Low-Intensity Pulsed Ultrasound Prevents Development of Bisphosphonate-Related Osteonecrosis of the Jaw-Like Pathophysiology in a Rat Model.

We developed a rat model of bisphosphonate-related osteonecrosis of the jaw (BRONJ) by removing a maxillary molar tooth (M1) from ovariectomized rats after treatment with alendronate. To mimic periodontitis, some of the rats were administered Porphyromonas gingivalis (p. gingivalis) at the M1 site every 2 to 3 d for 2 wk. Rats pretreated with alendronate plus p. gingivalis showed delayed healing of socket epithelia, periosteal reaction of alveolar bone formation and lower bone mineral density in the alveolus above adjacent M2 teeth. These abnormalities were prevented by tooth socket exposure to 20 min/d low-intensity pulsed ultrasound (LIPUS), which restored diminished expression of RANKL, Bcl-2, IL-6, Hsp70, NF-κB and TNF-α messenger ribonucleic acids in remote bone marrow, suggesting LIPUS prevented development of BRONJ-like pathophysiology in rat by inducing systemic responses for regeneration, in addition to accelerating local healing. Non-invasive treatment by LIPUS, as well as low-level laser therapy, may be useful for medication-related osteonecrosis of the jaw patients.

C-X-C Motif Chemokine Ligand 14 is a Unique Multifunctional Regulator of Tumor Progression.

Cancer is a leading cause of death and disease worldwide, with a tremendous financial impact. Thus, the development of cost-effective novel approaches for suppressing tumor growth and progression is essential. In an attempt to identify the mechanisms responsible for tumor suppression, we screened for molecules downregulated in a cancer progression model and found that the chemokine CXCL14, also called BRAK, was the most significantly downregulated. Increasing the production of CXCL14 protein by transfecting tumor cells with a CXCL14 expression vector and transplanting the cells into the back skin of immunodeficient mice suppressed tumor cell growth compared with that of parental tumor cells, suggesting that CXCL14 suppressed tumor growth in vivo. However, some studies have reported that over-expression of CXCL14, especially in stromal cells, stimulated the progression of tumor formation. Transgenic mice expressing 10-fold more CXCL14 protein than wild-type C57BL/6 mice showed reduced rates of chemical carcinogenesis, transplanted tumor growth, and metastasis without apparent side effects. CXCL14 also acts as an antimicrobial molecule. In this review, we highlight recent studies involving the identification and characterization of CXCL14 in cancer progression and discuss the reasons for the context-dependent effects of CXCL14 on tumor formation.

C-terminal α Domain of p63 Binds to p300 to Coactivate ß-Catenin.

TP63 (p63), a member of the tumor suppressor TP53 (p53) gene family, is essential for ectodermal tissue development and suppresses malignant progression of carcinomas. The most abundant isoform, ΔNp63α (referred to as p63), lacks the N-terminal transactivation (TA) domain, and was originally characterized as a dominant-negative type suppressor against p53 family proteins. It also binds to TCF/LEF to inhibit ß-catenin. Nevertheless, transcriptional activation by p63 has also been observed in varied systems. To understand the puzzling results, we analyzed the structure-function relationship of p63 in the control of ß-catenin-dependent transcription. p63 acted as a suppressor of moderately induced ß-catenin. However, when nuclear targeted S33Y ß-catenin was applied to cause the maximum enhancer activation, p63 displayed a ß-catenin-coactivating function. The DNA-binding domain of p63 and the target sequence facilitated it. Importantly, we newly found that, despite the absence of TA domain, p63 was associated with p300, a general adaptor protein and chromatin modifier causing transcriptional activation. C-terminal α domain of p63 was essential for p300-binding and for the coactivator function. These results were related to endogenous p63-p300 complex formation and Wnt/ß-catenin-responsive gene regulation by p63 in squamous cell carcinoma lines. The novel p63-p300 interaction may be involved in positive regulation of gene expression in tissue development and carcinogenesis.

TRPM5 mediates acidic extracellular pH signaling and TRPM5 inhibition reduces spontaneous metastasis in mouse B16-BL6 melanoma cells.

Extracellular acidity is a hallmark of solid tumors and is associated with metastasis in the tumor microenvironment. Acidic extracellular pH (pH e ) has been found to increase intracellular Ca2+ and matrix metalloproteinase-9 (MMP-9) expression by activating NF-κB in the mouse B16 melanoma model. The present study assessed whether TRPM5, an intracellular Ca2+-dependent monovalent cation channel, is associated with acidic pH e signaling and induction of MMP-9 expression in this mouse melanoma model. Treatment of B16 cells with Trpm5 siRNA reduced acidic pH e -induced MMP-9 expression. Enforced expression of Trpm5 increased the rate of acidic pH e -induced MMP-9 expression, as well as increasing experimental lung metastasis. This genetic manipulation did not alter the pH e critical for MMP-9 induction but simply amplified the percentage of inducible MMP-9 at each pH e . Treatment of tumor bearing mice with triphenylphosphine oxide (TPPO), an inhibitor of TRPM5, significantly reduced spontaneous lung metastasis. In silico analysis of clinical samples showed that high TRPM5 mRNA expression correlated with poor overall survival rate in patients with melanoma and gastric cancer but not in patients with cancers of the ovary, lung, breast, and rectum. These results showed that TRPM5 amplifies acidic pH e signaling and may be a promising target for preventing metastasis of some types of tumor.

13. Effect of Low-Intensity Pulsed Ultrasound (LIPUS) on Remote Bone Marrow in Rats With Healing Socket.

OBJECTIVE: We reported at the previous annual meeting that LIPUS treatment of the molar tooth sockets of retired breeder rats accelerated alveolar bone healing, and that associated humoral effects were seen with elevated blood flow. Namely, LIPUS induced VEGF/angiogenesis along with elevated baseline blood flow rate, which was further associated with a sudden depression of blood flow rate in the socket immediately after cessation of LIPUS treatment. Prior injection with EP4 PGE2 receptor antagonist, but not EP3 antagonist, abolished this LIPUS-induced depression, and topical application of PGE2 to the socket epithelium mimicked the LIPUS-induced depression. In fact, the serum level of PGE2 increased after LIPUS treatment, and significantly increased in the blood flow rate at remote sites on the foot dorsum and tail after 20 minutes. Therefore, in the current study, we examined the tibia bone marrow, which is likely to respond to circulating PGE2. METHODS: Right maxillary first molars were removed from retired female breeder rats in both the LIPUS and the control groups. LIPUS was applied extrabuccally to the socket every 24 hours for 2 weeks starting one day after extraction. Removed bone samples were fixed with 4% formaldehyde to prepare undecalcified frozen sections using Kawamoto's method for immunohistochemical or histochemical staining. Bone marrow samples dissected from the tibia were treated with RNAlater (Ambion) for later RT-PCR analysis. RESULTS AND DISCUSSION: Chemokine receptor CXCR4-positive bone marrow cells increased in the tibia of the LIPUS-treated rat. Together with ubiquitously expressed CXCL12(SDF-1), it is suggested that PGE2 released from the exposed socket is responsible for the recruitment, proliferation and mobilization of the precursors of bone forming cells. LIPUS is thought to exert humoral effects by recruiting bone marrow cells into the healing socket along with VEGF/angiogenesis induced by PGE2.

Porphyromonas gingivalis infection modifies oral microcirculation and aortic vascular function in the stroke-prone spontaneously hypertensive rat (SHRSP).

The functional modulation of vascular endothelial cells associated with stroke and periodontal disease has not yet been clarified. The objective of this study is to analyze the vascular endothelial function of periodontitis and stroke animal models. We examined endothelial function and gingival blood flow in oral microcirculation in vivo and measured the isometric tension in vitro of the aorta in animal models for lifestyle-related diseases, such as periodontitis and stroke. Gingival reactive hyperemia (GRH) was measured using laser Doppler flowmetry. Wistar Kyoto rats (WKY) were used as control animals; Porphyromonas gingivalis (P. gingivalis) infected WKY (WKY + Pg) as the periodontitis model; stroke-prone spontaneously hypertensive rat (SHRSP) as the stroke model; and a final group consisting of P. gingivalis infected SHRSP (SHRSP + Pg). Furthermore, for each group, the relaxation of descending aortic ring preparations was measured using a force transducer. The GRH was estimated by maximum response (peak), time taken for the maximum response to fall to one half (T1/2), and increased total amount of blood flow (mass). The relative change in T1/2 and mass increased in SHRSP + Pg compared to WKY. However, mass significantly increased in WKY (758.59 ± 88.21 ml/min/100 g s to 1755.55 ± 226.10 ml/min/100 g s) and SHRSP (1214.87 ± 141.61 ml/min/100 g s to 2674.32 ± 675.48 ml/min/100 g s) after treatment with acetylcholine. In addition, T1/2 and mass significantly increased in WKY + Pg (624.18 ± 96.36 ml/min/100 g s to 2629.90 ± 612.01 ml/min/100 g s) and SHRSP + Pg (1116.36 ± 206.24 ml/min/100 g s to 1952.76 ± 217.39 ml/min/100 g s) after treatment with nitroglycerin. Furthermore, the endothelium-dependent relaxation of ring preparations, evoked by acetylcholine, was attenuated in SHRSP compared with WKY, but not in SHRSP + Pg. This attenuation effect in SHRSP could be prevented by superoxide dismutase pretreatment. Our results suggest altered endothelial function may occur in gingival tissue in animal models experiencing both periodontitis and stroke. Therefore, these results indicate the disruption of vascular function in oral microcirculation may be caused by the interaction between the oxidative stress induced by periodontitis and nitric oxide in periodontitis, similar to the interactions present in stroke cases.

Protective effects of (6R)-5,6,7,8-tetrahydro-l-biopterin on local ischemia/reperfusion-induced suppression of reactive hyperemia in rat gingiva.

We herein investigated the regulatory mechanism in the circulation responsible for rat gingival reactive hyperemia (RH) associated with ischemia/reperfusion (I/R). RH was analyzed using a laser Doppler flowmeter. RH and I/R were elicited by gingival compression and release with a laser Doppler probe. RH increased in a time-dependent manner when the duration of compression was between 30 s and 20 min. This increase was significantly suppressed by N (ω)-nitro-l-arginine-methyl-ester (l-NAME), 7-nitroindazole (7-NI), and 2,4-diamino-6-hydroxypyrimidine (DAHP). However, RH was markedly inhibited following 60 min of compression. This inhibition was significantly decreased by treatments with superoxide dismutase (SOD), (6R)-5,6,7,8-tetrahydro-l-biopterin (BH4), and sepiapterin. The luminescent intensity of superoxide anion (O2 (•-))-induced 2-methyl-6-(4-methoxyphenyl)-3,7-dihydroimidazo-[1,2-a] pyrazine-3-one (MCLA) was markedly decreased by SOD and BH4, but only slightly by sepiapterin. BH4 significantly decreased O2 (•-) scavenging activity in a time-dependent manner. These results suggested that nitric oxide (NO) secreted by the nitrergic nerve played a role in regulating local circulation in rat gingiva. This NO-related regulation of local circulation was temporarily inhibited in the gingiva by the I/R treatment. The decrease observed in the production of NO, which was caused by suppression of NO synthase (NOS) activity subsequent to depletion of the NOS co-factor BH4 by O2 (•-), played a partial role in this inhibition.

Suppressed rate of carcinogenesis and decreases in tumour volume and lung metastasis in CXCL14/BRAK transgenic mice.

Cancer progression involves carcinogenesis, an increase in tumour size, and metastasis. Here, we investigated the effect of overexpressed CXC chemokine ligand 14 (CXCL14) on these processes by using CXCL14/BRAK (CXCL14) transgenic (Tg) mice. The rate of AOM/DSS-induced colorectal carcinogenesis in these mice was significantly lower compared with that for isogenic wild type C57BL/6 (Wt) mice. When tumour cells were injected into these mice, the size of the tumours that developed and the number of metastatic nodules in the lungs of the animals were always significantly lower in the Tg mice than in the Wt ones. Injection of anti-asialo-GM1 antibodies to the mice before and after injection of tumour cells attenuated the suppressing effects of CXCL14 on the tumor growth and metastasis, suggesting that NK cell activity played an important role during CXCL14-mediated suppression of tumour growth and metastasis. The importance of NK cells on the metastasis was also supported when CXCL14 was expressed in B16 melanoma cells. Further, the survival rates after tumour cell injection were significantly increased for the Tg mice. As these Tg mice showed no obvious abnormality, we propose that CXCL14 to be a promising molecular target for cancer suppression/prevention.

Medical-grade collagen peptide in injectables provides antioxidant protection.

Medical-grade collagen peptide is used as an additive agent in pharmaceutical formulations; however, it is unknown as to whether the compound exerts antioxidant effects in vitro. In this study, we evaluated the antioxidant effects of medical-grade collagen peptide on reactive oxygen species such as hydroxyl radical, superoxide anion radical and singlet oxygen using electron spin resonance and spin trapping. We confirmed that medical-grade collagen peptide directly inhibited hydroxyl radical generated by the Fenton reaction or by ultraviolet irradiation of hydrogen peroxide, and singlet oxygen. In addition, an antioxidant effect of medical-grade collagen peptide on singlet oxygen was observed in peptide fractions 12-22. The total amount of antioxidant amino acids (Gly, Hyp, Glu, Ala, Cys, Met and His) constituted more than half of the total amino acids in these fractions. These results suggest that the observed antioxidant properties of medical-grade collagen peptide are due to the compound containing antioxidant amino acids. Medical-grade collagen peptide, which is used in pharmaceuticals, and especially in injectables, could provide useful antioxidant properties to protect the active ingredient from oxidation.

Reactive oxygen species scavenging activity of Jixueteng evaluated by electron spin resonance (ESR) and photon emission.

Jixueteng, the dried stem of Spatholobus suberectus Dunn (Leguminosae), is a traditional Chinese herbal medicine that is commonly classified as a herb that promotes blood circulation and can be used to treat blood stasis. The aim of this study was to examine the reactive oxygen species (ROS) scavenging activity of Jixueteng and other herbal medicines. The ROS scavenging activities of the water extracts of Jixueteng, Cnidium officinale and Salvia miltiorrhiza were examined using an electron spin resonance (ESR) technique and faint luminescence measurement. The ESR signal intensities of the superoxide anion (O2·) and hydroxyl radical (HO·) were reduced more by Jixueteng than the other herbal medicines we tested. High photon emission intensity to hydrogen peroxide (H202) and HO· was observed in Jixueteng using the XYZ chemiluminescence system that was used as faint luminescence measurement and analysis. The results of the present study revealed that the ROS scavenging activity of 8% Jixueteng was the strongest among the herbal medicines we tested. It has been reported that Jixueteng includes various polyphenols. In the ROS scavenging activity by Jixueteng, it is supposed that the antioxidant activity caused by these polyphenols would contribute greatly. In conclusion, a water extract component of Jixueteng had potent free radical scavenging activity and an antioxidative effect that inhibited the oxidative actions of O2·â», H2O2 and HO·. Therefore, Jixueteng represents a promising therapeutic drug for reactive oxygen-associated pathologies.

Fasudil, a Rho kinase inhibitor, suppresses tumor growth by inducing CXCL14/BRAK in head and neck squamous cell carcinoma.

CXCL14/BRAK (BRAK) is a secreted chemokine with anti-tumor activity, and its expression is suppressed in tumor cells. We previously reported the anti-tumor activity of BRAK in cell lines of head and neck squamous cell carcinoma (HNSCC) and the suppression of BRAK secretion in these cells. BRAK secretion in fibrosarcoma cells is restored by Fasudil, which is a Rho-kinase (ROCK) inhibitor. In this study, we examined the anti-tumor effect of BRAK by evaluating its gene expression and protein secretion in HNSCC cell lines. We found that BRAK mediated the suppressive effect of Fasudil against HNSCC cells. Tumor development in female BALB/cAJclnu/nu mice was suppressed by Fasudil. Also secretion of BRAK protein by tumor cell lines in vitro was significantly stimulated by Fasudil treatment. Similarly, the production of BRAK protein was significantly increased by the addition of Fasudil to cultured tumor cells. Furthermore Fasudil significantly increased BRAK gene expression at the mRNA level in HNSCC cell line. Inhibition of the RhoA/ROCK pathway by siRNAs significantly stimulated BRAK gene expression. These results show that the tumor-suppressive effect of Fasudil was mediated by BRAK, suggesting that Fasudil may therefore be useful for the treatment of HNSCC.

Reactive oxygen species production in mitochondria of human gingival fibroblast induced by blue light irradiation.

In recent years, it has become well known that the production of reactive oxygen species (ROS) induced by blue-light irradiation causes adverse effects of photo-aging, such as age-related macular degeneration of the retina. Thus, orange-tinted glasses are used to protect the retina during dental treatment involving blue-light irradiation (e.g., dental resin restorations or tooth bleaching treatments). However, there are few studies examining the effects of blue-light irradiation on oral tissue. For the first time, we report that blue-light irradiation by quartz tungsten halogen lamp (QTH) or light-emitting diode (LED) decreased cell proliferation activity of human gingival fibroblasts (HGFs) in a time-dependent manner (<5 min). Additionally, in a morphological study, the cytotoxic effect was observed in the cell organelles, especially the mitochondria. Furthermore, ROS generation induced by the blue-light irradiation was detected in mitochondria of HGFs using fluorimetry. In all analyses, the cytotoxicity was significantly higher after LED irradiation compared with cytotoxicity after QTH irradiation. These results suggest that blue light irradiation, especially by LED light sources used in dental aesthetic treatment, might have adverse effects on human gingival tissue. Hence, this necessitates the development of new dental aesthetic treatment methods and/or techniques to protect HGFs from blue light irradiation during dental therapy.

Acidic extracellular microenvironment and cancer.

Acidic extracellular pH is a major feature of tumor tissue, extracellular acidification being primarily considered to be due to lactate secretion from anaerobic glycolysis. Clinicopathological evidence shows that transporters and pumps contribute to H+ secretion, such as the Na+/H+ exchanger, the H+-lactate co-transporter, monocarboxylate transporters, and the proton pump (H+-ATPase); these may also be associated with tumor metastasis. An acidic extracellular pH not only activates secreted lysosomal enzymes that have an optimal pH in the acidic range, but induces the expression of certain genes of pro-metastatic factors through an intracellular signaling cascade that is different from hypoxia. In addition to lactate, CO2 from the pentose phosphate pathway is an alternative source of acidity, showing that hypoxia and extracellular acidity are, while being independent from each other, deeply associated with the cellular microenvironment. In this article, the importance of an acidic extracellular pH as a microenvironmental factor participating in tumor progression is reviewed.

Increased oxidative stress biomarkers in the saliva of Down syndrome patients.

OBJECTIVE: The DNA oxidation byproduct 8-hydroxy-2'-deoxyguanosine (8-OHdG) is a well-known biomarker used to evaluate oxidative stress. We previously reported that the generation of reactive oxygen species (ROS) is increased in cultured gingival fibroblasts (GF) from patients with Down syndrome (DS). Thus, the aim of this study was to evaluate 8-OHdG as a marker of oxidative stress in saliva of DS patients. MATERIALS AND METHODS: The study group consisted of DS patients (66 patients; age range 1-62 years) and systemically healthy control subjects (71 subjects; age range 4-58 years). Periodontal status was judged based on standard measurements of probing depth (PD) and gingival index (GI). The salivary levels of 8-OHdG were determined using an enzyme-linked immunosorbent assay. RESULTS: The mean of PD and GI values were not significantly different between young (1-12 years) patients with DS (DS-1) and controls (C-1) or between adult (30-62 years) patients with DS (DS-2) and controls (C-2). There were statistically significant positive correlations between the salivary 8-OHdG levels and GI in the DS-1, DS-2 and C-2 groups, but not in the C-1. There were also statistically significant positive correlations between salivary 8-OHdG levels and PD in the DS-2 and C-2 groups, but not in the DS-1 or C-1 groups. The salivary levels of 8-OHdG of DS-1 and DS-2 groups were significantly higher than in the C-l and C-2 groups, respectively. CONCLUSIONS: These results suggest that progressive oxidative stress occurred in DS patients. Oxidative stress may contribute to the clinical features of DS, particularly to the progressive periodontitis characteristic of early ageing.

Fasudil suppresses fibrosarcoma growth by stimulating secretion of the chemokine CXCL14/BRAK.

We previously reported that chemokine CXCL14/BRAK (BRAK) has antitumor activity in several carcinoma cells indicating that BRAK secretion suppresses carcinoma cells. Ras-homologous small GTPase (RhoA) and Rho-associated coiled-coil-containing protein kinase (ROCK) are important regulators of secretory processes, and activation of the RhoA/ROCK signaling pathway stimulates tumor invasion and metastasis. We investigated the effects of fasudil, a specific ROCK inhibitor, on BRAK secretion and tumor progression in mesenchymal fibrosarcoma cells (MC57). We demonstrated the antitumor activity of secreted BRAK using MC57 transplantation of BRAK in overexpressed transgenic mice. Further, to eliminate the influence of change in the mRNA expression of endogenous BRAK, we produced stable MC57 cell lines expressing BRAK (MC57-BRAK) or mock vector (MC57-MOCK). Fasudil significantly increased BRAK secretion by MC57-BRAK cells in a dose-dependent manner. To determine the effect of fasudil on tumor growth, MC57-BRAK and MC57-MOCK cells were transplanted into wild-type mice. Fasudil treatment suppressed tumor growth only in mice that had received MC57-BRAK cell transplants. These results indicate that fasudil inhibits fibrosarcoma growth by stimulating BRAK secretion and suggests that fasudil therapy might have clinical efficacy.

Dental resin curing blue light induced oxidative stress with reactive oxygen species production.

Dental resin curing blue light has been used in the treatment of tooth bleaching and to restore teeth with resin-based composite fillings. However, there has been little consideration of its effect on oral tissues such as dental pulp and oral mucosa. The aim of this study was to investigate whether dental resin curing blue light irradiation affects the dental pulp, especially the blood vessels that are known as the first target of reactive oxygen species (ROS), which play an important role in vascular reactivity. We found that blue light irradiation increased the level of lipid peroxidation in isolated rat aorta blood vessels by measuring malondialdehyde. Furthermore, cell proliferative activity was decreased in a time-dependent manner and apoptosis of human aorta vascular smooth muscle cells (VSMCs) was induced. These results indicated that (ROS) such as hydrogen peroxide and hydroxyl radicals were generated in VSMCs by irradiation with blue light, and they induced cytotoxicity associated with oxidative stress, which increased lipid peroxidation and apoptosis. In addition, N-acetyl-l-cysteine, which is a typical intracellular antioxidant, protected VSMCs against cytotoxicity associated with oxidative stress. These findings suggested that antioxidants may be used to prevent oxidative stress in dental pulp by repeated and/or multiple treatments with blue light irradiation in future dental treatments.

Calcium-calmodulin signaling induced by epithelial cell differentiation upregulates BRAK/CXCL14 expression via the binding of SP1 to the BRAK promoter region.

The chemokine BRAK/CXCL14 (BRAK) is expressed in normal squamous epithelium, but is not expressed or is expressed at negligible levels in head and neck squamous cell carcinoma. Malignant cells are known to be dedifferentiated compared with normal epithelial cells, suggesting a role for differentiation cues in the expression of BRAK. Thus, we examined the relationship between BRAK expression and stages of differentiation level in epithelial cells. Immunohistochemical analysis showed that BRAK protein was expressed in cells above the spinous cell layer in normal epithelia. In HSC-3 cells in culture, expression of BRAK mRNA was significantly upregulated by cell contact in a cell density-dependent manner, and mRNA expression of cell differentiation markers such as involucrin, cystatin-A, TGM1, TGM3, and TGM5 was concomitantly augmented. Furthermore, the upregulation of BRAK induced by cell contact was suppressed by chlorpromazine, a specific inhibitor of calmodulin. We previously reported that GC boxes and a TATA-like sequence in the BRAK promoter region are associated with the expression of BRAK. Using a promoter assay and ChIP, we demonstrated that binding of the stimulating protein-1 (SP1) transcription factor to a GC box upstream of the BRAK transcription start site was necessary for cell density-dependent upregulation of BRAK. These results indicated that upregulation of BRAK was accompanied by differentiation of epithelial cells induced by calcium/calmodulin signaling, and that SP1 binding to the BRAK promoter region played an important role in this signaling.

Assessments of salivary antioxidant activity using electron spin resonance spectroscopy.

OBJECTIVE: In recent years, the function of saliva has been focused on evaluation of general status. The relationship between salivary antioxidant activity and periodontal disease progression is unclear. The aim of this study is to assess the relationship between periodontal disease and salivary antioxidant activity towards various reactive oxygen species (ROS) using electron spin resonance (ESR) technique. METHODS: We demonstrated that whole saliva derived rats or human subjects scavenged ROS such as superoxide (O(2)(·-)) and hydroxyl radical (HO(·)) using ESR spectroscopy with spin trapping agent. In addition, we assessed the relationship between antioxidants activity towards ROS and periodontal index with superoxide dismutase (SOD) activity in human subject saliva. RESULTS: Antioxidant activity towards O(2)(·-) was increased by Porphyromonas gingivalis (P. gingivalis) infection in rat, although antioxidant activity towards HO(·) was not changed. In human, a strong correlation (r = 0.88, p < 0.01) recognized between salivary antioxidant activity towards O(2)(·-) and probing pocket depth (PPD). In addition, the intensity of salivary antioxidant activity depended on SOD activity level. SOD activity was also correlated with PPD. CONCLUSIONS: Rat salivary antioxidant activity towards O(2)(·-) was up-regulated by the inflammatory response caused by P. gingivalis infection. Similar response was recognized in human saliva with periodontal index. Additionally, a linear correlation between antioxidant activity towards O(2)(·-) and SOD activity was verified by ESR technique. Therefore, evaluation of the salivary antioxidant activity towards O(2)(·-) might be an effective parameter for the objective assessment of periodontal disease progression.

Direct assessment of the antioxidant properties of midazolam by electron spin resonance spectroscopy.

Some antioxidant anesthetics directly inhibit lipid peroxidation mediated via the generation of reactive oxygen species (ROS). To date, the scavenging effects of midazolam on ROS have not been directly assessed. We investigated the inhibitory effect of midazolam on ROS [hydroxyl radical (HO(·)) and superoxide (O (2) (·-) )] by in vitro X-band electron spin resonance with the spin-trapping agent 5,5-dimethyl-1-pyrroline-N-oxide. Our results indicated that HO(·) and O (2) (·-) were not affected by midazolam at clinically relevant concentrations, but were directly scavenged by midazolam at high concentrations (i.e., >4.6 and >1.5 mM, respectively).