Limits...
Protective effect of chitosan oligosaccharide lactate against DNA double-strand breaks induced by a model methacrylate dental adhesive.

Szczepanska J, Pawlowska E, Synowiec E, Czarny P, Rekas M, Blasiak J, Szaflik JP - Med. Sci. Monit. (2011)

Bottom Line: Monomers of methacrylates used in restorative dentistry have been recently reported to induce DNA double-strand breaks (DSBs) in human gingival fibroblasts (HGFs) in vitro.Because such monomers may penetrate the pulp and oral cavity due to the incompleteness of polymerization and polymer degradation, they may induce a similar effect in vivo.Therefore, the protection against DSBs induced by methacrylate monomers released from dental restorations is imperative.

View Article: PubMed Central - PubMed

Affiliation: Department of Developmental Dentistry, Medical University of Lodz, Lodz, Poland.

ABSTRACT

Background: Monomers of methacrylates used in restorative dentistry have been recently reported to induce DNA double-strand breaks (DSBs) in human gingival fibroblasts (HGFs) in vitro. Because such monomers may penetrate the pulp and oral cavity due to the incompleteness of polymerization and polymer degradation, they may induce a similar effect in vivo. DSBs are the most serious type of DNA damage and if misrepaired or not repaired may lead to mutation, cancer transformation and cell death. Therefore, the protection against DSBs induced by methacrylate monomers released from dental restorations is imperative.

Material/methods: We examined the protective action of chitosan oligosaccharide lactate (ChOL) against cytotoxic and genotoxic effects induced by monomers of the model adhesive consisting of 55% bisphenol A-diglycidyl dimethacrylate (Bis-GMA) and 45% 2-hydroxyethyl methacrylate (HEMA). We evaluated the extent of DSBs by the neutral comet assay and the phosphorylation of the H2AX histone test.

Results: ChOL increased the viability of HGFs exposed to Bis-GMA/HEMA as assessed by flow cytometry. ChOL decreased the extent of DSBs induced by Bis-GMA/HEMA as evaluated by neutral comet assay and phosphorylation of the H2AX histone. ChOL did not change mechanical properties of the model adhesive, as checked by the shear bond test. Scanning electron microscopy revealed a better sealing of the dentinal microtubules in the presence of ChOL, which may protect pulp cells against the harmful action of the monomers.

Conclusions: ChOL can be considered as an additive to methacrylate-based dental materials to prevent DSBs induction, but further studies are needed on its formulation with the methacrylates.

Show MeSH

Related in: MedlinePlus

DNA double strand breaks (DSBs) in human gingival fibroblasts exposed for 6 h at 37°C to the mixture of methacrylate monomers containing 55% bisphenol A-diglycidyl dimethacrylate and 45% 2-hydroxyethyl methacrylate (w/w) (Bis-GMA/HEMA) at 0.2 Bis-GMA with (+ ChOL) or without (– ChOL) 0.2% chitosan oligosaccharide lactate (ChOL) evaluated by the phosphorylation of the H2AX histone assay and compared with unexposed control (C, white bar). The intensity of fluorescence of the phosphorylated histone, γ-H2AX, is plotted and this quantity is positively correlated with the number of DSBs. Hydrogen peroxide at 1 mM was used as a positive control. The cells were incubated with appropriate antibodies, stained with Alexa Fluor and propidium iodine and analyzed by flow cytometry (upper diagrams). Error bars denote SEM, * p<0.05, ** p<0.01,*** p<0.001 as compared with unexposed control, an asterisk above the last pair of bars indicates the significance of the difference between effects induced by Bis-GMA/HEMA with and without ChOL.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3539618&req=5

f5-medscimonit-17-8-br201: DNA double strand breaks (DSBs) in human gingival fibroblasts exposed for 6 h at 37°C to the mixture of methacrylate monomers containing 55% bisphenol A-diglycidyl dimethacrylate and 45% 2-hydroxyethyl methacrylate (w/w) (Bis-GMA/HEMA) at 0.2 Bis-GMA with (+ ChOL) or without (– ChOL) 0.2% chitosan oligosaccharide lactate (ChOL) evaluated by the phosphorylation of the H2AX histone assay and compared with unexposed control (C, white bar). The intensity of fluorescence of the phosphorylated histone, γ-H2AX, is plotted and this quantity is positively correlated with the number of DSBs. Hydrogen peroxide at 1 mM was used as a positive control. The cells were incubated with appropriate antibodies, stained with Alexa Fluor and propidium iodine and analyzed by flow cytometry (upper diagrams). Error bars denote SEM, * p<0.05, ** p<0.01,*** p<0.001 as compared with unexposed control, an asterisk above the last pair of bars indicates the significance of the difference between effects induced by Bis-GMA/HEMA with and without ChOL.

Mentions: We observed a significant increase in the tail DNA in the comet of the neutral comet assay for all concentrations of Bis-GMA/HEMA monomers mixture (Figure 4). These results prompted us to employ the H2AX histone phosphorylation assay as a more reliable method to detect and quantify DSBs and assess the protective potential of ChOL against DSBs induced by the Bis-GMA/HEMA monomers. The results confirmed the ability of the methacrylates of the model adhesive at 0.2 mM Bis-GMA to induce DSBs and the protective action of 0.2% ChOL, which decreased DSBs-inducing effect of Bis-GMA/HEMA monomers (p<0.05) (Figure 5).


Protective effect of chitosan oligosaccharide lactate against DNA double-strand breaks induced by a model methacrylate dental adhesive.

Szczepanska J, Pawlowska E, Synowiec E, Czarny P, Rekas M, Blasiak J, Szaflik JP - Med. Sci. Monit. (2011)

DNA double strand breaks (DSBs) in human gingival fibroblasts exposed for 6 h at 37°C to the mixture of methacrylate monomers containing 55% bisphenol A-diglycidyl dimethacrylate and 45% 2-hydroxyethyl methacrylate (w/w) (Bis-GMA/HEMA) at 0.2 Bis-GMA with (+ ChOL) or without (– ChOL) 0.2% chitosan oligosaccharide lactate (ChOL) evaluated by the phosphorylation of the H2AX histone assay and compared with unexposed control (C, white bar). The intensity of fluorescence of the phosphorylated histone, γ-H2AX, is plotted and this quantity is positively correlated with the number of DSBs. Hydrogen peroxide at 1 mM was used as a positive control. The cells were incubated with appropriate antibodies, stained with Alexa Fluor and propidium iodine and analyzed by flow cytometry (upper diagrams). Error bars denote SEM, * p<0.05, ** p<0.01,*** p<0.001 as compared with unexposed control, an asterisk above the last pair of bars indicates the significance of the difference between effects induced by Bis-GMA/HEMA with and without ChOL.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3539618&req=5

f5-medscimonit-17-8-br201: DNA double strand breaks (DSBs) in human gingival fibroblasts exposed for 6 h at 37°C to the mixture of methacrylate monomers containing 55% bisphenol A-diglycidyl dimethacrylate and 45% 2-hydroxyethyl methacrylate (w/w) (Bis-GMA/HEMA) at 0.2 Bis-GMA with (+ ChOL) or without (– ChOL) 0.2% chitosan oligosaccharide lactate (ChOL) evaluated by the phosphorylation of the H2AX histone assay and compared with unexposed control (C, white bar). The intensity of fluorescence of the phosphorylated histone, γ-H2AX, is plotted and this quantity is positively correlated with the number of DSBs. Hydrogen peroxide at 1 mM was used as a positive control. The cells were incubated with appropriate antibodies, stained with Alexa Fluor and propidium iodine and analyzed by flow cytometry (upper diagrams). Error bars denote SEM, * p<0.05, ** p<0.01,*** p<0.001 as compared with unexposed control, an asterisk above the last pair of bars indicates the significance of the difference between effects induced by Bis-GMA/HEMA with and without ChOL.
Mentions: We observed a significant increase in the tail DNA in the comet of the neutral comet assay for all concentrations of Bis-GMA/HEMA monomers mixture (Figure 4). These results prompted us to employ the H2AX histone phosphorylation assay as a more reliable method to detect and quantify DSBs and assess the protective potential of ChOL against DSBs induced by the Bis-GMA/HEMA monomers. The results confirmed the ability of the methacrylates of the model adhesive at 0.2 mM Bis-GMA to induce DSBs and the protective action of 0.2% ChOL, which decreased DSBs-inducing effect of Bis-GMA/HEMA monomers (p<0.05) (Figure 5).

Bottom Line: Monomers of methacrylates used in restorative dentistry have been recently reported to induce DNA double-strand breaks (DSBs) in human gingival fibroblasts (HGFs) in vitro.Because such monomers may penetrate the pulp and oral cavity due to the incompleteness of polymerization and polymer degradation, they may induce a similar effect in vivo.Therefore, the protection against DSBs induced by methacrylate monomers released from dental restorations is imperative.

View Article: PubMed Central - PubMed

Affiliation: Department of Developmental Dentistry, Medical University of Lodz, Lodz, Poland.

ABSTRACT

Background: Monomers of methacrylates used in restorative dentistry have been recently reported to induce DNA double-strand breaks (DSBs) in human gingival fibroblasts (HGFs) in vitro. Because such monomers may penetrate the pulp and oral cavity due to the incompleteness of polymerization and polymer degradation, they may induce a similar effect in vivo. DSBs are the most serious type of DNA damage and if misrepaired or not repaired may lead to mutation, cancer transformation and cell death. Therefore, the protection against DSBs induced by methacrylate monomers released from dental restorations is imperative.

Material/methods: We examined the protective action of chitosan oligosaccharide lactate (ChOL) against cytotoxic and genotoxic effects induced by monomers of the model adhesive consisting of 55% bisphenol A-diglycidyl dimethacrylate (Bis-GMA) and 45% 2-hydroxyethyl methacrylate (HEMA). We evaluated the extent of DSBs by the neutral comet assay and the phosphorylation of the H2AX histone test.

Results: ChOL increased the viability of HGFs exposed to Bis-GMA/HEMA as assessed by flow cytometry. ChOL decreased the extent of DSBs induced by Bis-GMA/HEMA as evaluated by neutral comet assay and phosphorylation of the H2AX histone. ChOL did not change mechanical properties of the model adhesive, as checked by the shear bond test. Scanning electron microscopy revealed a better sealing of the dentinal microtubules in the presence of ChOL, which may protect pulp cells against the harmful action of the monomers.

Conclusions: ChOL can be considered as an additive to methacrylate-based dental materials to prevent DSBs induction, but further studies are needed on its formulation with the methacrylates.

Show MeSH
Related in: MedlinePlus