Extracellular environment as one mediator of blue light-induced mitochondrial suppression.

OBJECTIVES: The current study tested the hypothesis that the extracellular environment mediates mitochondrial suppression of oral epithelial cells and fibroblasts by blue light. METHODS: We exposed Balb fibroblasts (Balb), normal human epidermal keratinocytes (NHEK), and oral squamous carcinoma cells (OSC2) to blue light (30-120J/cm2) in different cell-culture media and in phosphate buffered saline (PBS). Mitochondrial activity (MTT method) was used to assess cellular response 72 h post-light exposure. Cell-culture media were replaced or supplemented before or after light exposure to assess the variables of exposure time and medium degradation as mediators of blue light-induced effects. RESULTS: Mitochondrial activity of NHEK was not suppressed by exposure to blue light regardless of extracellular conditions. The mitochondrial activity of OSC2 and Balb cells was suppressed most when cells were exposed to light in cell-culture medium (versus PBS). Blue light suppressed mitochondrial activity more when irradiated medium remained in contact with the cells at least 1h, indicating a time-dependence of the medium effects. Neither a replacement nor a supplementation of medium components reduced blue light-induced mitochondrial suppression. SIGNIFICANCE: Our results suggest that tissue environments influence cellular responses to blue light and that these environments should be considered when assessing any biological effects of blue light during the photopolymerization of restorative resins.

Gold-induced reactive oxygen species (ROS) do not mediate suppression of monocytic mitochondrial or secretory function.

UNLABELLED: The toxicity of anti-rheumatic gold compounds has limited their use and development, yet both the toxicological and therapeutic actions of these compounds remain unclear. In the current study, we tested the hypothesis that intracellular reactive oxygen species (ROS) induced by Au(I) or Au(III) compounds mediate their ability to suppress mitochondrial activity. METHODS: Human THP1 monocytes were exposed to HAuCl(4) x 3H(2)O (Au(III)), or the anti-rheumatic compounds auranofin (AF) or gold sodium thiomalate (GSTM) for 6-72 h, after which mitochondrial activity (succinate dehydrogenase) was measured. To assess the role of cellular redox status as a mediator of mitochondrial suppression, monocytes were pre-treated with a pro-oxidant (t-butyl hydroquinone, t-BHQ) or antioxidant (N-acetyl cysteine, NAC ). ROS levels were measured 0-24h post-gold addition to determine their role as mediators of mitochondrial activity suppression. RESULTS: AF was the most potent inhibitor of mitochondrial activity, followed by Au(III) and GSTM. Only Au(III) induced intracellular ROS; no ROS formation was observed in response to AF or GSTM exposure. Although anti- and pro-oxidants had some effects on mitochondrial suppression of Au compounds, collectively the data do not support redox effects or ROS formation as major mediators of Au-compound mitochondrial suppression. CONCLUSIONS: Our results do not indicate that ROS and redox effects play major roles in mediating the cytotoxicity of AF, GSTM or Au(III).

Corrosion rates of stainless steel under shear stress measured by a novel parallel-plate flow chamber.

A unique parallel-plate flow chamber has been engineered to assess the corrosion properties of implant materials in biological environments under shear flow. This parallel-plate flow chamber provides a novel approach to investigate hypotheses regarding cellular-material-mechanical-force interactions that influence the success or failure of implant devices. The results of the current study demonstrated that physiological stresses (0.5-50 dynes/cm2) from laminar flow from cell culture media did not significantly alter corrosion rates of stainless steel, providing baseline information for an extensive study of the cellular-material-mechanical-force interactions. Furthermore, this study demonstrated that this device is electrochemically stable and provides reproducible results within test parameters. In addition, the results were not significantly different from corrosion tests on bulk samples. Therefore, this system will be useful for investigating cell-material interactions under shear stress for implant alloys or other opaque materials. This information is currently lacking. The results of the present study also support further development of this test system to assess cellular responses to these materials under shear stresses.

Mercury (II) alters mitochondrial activity of monocytes at sublethal doses via oxidative stress mechanisms.

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.

Blue light generates reactive oxygen species (ROS) differentially in tumor vs. normal epithelial cells.

OBJECTIVES: Blue light of high intensity is commonly used in dentistry to activate polymerization of resin restorative materials. Other than its effects on the retina, the biological effects of blue light (380-500nm wavelengths) are poorly studied. Limited evidence suggests that blue light acts by forming intracellular reactive oxygen species (ROS) that then affect critical cell functions. If the biological effects of blue light are redox-mediated, antioxidants might be used to mitigate unwanted side effects of blue light during clinical use, or blue light might be used therapeutically to modulate redox-sensitive cell signaling responses. METHODS: Intracellular ROS were estimated using HFLUOR-DA (dihydrofluorescein diacetate), a vital fluorescein-based, redox-sensitive dye. ROS were measured in normal (NHEK) and oral squamous carcinoma (OSC2) epithelial cells, shown previously to respond differentially to blue light irradiation. Two-hour cumulative levels of ROS and approximate ROS lifetimes were measured after irradiation doses of 5-30 J/cm(2). The blue light-induced generation of ROS was further tested by the ability of the antioxidants N-acetylcysteine (NAC) and vitamin E to mitigate intracellular ROS levels. RESULTS: Dose-dependent ROS levels were generated in both NHEK and OSC2 cells, but cumulative levels were higher and persisted longer in the OSC2 cells. Both vitamin E and NAC significantly reduced blue-light-induced levels of ROS, but were more effective in the OSC2 cells. SIGNIFICANCE: The induction of intracellular ROS by blue light implies that redox effects may mediate cellular responses to blue light. This result suggests the opportunity to mitigate any effects of direct or coincident exposure during dental treatment via antioxidants, and the opportunity to exploit differences in redox processing among cells for possible treatment of epithelial cancer or wound healing.