Effects of HEMA on Nrf2-related gene expression using a newly developed 3D co-culture model of the oral mucosa. Issue 9 (September 2019)

• Record Type: Journal Article
• Authors: Perduns, Renke; Volk, Joachim; Plum, Melanie; Gutzki, Frank; Kaever, Volkhard; Geurtsen, Werner
• Is Part Of: Dental materials. Volume 35:Issue 9(2019)
• Page Start: 1214
• Page End: 1226
• Publication Date: 2019-09
• Subjects: HEMA -- Oral keratinocytes -- Gingival fibroblasts -- 3D co-culture -- Redox-regulated gene expression; Dentistry -- Periodicals; Dental materials -- Periodicals; 617.695
• DOI: 10.1016/j.dental.2019.05.006
• Languages: English
• ISSNs: 0109-5641

Abstract

Highlights: Characterization of a 3D co-culture model consisting of oral keratinocytes and HGFs. HEMA is able to penetrate a confluent layer of oral keratinocytes on culture inserts. HEMA induces Nrf2 target genes in both cell types in 3D co-culture models. Abstract: Objective: 2-Hydroxyethyl methacrylate (HEMA) is a component of many resin-modified materials and elutes from dental restorations into the oral cavity. Objective of our investigation was to determine the impact of HEMA on oral keratinocytes (OKF6/TERT2) and gingival fibroblasts (HGFs) in a newly established 3D co-culture model (3D-CCM) and to analyze the permeability of OKF6/TERT2 cells for HEMA. Methods: Well-characterized 3D-CCMs, consisting of confluent OKF6/TERT2 cells on cell culture inserts above HGF-containing collagen gels, were treated supra-epithelial with HEMA. Mass spectrometry was used to measure the supra- and sub-epithelial distribution of HEMA after 24 h. The impact of HEMA on nuclear factor erythroid 2-related factor 2 (Nrf2) target genes was measured by qRT-PCR and western blot analysis. Results: Mass spectrometry showed that HEMA was evenly distributed above and below the keratinocyte layer after 24 h. Analyzed target genes of Nrf2 were induced in both cell types on the mRNA-level but less pronounced in HGFs. On the protein-level, both cell types showed similar effects: At 5 mM HEMA, heme oxygenase-1 was induced 5.1-fold in OKF6/TERT2 cells and 4.1-fold in HGFs. NAD(P)H quinone dehydrogenase-1 was approximately induced 1.85-fold in both cell types. Significance: Our 3D-CCM is suitable to analyze the biocompatibility of dental materials due to an improved simulation of the oral mucosa compared to monolayer cultures. Our results indicate that HEMA is able to penetrate a dense layer of keratinocytes and to activate the cellular oxidative defense response. This may be due to the activation of the Nrf2-pathway in both cell types.