K Debari

Showa University, Shinagawa, Tōkyō, Japan

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Publications (35)54.56 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: We report the effects of specific and potent inhibitors of vacular-type H(+)-ATPase and lysosomal cysteine proteinases, cathepsins, on the ultrastructure, expression of these enzymes, and resorptive functions of cultured osteoclasts. Osteoclasts were formed by co-culture of marrow cells and calvarial primary osteoblasts of ddY mice. Formed osteoclasts were cultured on dentine slices for 6-48 hr with either an H(+)-ATPase inhibitor, bafilomycin A1, or a cysteine proteinase inhibitor, E-64. In control cultures with no additive, osteoclasts were structurally characterized by the development of ruffled borders and clear zones, and formed many resorption lacunae on dentine slices. Both H(+)-ATPase and cathepsin K were strongly expressed in the ruffled borders of these osteoclasts. In bafilomycin A1-treated cultures, osteoclasts lacked ruffled borders, and resorption lacuna formation was markedly diminished. This effect of bafilomycin A1 on osteoclast structure was reversible by removal of the compound. Bafilomycin A1 treatment altered the subcellular localization and decreased the expression of H(+)-ATPase molecules. H(+)-ATPase expression was observed throughout the cytoplasm, but not along the plasma membranes facing dentine slices. On the other hand, E-64 treatment did not affect the ultrastructure of osteoclasts and the expression of enzyme molecules. Although E-64 showed no effect on demineralization of dentine slices, it dose-dependently reduced resorption lacuna formation. Our results suggest that 1) bafilomycin A1 dose-dependently inhibits resorption lacuna formation via inhibition of ruffled border formation, 2) H(+)-ATPase expression is closely associated with the cytoskeleton of osteoclasts, and 3) E-64 treatment decreases the depth of resorption lacunae, by inhibition of secreted cathepsin K activity, but does not impair ruffled border formation and the associated expression of H(+)-ATPase and cathepsin K in osteoclasts.
    The Anatomical Record Part A Discoveries in Molecular Cellular and Evolutionary Biology 03/2003; 270(2):152-61.
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    ABSTRACT: The effects of enamel matrix derivative (EMD; Emdogain) on new trabecular bone induction after pure bioinert titanium (Ti) implantation in the rat femur were examined by means of routine light and transmission electron microscopy, immunohistochemistry, and backscattered electron image analysis. Newly designed mini-Ti implants (3.5 mm in length and 1.6 mm in diameter) were placed in the corticotrabecular area of the femur with either EMD or its carrier, propylene glycol alginate, as control. On post-implantation days 4, 7, 14, and 30, the dissected femur was examined in the transverse direction through Ti implants. In both control and EMD-applied femurs, trabecular bone formation was recognized over the implant surfaces and within medullary cavities even at 4 days post-implantation. These newly formed bone trabeculae around the Ti implants were immunoreactive for bone sialoproteins as a bone matrix marker, and osteoclastic bone resorption became evident in these bone trabeculae after 7 days post-implantation. Although trabecular bone area around the implants was markedly decreased at 30 days post-implantation compared with those at 14 days, the trabecular bone areas in EMD-applied femurs were significantly greater than those in propylene glycol alginate-applied femurs at both 14 and 30 days post-implantation. Our results suggest that EMD is an effective biological matrix for enhancing new trabecular bone induction and resulting attachment of orthopedic prostheses to the recipient bone.
    Journal of Biomedical Materials Research 06/2002; 60(2):269-76.
  • Tetsuo Kodaka, Kazuhiro Debari
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    ABSTRACT: We examined afibrillar cementum (AFC) and cementicle-like structures (CLS) in human teeth by scanning electron microscopy and energy-dispersive X-ray microanalysis. The AFC showed a spur- or island-, plate- and mass-like structure with appositional laminations, while large masses in the enamel fissures enclosed CLS showing concentric appositional rings. Such AFC was observed in enamel fissures, an abnormal enamel pit, dens invaginatus and root furcations with enamel droplets, as well as on the cervical enamel surfaces, where ameloblasts are differentiated at the later or last stage of enamel formation. Cementicle-like structures were occasionally found independent from AFC and some CLS contained epithelial cell-like or ameloblast-like remnants in the core, surrounded by a few or many concentric rings. In addition, cementicles (CEC) in the root furcations also contained the remnants of Malassez's epithelial-rest cells surrounded by a few concentric rings. In some areas, AFC was mixed with enamel structures. These results show that the organic material in some parts of AFC and CLS may be derived from epithelial cells similar to that of CEC. Calcification values of AFC and CLS were significantly higher than that of fibrillar cementum, and the minute crystals are probably apatite.
    Journal of Electron Microscopy 02/2002; 51(5):327-35. · 1.44 Impact Factor
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    ABSTRACT: To elucidate the effects of enamel matrix derivative (EMD: Emdogain) on bone regeneration in rat femurs after drill-hole injury, defects in bone were filled with either EMD or its carrier, PGA, as control. On postoperative days 4 to 28, dissected femurs were examined by means of various morphological approaches. In both experimental groups, formation of trabecular bone, which was immunostained for bone sialoproteins (BSP), had occurred in the medullary cavities at cylindrical bone defects on Day 7 postoperatively. Cuboidal osteoblasts were clearly observed on these newly-formed BSP-positive bone trabeculae. On Days 7 and 14, many multinucleated giant cells, which strongly expressed cathepsin K, had appeared on these bone trabeculae, indicating active bone remodeling. In these bone trabeculae, Ca and P weight % and Ca/P ratio were similar to those of cortical bone, and there was no significant difference between the PGA- and EMD-applied groups. Bone volume fraction of newly-formed bone trabeculae on Day 7 postoperatively was significantly higher in the EMD-applied group than in the PGA-applied controls. Because of active bone remodeling and the marked decrease of bone volume, on Days 14 and 28 postoperatively, however, there was no longer a significant difference in trabecular bone volume fraction between the experimental groups. Our results suggest that EMD possesses an osteo-promotive effect on bone and medullary regeneration during wound healing of injured long bones.
    The Anatomical Record 01/2002; 264(4):438-46.
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    ABSTRACT: Acute effects of ovariectomy on the bone wound healing processes after maxillary molar extraction in aged rats were examined by means of quantitative scanning electron microscopy (SEM), backscattered electron image (BSE) analysis and energy-dispersive X-ray (EDX) microanalysis. Six-month-old female rats underwent either sham operation or bilateral ovariectomy, and 7 days postoperatively, the maxillary first molars were extracted. On post-extraction days 7, 30 and 60, the dissected maxillary bone surfaces were examined by SEM to reveal the bone formative and resorptive areas around the extracted alveolar sockets. In addition, the resin-embedded maxillae were micromilled in the transverse direction through the extracted alveolar sockets, and the newly-formed bone mass on the buccal bone surfaces and within the extracted sockets was examined by BSE analysis. Compared with sham-operated controls, the extent of newly-formed bone mass on the buccal bone surfaces in OVX rats was significantly decreased, due to increased bone resorption. On the other hand, new bone formation within the extracted sockets was similar in the experimental groups. In EDX microanalysis of these newly-formed bone matrices, both Ca and P weight % and Ca/P molar ratio were similar in the experimental groups. Our results suggest that 1) acute estrogen deficiency induced by ovariectomy stimulates sustained bone resorption, but has less effect on bone formation, and 2) bone wound healing after maxillary molar extraction within extracted alveolar sockets is not significantly delayed by ovariectomy, but bony support by newly-formed bone mass on the maxillary bone surfaces at the buccal side of the extracted sockets is significantly decreased, due to increased bone resorption.
    The Anatomical Record 03/2001; 262(2):203-12.
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    ABSTRACT: Acute effects of ovariectomy on the bone wound healing processes after maxillary molar extraction in aged rats were examined by means of quantitative scanning electron microscopy (SEM), backscattered electron image (BSE) analysis and energy-dispersive X-ray (EDX) microanalysis. Six-month-old female rats underwent either sham operation or bilateral ovariectomy, and 7 days postoperatively, the maxillary first molars were extracted. On post-extraction days 7, 30 and 60, the dissected maxillary bone surfaces were examined by SEM to reveal the bone formative and resorptive areas around the extracted alveolar sockets. In addition, the resin-embedded maxillae were micromilled in the transverse direction through the extracted alveolar sockets, and the newly-formed bone mass on the buccal bone surfaces and within the extracted sockets was examined by BSE analysis. Compared with sham-operated controls, the extent of newly-formed bone mass on the buccal bone surfaces in OVX rats was significantly decreased, due to increased bone resorption. On the other hand, new bone formation within the extracted sockets was similar in the experimental groups. In EDX microanalysis of these newly-formed bone matrices, both Ca and P weight % and Ca/P molar ratio were similar in the experimental groups. Our results suggest that 1) acute estrogen deficiency induced by ovariectomy stimulates sustained bone resorption, but has less effect on bone formation, and 2) bone wound healing after maxillary molar extraction within extracted alveolar sockets is not significantly delayed by ovariectomy, but bony support by newly-formed bone mass on the maxillary bone surfaces at the buccal side of the extracted sockets is significantly decreased, due to increased bone resorption. Anat Rec 262:203–212, 2001. © 2001 Wiley-Liss, Inc.
    The Anatomical Record 01/2001; 262(2):203 - 212.
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    ABSTRACT: We examined the effects of a nonantimicrobial tetracycline analogue, CMT-8, on bone loss and osteoclasts in ovariectomized (OVX) rats. Three-month-old female rats were OVX, and, one week later, distributed into three groups: sham-operated non-OVX controls, untreated OVX controls, and CMT-8-treated OVX rats. After 145 days of daily drug administration (p.o.), the femurs were dissected and examined histologically. Ovariectomy markedly decreased trabecular and cortical bone volume in the metaphyses compared to sham-operated controls. Treating the OVX rats with CMT-8 produced a significant inhibition of trabecular and cortical bone loss and induced new bone formation, in which connectivity of the trabecular struts was increased by bridging the adjacent longitudinal bone trabeculae. Ultrastructurally, CMT-8 reduced ruffled border formation in osteoclasts, while it caused no structural impairment in osteoblasts. To further evaluate the effects of CMT-8 on the resorbing activity of osteoclasts, osteoclasts were cultured on dentine slices pretreated with CMT-8 at concentrations of 2, 10, or 50 micrograms/ml, and resorption lacuna formation on the dentine surface was found to be reduced, dose-dependently, by the bound CMT-8. Our results suggest that CMT-8 therapy effectively inhibits post-ovariectomy bone loss not only by inducing new bone formation, but also by inhibiting osteoclastic bone resorption, and that CMT-8 binding to bone may provide a prolonged release delivery of this anti-resorptive therapy.
    Annals of the New York Academy of Sciences 07/1999; 878:347-60. · 4.38 Impact Factor
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    ABSTRACT: Postmenopausal oestrogen deficiency results in bone loss (osteoporosis) in humans and experimental animals. The loss of trabecular bone in the ovariectomized (OVX) rat provides a useful experimental model of post-menopausal osteoporosis. At 5 months after ovariectomy of 3-month-old female rats, the mid and distal femurs and maxillae were dissected and processed for quantitative backscattered electron microscopic examinations. Histomorphometric analysis of femurs in OVX rats showed significant loss in metaphyseal trabecular bone areas compared with sham-operated controls; no significant bone loss was observed in the cortical bone areas of mid-diaphyses in OVX rats. Net bone areas in the maxillae of OVX rats was similar to that of sham-operated controls. Bone structure of maxillae in OVX rats was also similar to that in controls. Our results suggest that, in this animal model of osteoporosis, prominent bone loss occurs mainly in the bone areas formed by endochondral ossification such as distal femurs, but those areas formed by intramembranous ossification such as mid-femurs and maxillae sustained less effects by OVX.
    Journal of Electron Microscopy 02/1999; 48(4):465-9. · 1.44 Impact Factor
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    ABSTRACT: The cellular mechanisms of cartilage-bone replacement in endochondral bone formation, in mandibular condylar heads, are poorly understood. In particular, there is no definitive evidence indicating whether cartilage is resorbed by so-called chondroclasts. Using 3-week-old male beagle dogs, we examined the cartilage-bone replacement processes in mandibular condylar heads by means of light and electron microscopy. Calcification of the cartilage matrix occurred in the central area of the longitudinal septa but not in thin transverse septa. Chondrocytic lacunae were opened by the removal of transverse septa by perivascular rough-surfaced endoplasmic reticulum (RER)-rich mononuclear cells. These cells also phagocytosed calcified cartilage fragments in the surface layer of longitudinal septa. Shortly thereafter, a thin bone layer was deposited on the remaining longitudinal septa by invading osteoblasts. Preosteoclastic multinucleated cells in lacunar canals developed neither ruffled borders nor clear zones in the cartilage matrix, but once the bone layer had been deposited on the remaining cartilage, these structures formed. Our results suggest that the cartilage-bone replacement in mandibular condylar heads involves four sequential processes: 1) degradation of the transverse septal cartilage by RER-rich mononuclear cells, 2) phagocytosis of calcified cartilage fragments in the longitudinal septa by these cells, 3) bone deposition of the remaining longitudinal septa, and 4) degradation of both bone and calcified cartilage by differentiated osteoclasts.
    Journal of Electron Microscopy 07/1996; 45(3):213-22. · 1.44 Impact Factor
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    ABSTRACT: Using 3-week-old male beagle dogs, we examined the cartilage-bone replacement processes in the mandibular condyle by means of light and electron microscopy. Calcification of the cartilage matrix occurred in the central area of the longitudinal septa, but not in the transverse septa. Perivascular mononuclear cells absorbed the transverse septa which initiated the opening of the chondrocytic lacunae. These cells phagocytosed septal cartilage fragments. Shortly thereafter, a thin bone layer was deposited on the remaining longitudinal septa by invading osteoblasts. Osteoclasts in lacunae developed neither ruffled borders nor clear zones in the cartilage matrix, but once the bone layer has been deposited in the remaining cartilage, these structures formed. Our results suggest that the cartilage-bone replacement in mandibular condyle involves three sequential processes: 1) degradation and phagocytosis of cartilage fragments in the transverse septa by mononuclear cells, 2) bone deposition over the remaining longitudinal septa, and 3) degradation of both calcified cartilage and bone by osteoclasts.
    Kaibogaku Zasshi 05/1996; 71(2):106-14.
  • T Kodaka, R Mori, T Sano, K Debari
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    ABSTRACT: By energy-dispersive X-ray microanalysis (EDX) in scanning electron microscopy, we studied the calcification of the inner-basic lamellas of a cow bone and the coronal cementum of a horse tooth treated with sodium hypochlorite (NaOCl). These tissues were divided into 4 groups with a combination of NaOCl treatment and polyester-resin embedding including, [A]: non-NaOCl + Resin, [B]: non-NaOCl + non-resin, [C]: NaOCl + Resin, and [D]: NaOCl + non-Resin. From the Ca and P values by EDX analysis, it was suggested that the natural porous spaces of [B] were higher than those of [A], and both the natural and NaOCl-soluble porous spaces were highest in [D]. However, [A] had the lowest porous spaces in both the tissues because the micropores formed 3-dimensionally by NaOCl treatment might be incompletely filled with the resin. The backscattered electron microscopy and the difference of the Ca/P ratios indicated that the NaOCl treatment of the calcified tissues caused some minerals besides organic materials to dissolve. Thus, the sample preparation of [B] is suitable for the quantitative EDX of calcified tissues, whereas the data of [C] except for the Ca/P ratio may be used to approximate the Ca and P contents.
    Journal of Electron Microscopy 11/1995; 44(5):289-94. · 1.44 Impact Factor
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    ABSTRACT: We examined 15 variably-sized gallstones, taken from an elderly male, by backscattered electron imaging and energy-dispersive X-ray microanalysis to learn the structural and distribution patterns of gallstone calcium (Ca-) salts. Of the 13 cholesterol-rich stones, nine stones had peripheral concentric layers of Ca-carbonate, whereas 2 stones had peripheral layers of Ca-phosphate. No Ca-salts were detected from 2 cholesterol-rich stones. The 2 stones containing Ca-phosphate had no Ca-salt cores, whereas the stones containing Ca-carbonate were separated into 3 different types: two stones with a Ca-carbonate core, four stones with several Ca-bilirubinate cores of glass-like structure, and 3 stones lacking Ca-salt cores. A closer view of the Ca-salt layers, which may be occasionally coexistent with Ca-bilirubinate, mainly showed either laminate deposits or numerous globules with a few laminae. Of the 2 cholesterol-poor stones, one had dispersed particles mainly of Ca-phosphate, and the other had loosely dispersed particles with small amounts of Ca-phosphate, bilirubinate, and/or palmitate. Some relationship between the size and Ca-salt species of these gallstones was suggested. Gallstones collected from the same individual showed a considerable heterogeneity of Ca-salts.
    Scanning microscopy 10/1995; 9(3):907-20; discussion 920-4.
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    ABSTRACT: This study was designed to evaluate the effects of specific and potent cathepsin inhibitors on osteoclastic resorptive functions in vitro by means of a novel ultrastructural assay system. Mouse bone marrow cell-derived osteoclasts were suspended on dentine slices and cultured for 48 hours in the presence of either E-64 (a generalized cysteine proteinase inhibitor) or Z-Phe-Phe-CHN2 (a selective cathepsin L inhibitor). After the removal of cultured osteoclasts, co-cultured dentine slices were examined using electron microscopy: backscattered (BSEM), scanning (SEM), and atomic force (AFM). In morphometric analyses of BSEM images, there were no significant differences in the areas of demineralized dentine surfaces between control and inhibitor-treated groups, suggesting that cathepsin inhibitors had no effect on dentine demineralization by cultured osteoclasts. However, in SEM and AFM observations, both inhibitors remarkably reduced to the same extent, the formation of deep resorption lacunae on dentine slices that had resulted from degradation of matrix collagen. In addition, Z-Phe-Phe-CHN2 treatment produced deeper, ring-like grooves with little collagen exposure in shallow resorption lacunae. These results strongly suggest that (1) cathepsins released by osteoclasts are involved in the formation of deep resorption lacunae, and (2) cathepsin L plays a key role in bone resorption.
    Calcified Tissue International 07/1995; 56(6):566-70. · 2.50 Impact Factor
  • T Kodaka, R Mori, K Debari, M Yamada
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    ABSTRACT: The calcareous concretions of human pineal bodies were investigated with scanning electron microscopy and electron probe microanalysis. The initial concretions measuring 5-7 microns in diameter may have started at the calcified pinealocytes. They grew appositionally forming concentric laminations, and then the simple calcospherulites over 20 microns occasionally aggregated with each other. Some of them became numerous spherulite-aggregated concretions. Others individually grew with scallop-shaped concentric laminations at intervals of 0.05-1 microns and became lobated calcospherulites up to 0.5 mm. The concretions over 0.5 mm were formed by their attachments. The major elements were Ca and P, while traces of S, Mg, and Na were detected. In the calcification and crystallization values, the center of the concretions over 50 microns was significantly higher than the periphery, while there were no differences among the centers and also among the peripheries. The Ca and P amounts in the center were 30.8% and 14.2% by weight and the Ca/P molar ratio was 1.68; thereby the sand-grain-shaped crystals may be nearly hydroxyapatite, as reported previously.
    Journal of Electron Microscopy 11/1994; 43(5):307-17. · 1.44 Impact Factor
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    ABSTRACT: The dentin bonding efficacies of two commercial dentin bonding systems and experimental self-etching dentin primers composed of methacryloxyethyl hydrogen phenyl phosphate (Phenyl-P) and either hydroxyethyl methacrylate (HEMA) or glyceryl methacrylate (GM,2,3-dihydroxypropyl methacrylate) were examined. The wall-to-wall polymerization contraction gap width of a commercial light-activated resin composite in a cylindrical dentin cavity and the tensile bond strength to a flat dentin surface were measured. Changes in dentin hardness were determined by Micro Vickers Hardness measurement, and an SEM observation was performed after priming. Formation of a contraction gap was completely prevented by the application of Phenyl-P diluted in HEMA or GM solution combined with a commercial dentin bonding agent, although gap formation was evident in nearly half of the specimens with both commercial dentin bonding systems. The mean tensile bond strengths of the tested groups varied from 16.3 to 20.7 MPa, and there were no significant differences between groups. Based on the measurement of Micro Vickers Hardness and SEM observation after priming, a slight reduction in dentin hardness was observed. However, this reduction in dentin hardness due to self-etching priming did not significantly correlate with either contraction gap width or tensile bond strength.
    Journal of Dental Research 06/1994; 73(5):1088-95. · 3.83 Impact Factor
  • T Kodaka, K Debari, T Sano, M Yamada
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    ABSTRACT: Human calcium phosphate calculi: two sialoliths, a urolith, a rhinolith, and a tonsillolith were investigated by scanning electron microscopy (SEM) and energy-dispersive X-ray analysis (EDX). The sialoliths and urolith had appositional shells with thick cortices, respectively, around several nuclei composed of calcospherulites and a rubber-film fragment. The rhinolith had a thin cortex with appositional laminations around a glomerulus-like mass of calcified cotton-like strings. The tonsillolith had a rough cortex with appositional laminations. Its porous interior was composed of numerous calcified conglomerates with microorganisms and calcified masses with fine appositional laminations around the conglomerates. The major crystals were identified as biological apatites (AP) with a sand-grain rather than a needle-like shape, and plate-shaped octacalcium phosphate (OCP). The AP deposits of the rhinolith probably were associated with magnesium (Mg) phosphates or contained Mg. No OCP was found in the rhinolith. The AP deposits were mainly formed by extracellular calcification. Hexahedral crystals, identified as Mg-containing whitlockite (WH), were precipitated in the internal spaces of the AP and OCP deposits. The rhinolith nucleus consisted of WH crystal deposits only.
    Scanning microscopy 02/1994; 8(2):241-56; discussion 256-7.
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    ABSTRACT: The effect of delayed light curing of resin composite on marginal adaptation has been examined by measuring the wall-to-wall polymerization contraction gap when using a commercial resin composite together with experimental dentine bonding systems to restore cylindrical preparations in dentine. Morphological changes in dentine during dentine bonding procedures were observed using a scanning electron microscope. In a previous report, the contraction gap width for a resin composite increased when irradiation of the resin system was delayed, despite the use of a dentine bonding system considered to be 'contraction' gap free. Such deterioration in marginal adaptation was minimized by use of an experimental dentine primer, 40% erythritol methacrylate aqueous solution (EM), followed by the use of a commercial dual- or autocured dentine bonding agent. Under scanning electron microscopy, the dentine surface microstructure became unclear after EM priming, and a polymer film was detected after polymerization of the dual-cured dentine bonding agent. The hydrogelled primer and the formation of a polymer network on the dentine surface may prevent the flow of fluid from the pulp through the dentine tubules, and maintain marginal integrity if there is delay in light curing of light-activated resin composite systems.
    Journal of Dentistry 01/1994; 21(6):344-9. · 3.20 Impact Factor
  • T Sasaki, K Debari, M Hasemi
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    ABSTRACT: We have developed a novel ultrastructural assay system for osteoclastic resorptive function. After osteoclasts had been co-cultured on dentine slices for 48 hr, the slices were fixed with glutaraldehyde and examined by means of backscattered electron, scanning electron, and scanning probe microscopies. Backscattered electron images showed areas of low mineralization on dentine surfaces, which, by superimposition of concave-convex images, corresponded to resorption lacunae. The measurement of such resorption lacunae by scanning probe microscopy revealed 3-dimensional topography and their exact depths and volumes. Analysis based on this system provides reliable qualitative and quantitative assessment of osteoclastic resorption.
    Journal of Electron Microscopy 11/1993; 42(5):356-9. · 1.44 Impact Factor
  • T Sasaki, K Debari, N Udagawa
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    ABSTRACT: The osteoclastic cytoskeleton has been demonstrated to be composed of microfilaments. Osteoclastic multinucleated cells were suspended on dentine slices and cultured for 24 hours in the presence or absence of cytochalasin D (CD), a specific and potent inhibitor of actin filament elongation to determine the role of this cytoskeleton. Cultured cells and co-cultured dentine slices were examined ultrastructurally. Unlike those in control cultures without CD, osteoclasts in CD-treated cultures became spherical in shape and lacked microvilli on their basolateral cell surfaces. Most importantly, CD treatment induced a complete disappearance of the ruffled border-clear zone complexes in osteoclasts, which resulted in loss of osteoclast-cytoplasmic polarity. Morphometric analysis of backscattered electron micrographs of co-cultured dentine slices revealed that CD treatment strongly inhibited the formation of resorption lacunae in a dose-dependent manner. These results suggest that the cytoarchitecture, as well as the bone-resorbing function, of the osteoclast is highly regulated by the F-actin-containing microfilamentous cytoskeleton in the ruffled border-clear zone complex.
    Calcified Tissue International 10/1993; 53(3):217-21. · 2.50 Impact Factor
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    ABSTRACT: The environmental SEM (E-SEM) can be used unfixed biological samples under a low vacuum and wet condition. In this study, the fractured dentin of unfixed human teeth was treated with a 30% hydrogen peroxide solution (H2O2) for the examination of tooth-bleaching prior to the E-SEM and a conventional SEM. The peritubular matrix (PM) always showed a few cracks along the long axis of a dentinal tubule, and the ends of fine fibrils rose to the smoothly changed surface of the intertubular matrix (IM). The E-SEM with non-fixation and the conventional SEM following fixation indicated that the hydrogen peroxide solution easily permeated the PM and dissolved the non-fibrillar substance including the cracks of the PM by the constriction. In the IM, the solution may partially dissolve the organic parts within mineralized fibrils as well as non-fibrillar substance between the fibrils, although these remnants might precipitate again there.
    Journal of Electron Microscopy 11/1992; 41(5):381-6. · 1.44 Impact Factor