DOI: 10.5061/dryad.51c59zwmf
title: DNA adducts form in mouse lung and liver after oral naphthalene exposure

description: 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. 
description: 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 <sup>14</sup>C-naphthalene. Dosing
 solutions of <sup>14</sup>C-naphthalene were prepared by
 diluting stock <sup>14</sup>C-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
 <sup>14</sup>C-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.   
description: # DNA adducts form in mouse lung and liver after oral naphthalene exposure
 Dataset DOI: [10.5061/dryad.51c59zwmf](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.