Carol A. Lefebvre

Effect of dentin bonding agents on the secretion of inflammatory mediators from macrophages.

Dentin bonding agents (DBAs) have been proposed as substitutes for amalgam as root-end filling materials. The current study tested the hypothesis that certain components of DBAs could alter the secretion of cytokines from macrophages. Such alteration would likely be undesirable for healing of the periapical tissues. Human THP-1 macrophages were exposed to 2-hydroxyethyl methacrylate, 4-methacryloxyethyl trimelliate anhydride, bisphenol-gycidylmethacrylate, and urethane dimethacrylate. The secretion of interleukin-1 beta (IL-1 beta) and tumor necrosis factor-alpha (TNF-alpha) were measured with or without challenge by lipopolysaccharide (LPS). Results showed that all DBA components completely suppressed LPS-induced IL-1 beta and TNF-alpha secretion at concentrations that suppressed mitochondrial activity by 50%. In addition, 4-methacryloxyethyl trimelliate anhydride induced secretion of IL-1 beta, but not TNF-alpha, without the LPS challenge. These results indicate that DBA components may alter normal macrophage-directed inflammatory responses if the macrophages are exposed to sufficiently high concentrations of these components.

Cytotoxicity of eluates from light-polymerized denture base resins

This study examined the metabolic effects of eluates from four light-polymerized denture base resins and one heat-polymerized denture base resin on oral epithelial cells in vitro. The eluate was cell culture medium that contained either or both of apparently nonpolymerized components and reaction products that diffused out of the resin samples. Eluates were prepared by daily transfer of sample disks in a cell culture medium over 10 days. Toxicity of eluates was tested immediately after transfer (fresh) and after storage for 30 days (aged) by use of radioisotope incorporation and cell viability studies. The fresh eluates inhibited cell metabolism, whereas the aged eluates stimulated then inhibited the responses. Results suggest that the components that leach out of the tested materials do so at different rates and have prolonged toxic effects on cells. Thus soaking prosthesis in water before insertion may be beneficial.

Biocompatibility of visible light-cured resin systems in prosthodontics

Frequently dental products are introduced that have had little or no biologic testing. Cell culture systems that traditionally have been used for the study of cellular responses have recently been used to assess biocompatibility. This article reviews various cellular toxicity assays and their application to the resin systems used in clinical prosthodontics.

Effects of dentin bonding agents on macrophage mitochondrial activity.

Dentin bonding agents (DBA) have been considered for use as root-end fillings. Previous studies have documented the release of DBA components in vivo and in vitro, but the biological implications are not clear. The macrophage is important in wound healing, and likely to be important in any inflammatory response. Therefore, this study determined the concentrations of the components of DBAs that suppress the mitochondrial activity of human macrophages in vitro. THP-1 macrophages were cultured in the presence of four DBA components (2-hydroxyethyl methacrylate (HEMA), 4-methacryloxyethyl trimellitate anhydride (4-META), bisphenol-glycidylmethacrylate (Bis-GMA), and urethane dimethacrylate (UDMA)) at various concentrations and for varying durations. Residual effects were also measured after the resins were removed. Controls received only the vehicle solution, ethanol or water. THP-1 mitochondrial activity was estimated using the MTT assay, and the 50% toxicity concentrations (TC50) were determined graphically. Resin components suppressed the mitochondrial activity of macrophages at different concentrations (TC50 values for HEMA (10,000 mumol/L), 4-META (3,800 mumol/L), Bis-GMA (130 mumol/L), and UDMA (110 mumol/L) at 24 h, and the effect was time-dependent. Residual effects were observed for all resins.

Burning mouth syndrome: A review of etiologies

STATEMENT OF PROBLEM Dental practitioners occasionally have patients present clinically with a history of chief complaint of burning and painful sensations in the oral cavity. Often the patient demonstrates clinically normal mucosa, which can make formulating a diagnosis challenging. This scenario, has been referred to as burning mouth syndrome, a multifactorial syndrome. PURPOSE The purpose of this article is to present a review of etiologic factors and clinical implications related to the condition of burning mouth syndrome.

Effects of long-term sub-lethal concentrations of dental monomers on THP-1 human monocytes

Studies have shown that monomers from dental resins are acutely cytotoxic, but little is known of their long-term effects at sub-lethal concentrations. The current study determined the long-term effects of sub-lethal concentrations of TEGDMA (triethyleneglycol dimethacrylate) and Bis-GMA (bisphenol-glycidylinethacrylate), two common dental monomers, on the in vitro cellular proliferation, succinic dehydrogenase activity, and total cellular protein production of monocytes. Human THP-1 monocytes were exposed to concentrations of 100, 200, and 400 micromol l(-1) of TEGDMA or 1, 5, and 25 micromol l(-1) Bis-GMA for 5 weeks. Controls received only vehicle solutions of ethanol. Each week cellular proliferation (hemocytometer), succinic dehydrogenase (SDH) activity (MTT) and total cellular protein (bicinchoninic acid) were assessed. The results were compared with ANOVA and Tukey intervals (alpha = 0.05). TEDGMA had no proliferative or cellular protein effects, but increased SDH activity 20-60% in week 1 (p < 0.05). SDH activity then decreased 40% in week 2, followed by a gradual increase of 30-40% over week 3-5 (p < 0.05). Bis-GMA reduced proliferation by 40-60% from 1-5 weeks exposure (p < 0.05). However, SDH activity and total protein per cell were not affected. There was some indication of increased SDH activity after 5 weeks (20-30%, p < 0.05). Sub-lethal concentrations of TEGDMA and Bis-GMA have significant long-term effects on monocytes at low-dose 5-week exposures in vitro. Each monomer acted differently.

Effects of an aminomethacrylate on epithelial cell lipid metabolism

Methacrylates can affect cell functions by surfactant-like effects or by altering cell lipid composition. Dimethylaminoethyl methacrylate (DMAEMA), an activator widely used in visible-light polymerized dental resins has been shown to elute readily into aqueous environments. The current study examined the metabolism of this material by oral epithelial cells (HCP) and its subsequent effects on cell lipids. Cells were plated in culture medium, then exposed to DMAEMA in the presence of 14C-acetate, a precursor which labeled the cell lipids. Other cultures were prelabeled with radioisotope, then exposed to DMAEMA. After incubation, the cell lipids were extracted and separated by TLC. Radioactive lipids were located and quantitated. Exposure of the cells to DMAEMA resulted in decreased synthesis of cholesterol with a concomitant increase in sterol precursors. Cholesterol esters and triacylglycerides also increased. Among the polar lipids, phosphatidyl choline (PC) and phosphatidyl ethanolamine (PE) decreased in response to DMAEMA. However, dimethylphosphatidyl ethanolamine (DMPE), a precursor of PC not detectable in control cultures, accumulated to a significant extent in cells exposed to DMAEMA. Furthermore, changes in PC and DMPE levels persisted in the cells for at least 48 h after removal of the DMAEMA. The results indicate that DMAEMA produces alterations in the relative amounts of several cellular neutral and polar lipids. Such alterations, especially of the normal phospholipid composition, along with an alteration in cellular cholesterol, could result in altered membrane-associated cell functions.

Effect of treatment concentration on lipopolysaccharide affinity for two alloys

OBJECTIVE This study compared gram-negative bacterial lipopolysaccharide (LPS) adherence to and elution from a Type III gold and a Ni-Cr-Be alloy using Escherichia coli LPS. METHOD One-half of the specimens of each alloy were pre-treated with 500 micrograms non-radiolabeled E. coli LPS for 24 h at 37 degrees C. All disks were then incubated with 0.15, 15 or 150 micrograms radiolabeled E. coli LPS for 24 h at 37 degrees C. To evaluate radiolabeled LPS elution, specimens were transferred to LPS-free water and incubated for 24 h at 37 degrees C. The elution scheme, which consisted of 24 h incubations and subsequent transfer to new LPS-free water, continued for up to 96 h total elution. Radiolabeled LPS adherence and elution was determined through liquid scintillation spectrometry. Control disks not treated with LPS were evaluated throughout the study with an enzymatic assay to ensure that extraneous LPS contamination did not occur. A multifactor ANOVA (p = 0.05) was used to evaluate differences in adherence to alloy specimens based upon alloy type, pretreatment status and [3H]LPS concentration. A repeated measures analysis ANOVA (p = 0.05) was used to evaluate differences in elution patterns among groups over time. Least square means were compared in case of significant effects. RESULTS Toxin uptake at each treatment concentration was significantly different from the other treatment concentrations. In addition, significantly greater amounts of [3H]LPS eluted from the non-pretreated Ni-Cr-Be alloy following the 0.15 and 15 micrograms radiolabeled [3H]LPS treatment, whereas no difference in elution was found among experimental groups following the 150 micrograms [3H]LPS treatment. SIGNIFICANCE E. coli LPS, an LPS type representative of enteric bacteria common to the gingival sulcus, has differing affinities for the alloys. This affinity difference could influence periodontal inflammatory processes, thereby resulting in differing tissue responses adjacent to dental restorations fabricated from these materials. The interaction of other LPS types with these alloys could differ.

Response of oral mucosal cells to glass ionomer cements

Although glass ionomer cements are generally considered to be tissue-compatible, it has been suggested that unreacted components or setting reaction by-products can affect cell metabolism. The current study examined the effects of constituents leached out of three glass ionomer cements on growth and metabolism of oral epithelial cells. Aseptically prepared discs of Ketac-Cem Radiopaque (KCR), Ketac-Cem Maxicap (KCM) and Fuji I were incubated in Dulbeccos medium for 10 d, with daily medium changes. Cultures of hamster cheek pouch (HCP) cells, a line of hamster buccal pouch epithelial cells, were incubated in control or eluate-containing media for 24 h. Viable cell numbers were determined by the colorimetric MTS assay, and DNA and RNA syntheses were assessed using [3H]thymidine and [3H]uridine incorporation, respectively. Responses to materials were determined by comparison of cell numbers and radioisotope incorporation (counts per minute (cpm) per 1000 cells). Results were analysed by ANOVA and Duncans multiple range test, then converted to percent control for comparison. The eluates of all three materials from the first 24 h of soaking inhibited HCP cell growth. The number of cells in cultures exposed to Fuji were 88% of control cultures, while those exposed to KCR and KCM were 58% and 59% of control, respectively. The difference between Fuji-exposed and control cultures was significant (P < 0.05). The two Ketac cements were different from Fuji-exposed and control cultures (P < 0.05) but not from each other. All of the materials caused significant increases in labelling of DNA compared to control cultures (P < 0.05) when calculated on a per cell basis, but the materials did not differ from each other. Both Ketac cements also significantly stimulated labelling of RNA per cell compared to control cultures (P < 0.05). All effects of the material decreased over time. Results suggest that leachable components of the materials may affect the rate of progression of HCP cells through the cell cycle, rather than overt toxicity that results in cell death.

Alterations in cell lipid metabolism by glycol methacrylate (HEMA)

Components of dental resins such as dimethylaminoethyl methacrylate (DMAEMA) can alter cell lipid composition, presumably by esterase-mediated hydrolysis. The resulting dimethylethanolamine is incorporated into cell phospholipids, while the methacrylic acid may alter several metabolic pathways. We hypothesize that HEMA is cleaved in a similar manner and the released ethylene glycol is incorporated into cell lipids, yielding phosphatidylethylene glycol (PtEG), and the methacrylic acid alters other lipid pathways in a manner similar to that of methacrylic acid released from hydrolysis of DMAEMA. Cultures of hamster buccal pouch (HCP) and rabbit kidney (RK13) epithelial cells were exposed to subtoxic concentrations of HEMA in the presence of [14C]-acetate or [3H]-oleic acid. Other cultures were prelabeled with [14C]-acetate followed by exposure to various concentrations of HEMA. Cell lipids were extracted by the method of Bligh and Dyer and separated by thin layer chromatography on silica gel K-6 plates or SG-81 silica gel loaded chromatography paper. The fate of the ethylene glycol was traced using [14C]-ethylene glycol. Radioactive lipids were located using autoradiography and known standard lipids and quantitated by liquid scintillation spectrometry. In the presence of HEMA several classes of lipids were altered. Among the neutral lipids, the most notable changes involved sterol precursors, triglycerides, fatty acids, and cholesterol esters, while phosphatidylcholine was affected among the phospholipids. The results differed quantitatively between the two cell types. Results also suggest that EG, including that released by hydrolysis of HEMA, is incorporated into cell phospholipids, producing PtEG. The changes in neutral lipid labeling may occur by alteration of lipid synthetic pathways utilizing acetyl Co-A as well as inhibition of enzymes involved in synthesis of cholesterol from sterol precursors and hydrolysis of cholesterol esters. Synthesis of PtEG may take place via phospholipase D-mediated headgroup exchange. Alterations in the cellular lipids may affect cell membrane properties and associated cell functions.