Penetration rates into the stratum corneum layer: A novel quantitative indicator for assessing skin barrier function.

BACKGROUND: The stratum corneum (SC), the outermost layer of the skin epidermis, acts as an effective bi-directional barrier, preventing water loss (inside-outside barrier) and entry of foreign substances (outside-inside barrier). Although transepidermal water loss (TEWL) is a widely-used measure of barrier function, it represents only inside-outside protection. Therefore, we aimed to establish a non-invasive method for quantitative evaluation of the outside-inside barrier function and visually present a skin barrier model. MATERIALS AND METHODS: Skin barrier damage was induced by applying a closed patch of 1% sodium dodecyl sulfate to the forearms of eight participants; they were instructed to apply a barrier cream on a designated damaged area twice daily for 5 days. The SC barrier was evaluated by measuring TEWL and fluorescein sodium salt penetration rate before, immediately after, and 5 days after damage. The penetration rate was assessed using tape-stripping (TS) technique and fluorescence microscopy. RESULTS: The rates of fluorescein sodium salt penetration into the lower layers of SC differed significantly based on the degree of skin barrier damage. The correlation between penetration rate and TEWL was weak after two rounds of TS and became stronger after subsequent rounds. Five days after skin barrier damage, the penetration rate of all layers differed significantly between areas with and without the barrier cream application. CONCLUSION: Our findings demonstrated that the penetration rate was dependent on skin barrier conditions. The penetration rate and corresponding fluorescence images are suitable quantitative indicators that can visually represent skin barrier conditions.

Microplastic contamination in groundwater on a volcanic Jeju Island of Korea.

Microplastic (MPs) contamination in groundwater has received massive attention since plastic waste has been released directly into the environment. This study investigates MPs contamination in groundwater on the Jeju volcanic Island, Korea. To the best of our knowledge, this is the first study to identify MPs in groundwater from volcanic islands. A total of 21 sites were sampled for groundwater wells and springs in July and September (2021). Sampling was performed without cross-contamination through quality assurance and quality control. The results showed that MPs abundance ranged from 0.006 to 0.192 particles/L in groundwater samples. Additionally, MPs were detected in deep groundwater wells where the groundwater level was 143 m below ground surface. Eight MPs polymer types, including polypropylene, polyethylene, polyethylene terephthalate, polyvinyl chloride, polystyrene, polyamide, acrylonitrile butadiene styrene, and polyurethane, were detected using Micro-Fourier Transform Infrared Spectroscopy (µ-FT-IR). Most of the detected MPs size ranged from 20 to 100 µm, accounting for 95% of the total. Fragments and fiber shaped MPs were detected, with the majority of them being fragmented in groundwater samples. The concentrations of MPs and major ions in groundwater showed a positive correlation. A negative correlation was observed between MPs concentration and topographic elevation (r = -0.59, p = 0.01). The source of MPs contamination is most likely attributed to agricultural activities, such as plastic mulching and greenhouses, which account for most of the land use in the study area. In this study, MPs entered the aquifer through the soil at the surface and seeped through cracks in fractured rock on basalt with sealed groundwater wells. This study takes 500 L of samples to prevent sample bias, reveal plastic contamination in groundwater, and indicating the characteristics and sources of contaminated plastics.

The forgotten impacts of plastic contamination on terrestrial micro- and mesofauna: A call for research.

Microplastics (MP) and nanoplastics (NP) contamination of the terrestrial environment is a growing concern worldwide and is thought to impact soil biota, particularly the micro and mesofauna community, by various processes that may contribute to global change in terrestrial systems. Soils act as a long-term sink for MP, accumulating these contaminants and increasing their adverse impacts on soil ecosystems. Consequently, the whole terrestrial ecosystem is impacted by microplastic pollution, which also threatens human health by their potential transfer to the soil food web. In general, the ingestion of MP in different concentrations by soil micro and mesofauna can adversely affect their development and reproduction, impacting terrestrial ecosystems. MP in soil moves horizontally and vertically because of the movement of soil organisms and the disturbance caused by plants. However, the effects of MP on terrestrial micro-and mesofauna are largely overlooked. Here, we give the most recent information on the forgotten impacts of MP contamination of soil on microfauna and mesofauna communities (protists, tardigrades, soil rotifers, nematodes, collembola and mites). More than 50 studies focused on the impact of MP on these organisms between 1990 and 2022 have been reviewed. In general, plastic pollution does not directly affect the survival of organisms, except under co-contaminated plastics that can increase adverse effects (e.g. tire-tread particles on springtails). Besides, they can have adverse effects at oxidative stress and reduced reproduction (protists, nematodes, potworms, springtails or mites). It was observed that micro and mesofauna could act as passive plastic transporters, as shown for springtails or mites. Finally, this review discusses how soil micro- and mesofauna play a key role in facilitating the (bio-)degradation and movement of MP and NP through soil systems and, therefore, the potential transfer to soil depths. More research should be focused on plastic mixtures, community level and long-term experiments.

Methylmercury directly modifies the 105th cysteine residue in oncostatin M to promote binding to tumor necrosis factor receptor 3 and inhibit cell growth.

We previously found that methylmercury induces expression of oncostatin M (OSM), which is released extracellularly and binds to tumor necrosis factor receptor 3 (TNFR3), possibly enhancing its own toxicity. However, the mechanism by which methylmercury causes OSM to bind to TNFR3 rather than to its known receptors, OSM receptor and LIFR, is unknown. In this study, we aimed to elucidate the effect of methylmercury modification of cysteine residues in OSM on binding to TNFR3. Immunostaining of TNFR3-V5-expressing cells suggested that methylmercury promoted binding of OSM to TNFR3 on the cell membrane. In an in vitro binding assay, OSM directly bound to the extracellular domain of TNFR3, and this binding was promoted by methylmercury. Additionally, the formation of a disulfide bond in the OSM molecule was essential for the binding of both proteins, and LC/MS analysis revealed that methylmercury directly modified the 105th cysteine residue (Cys105) in OSM. Next, mutant OSM, in which Cys105 was replaced by serine or methionine, increased the binding to TNFR3, and a similar effect was observed in immunoprecipitation using cultured cells. Furthermore, cell proliferation was inhibited by treatment with Cys105 mutant OSMs compared with wildtype OSM, and this effect was cancelled by TNFR3 knockdown. In conclusion, we revealed a novel mechanism of methylmercury toxicity, in which methylmercury directly modifies Cys105 in OSM, thereby inhibiting cell proliferation via promoting binding to TNFR3. This indicates a chemical disruption in the interaction between the ligand and the receptor is a part of methylmercury toxicity.

Hydrogeological characteristics and water chemistry in a coastal aquifer of Korea: implications for land subsidence.

Land subsidence is the gradual or sudden dropping of the ground surface developed by increasing the total stress. Most studies have discussed the relationship between land subsidence with groundwater level. However, there is a lack of discussion on groundwater environmental changes after occurring land subsidence. This study aimed to evaluate the hydrogeological and water chemistry characteristics of construction sites with land subsidence. Land subsidence in the Yangyang coastal area occurred suddenly on August 3, 2022, when the retaining wall of the construction collapsed. The groundwater level was measured three times, and water samples were collected twice between August 5, 2022, and September 5, 2022, for laboratory analysis. After land subsidence occurred, the average groundwater level was - 19.91 m ground level (GL) on August 9, 2022, and finally decreased to - 19.21 m GL on September 05, 2022. The groundwater levels surrounding the construction site gradually increased for a month. The electrical conductivity value measured at the monitoring wells ranged from 89 to 7800 µS/cm, and four wells exceeded the measurement limit near the groundwater leaked points. The highest mixing ratio of leaked water samples, collected on August 9, 2022, was 27.6%. Furthermore, the fresh groundwater-saltwater interface depth was estimated to be above the construction bottom. Although groundwater levels recovered, the groundwater quality continuously is affected by saltwater. This finding could contribute to understanding the hydrogeological characteristics surrounding construction sites with land subsidence and provide insight into the hydrochemical evolution process during declined groundwater levels in coastal aquifers.

Cadmium toxicity mediated by the inhibition of SLC2A4 expression in human proximal Tubule cells.

Cadmium (Cd) is an environmental contaminant that causes renal toxicity. We have previously demonstrated that Cd induces renal toxicity by altering transcriptional activities. In this study, we show that Cd markedly inhibited the activity of transcription factor MEF2A in HK-2 human proximal tubule cells, which generated significant cytotoxicity in the cells. This reduction in the nuclear levels of MEF2A protein may be involved in the Cd-induced inhibition of MEF2A activity. We also demonstrate that one of the glucose transporters, GLUT4, was downregulated not only by Cd treatment but also by MEF2A knockdown. Knockdown of SLC2A4, encoding GLUT4, eliminated both cell viability and Cd toxicity. Cd treatment or SLC2A4 deficiency reduced the cellular concentration of glucose. Therefore, the suppression of SLC2A4 expression, which mediates the reduction in cellular glucose, is involved in Cd toxicity. The Cd toxicity induced by the reduction in GLUT4 may be associated with a reduction of cellular ATP levels in HK-2 cells. The levels of Slc2a4 mRNA in the kidney of mice exposed to Cd for 6 or 12 months were significantly lower than those in the control group. These results demonstrate that Cd exerts its cytotoxicity through the suppression in SLC2A4 expression and the subsequent inhibition of MEF2A transcriptional activity. Cd-induced suppression of SLC2A4 expression also reduces cellular ATP levels, partly by reducing glucose levels. This study suggests that the glucose transporter plays an important role in the renal toxicity of Cd, and provides a crucial breakthrough in our understanding of the mechanism of Cd toxicity.

Cadmium Toxicity Is Regulated by Peroxisome Proliferator-Activated Receptor δ in Human Proximal Tubular Cells.

Cadmium (Cd) is a toxic heavy metal that is widely present in the environment. Renal proximal tubule disorder is the main symptom of Cd chronic poisoning. Our previous study demonstrated that Cd inhibits the total activities of peroxisome proliferator-activated receptor (PPAR) transcription factors in human and rat proximal tubular cells. In this study, we investigated the involvement of PPAR in Cd renal toxicity using the HK-2 human proximal tubular cell line. Among PPAR isoform genes, only PPARD knockdown significantly showed resistance to Cd toxicity in HK-2 cells. The transcriptional activity of PPARδ was decreased not only by PPARD knockdown but also by Cd treatment. DNA microarray analysis showed that PPARD knockdown changed the expression of apoptosis-related genes in HK-2 cells. PPARD knockdown decreased apoptosis signals and caspase-3 activity induced by Cd treatment. PPARD knockdown did not affect the intracellular Cd level after Cd treatment. These results suggest that PPARδ plays a critical role in the modification of susceptibility to Cd renal toxicity and that the apoptosis pathway may be involved in PPARδ-related Cd toxicity.

Novel Technique for the Measurement of Fetal Right Modified Myocardial Performance Index Using Synchronized Images of Right Ventricular Inflow and Outflow and Clinical Application to Twin-to-Twin Transfusion Syndrome.

OBJECTIVE: To evaluate the reproducibility of the fetal right modified myocardial performance index (Mod-MPI) obtained by synchronizing the inflow and outflow images of the right ventricle (RV) and to evaluate its feasibility through clinical application to twin-to-twin transfusion syndrome (TTTS). METHODS: We prospectively evaluated 77 normal fetuses. Two experienced operators individually measured the right Mod-MPI using two different methods: (1) separate recording of the RV inflow and outflow using pulsed-wave Doppler (conventional method) and (2) synchronization of RV inflow and outflow images using the MPI + ™ function based on the closure click of the pulmonary valve (new method). To evaluate the clinical utility of the new method, we measured the right Mod-MPI in 33 TTTS fetuses using the new method before and after the fetoscopic laser coagulation (FLC). RESULTS: There was no statistical difference in Mod-MPI values between the two methods (p = .242). For both operators, the intra- and interoperator reproducibility of Mod-MPI was high with both methods (ICCs >0.950). Among the components of Mod-MPI measured using the new method, ejection time showed the highest reproducibility, whereas isovolumetric relaxation time demonstrated the lowest reproducibility. In TTTS fetuses, the Mod-MPI significantly decreased after FLC in recipients, and there was no difference in MPI values before and after FLC in donors. CONCLUSION: Right Mod-MPI measurement after the synchronization of RV inflow and outflow images is a reliable technique for evaluating fetal right cardiac function. This novel method can also independently evaluate the systolic and diastolic functions of the right heart.

Feasibility and accuracy of a novel automated three-dimensional ultrasonographic analysis system for abdominal aortic aneurysm: comparison with two-dimensional ultrasonography and computed tomography.

BACKGROUND: Accurate measurement of the maximum aortic diameter (Dmax) is crucial for patients with abdominal aortic aneurysm (AAA). Aortic computed tomography (CT) provides accurate Dmax values by three-dimensional (3-D) reconstruction but may cause nephrotoxicity because of contrast use and radiation hazard. We aimed to evaluate the accuracy of a novel semi-automated 3-D ultrasonography (3-D US) system compared with that of CT as a reference. METHODS: Patients with AAA (n = 59) or individuals with normal aorta (n = 18) were prospectively recruited in an outpatient setting. Two-dimensional ultrasonography (2-D US) and 3-D US images were acquired with a single-sweep volumetric transducer. The analysis was performed offline with a software. Dmax and the vessel area of the Dmax slice were measured with 2-D US, 3-D US, and CT. The lumen and thrombus areas of the Dmax slice were also measured in 40 patients with intraluminal thrombus. Vessel and thrombus volumes were measured using 3-D US and CT. RESULTS: The Dmax values from 3-D US demonstrated better agreement (R2 = 0.984) with the CT values than with the 2-D US values (R2 = 0.938). Overall, 2-D US underestimated Dmax compared with 3-D US (32.3 ± 12.1 mm vs. 35.1 ± 12.0 mm). The Bland-Altman analysis of the 3-D US values, revealed better agreement with the CT values (2 standard deviations [SD], 2.9 mm) than with the 2-D US values (2 SD, 5.4 mm). The vessel, lumen, and thrombus areas all demonstrated better agreement with CT than with 2-D US (R2 = 0.986 vs. 0.960 for the vessel, R2 = 0.891 vs. 0.837 for the lumen, and R2 = 0.977 vs. 0.872 for the thrombus). The thrombus volume assessed with 3-D US showed good correlation with the CT value (R2 = 0.981 and 2 SD in the Bland-Altman analysis: 13.6 cm3). CONCLUSIONS: Our novel semi-automated 3-D US analysis system provides more accurate Dmax values than 2-D US and provides precise volumetric data, which were not evaluable with 2-D US. The application of the semi-automated 3-D US analysis system in abdominal aorta assessment is easy and accurate.

Repeated measurements of facial skin characteristics using the Janus-â ¢ measurement system.

BACKGROUND: For personalized skin care, noninvasive quantitative methods to evaluate facial skin characteristics are important. Janus-III is one of the most widely used imaging analysis devices in the skin care industry in Korea. Janus-III generates values for a range of skin characteristics. Due to the convenience of obtaining results for a variety of skin characteristics in a single measurement, the use of Janus-III in cosmetic stores and research institutes has been recently increasing. However, the consistency of skin measurements of Janus-III has not been elucidated yet. MATERIALS AND METHODS: In this study, we repeated skin measurements three times for 70 different subjects and compared each numerical value in order to assess the consistency of the Janus-III. For this purpose, we compared between-sample distances and within-sample distances. RESULTS: We found important patterns for future analyses in terms of consistency. First, the average values of skin measurement categories were more reliable than individual part values of facial segments. Second, center part values such as forehead and nose were more reliable than side part values such as left and right part segments. CONCLUSION: If researchers who use Janus-III for studies of facial characteristics analyze average and center part values first, they can obtain relatively reliable patterns of facial skin characteristics.

Methylmercury toxic mechanism related to protein degradation and chemokine transcription.

Methylmercury is an environmental pollutant that causes neurotoxicity. Recent studies have reported that the ubiquitin-proteasome system is involved in defense against methylmercury toxicity through the degradation of proteins synthesizing the pyruvate. Mitochondrial accumulation of pyruvate can enhance methylmercury toxicity. In addition, methylmercury exposure induces several immune-related chemokines, specifically in the brain, and may cause neurotoxicity. This summary highlights several molecular mechanisms of methylmercury-induced neurotoxicity.

Imaging of transition charge densities involving carbon core excitations by all X-ray sum-frequency generation.

X-ray diffraction signals from the time-evolving molecular charge density induced by selective core excitation of chemically inequivalent carbon atoms are calculated. A narrowband X-ray pulse selectively excites the carbon K-edge of the -CH3 or -CH2F groups in fluoroethane (CH3-CH2F). Each excitation creates a distinct core coherence which depends on the character of the electronic transition. Direct propagation of the reduced single-electron density matrix, using real-time time-dependent density functional theory, provides the time-evolving charge density following interactions with external fields. The interplay between partially filled valence molecular orbitals upon core excitation induces characteristic femtosecond charge migration which depends on the core-valence coherence, and is monitored by the sum-frequency generation diffraction signal. This article is part of the theme issue 'Measurement of ultrafast electronic and structural dynamics with X-rays'.

Transcription Factors and Downstream Genes in Cadmium Toxicity.

Cadmium (Cd) is a harmful heavy metal widely present in the environment which can cause severe kidney damage. The proximal tubular cells are the main target of renal Cd toxicity. The consequences of Cd cytotoxicity involve apoptosis and necrosis. Recently, we and others have focused on how Cd affects transcription factors and the regulation of their target genes. Those studies showed that transcription factors initiate numerous pathways upon Cd exposure, leading to apoptosis, autophagic cell death, disruption of cell-cell adhesion, and generation of mitochondrial reactive oxygen species. Of particular note, Cd induces endoplasmic reticulum stress, resulting in not only apoptosis but also autophagic dysregulation, which can trigger cell damage. In some cases, however, Cd-regulated transcription factors can induce cell survival signaling. This review centers on our own research to elucidate the transcription factor-downstream gene cascades that are central to Cd-induced renal toxicity.

Comparing the feasibility and accuracy of three-dimensional ultrasound to two-dimensional ultrasound and computed tomography angiography in the assessment of carotid atherosclerosis.

AIMS: Two-dimensional ultrasound (2D-US) is the mainstay imaging technique used to evaluate carotid atherosclerosis. An automated single sweep three-dimensional ultrasound (3D-US) technique became available. We evaluated the feasibility and accuracy of 3D-US in the assessment of carotid plaques compared to those of 2D-US. Carotid computed tomography angiography (CTA) was used as a reference. METHODS AND RESULTS: Among 126 stroke patients who underwent carotid 2D-US, 73 underwent 3D-US and carotid CTA. 3D-US was pursued when there were carotid plaques or when area stenosis was ≥ 20% by 2D-US. Both 2D- and 3D-US images of the carotid arteries were acquired using a dedicated ultrasound system that was equipped with the single sweep volumetric transducer. In total, 266 arteries from 73 patients were selected for comparison of the detection rate of carotid plaques between 2D- and 3D-US. Among the 73 patients, carotid CTA detected 139 plaques. 3D-US demonstrated a higher detection rate of carotid plaques than did 2D-US (108 plaques (77.9%) vs. 70 plaques (50.4%)) when using carotid CTA as a reference standard. Carotid plaque volume (PV) of 133 vessels from 73 patients were quantitatively evaluated using both 3D-US and carotid CTA. Plaque volume of carotid artery was comparable between 3D-US and CTA (148.5 ± 133.0 mm3 vs. 154.1 ± 134.6 mm3 , P = .998, R: 0.9825, P-value for r < .001). CONCLUSION: 3D-US using a single sweep technique was a feasible and accurate method of detecting arterial plaques and assessing plaque volume.

Activation of ornithine decarboxylase protects against methylmercury toxicity by increasing putrescine.

We previously reported significantly increased level of putrescine, a polyamine, in the brains of mice administered methylmercury. Moreover, addition of putrescine to culture medium reduced methylmercury toxicity in C17.2 mouse neural stem cells. In this study, the role of ornithine decarboxylase (ODC), an enzyme involved in putrescine synthesis, in response to methylmercury toxicity was investigated. Methylmercury increased ODC activity in mouse cerebrum and cerebellum, but this increase was hardly observed in the kidney and liver, where methylmercury accumulated at a high concentration. In the cerebrum and cerebellum, increased putrescine was observed with methylmercury administration. Methylmercury increased ODC activity in C17.2 cells, but this was almost completely abolished in the presence of an ODC inhibitor. Methylmercury also increased the level of ODC protein in mouse brain and C17.2 cells. In addition, C17.2 cells pretreated with ODC inhibitor showed higher methylmercury sensitivity than control cells. These results suggest that the increased ODC activity by methylmercury is involved in the increase in putrescine level, and ODC plays an important role in the reduction of methylmercury toxicity. This is the first study to provide evidence that increased ODC activity may be a protective response against methylmercury-induced neurotoxicity.

Endothelial function estimated by digital reactive hyperemia in patients with atherosclerotic risk factors or coronary artery disease.

Due to the complex profile of atherosclerotic risk factors, an integrated analysis of a specific individual is difficult. Endothelial function reflects a composite of various risk factors, and may be used as an optimal tool to estimate the overall atherosclerotic risk. In this study, we investigated the value of digital Reactive Hyperemia Index (RHI) in an actual population with multiple atherosclerotic risk factors or coronary artery disease (CAD). A total of 417 patients from the Seoul National University Boramae Medical Center RHI registry were enrolled in this study. Patients were enrolled from January, 2013 to July, 2016. RHI was measured using the EndoPAT2000 system (Itamiar Medical Inc. Israel). The mean value of RHI was 1.67 ± 0.48 in total study population. Among various atherosclerotic risk factors, RHI was significantly lower in older (> 60 years) patients, diabetics, smokers' patients on statin therapy, and those with coronary or cerebrovascular disease. Most of these findings were consistent after adjustment with multiple regression analysis. RHI was significantly associated with CAD, with a hazard ratio of 1.80 (95% confidence interval 1.15-2.80, p = 0.010). In the subgroup with CAD, current smoking was the only finding showing a lower RHI. Brachial-ankle pulse wave velocity, which is a surrogate marker of arterial atherosclerotic change, was not correlated with RHI in patients with clinically significant atherosclerotic disease. RHI was significantly reduced by major atherosclerotic risk factors and in clinical atherosclerotic disease. RHI may reflect a composite effect of risk factors pertaining to the endothelial function.

Assessment of multidirectional movements of the common carotid artery in atherothrombotic stroke using dimensional speckle tracking carotid ultrasonography: A prospective, controlled cohort study.

AIM: We aimed to explore the relevance of multidirectional movements of the common carotid artery in atherothrombotic stroke. METHODS AND RESULTS: We prospectively enrolled 69 patients with atherothrombotic stroke (stroke group) and 69 age-matched and sex-matched controls (control group) who underwent carotid ultrasonography. Based on the speckle tracking technique, circumferential and radial movements of the common carotid artery were analyzed from transverse and longitudinal B-mode images of the common carotid artery. Peak longitudinal and radial displacements, strain, and strain rate were measured. Mean age of the overall population was 64 ± 11 years, and 57% (78 patients) were men. In the transverse image, circumferential strain was significantly lower in the stroke group than in the control group (5.6 ± 1.6 vs 4.2 ± 1.7, P < .001). In multivariate logistic regression analysis, circumferential strain was independently associated with stroke (odds ratio: 0.60, P = .001). Compared with conventional risk factors, as well as carotid intima-media thickness and carotid plaque, the addition of a strain parameter appeared to improve discrimination of a stroke event (area under the receiver operating characteristic curve: 0.65 and 0.75 vs 0.84, respectively; P < .05 for both). CONCLUSIONS: Circumferential strain of the common carotid artery might serve as a novel surrogate marker of atherothrombotic stroke. Multidirectional strain imaging of the common carotid artery may provide more information than conventional carotid ultrasonography and identify clinical risk factors for risk stratification of an acute stroke event.

Necdin modulates osteogenic cell differentiation by regulating Dlx5 and MAGE-D1.

Osteoblasts originate from mesenchymal stem cells that also differentiate into adipocytes, myoblasts, chondrocytes and fibroblasts. Osteogenic differentiation involves diverse regulatory proteins, including transcription and growth factors. Neurally differentiated embryonal carcinoma-derived protein (Necdin) has been identified as a key regulator of cell differentiation in various tissues, including neuronal, adipose, and muscular tissues; although its role in bone tissue remains to be established. Here, we investigated the potential involvement of Necdin in osteogenic differentiation. Our experiments revealed high expression of Necdin during osteoblast differentiation. Moreover, both transient and stable expression of Necdin induced osteoblast-specific markers in an osteogenic cell line through formation of a complex with melanoma-associated antigen D1 (MAGE-D1) and distal-less Homeobox 5 (Dlx5) and Runx2 promoter activation. Necdin expression was further associated with suppression of both cell proliferation and death in osteoblasts. Our results suggest that Necdin plays roles in cellular differentiation, proliferation and death in bone tissue.

In Vitro Effects of Polyphosphate against Prevotella intermedia in Planktonic Phase and Biofilm.

Polyphosphate (polyP) has gained a wide interest in the food industry due to its potential as a decontaminating agent. In this study, we examined the effect of sodium tripolyphosphate (polyP3; Na5P3O10) against planktonic and biofilm cells of Prevotella intermedia, a major oral pathogen. The MIC of polyP3 against P. intermedia ATCC 49046 determined by agar dilution method was 0.075%, while 0.05% polyP3 was bactericidal against P. intermedia in time-kill analysis performed using liquid medium. A crystal violet binding assay for the assessment of biofilm formation by P. intermedia showed that sub-MICs of polyP3 significantly decreased biofilm formation. Under the scanning electron microscope, decreased numbers of P. intermedia cells forming the biofilms were observed when the bacterial cells were incubated with 0.025% or higher concentrations of polyP3. Assessment of biofilm viability with LIVE/DEAD staining and viable cell count methods showed that 0.05% or higher concentrations of polyP3 significantly decreased the viability of the preformed biofilms in a concentration-dependent manner. The zone sizes of alpha-hemolysis formed on horse blood agar produced by P. intermedia were decreased in the presence of polyP3. The expression of the genes encoding hemolysins and the genes of the hemin uptake (hmu) locus was downregulated by polyP3. Collectively, our results show that polyP is an effective antimicrobial agent against P. intermedia in biofilms as well as planktonic phase, interfering with the process of hemin acquisition by the bacterium.

Single-molecule visualization of ROS-induced DNA damage in large DNA molecules.

We present a single molecule visualization approach for the quantitative analysis of reactive oxygen species (ROS) induced DNA damage, such as base oxidation and single stranded breaks in large DNA molecules. We utilized the Fenton reaction to generate DNA damage with subsequent enzymatic treatment using a mixture of three types of glycosylases to remove oxidized bases, and then fluorescent labeling on damaged lesions via nick translation. This single molecule analytical platform provided the capability to count one or two damaged sites per λ DNA molecule (48.5 kb), which were reliably dependent on the concentrations of hydrogen peroxide and ferrous ion at the micromolar level. More importantly, the labeled damaged sites that were visualized under a microscope provided positional information, which offered the capability of comparing DNA damaged sites with the in silico genomic map to reveal sequence specificity that GTGR is more sensitive to oxidative damage. Consequently, single DNA molecule analysis provides a sensitive analytical platform for ROS-induced DNA damage and suggests an interesting biochemical insight that the genome primarily active during the lysogenic cycle may have less probability for oxidative DNA damage.