title: In vitro profiling of epigenetic modifications underlying heavy metal toxicity of tungsten-alloy and its components.
abstract: Tungsten-alloy has carcinogenic potential as demonstrated by cancer development in rats with intramuscular implanted tungsten-alloy pellets. This suggests a potential involvement of epigenetic events previously implicated as environmental triggers of cancer. Here, we tested metal induced cytotoxicity and epigenetic modifications including H3 acetylation, H3-Ser10 phosphorylation and H3-K4 trimethylation. We exposed human embryonic kidney (HEK293), human neuroepithelioma (SKNMC), and mouse myoblast (C2C12) cultures for 1-day and hippocampal primary neuronal cultures for 1-week to 50-200 μg/ml of tungsten-alloy (91% tungsten/6% nickel/3% cobalt), tungsten, nickel, and cobalt. We also examined the potential role of intracellular calcium in metal mediated histone modifications by addition of calcium channel blockers/chelators to the metal solutions. Tungsten and its alloy showed cytotoxicity at concentrations > 50 μg/ml, while we found significant toxicity with cobalt and nickel for most tested concentrations. Diverse cell-specific toxic effects were observed, with C2C12 being relatively resistant to tungsten-alloy mediated toxic impact. Tungsten-alloy, but not tungsten, caused almost complete dephosphorylation of H3-Ser10 in C2C12 and hippocampal primary neuronal cultures with H3-hypoacetylation in C2C12. Dramatic H3-Ser10 dephosphorylation was found in all cobalt treated cultures with a decrease in H3 pan-acetylation in C2C12, SKNMC and HEK293. Trimethylation of H3-K4 was not affected. Both tungsten-alloy and cobalt mediated H3-Ser10 dephosphorylation were reversed with BAPTA-AM, highlighting the role of intracellular calcium, confirmed with 2-photon calcium imaging. In summary, our results for the first time reveal epigenetic modifications triggered by tungsten-alloy exposure in C2C12 and hippocampal primary neuronal cultures suggesting the underlying synergistic effects of tungsten, nickel and cobalt mediated by changes in intracellular calcium homeostasis and buffering.
authors.affiliation: Anatomy, Physiology and Genetics, Uniformed Services University of the Health Sciences, Bethesda, MD 20814, USA. Ranjana.Verma.CTR@usuhs.mil
authors.forename: Ranjana
authors.initials: R
authors.lastname: Verma
authors.forename: Xiufen
authors.initials: X
authors.lastname: Xu
authors.forename: Manoj K
authors.initials: MK
authors.lastname: Jaiswal
authors.forename: Cara
authors.initials: C
authors.lastname: Olsen
authors.forename: David
authors.initials: D
authors.lastname: Mears
authors.forename: Giuseppina
authors.initials: G
authors.lastname: Caretti
authors.forename: Zygmunt
authors.initials: Z
authors.lastname: Galdzicki
mesh_terms: D000497:Alloys / Q000633:toxicity*; D000818:Animals; D020746:Calcium Channels, L-Type / Q000502:physiology; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D044127:Epigenesis, Genetic; D006657:Histones / Q000378:metabolism; D006801:Humans; D051379:Mice; D010766:Phosphorylation; D014414:Tungsten / Q000633:toxicity*
chemical_list: D000497:Alloys; D020746:Calcium Channels, L-Type; D002614:Chelating Agents; D006657:Histones; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D004533:Egtazic Acid; D014414:Tungsten