Search

Previous studies report that microsized monosodium titanates (MSTs) deliver metal ions and species to mammalian cells and bacteria with cell-specific and metal-specific effects. In this study, we explored the use of MST and a new synthesized nanosized monosodium titanate (nMST) to deliver gold(III), cisplatin, or platinum(IV) to two human cell lines with different population doubling times, in vitro. The effect was measured using a fluorescent mitochondrial activity assay (CellTiter-Blue(®) Assay). This fluorescence assay was implemented to mitigate optical density measurement errors owing to particulate titanate interference and allowed for the studies to be extended to higher titanate concentrations than previously possible. Overall, native MST significantly (p 0.05) decreased mitochondrial activity of both cell types by 50% at concentrations of50 mg/L. Native nMST significantly suppressed the rapidly dividing cell line (by 50%) over untreated cultures, but had no effect on the more slowly dividing cells. For both cell types, increased titanate concentrations resulted in increased effects from delivered metals. However, there was no difference in the effect of metal delivered from micro- versus nano-sized MST.

The in vitro cytotoxic response to endodontic sealers was assessed for one year. AH-Plus (AHP), Epiphany (EPH), EndoRez (ER), Guttaflow (GF), InnoEndo (IN), and Pulp Canal Sealer (PCS) were exposed to mouse osteoblasts and human monocytes after curing, 52 weeks of aging, and after resurfacing post-aging; cellular response was estimated by succinate dehydrogenase (SDH) activity. The effect of materials on TNFα secretion from activated (LPS) and inactivated monocytes also was measured. Cell responses were compared with ANOVA and Tukey post hoc analysis (α = 0.05). Initially, all materials except GF suppressed osteoblastic SDH activity compared with Teflon (Tf) controls. SDH activity in cells exposed to some aged sealers improved significantly; but IN and ER remained cytotoxic. When aged materials were resurfaced then tested, AHP, ER, GF, and IN did not change. EPH and PCS were more toxic. Monocytes responded similarly to the osteoblasts. No endodontic sealer activated monocytic TNFα secretion (p > 0.05 vs. −LPS Tf-controls). LPS-activated monocytes exposed to unresurfaced AHP and IN significantly suppressed TNFα secretion. When activated monocytes were exposed to the resurfaced sealers, differential suppression of TNFα secretion was observed for three of the four sealers tested (EPH, IN, and PCS). The results suggest that long-term aging may be a useful adjunct to in vitro assessment of these materials. © 2012 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2012.

Nickel-based alloys are used for dental restorations because of their strength, high moduli, and relatively low cost. However, these alloys corrode significantly in use, particularly in lower pH environments that are common under oral biofilms. Ni(II) corrosion products increase inflammatory cytokine secretion from activated monocytes, suggesting that nickel alloys may exacerbate inflammatory responses in adjacent periodontal tissues caused by dental plaque. Because inflammatory cytokine secretion is regulated in part by the NFκB signaling pathway, our goal in the current work was to determine whether Ni(II) altered cellular levels or nuclear localization of NFκB-family subtypes. THP1 monocytes were exposed to Ni(II) for 72 h, and activated with lipopolysaccharide for the last 30 min to 6 h. Secretion of IL6 and TNFα were measured using ELISA, and NFκB levels and localization was measured using SDS-PAGE with immunoblots and digital analysis. We observed that Ni(II) did not alter the levels of secreted TNFα from activated monocytes, but increased secreted IL6 levels about 30% over controls. Ni(II) did not alter whole-cell levels of any NFκB subtype, but increased nuclear persistance of p65 and c-Rel. Our results suggest that Ni(II) may increase inflammatory cytokine secretion by increasing nuclear localization of some NFκB subtypes. Further studies should be done to determine the prominence of this mechanism in clinical environments. © 2012 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2012.

Metal-based drugs are largely undeveloped in pharmacology. One limiting factor is the systemic toxicity of metal-based compounds. A solid-phase, sequestratable delivery agent for local delivery of metals could reduce systemic toxicity, facilitating new drug development in this nascent area. Amorphous peroxotitanates (APT) are ion-exchange materials with high affinity for several heavy metal ions and have been proposed to deliver or sequester metal ions in biological contexts. In the current study, we tested a hypothesis that APTs are able to deliver metals or metal compounds to cells. We exposed fibroblasts (L929) or monocytes (THP1) to metal-APT materials for 72 h in vitro and then measured cellular mitochondrial activity (SDH-MTT method) to assess the biological impact of the metal-APT materials versus metals or APT alone. APT alone did not significantly affect cellular mitochondrial activity, but all metal-APT materials suppressed the mitochondrial activity of fibroblasts (by 30-65% of controls). The concentration of metal-APT materials required to suppress cellular mitochondrial activity was below that required for metals alone, suggesting that simple extracellular release of the metals from the metal-APT materials was not the primary mechanism of mitochondrial suppression. In contrast to fibroblasts, no metal-APT material had a measurable effect on THP1 monocyte mitochondrial activity, despite potent suppression by metals alone. This latter result suggested that "biodelivery" by metal-APT materials may be cell type-specific. Therefore, it appears that APTs are plausible solid-phase delivery agents of metals or metal compounds to some types of cells for potential therapeutic effect.

The effects of hyperglycemia, altered cell function, or inflammatory mediators on implant corrosion are not well studied; yet, these effects are critical to implant biocompatibility and osseointegration. Because implant placement is burgeoning, patients with medically compromising systemic conditions such as diabetes are increasingly receiving implants, and the role of other inflammatory diseases on implant corrosion also needs investigation. In the current study, the corrosion properties of commercially available, machined titanium implants were studied in blood, cultures of monocytic cells, and solutions containing elevated dextrose concentrations. Implant corrosion was estimated by open circuit potentials, linear polarization resistance, and electrical impedance spectroscopy (EIS) for 26 h. In selected samples, THP1 monocytic cells were activated for 2 h with Lipopolysaccharide prior to implant exposure, and IL-1beta secretion was measured to assess the affect of the implants on monocyte activation. Implants under conditions of inflammatory stress exhibited more negative E(corr) values, suggesting an increased potential for corrosion. Linear polarization measurements detected increased corrosion rates in the presence of elevated dextrose conditions over PBS conditions. EIS measurements suggested that implants underwent surface passivation reactions that may have limited corrosion over the short term of this test. This result was supported by cyclic polarization tests. IL-1beta secretion was not altered under conditions of corrosion or implant exposure. The results suggest that inflammatory stress and hyperglycemia may increase the corrosion of dental endosseous titanium-based implants, but that longer, more aggressive electrochemical conditions may be necessary to fully assess these effects.

Monosodium titanate (MST) particles effectively bind specific metals and are therefore promising compounds for delivery or sequestration of metals in biological contexts. Yet, the biological properties of MST are largely unexplored. Our previous study showed that the cytotoxicity of these compounds was mild, but the nature of the dose response curves suggested that residual titanates in culture may have interfered with the assay. In the current study, we assessed the importance of these artifacts, and extended our previous results using fibroblasts for biological evaluation. We also assessed the biological response to a new type of titanate (referred to as amorphous peroxo-titanate or APT) that shows more promising metal binding properties than MST. Methods: The degree of titanate-induced interference in the MTT (mitochondrial activity assay) was estimated by means of cell-free assays with and without a final centrifugation step to remove residual titanate particulate. Cytotoxic responses to titanates were assessed by measuring succinate dehydrogenase activity (by MTT) in THP1 monocytes or L929 fibroblasts after 24–72 h exposures. Monocytic activation by APT was assessed by TNFα secretion (ELISA) from monocytes with or without lipopolysaccharide (LPS) activation. Results: We confirmed that residual titanate particulates may alter the SDH activity assay, but that this effect is eliminated by adding a final centrifugation step to the standard MTT procedure. Addition of MST or APT at concentrations up to 100 mg/L altered succinate dehydrogenase activity by < 25% in both monocytes and fibroblasts. Fibroblasts displayed time-dependent adaptation to the MST. APT did not trigger TNFα secretion or modulate LPS-induced TNFα secretion from monocytes. Conclusions: Although further in vitro and in vivo assessment is needed, MST and APT exhibit biological properties that are promising for their use as agents to sequester or deliver metals in biological systems. © 2007 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2007

We assessed the biological response to several novel titanium alloys that have promising physical properties for biomedical applications. Four commercial titanium alloys [Super-TIX(R) 800, Super-TIX(R) 51AF, TIMETAL(R) 21SRx, and Ti-6Al-4V (ASTM grade 5)] and three experimental titanium alloys [Ti-13Cr-3Cu, Ti-1.5Si and Ti-1.5Si-5Cu] were tested. Specimens (n = 6; 5.0 x 5.0 x 3.0 mm(3)) were cast in a centrifugal casting machine using a MgO-based investment and polished to 600 grit, removing 250 mum from each surface. Commercially pure titanium (CP Ti: ASTM grade 2) and Teflon (polytetrafluoroethylene) were used as positive controls. The specimens were cleaned and disinfected, and then each cleaned specimen was placed in direct contact with Balb/c 3T3 fibroblasts for 72 h. The cytotoxicity [succinic dehydrogenase (SDH) activity] of the extracts was assessed using the MTT method. Cytotoxicity of the metals tested was not statistically different compared to the CP Ti and Teflon controls (p > 0.05). These novel titanium alloys pose cytotoxic risks no greater than many other commonly used alloys, including commercially pure titanium. The promising short-term biocompatibility of these Ti alloys is probably due to their excellent corrosion resistance under static conditions, even in biological environments.

Monosodium titanate (MST) is an inorganic sorbent/ion exchanger developed for the removal of radionuclides from nuclear wastes. We investigated the ability of MST to bind Cd(II), Hg(II), Au(III), or the Au-organic compound auranofin to establish the utility of MST for applications in environmental decontamination or medical therapy (drug delivery). Adsorption isotherms for MST were determined at pH 7-7.5 in water or phosphate-buffered saline. The extent of metal binding was determined spectroscopically by measuring the concentrations of the metals in solution before and after contact with the MST. Cytotoxic responses to MST were assessed using THP1 monocytes and succinate dehydrogenase activity. Monocytic activation by MST was assessed by TNFalpha secretion (ELISA) with or without lipopolysaccharide (LPS) activation. MST adsorbed Cd(II), Hg(II), and Au(III) under conditions similar to those in physiological systems. MST exhibited the highest affinity for Cd(II) followed by Hg(II) and Au (III). MST (up to 100 mg/L) exhibited only minor (

The perennial controversy about the safety of mercury in dental amalgams has adversely affected the availability and the quality of dental care. Chronic Hg(II) blood concentrations above 300 nM are known to alter function of the nervous system and the kidney. However, the effects of blood concentrations of 10 to 75 nM, far more common in the general population, are not clear and mechanisms of any effects are not known. The monocyte is an important potential target of Hg(II) because of its critical role in directing inflammatory and immune responses. In the current study we tested the hypothesis that concentrations of Hg(II) of 10 to 300 nM alter monocyte activity via a redox-dependent mechanism. Mitochondrial activity was used to establish inhibitory concentrations of Hg(II) following 6 to 72 h of exposures to THP1 human monocytic cells. Then subinhibitory concentrations were applied, and total glutathione levels and reactive oxygen species (ROS) were measured. Antioxidants [N-acetyl cysteine, (NAC); Na2SeO3, (Se)] and a pro-oxidant (tert-butylhydroquinone, tBHQ) were used to support the hypothesis that Hg(II) effects were redox-mediated. After 72 h of exposure, 20 microM of Hg(II) inhibited monocytic mitochondrial activity by 50%. NAC mitigated Hg(II)-induced mitochondrial suppression only at concentrations of greater than 10 microM, but Se had few effects on Hg-induced mitochondrial responses. tBHQ significantly enhanced mitochondrial suppression at higher Hg(II) concentrations. Hg(II) concentrations of 75 and 300 nM (0.075 and 0.30 microM, respectively) significantly increased total glutathione levels, and NAC mitigated these increases. Se plus Hg(II) significantly elevated Hg-induced total cellular glutathione levels. Increased ROS levels were not detected in monocytes exposed to mercury. Hg(II) acts in monocytic cells, at least in part, through redox-mediated mechanisms at concentrations below those commonly associated with chronic mercury toxicity, but commonly occurring in the blood of some dental patients.

The use of hydrophilic dental monomers in dentin bonding agents has vastly improved resin–dentin bond strengths, but incomplete polymerization of these monomers and their leaching into adjacent (pulpal) oral tissues has raised concerns about their biocompatibility. The sublethal effects of these resins are virtually unknown, but their electrophilic nature led to the hypothesis that they may alter cellular oxidative stress pathways. Glutathione balance between reduced (GSH) and oxidized (GSSG) is a major mechanism by which cells maintain redox balance and was therefore the focus of the current investigation. THP-1 human monocytic cells were exposed to hydroxyethyl methacrylate (HEMA), benzoyl peroxide (BPO), camphorquinone (CQ), or triethyelene glycol dimethacrylate (TEGDMA) for 24 h at sublethal doses, then GSH and GSSG levels were measured by means of Ellman's method adapted for cell culture. The results indicate that these dental resin compounds act at least partly via oxidative stress by increasing GSH levels at sublethal concentrations. However, the GSH-GSSG ratio was relatively unaffected. Only BPO altered the GSH-GSSG ratio at 24 h, again at sublethal levels (7.5–15 μmol/L). The results support the hypothesis that resin monomers act, at least in part, via oxidative stress, and that oxidative-stress pathways should be one focus of future investigations of monomer biocompatibility. © 2005 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater

Blue light (lambda = 380-500 nm) historically has been used to initiate polymerization of biomaterials and recently has been proposed as a therapeutic agent. New evidence suggests that cell-type-specific responses result from redox changes induced by exposure to blue light. Cultured cells were exposed to defined doses of blue light, equivalent to exposure times of 10 s and 2 min, to achieve energies of 5 J/cm2 and 60 J/cm2, respectively, after which (a) viable cell number, (b) cellular protein profiles, (c) mitochondrial succinate dehydrogenase (SDH) activity, (d) total reactive oxygen species (ROS), and (e) induction of apoptosis were compared to that of nonexposed control cultures. Results showed that blue-light exposure arrested monocyte cell growth and increased levels of peroxiredoxins. SDH activity of normal epidermal keratinocytes (NHEK) was slightly enhanced by blue light, whereas identical treatment of OSC2 oral tumor cells resulted in significant suppression of SDH activity. Blue-light exposure generally induced higher levels of total ROS in OSC2 cells than in NHEK. Finally, only OSC2 cells exhibited signs of apoptosis via Annexin V staining following exposure to blue light. These data support the central hypothesis that blue light induces an oxidative stress response in cultured cells resulting in cell-type-specific survival outcomes. The identification of oxidative stress as a mediator of the effects of blue light is a critical first step in defining its biological risks and therapeutic opportunities.

Many transition metals have been viewed collectively as nonspecific biological toxins in cells, which has limited investigation into their possible therapeutic effects. In the current study, the effects of Au(III), Ni(II), and Pd(II) on the differential secretion of cytokines from monocytes has been investigated. This is critical to understanding any therapeutic potential of these metals, their allergenicity, or the clinical effects of current metal therapies such as chrysotherapy. Lethal concentrations (defined as > 50% suppression of mitochondrial succinate dehydrogenase (SDH) activity) of metals were determined by dose-response curves with the use of 72 h exposures to human THP-1 monocytes. Then, secretion of TNFalpha, IL1beta, and IL6 were measured after the monocytes were exposed to sublethal concentrations of metals, with or without stimulation by lipopolysaccharide. The concentrations of Au(III), Pd(II), and Ni(II) required to suppress SDH activity by 50% were found to be 255, 270, and 90 microM, respectively. No sublethal concentration of any metal alone caused secretion of the cytokines. However, LPS-induced cytokine secretion was significantly and differentially altered by sublethal concentrations of each metal. Differential responses were highly dependent on metal concentration and involved both suppression and potentiation of the LPS activation. In the case of Ni(II), potentiation of TNFalpha, IL1beta, and IL6 ranged from 200% for TNFalpha to over 1200% for IL6. Metals such as Au(III), Pd(II), and Ni(II) differentially alter cytokine expression from monocytes. These results imply that metals have more specific effects on cell signaling than previously assumed. These results also are important in explaining multiple clinical effects often seen with chrysotherapy, identifying potential new avenues for metal therapy, and understanding the inflammatory effects of metals such as nickel.

The release of elements from dental alloys has been linked to alloy biocompatibility. Much of the research measuring elemental release has been done in vitro under passive conditions. The current study supplements a previous report that measured elemental release from dental alloys during and after the equivalent of 1 week of toothbrushing. In the current study, toothbrushing times were extended to the equivalent of 2 years, and elemental release was measured during and after brushing, with and without toothpaste. The results showed that for the major classes of dental alloys, brushing alone caused no significant elemental release during the brushing, and only minor increases after brushing. Brushing with toothpaste caused significant increases in elemental release for all elements of all alloys, but the largest increases were for the two nickel-based alloys. Nickel released during brushing with toothpaste reached 600-800 microg/cm(2) of alloy surface. Both beryllium-containing and non-beryllium-containing nickel-based alloys behaved similarly, refuting claims that non-beryllium alloys are superior in this regard. Thus, brushing with toothpaste under these extended in vitro conditions appears to increase the biological liabilities from elemental release for all alloys, but primarily for nickel-based alloys.