title: The influence of glutathione on redox regulation by antioxidant proteins and apoptosis in macrophages exposed to 2-hydroxyethyl methacrylate (HEMA).
abstract: Resin monomers like 2-hydroxyethyl methacrylate (HEMA) disturb cell functions including responses of the innate immune system, mineralization and differentiation, or induce cell death via apoptosis. These phenomena are associated with oxidative stress and a reduction in the concentration of the antioxidant glutathione (GSH), resulting in imbalanced redox homeostasis. Thus far, the precise mechanism of how resin monomers interfere with cellular redox regulation is unknown. The present study provides insight into the induction of apoptosis and the differential expression of antioxidant enzymes depending on the availability of GSH. Buthionine sulfoximine (BSO) was used to inhibit GSH synthesis, while 2-oxothiazolidine-4-carboxylate (OTC), and N-acetylcysteine (NAC) as prodrugs supported GSH synthesis in RAW264.7 mouse macrophages exposed to HEMA (0-8 mm) for 24 h. The level of GSH was significantly decreased after cells were preincubated with BSO, and the formation of reactive oxygen species (ROS) increased in cultures subsequently exposed to HEMA. Apoptosis was drastically increased by BSO in HEMA-exposed cell cultures as well, but OTC and NAC retracted HEMA-induced cell death. These results show that dental monomer-induced apoptosis is causally related to the availability of GSH. The hydrogen peroxide decomposing enzymes glutathione peroxidase (GPx1/2) and catalase were differentially regulated in HEMA-exposed cultures. Expression of GPx1/2 was inhibited by HEMA and further reduced in the presence of BSO. SOD1 (superoxide dismutase) expression was inhibited in the presence of HEMA, and was decreased to an even greater extent by BSO, possibly due to H(2)O(2)-feedback inhibition. The expression of catalase was considerably up-regulated in HEMA-exposed cultures, implying that H(2)O(2) is the type of ROS that is significantly increased in monomer-exposed cells. OTC and NAC counteracted the effect of HEMA on GPx1/2, SOD1, and catalase expression. HO-1 (heme oxygenase) expression was strongly enhanced by HEMA, suggesting the need for further antioxidants like bilirubin to support enzyme activities that directly regulate H(2)O(2) equilibrium. Expression of the oxidoreductase thioredoxin (TRX1), the second major thiol-dependent antioxidant system in eukaryotic cells, was slightly reduced, while the oxygen-sensing protein HIF-1α was downregulated in HEMA-exposed cell cultures. These results indicate that cells and tissues actively respond to monomer-induced oxidative stress by the differential expression of enzymatic antioxidants.
authors.affiliation: Department of Operative Dentistry and Periodontology, University Hospital Regensburg, D-93042 Regensburg, Germany.
authors.forename: Stephanie
authors.initials: S
authors.lastname: Krifka
authors.forename: Karl-Anton
authors.initials: KA
authors.lastname: Hiller
authors.forename: Gianrico
authors.initials: G
authors.lastname: Spagnuolo
authors.forename: Anahid
authors.initials: A
authors.lastname: Jewett
authors.forename: Gottfried
authors.initials: G
authors.lastname: Schmalz
authors.forename: Helmut
authors.initials: H
authors.lastname: Schweikl
mesh_terms: D000818:Animals; D000975:Antioxidants / Q000378:metabolism*; D017209:Apoptosis / Q000187:drug effects*; D002478:Cells, Cultured; D005978:Glutathione / Q000494:pharmacology*; D008264:Macrophages / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D008689:Methacrylates / Q000494:pharmacology*; D051379:Mice; D008954:Models, Biological; D009336:Necrosis; D010084:Oxidation-Reduction / Q000187:drug effects; D011506:Proteins / Q000378:metabolism*; D017382:Reactive Oxygen Species / Q000378:metabolism; D012117:Resins, Synthetic / Q000494:pharmacology
chemical_list: D000975:Antioxidants; D008689:Methacrylates; D011506:Proteins; D017382:Reactive Oxygen Species; D012117:Resins, Synthetic; C005044:hydroxyethyl methacrylate; D005978:Glutathione