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Metallothionein blocks oxidative DNA damage in vitro

  • Immunotoxicology
  • Published:
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Abstract

The role of metallothionein (MT) in mitigation of oxidative DNA damage (ODD) induced by either cadmium (Cd) or the direct oxidant hydrogen peroxide (H2O2) was systematically examined using MT-I/II double knockout (MT-null) or MT-competent wild-type (WT) cells. Both toxicants were much more lethal to MT-null cells (Cd LC50 = 6.6 μM; H2O2 LC50 = 550 μM) than to WT cells (Cd LC50 = 16.5 μM; H2O2 LC50 = 930 μM). Cd induced concentration-related MT increases in WT cells, while the basal levels were undetectable and not increased by Cd in MT-null cells. ODD, measured by the immuno-spin trapping method, was minimally induced by sub-toxic Cd levels (1 or 5 μM; 24 h) in WT cells, but markedly increased in MT-null cells (>430 %). Similarly, ODD was induced to higher levels by lower concentrations of H2O2 in MT-null cells than WT cells. Transfection of MT-I into MT-null cells reduced both Cd- and H2O2-induced cytolethality and ODD. Cd increased the expression of the oxidant defense genes, HO-1, and GSTa2 to a much greater extent in MT-null cells than in WT. Cd or H2O2 exposure increased the expression of key transport genes, Mrp1 and Mrp2, in WT cells but not in MT-null cells. MT protects against Cd- and H2O2-induced ODD in MT-competent cells possibly by multiple mechanisms, potentially including direct metal ion sequestration and sequestration of oxidant radicals by MT. MT-deficient cells appear to adapt to Cd primarily by turning on oxidant response systems, while MT-competent cells activate MT and transport systems.

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Abbreviations

Cd:

Cadmium

CdCl2 :

Cadmium chloride

DMPO:

5,5-dimethyl-1-pyrroline N-oxide

GST:

Glutathione S-transferase

GST-π:

Glutathione S-transferase pi 1

GSTα2:

Glutathione S-transferase-α2

HO-1:

Heme oxygenase 1

H2O2 :

Hydrogen peroxide

IST:

Immuno-spin trapping

LC50 :

Lethal concentration 50 %

Mrp1:

Multidrug resistance-related protein 1

Mrp2:

Multidrug resistance-related protein 2

MT:

Metallothionein

MT-null:

MT-I/II knockout

ODD:

Oxidative DNA damage

RT-PCR:

Reverse transcription-polymerase chain reaction

ROS:

Reactive oxygen species

WT:

Wild type

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Acknowledgments

The authors thank Drs. Rachel Person, Katie Pelch, John Bucher, and Nigel Walker for critical review of this manuscript. The authors also thank Matthew W. Bell for aid with the graphics. This article may be the work product of an employee or a group of employees of the NIEHS, NIH; however, the statements contained herein do not necessarily represent the statements, opinions, or conclusions of the NIEHS, NIH. or the United States Government. The content of this publication does not necessarily reflect the views or the policies of the Department of Health and Human Services, nor does mention of trade names, commercial products, or organizations imply endorsement by the U.S. Government.

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Authors and Affiliations

  1. Inorganic Toxicology Group, National Toxicology Program Laboratory, Division of the National Toxicology Program, National Institute of Environmental Health Sciences, PO Box 12233, Mail Drop E1-07, 111 Alexander Drive, Research Triangle Park, NC, 27709, USA

    Wei Qu & Michael P. Waalkes

  2. Division of Translational Biology, The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC, 27709, USA

    Jingbo Pi

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  1. Wei Qu
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Correspondence to Michael P. Waalkes.

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Qu, W., Pi, J. & Waalkes, M.P. Metallothionein blocks oxidative DNA damage in vitro. Arch Toxicol 87, 311–321 (2013). https://doi.org/10.1007/s00204-012-0927-y

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