DNA adducts form in mouse lung and liver after oral naphthalene exposure
Data files
Jul 14, 2025 version files 18.27 KB
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NA-DNA_invivo_WTLiver_dryad_csv.csv
6.11 KB
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NA-DNA_invivo_WTLung_dryad_csv.csv
5.40 KB
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README.md
6.76 KB
Abstract
Naphthalene is a ubiquitous environmental contaminant and suspected carcinogen that forms DNA adducts in tissue explants, which is a mechanism of concern for genotoxicity, but it was not known if naphthalene could form DNA adducts in vivo. Naphthalene-DNA adducts are small and difficult to detect, so we utilized accelerator mass spectrometry to detect adducts with high sensitivity. Wild-type C57BL/6 mice were orally exposed to 50 mg/kg 14C-naphthalene, lung and liver were collected 2, 4, 24, or 72 hours post-exposure, then DNA was isolated from the tissue. DNA was then processed for accelerator mass spectrometry analysis following standard protocols, explained in detail in Domanico et al. (2025)(doi.org/10.1093/toxsci/kfaf017). Resulting measurements of 14C signal in these DNA samples comprise this dataset, and can be used to calculate the level of DNA adduct formation in tissue. We detected naphthalene-DNA adducts at various timepoints post-exposure, with levels decreasing over time, though a subset of DNA adducts still persisted to 72 hours post-exposure in both lung and liver. Evidence of naphthalene-DNA adduct formation in vivo provides a foundation for further studies into the potential genotoxic mechanism of naphthalene.
Dataset DOI: 10.5061/dryad.51c59zwmf
Description of the data and file structure
Accelerator mass spectrometry (AMS) was used to detect stable DNA adducts formed with murine lung and liver DNA following an in vivo oral exposure to radiolabeled 14C-naphthalene. Dosing solutions of 14C-naphthalene were prepared by diluting stock 14C-naphthalene (Moravek Inc.) with unlabeled naphthalene (Sigma-Aldrich) to reach a lower specific activity acceptable for use with AMS. Wild-type C57BL/6 mice were exposed to the 14C-naphthalene dosing solution or a corn oil vehicle via oral gavage; mouse lung lobes and liver were then collected 2, 4, 24, or 72 hours post-exposure. Tissue was homogenized and DNA was extracted via silica adsorption (DNeasy Kit; Qiagen), with particular care taken to remove proteins, as naphthalene also forms protein-DNA adducts. DNA purity was confirmed by Nanodrop before proceeding with AMS processing. DNA samples were then dried in a quartz vial, combined with tributyrin carbon carrier, and combusted to yield filamentous carbon. AMS was used to measure Carbon-14 isotope ratios per sample, which can be used to calculate the quantity of naphthalene-DNA adducts in each sample. Further detail on methodology and adduct calculations can be found in Domanico et al. (2025) (doi.org/10.1093/toxsci/kfaf017).
Files and variables
File: NA-DNA_invivo_WTLung_dryad_csv.csv
Description: Information and measurements for lung DNA samples from C57BL/6 mice exposed to 14C-naphthalene (50 mg/kg oral). Values include specific activity of the dosing solution, DNA quantity, and fraction modern (14C signal), which are required to calculate total DNA adduct levels, as explained in the Supplement of DOI: 10.1093/toxsci/kfaf017.
Variables
- Sample ID: a unique identifier given to every experimental animal in the Laura Van Winkle Lab, and is recorded in a physical logbook for reference.
- Group: strain, sex, and treatment of each sample, deciphered as follows: “Strain_Sex_TreatmentHours”. WT = wildtype C57BL/6 mice. M = male, F = female. NA = 14C-naphthalene, CO = corn oil vehicle; samples collected at 02 = 2 hours post-exposure, 04 = 4 hours post-exposure, 24 = 24 hours post-exposure, 72 = 72 hours post-exposure. Example: “WT_M_NA72” = wild-type male exposed to 14C-naphthalene and collected 72 hours later. TB_batch_Letter used for tributyrin standards, which were not experimental samples and acted only as a carbon carrier. Use of tributyrin is explained in the Supplement of DOI: 10.1093/toxsci/kfaf017.
- AMS Batch: letter to designate which samples were run in the same AMS batch (only a certain number can fit in the sample wheel inserted into the accelerator setup).
- Sample Type: tributyrin (carbon carrier, no DNA) or lung DNA (DNA extracted from whole lung lobes).
- Sample Amount (ug DNA): quantity of DNA in the sample, in micrograms.
- Specific Activity (14C-NA; nCi/nmol): specific activity of the 14C-naphthalene solution used to expose the mice, in nCi/nmol. Vehicle = corn oil vehicle samples (which only contain endogenous 14C) and Carrier = tributyrin carbon carrier samples.
- AMS ID: another unique identifier for each sample, assigned prior to the accelerator mass spectrometry (AMS) run.
- TB Quantity (uL): tributyrin quantity, added as a carbon carrier to all experimental samples prior to sample combustion (see Supplement of DOI: 10.1093/toxsci/kfaf017 for more details).
- FRAC. MOD.: fraction modern is the representative unit of 14C signal in the sample (see Supplement of DOI: 10.1093/toxsci/kfaf017 for more details). Control (corn oil vehicle) DNA samples will have a low but measurable fraction modern, representative of the endogenous 14C present in the sample. Tributryin samples will also have a very low but measurable fraction modern for the same reason.
File: NA-DNA_invivo_WTLiver_dryad_csv.csv
Description: Information and accelerator mass spectrometry measurements for liver DNA samples from C57BL/6 mice exposed to 14C-naphthalene (50 mg/kg oral). Values include specific activity of the dosing solution, DNA quantity, and fraction modern (14C signal), which are required to calculate total DNA adduct levels, as explained in the Supplement of DOI: 10.1093/toxsci/kfaf017.
Variables
- Sample ID: a unique identifier given to every experimental animal in the Laura Van Winkle Lab, and is recorded in a physical logbook for reference.
- Group: strain, sex, and treatment of each sample, deciphered as follows: “Strain_Sex_TreatmentHours”. WT = wildtype C57BL/6 mice. M = male, F = female. NA = 14C-naphthalene, CO = corn oil vehicle; samples collected at 02 = 2 hours post-exposure, 04 = 4 hours post-exposure, 24 = 24 hours post-exposure, 72 = 72 hours post-exposure. Example: “WT_M_NA72” = wild-type male exposed to 14C-naphthalene and collected 72 hours later. TB_batch_Letter used for tributyrin standards, which were not experimental samples and acted only as a carbon carrier. Use of tributyrin is explained in the Supplement of DOI: 10.1093/toxsci/kfaf017.
- AMS Batch: letter to designate which samples were run in the same AMS batch (only a certain number can fit in the sample wheel inserted into the accelerator setup).
- Sample Type: tributyrin (carbon carrier, no DNA) or liver DNA (DNA extracted from homogenized whole liver).
- Sample Amount (ug DNA): quantity of DNA in the sample, in micrograms.
- Specific Activity (14C-NA; nCi/nmol): specific activity of the 14C-naphthalene solution used to expose the mice, in nCi/nmol. Vehicle = corn oil vehicle samples (which only contain endogenous 14C) and Carrier = tributyrin carbon carrier samples.
- AMS ID: another unique identifier for each sample, assigned prior to the accelerator mass spectrometry (AMS) run.
- TB Quantity (uL): tributyrin quantity, added as a carbon carrier to all experimental samples prior to sample combustion (see Supplement of DOI: 10.1093/toxsci/kfaf017 for more details).
- FRAC. MOD.: fraction modern is the representative unit of 14C signal in the sample (see Supplement of DOI: 10.1093/toxsci/kfaf017 for more details). Control (corn oil vehicle) DNA samples will have a low but measurable fraction modern, representative of the endogenous 14C present in the sample. Tributryin samples will also have a very low but measurable fraction modern for the same reason.
Code/software
Data can be viewed and analyzed using multiple software options that can open .csv files, including R, Microsoft Excel, or another spreadsheet program.
Access information
All data was generated by the research team.
Accelerator mass spectrometry (AMS) was used to detect stable DNA adducts formed with murine lung and liver DNA following an in vivo oral exposure to radiolabeled 14C-naphthalene. Dosing solutions of 14C-naphthalene were prepared by diluting stock 14C-naphthalene (Moravek Inc.) with unlabeled naphthalene (Sigma-Aldrich) to reach a lower specific activity acceptable for use with AMS. Wild-type C57BL/6 mice were exposed to the 14C-naphthalene dosing solution or a corn oil vehicle via oral gavage; mouse lung lobes and liver were then collected 2, 4, 24, or 72 hours post-exposure. Tissue was homogenized and DNA was extracted via silica adsorption (DNeasy Kit; Qiagen), with particular care taken to remove proteins, as naphthalene also forms protein-DNA adducts. DNA purity was confirmed by Nanodrop before proceeding with AMS processing. DNA samples were then dried in a quartz vial, combined with tributyrin carbon carrier, and combusted to yield filamentous carbon. AMS was used to measure Carbon-14 isotope ratios per sample, which can be used to calculate the quantity of naphthalene-DNA adducts in each sample.