Biocompatibility of a titanium dioxide-coating method for denture base acrylic resin.

OBJECTIVES: Ease of denture cleaning is of paramount importance in geriatric patients and those with limited dexterity. We have previously investigated methods of coating dentures with titanium dioxide (TiO2 ) and reported the effects (self-cleaning and antibacterial) of such treatments in in vitro studies. This study was to verify the biocompatibility of a TiO2 -coated acrylic resin produced by the new coating method with spray-coating technique. METHODS: Specimens were prepared from denture base acrylic resin and polished up to grit #1000. The TiO2 -coating agent was sprayed onto the specimens using an airbrush gun. Specimens were then divided into 'polymethyl methacrylate (PMMA)', 'primer-coated PMMA' and 'TiO2 -coated PMMA' groups to be evaluated for biological safety using a hamster oral mucosa irritation test, a guinea pig skin sensitisation test and a rabbit intracutaneous test. The biological reaction was scored. RESULTS: Reaction scores were considerably <1.0, the acceptable limit set by the ISO, in all three tests. Indeed, in most samples, there was no deleterious effect at all. CONCLUSION: These results tested on animals demonstrate that denture base resin coated with TiO2 by this method does not cause irritation or sensitisation of the oral mucosa, skin or intracutaneous tissue and is therefore good biocompatibility for use in close proximity to oral mucosa and skin.

Molecular mechanisms of the antileukemia activities of retinoid and arsenic.

Acute promyelocytic leukemia (APL) is characterized by the occurrence of translocations between chromosomes 15 and 17, resulting in generation of a fusion protein of promyelocytic leukemia (PML) and retinoid A receptor (RAR) α. APL cells are unable to differentiate into mature granulocytes since PML-RARα functions as a strong transcriptional repressor for a gene involved in granulocyte differentiation. All-trans retinoic acid (ATRA) is the first agent that has been developed to target specific disease-causing molecules, i.e., ATRA suppresses abnormal functions of oncogenic proteins. Moreover, ATRA facilitates the differentiation of APL cells toward mature granulocytes by changing epigenetic modifiers from corepressor complexes to co-activator complexes on target genes after binding to the ligand-binding domain at the RARα moiety of the PML-RARα oncoprotein. On the other hand, arsenic trioxide (ATO), another promising agent used to treat APL, directly binds to the PML moiety of the PML-RARα protein, causing oxidation and multimerization. ATO enhances the conjugation of small ubiquitin-like modifiers to PML-RARα, followed by ubiquitination and degradation, relieving the genes associated with granulocytic differentiation from suppressive restraint by the oncoprotein. Recent clinical studies have demonstrated that combination therapy with both ATRA and ATO is useful to achieve remission.

Proteomic analysis of lipopolysaccharide-treated submandibular gland in rat.

We investigated changes in the protein profile of submandibular gland (SMG) with inflammation induced by exposure to lipopolysaccharide (LPS) with the aim of identifying potential molecular markers of injured gland. Lipopolysaccharide (2.5µg) was directly administered into rat SMG unilaterally by retrograde ductal injection. At 12hr after treatment, the gland was excised and the proteins identified by two-dimensional difference gel electrophoresis and matrix-assisted laser desorption/ionization-time of flight mass spectrometry. Many proteins in the LPS-treated gland showed a marked change compared to those in the contralateral gland. Of particular note were increases in ubiquitin, a highly-conserved small regulatory protein and in calgranulin B, which has an immunological function in inflammation. Proteins related to apoptosis and stress also showed change in the inflamed gland. The results of this study suggest that the ubiquitin system of protein modification is involved in LPS-induced inflammation in salivary gland, and that a number of specific proteins might be applicable as molecular markers in the monitoring of inflamed or injured gland.

Modulation of benzodiazepine receptor, adrenoceptor and muscarinic receptor by diazepam in rat parotid gland.

This study investigated the influence of diazepam on the binding characteristics of adrenoceptor, muscarinic and benzodiazepine receptors in rat parotid gland membrane using a radioligand binding assay. At a concentration of >10(-6)M, diazepam competed with [(3)H]dihydroalprenolol for ß-adrenoceptor, but not [(3)H]prazosin for α-adrenoceptor or [(3)H]quinuclidinyl benzilate for muscarinic receptor. Continuous administration of diazepam at doses of 0.4mg/kg/day, i.p. for 7days in rat significantly decreased pilocarpine (4.0mg/kg, i.p.)-induced parotid salivary flow. Diazepam also produced a significant increase in the dissociation constant (Kd) value for [(3)H]dihydroalprenolol binding, but no change in the maximal binding capacity (Bmax) value, and a decrease in the Kd value for [(3)H]diazepam binding to benzodiazepine receptors, but no change in the Kd or Bmax values for [(3)H]prazosin or [(3)H]quinuclidinyl benzilate binding. These results suggest that continuous administration of diazepam modifies affinity for ß-adrenoceptor and benzodiazepine receptor binding sites in parotid gland membrane and that changes in these binding sites may be closely related to diazepam-induced suppression of salivary secretion.

Diazepam enhances production of diazepam-binding inhibitor (DBI), a negative saliva secretion regulator, localized in rat salivary gland.

Peripheral-type benzodiazepine receptor (PBR) and central-type benzodiazepine receptor (CBR) in salivary gland play a role in the inhibitory regulation of salivary secretion in rodents. Diazepam-binding inhibitor (DBI), an endogenous ligand for PBR, produces neurosteroids, which modulate CBR activity. In this study, we investigated the effect of repetitive administration of diazepam (DZP) on salivary secretion and expression of DBI mRNA and peptide. Moreover, mRNA expression of PBR and pituitary adenylate cyclase-activating polypeptide (PACAP), a transcriptional regulator for DBI promoter, was evaluated after repetitive administration of DZP. Repetitive administration, but not single administration, of 0.4 mg/kg DZP caused inhibition of salivary secretion and enhanced expression of DBI, PACAP, and PBR mRNA in rat salivary gland, with an increase in production of DBI peptide. These results suggest that repetitive administration of DZP stimulates DBI production, which may result in an increase in the suppressive effect of DZP on salivary secretion.

Diazepam Enhances Production of Diazepam-Binding Inhibitor (DBI), a Negative Saliva Secretion Regulator, Localized in Rat Salivary Gland.

Peripheral-type benzodiazepine receptor (PBR) and central-type benzodiazepine receptor (CBR) in salivary gland play a role in the inhibitory regulation of salivary secretion in rodents. Diazepam-binding inhibitor (DBI), an endogenous ligand for PBR, produces neurosteroids, which modulate CBR activity. In this study, we investigated the effect of repetitive administration of diazepam (DZP) on salivary secretion and expression of DBI mRNA and peptide. Moreover, mRNA expression of PBR and pituitary adenylate cyclase-activating polypeptide (PACAP), a transcriptional regulator for DBI promoter, was evaluated after repetitive administration of DZP. Repetitive administration, but not single administration, of 0.4 mg/kg DZP caused inhibition of salivary secretion and enhanced expression of DBI, PACAP, and PBR mRNA in rat salivary gland, with an increase in production of DBI peptide. These results suggest that repetitive administration of DZP stimulates DBI production, which may result in an increase in the suppressive effect of DZP on salivary secretion.

Paradoxical effect of 2,3-dimercapto-1-propanesulfonic acid (DMPS) on enhancing antitumor activity of cisplatin in ascites sarcoma 180 cells.

We investigated the enhancing effect of two metal-chelating compounds, 2,3-dimercapto-1-propanesulfonic acid (DMPS) and meso-2,3-dimercaptosuccinic acid (DMSA), on the antitumor activity of cisplatin (CDDP). In the in vivo experiments, DMPS showed a clear synergistic effect and significantly enhanced the antitumor activity of CDDP in terms of survival and life span in mice transplanted with ascites sarcoma 180 cells (S180 cells) at a dose of <100 micromol/kg, s.c., but not at a dose of >500 micromol/kg. On the other hand, DMSA did not enhance the antitumor activity of CDDP. DMPS (50 micromol/kg, s.c.) combined with CDDP also potently suppressed [(3)H]thymidine uptake in S180 cells implanted in mice, whereas DMSA did not. In the in vitro experiments, DMPS (10(-6) to 10(-5) M) produced a time- and dose-dependent decrease in intracellular Ca(2+) concentrations ([Ca(2+)](i)) in S180 cells and, in combination with CDDP, yielded a significant increase in intracellular platinum accumulation compared to that in cells treated with CDDP alone. These results indicate that DMPS used in combination with CDDP may be of considerable benefit in enhancing the cytotoxicity of CDDP in tumor cells, especially at a low dose. The results also suggest that the enhancing effect of DMPS is closely related to a decrease in [Ca(2+)](i) and that the suitable dose and adequate administrational time of DMPS are important for its effective action.

Immunohistochemical study on GABAergic system in salivary glands.

Gamma-aminobutyric acid (GABA) and its receptors are found in the central nervous system and several peripheral tissues. The purpose of this study was to determine the expression and distribution of GABA and glutamate decarboxylase (GAD), a GABA biosynthetic enzyme, in rat salivary gland. Western blot and real time quantitative RT-PCR revealed that GAD67 was the major isoform of GAD in the salivary glands. Furthermore, both GABA and GAD were detected around the acinar cells in the submandibular glands by immunohistochemical analysis. When both sympathetic and parasympathetic nerves related to the submandibular glands were denervated, the immunoreactivities of GABA and GAD were dramatically depressed, and levels of GAD67 and GABA significantly decreased. However, no morphological changes in the glands were observed after denervation. These results indicate that GAD67 is present around acinar cells in the salivary glands, and suggest that the GABAergic system in the glands is closely related to the autonomic nervous system.

Evaluation of high-performance liquid chromatography and mass spectrometry method for pharmacokinetic study of local anesthetic ropivacaine in plasma.

We studied the viability of high-performance liquid chromatography and mass spectrometry (LC/MS) as a selective and sensitive analytical method for measuring blood concentrations of the local anesthetic ropivacaine. Ropivacaine was effectively separated using a reverse-phase column and monitored at 275 m/z ion. The LC/MS method allowed measurement of concentrations of ropivacaine of lower than 75 ng/mL. The standard curve was linear and in the range of <1.5 microg/mL. Recovery of ropivacaine in plasma samples was over 90% after precipitation of plasma protein with trichloroacetic acid. The method was tested on the pharmacokinetics of plasma ropivacaine after single intravenous or subcutaneous administration in rabbits. The pharmacokinetic parameters showed a one-compartment model and a mean elimination half-life of 0.54+/-0.05 h and 2.83+/-0.51 h after administration at doses of 0.4 mg/kg, i.v. and 5 mg/kg, s.c., respectively. These values were in approximate agreement with previously obtained results in dogs. The results of the present study demonstrated that the LC/MS method was highly selective and sensitive for the measurement of ropivacaine, indicating that it offers a useful tool for monitoring the therapeutic effects and determining the pharmacokinetic parameters of this drug in blood.

Expression of BMP7 is associated with resistance to diabetic stress: comparison among mouse salivary glands.

We determined mRNA levels of bone morphogenetic protein 7 (BMP7), a growth and differentiation factor belonging to the transforming growth factor-beta superfamily, in the salivary glands of mice with streptozotocin (200 mg/kg, i.p.)-induced diabetes. We also examined the effects of BMP7 on secretion of saliva and degenerative change in salivary glands in diabetic mice. In normal mice, BMP7 mRNA levels were high in the submandibular gland and low in the parotid gland, while in diabetic mice, levels were significantly decreased in the parotid gland, but not in the submandibular gland. No significant difference was observed in mRNA levels of BMP receptors between normal and diabetic mice. In diabetic mice, pilocarpine (4 mg/kg, i.p.)-stimulated salivary secretion showed a remarkable decrease in both parotid and submandibular gland, although degree of reduction was smaller in the latter. Notable degeneration with vacuolation and atrophy was also found in parotid gland, whereas degeneration of submandibular gland was slight. Administration of BMP7 (50 and 100 microg/kg, i.v.) in diabetic mice induced a significant increase in salivary secretion, with rate of recovery higher in parotid gland than in submandibular gland. In diabetic mice, BMP7 also exhibited a powerful protective effect in degenerated salivary gland, especially in parotid gland. These results suggest that BMP7 acts to prevent diabetic damage in salivary gland, and that its cytoprotective effect is closely correlated with mRNA levels in tissue.

Hydroxyl radical scavenging effects of guaiacol used in traditional dental pulp sedation: reaction kinetic study.

Guaiacol, which is a phenolic compound with a methoxy group and used in traditional dental pulp sedation, has the property of inducing cell proliferation. To clarify these mechanisms of guaiacol, this study examined the hydroxyl radical (*OH) scavenging effects of guaiacol in vitro. Generation of *OH was carried out by the Fenton reaction using mixture of ascorbic acid, H2O2, and Fe(III)-EDTA, and *OH was detected by measuring the *OH-mediated production of degradation products of deoxyribose, which reacts with 2-thiobarbituric acid (TBA) and is relatively stable for a long time. At concentrations of 10(-10) M to 10(-3) M, guaiacol inhibited the TBA reactive substance (TBA-RS) formation in a dose-dependent manner. Phenol and formaldehyde were also found to inhibit the TBA-RS formation, but their inhibitory activities were lower than that of guaiacol. The concentrations of guaiacol, phenol, and formaldehyde needed to cause 50% inhibition of TBA-RS formation were approximately 5 x 10(-6), 5 x 10(-5), and 2 x 10(-3) M, respectively. In this reaction system, guaiacol showed no chelating reaction with ferrous ion and did not directly react with H2O2. Guaiacol also exhibited radical scavenging activity on 1,1-diphenyl-2-picrylhydrazyl (DPPH) stable free radical, but its scavenging activity was lower than that toward *OH. These results suggest that guaiacol is a potent scavenger of reactive oxygen radicals and that its radical scavenging activity may be associated with its effect on cell proliferation.

Inhibitory effect of diazepam on muscarinic receptor-stimulated inositol 1,4,5-trisphosphate production in rat parotid acinar cells.

1. This study examined the effect of diazepam (DZP) on phosphoinositide turnover, which plays an important role in the regulation of salivary secretion, in rat parotid acinar cells. 2. DZP (10(-9) M to 10(-5) M), a potent agonist of both central- and peripheral-type benzodiazepine receptors, dose-dependently decreased inositol 1,4,5-trisphosphate IP3 production stimulated by carbachol, a muscarinic receptor agonist, in the cells. 3. DZP produced a maximum inhibitory response at a concentration of 10(-5) M, with IP3 production decreased to 63% of maximal levels. The concentration inducing half maximal inhibition of IP3 production was approximately 3.5 x 10 (-8) M. 4. An inhibitory response to DZP was produced by a short-term pretreatment (<3 min) of the cells and prevented by antagonist and competing ligand for the central- and peripheral-type benzodiazepine receptors, flumazenil and PK 11195, respectively. 5. DZP showed a non-competitive inhibition of carbachol-stimulated IP3 production. It did not directly inhibit the activities of GTP-binding regulatory proteins and phosphatidylinositol 4,5-bisphosphate-specific phospholipase C (PLC) in the parotid gland membranes, though choline chloride inhibited PLC activity. 6. DZP (10(-5) M) attenuated the increase in the intracellular Ca2+ concentration ([Ca(2+)](i)) in the cells following stimulation of the muscarinic and alpha(1)-adrenoceptors. 7. These results suggest that in the parotid acinar cells, DZP inhibits muscarinic receptor-stimulated IP3 production through benzodiazepine receptors and that PLC activity which produces IP3 is inhibited by chloride. The decreases in IP3 and [Ca(2+)](i) in the cells may be connected with the suppression of salivary secretion induced by DZP.

Comparison of temporal changes in components of formalin guaiacol under several storage conditions.

This study examined the effects of storage conditions such as time course, temperature, fluorescent light, and darkness on the components and antibacterial activity of formalin guaiacol (FG) used in endodontic treatment. We measured the quantities of formaldehyde and guaiacol in FG and antibacterial activities against Staphylococcus aureus, Porphyromonas gingivalis, and Porphyromonas endodontalis. The components and antibacterial activity of FG in the brown or transparent tightly sealed containers were not affected by temperature or fluorescent light throughout the 4 week test. However, in the loosely sealed containers, formaldehyde and guaiacol in FG sample decreased remarkably within one week, not only in a temperature- and time-dependent manner, but also under fluorescent light at 20 degrees C. Furthermore, the antibacterial activities in the FG sample were significantly attenuated in parallel with the decrease in formaldehyde levels. Fluorescent light caused color changes and crystallization of FG samples in the transparent containers. These results suggest that it is important to replace fresh FG every 5 to 7 days for endodontic treatment and that, in the dental office, it is advisable to store fresh FG in tightly sealed containers every 2 weeks to maintain its efficacy.