikel enthält folgende Kapitel: Allgemeiner Wirkungscharakter Wirkungsmechanismus Toxikokinetik und Metabolismus Aufnahme, Verteilung, Ausscheidung Metabolismus Erfahrungen beim Menschen Einmalige Exposition Wiederholte Exposition Wirkung auf Haut und Schleimhäute Allergene Wirkung Reproduktionstoxizität Genotoxizität Kanzerogenität Tierexperimentelle Befunde und In‐vitro‐Untersuchungen Akute Toxizität Subakute, subchronische und chronische Toxizität Wirkung auf Haut und Schleimhäute Allergene Wirkung Reproduktionstoxizität Genotoxizität Kanzerogenität Bewertung

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DNA Strand breaks in testicular cells from humans and rats following in vitro exposure to 1,2-dibromo-3-chloropropane (DBCP)

Cited by 18 publications

References 24 publications

Reference cells and ploidy in the comet assay

Reference cells and ploidy in the comet assay

Gonadotoxic Effects of DBCP: A Historical Review and Current Concepts

1,2‐Dibrom‐3‐chlorpropan [MAK Value Documentation in German language, 2002]

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