Yoshimi Shigetani

A study of cavity preparation by Er:YAG laser. Effects on the marginal leakage of composite resin restoration.

The purpose of this study was to evaluate marginal leakage of composite resin restoration from cavities prepared by Er:YAG laser. The observation of the dentin surface after the application of laser irradiation was performed by LSM, the cutting surface showed a rough surface similar to scales, and exposed dentinal tubules were observed without striations or a smeared layer formation that were observed when using a rotary cutting device. Leakage tests revealed no significant differences in the marginal seal for both enamel and dentin between cavities prepared by Er:YAG laser irradiation and when using an air-turbine. In this study, the usefulness of cavity preparation by Er:YAG laser irradiation in composite resin restoration was suggested.

All-in-one self-etch model adhesives : HEMA-free and without phase separation

OBJECTIVE Aim of this study was to design HEMA-free all-in-one self-etch model adhesives without phase separation, and to investigate their efficiency on extracted human teeth. MATERIALS AND METHODS Compositions of adhesives in mass% (1): UDMA (25), 4-META (20), H(2)O (0, 1, 2, 5, 10, 20, 35, and 45), balance of acetone or ethanol. (2): UDMA (35), 4-META or 4-MET (28), H(2)O (0, 2, 4, 5, 6, and 8), balance of acetone. Phase separation was evaluated on samples exposed to ambient atmosphere. Conventional shear bond strengths (SBS, n=8) were determined on human enamel and dentin. Marginal adaptation (MGW, n=8) was assessed in cylindrical butt-joint dentin cavities. RESULTS Solutions (1) and (2) with 5 and 8% or less water content, respectively, showed no phase separation. SBSs on enamel were not different within the acetone- or ethanol-group and between the adhesive groups (1). Water content of adhesives (2) was a significant determinant of enamel SBSs, groups with 4-META or 4-MET were not different (p>0.05). Dentin SBSs with adhesives (1) were not different (p>0.05) within solvent groups, yet higher for acetone-dissolved adhesives (p<0.05). Dentin SBSs with adhesives (2) were different by water content and functional monomer (p<0.05). MGW for solutions (1) were smaller with acetone-dissolved than with ethanol-dissolved adhesives (p<0.001). Acetone solutions between 2 and 45% water content produced almost perfect marginal adaptation. Marginal adaptation of adhesives (2) was almost perfect at 5 through 8% water content. CONCLUSIONS Simplified HEMA-free self-etch adhesives without phase separation were prepared without compromises on bonding efficiency to enamel and dentin.

GaAlAs laser irradiation induces active tertiary dentin formation after pulpal apoptosis and cell proliferation in rat molars.

INTRODUCTION This study aimed to clarify pulpal responses to gallium-aluminum-arsenide (GaAlAs) laser irradiation. METHODS Maxillary first molars of 8-week-old rats were irradiated at an output power of 0.5 or 1.5 W for 180 seconds, and the samples were collected at intervals of 0 to 14 days. The demineralized paraffin sections were processed for immunohistochemistry for heat-shock protein (HSP)-25 and nestin in addition to cell proliferation assay using bromodeoxyuridine (BrdU) labeling and apoptosis assay using deoxynucleotidyl transferase deoxyuridine triphosphate nick end labeling (TUNEL). RESULTS Intense HSP-25 and nestin immunoreactivities in the odontoblast layer were weakened immediately after 0.5-W irradiation and recovered on day 1, resulting in slight tertiary dentin formation by day 14. On the contrary, 1.5-W irradiation immediately induced the loss of HSP-25 and nestin-immunoreactivities in the odontoblast layer. On day 1, numerous TUNEL-positive cells appeared in a degenerative zone that was surrounded by intense HSP-25 immunoreactivity. BrdU-positive cells occurred within the intensely HSP-25-immunopositive areas during days 2 through 5, whereas TUNEL-positive cells gradually decreased in number by day 5. HSP-25- and nestin-positive odontoblast-like cells were arranged along the pulp-dentin border by day 7, resulting in remarkable tertiary dentin formation on day 14. CONCLUSIONS The output energy determined pulpal healing patterns after GaAlAs laser irradiation; the higher energy induced the apoptosis in the affected dental pulp including odontoblasts followed by active cell proliferation in the intense HSP-25-immunoreactive areas surrounding the degenerative tissue, resulting in abundant tertiary dentin formation. Thus, the optimal GaAlAs laser irradiation elicited intentional tertiary dentin formation in the dental pulp.

Immunohistochemical analysis of subcutaneous tissue reactions to methacrylate resin-based root canal sealers

AIM To investigate subcutaneous tissue reactions to methacrylate resin-based root canal sealers by immunohistochemical assessment of inflammatory/immunocompetent cell infiltration. METHODOLOGY Silicone tubes containing freshly mixed Epiphany SE sealer, MetaSEAL, Super-Bond RC sealer, or a zinc oxide-eugenol sealer (Canals) were subcutaneously implanted into the backs of Wistar rats. Solid silicone rods implanted in different animals served as controls. After 7, 14 and 28 days, connective tissue surrounding the implants (n = 8, each) was processed for immunoperoxidase staining using OX6 (reactive to major histocompatibility complex class II molecules), ED1 (reactive to macrophages), and W3/13 (reactive primarily to neutrophils), and the number of positively stained cells within each field (1.2 × 0.8 mm) was enumerated. Statistical differences were analysed with Friedmans test and Scheffes test (comparisons between test materials) or Mann-Whitneys U-test (test-control comparisons). RESULTS Canals showed a significantly higher number of W3/13-positive cells (mostly neutrophils) than MetaSEAL at 28 days (P < 0.05). There were no significant differences in the numbers of OX6- or ED1-positive cells between each test material at any time point. Test-control comparisons revealed several significant differences for each antibody. This was most notable for ED1, where all the test materials at each time point, except for Epiphany SE at 28 days, showed significantly larger values than the corresponding controls. CONCLUSIONS All the methacrylate resin-based sealers tested showed a similar level of inflammatory/immunocompetent cell infiltration. MetaSEAL induced less-intense neutrophil infiltration than Canals. Controls exhibited milder infiltration of inflammatory/immunocompetent cells compared with all the test materials.

Impact of Streptococcus mutans on the generation of fluorescence from artificially induced enamel and dentin carious lesions in vitro

The purpose of this study was to examine whether Streptococcus mutans is implicated in the generation of fluorescence detected in carious lesions. Enamel surfaces and dentin cavities of extracted human teeth were subjected to artificial caries generation by exposing them either to a culture medium containing S. mutans or to a lactic acid buffer for 2 weeks. Fluorescence from the lesions was detected with confocal laser scanning microscopy or fluorescence microscopy at various excitation wavelengths, and maximum fluorescence radiance was computed using imageanalyzing software. Culture media of S. mutans were also examined for fluorescence generation. The results demonstrated that S. mutans-induced enamel and dentin lesions exhibited increased fluorescence in the red and green spectral regions, with the signal stronger in the red region. In the blue region, however, fluorescence signals in the corresponding area were below the background level. Significantly weaker or virtually no fluorescence was detected in lactic acid-demineralized lesions at all excitation wavelengths. Neither bacterial cells nor culture media generated any fluorescence. These results indicate that, although the presence of S. mutans may be a prerequisite for the emission of fluorescence from carious lesions, some interaction of S. mutans with exposed tooth matrix elements may also be required for the generation or unmasking of fluorophores.

M2 macrophages participate in the biological tissue healing reaction to mineral trioxide aggregate.

INTRODUCTION This study examined the protein and messenger RNA (mRNA) expression of molecules associated with M2 (wound healing) macrophages in mineral trioxide aggregate (MTA)-implanted rat subcutaneous tissue to elucidate the involvement of M2 macrophages in the connective tissue response to MTA. METHODS Silicone tubes containing freshly mixed MTA or a calcium hydroxide cement (Life; Kerr, Romulus, MI) were subcutaneously implanted into the backs of Wistar rats. Solid silicone rods implanted in different animals served as controls. The specimens were then double immunostained for ED1 (CD68, a general macrophage marker) and ED2 (CD163, an M2 macrophage marker). Immunostaining for CD34 (a marker for vascularization and wound healing) was also performed. Expression levels of CD34, CD163, and mannose receptor c type 1 (an M2 macrophage marker) mRNAs were determined with real-time polymerase chain reaction. RESULTS MTA-implanted subcutaneous tissues showed significant increases in the density of ED1+ED2+ macrophages beneath the implantation site and expression levels of CD163 and MMR mRNAs compared with Life-implanted and control tissues. MTA-implanted subcutaneous tissues also showed a significant increase of CD34-immunostained areas and up-regulation of CD34 mRNAs compared with Life-implanted and control tissues. CONCLUSIONS MTA implantation induced the accumulation of M2 macrophage marker (ED2)-expressing macrophages and enhanced the expression of M2 macrophage marker genes. MTA implantation also enhanced the expression of CD34, suggesting acceleration of the healing/tissue repair process. Taken together, biological connective tissue response to MTA may involve wound healing/tissue repair processes involving M2 macrophages.

Gene Expression Analysis of Membrane Transport Proteins in Normal and Lipopolysaccharide-inflamed Rat Dental Pulp

INTRODUCTION Membrane transport proteins (transporters) play a crucial role in the transmembrane uptake and/or efflux of various compounds such as inorganic ions, endogenous bioactive substances such as prostaglandins (PGs), and drugs such as nonsteroidal anti-inflammatory drugs. This study aimed to analyze mRNA expression of selected transporters related to drug disposition and PG transport in normal and lipopolysaccharide (LPS)-inflamed rat incisor pulp. METHODS Pulp tissues were subjected to reverse transcription-polymerase chain reaction (PCR) detection for transporter isoforms belonging to organic anion transporting polypeptide (Oatp), organic anion transporter (Oat), organic cation transporter (Oct), multidrug resistance-associated protein (Mrp), and multidrug resistance protein (Mdr) families. The levels of mRNA expression for PG transporters (Oatp1a5, Oatp1b2, Oatp2a1, Oatp2b1, and Oatp3a1) were compared in normal and LPS-inflamed pulps by using real-time PCR. RESULTS The pulp tissue expressed mRNAs for various transporters belonging to the Oatp, Oat, Oct, Mrp, and Mdr families. LPS inflammation caused significant up-regulation of Oatp2a1 (P < .01) and significant down-regulation of Oatp1a5, Oatp2b1 (P < .01), and Oatp3a1 (P < .05). CONCLUSIONS Rat incisor dental pulp expressed mRNAs for various transporter isoforms. The levels of mRNA expression for PG transporters were significantly up-regulated or down-regulated in LPS-inflamed dental pulp.

Prostaglandin Transporting Protein-mediated Prostaglandin E2 Transport in Lipopolysaccharide-inflamed Rat Dental Pulp

INTRODUCTION The prostaglandin transporter (Pgt) and multidrug resistance-associated protein (Mrp) 4 are membrane transport proteins that play crucial roles in the transmembrane uptake and/or efflux of prostaglandins (PGs). This study attempted to analyze the protein expression of Pgt and Mrp4 and their involvement in PGE2 efflux transport in lipopolysaccharide (LPS)-inflamed rat incisor pulp tissue. METHODS Pulpitis was induced in the upper incisors of Wistar rats by treating them with LPS for 24 hours. The protein expression levels of Pgt, Mrp4, and microsomal PGE synthase (mPGES) were analyzed with immunofluorescent staining. The amount of PGE2 released from the inflamed pulp tissue in the presence or absence of dipyridamole (an Mrp4 inhibitor) was assessed by using an enzyme-linked immunosorbent assay. RESULTS Double immunofluorescence staining revealed that the Pgt, Mrp4, and mPGES immunoreactivity co-localized in CD31-expressing endothelial cells. Moreover, the Mrp4 inhibitor caused a significant decrease in the amount of PGE2 released from the LPS-inflamed pulp (P < .01 at 24 hours). CONCLUSION Pgt, Mrp4, and mPGES expression was detected in the endothelial cells of normal and LPS-inflamed rat incisor pulp tissue, suggesting that these cells are associated with the biosynthesis and transmembrane transport of PGE2. The significant decrease in PGE2 release induced by the Mrp4 inhibitor suggests that Mrp4 contributes to the transport of PGE2 in the transmembrane efflux pathway.

Initial Transient Accumulation of M2 Macrophage–associated Molecule-expressing Cells after Pulpotomy with Mineral Trioxide Aggregate in Rat Molars

INTRODUCTION M2 (alternatively activated) macrophages are known to participate in wound healing and tissue repair. This study aimed to analyze the temporospatial changes in the distribution and density of M2 macrophage-associated molecule-expressing cells after pulpotomy with mineral trioxide aggregate (MTA) in rat molars to ascertain the role played by M2 macrophages in the healing of MTA-capped pulp tissue. METHODS The maxillary first molars of 8-week-old Wistar rats were pulpotomized and capped with MTA. After 1-14 days, the teeth were examined after hematoxylin-eosin staining or immunoperoxidase staining of CD68 (a general macrophage marker) and M2 macrophage markers (CD163 and CD204). The density of positively stained cells was enumerated in the surface and inner regions (0-100 μm and 300-400 μm, respectively, from the wound surface). RESULTS MTA capping initially caused mild inflammatory changes and the formation of a degenerative layer followed by progressive new matrix formation and calcified bridging. At 1-2 days, CD68-, CD163-, and CD204-positive cells started to accumulate beneath the degenerative layer, and the density of these cells was significantly higher in the surface region than in the inner region (P < .05). From 7 days onward, the 3 types of cells displayed an almost normal distribution beneath the newly formed dentinlike matrix. CONCLUSIONS After the pulpotomy of rat molars with MTA, M2 macrophage-associated molecule-expressing cells transiently accumulated beneath the degenerative layer under the MTA. This suggests that M2 macrophages participate in the initial phases of the healing of MTA-capped pulp tissue.