This study was carried out in order to develop a new implantable biomaterial Bone Morphogenetic Protein (BMP) possessing osteogenetic capability. It seems advisable to use some carrier material for BMP to work effectively. Hydroxypropyl cellulose (HPC) is known as a sustained-release drug. So, we prepared a BMP and HPC pellet, that exhibited a pattern of sustained release of BMP from the BMP-HPC pellets, and investigated its osteinductive potenital. BMP was extracted from bovine cortical bones. The ability the pellets of BMP to form new bone was ensured by implantation of into the muscle pouches of mice. The BMP -HPC pellets or BMP control were implanted in the muscle pouches of mice, and osteoinductive ability was examined at 7, 14 and 21 days after implantation. New bone formation was studied by means of roentogenographic and histologic observation. The amount of bone induced by the BMP-HPC pellets was determined by a computer supported image analysis system. In Both the BMP-HPC pellet and BMP alone groups., new bone formation was seen on roentgenogram, and new cartilage and bone were histologically observed. The osteoinductive capability of the BMP-HPC pellets was however greater than that of BMP alone. The sustained-release of BMP by HPC was found to enhance the osteoinductive capbility of BMP.

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I conducted a fundamental examination to apply an atelocollagen membrane chemically treated with cross-linking, to guided tissue regeneration technique (GTR), as a method of facilitating tissue regeneration after periodontal surgery. Twenty -eight mongrel adult dogs were used . The bony defect of dehiscence type was artificially prepared at the bilateral fourth premolars of the lower jaw on the buccal sides. After root planing, the wound healing was histopathologically examined at 4 and 6 weeks after surgery, in the three groups of the dogs with implantation of atelocollagen membranes cross linked chemically with different degrees (Col.× 0, Col.× 0.5, Col.× 10), and a group w (ithout implantation (Control). As a result, application of the atelocollagen membrane cross linked chemically (Col.× 0.5, Col.× 10) enabled the following: (1) to delay the time of membrane's dissolution in the body without prolongation of local inflammatory responses after membrane implantation, and 2) to suppress the epithelial downgrowth, and to increase the amounts of cementogenesis and osteogenesis, without causing excessive resorption of the roots of teeth. Thus, the GTR method using the atelocollagen membrane cross-linked chemically can be advantageous as a regenerative treatment in periodontal surgery.

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This study was designed to investigate the immunohistochemical localization of chondoroitin sulfate isomers (chondroitin-4 sulfate, -6 sulfate and dermatan sulfate) of proteoglycans (PGs) in dog periodontal tissues during experimental periodontitis (floss ligatures were placed for periods of 0, 3, 7, 21 and 60 days) using monoclonal anti-bodies (2-B-6, 3-B-3). The result demonstrated that on Day 0, chondroitin-4 sulfate and dermatan sulfate -PGs were widespread in gingival connective tissue and periodontal membrans, with an especially strong response of dermatan sulfate-PG along collagen fiber bundles. Chondroitin-6 sulfate PG was located at the junction between the gingival connective tissue and epithelium, some areas of connective tissue attachment and peripheral blood vessels in gingival and periodontal membranes. In alveolar bone, chondroitin-4 sulfate, -6 sulfate and dermatan sulfate-PGs were found inside of Haversian canals, Volkman's canals and lacunae. In the phase of acute inflammation (Days 3 and 7), the staining patterns or cnonaroitin-4 suirate, -o suiiate arm dermatan sulfate-PGs were barely observable in small areas of destroyed gingival connective tissue. In addition, positive staining of chondroitin-4 sulfate, -6 sulfate and dermatan sulfate-PGs was found along the periphery of cells like osteoclast and in the loose connective tissue of active alveolar bone resorption lacunae. In the pahse of chronic inflammation (days 21 and 60), in which destruction of gingival connective tissue occurs, chondroitin-4 sulfate, -6 sulfate and dermatan sulfate-PGs stained weakly in a widespread area. On days 21 and 60, the staining patterns of the three chondroitin sulfate isomer-PGs in bone resorption lacunae were weak compared with those on Day 3 and 7 even though bone resorption had advanced. These findings suggest that chondroitin-4 sulfate, -6 sulfate and dermatan sulfate-PGs may be degraded by inflammation in gingival connective tissue and may play an important role in the initial stage of bone resorption.

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The purpose of this study was to evaluate the correlation between destruction of periodontal component parts, measuring a biological width and a pathobiological width, in four different stages of inflammatory periodontal disease as assesed by extension of the inflammation into the periodontal tissue. The biological width represented the distance parallel to the axis of the tooth between the bottom of the gingival or periodontal pocket and the represented the distance parallel to the axis of the tooth between the bottom of the pocket and the coronal end of the alveolar bone area, where the Sharpey's fibers were attached.
Using step-serial undecalcified sections, 129 interproximal sites of 94 teeth in 30 human jaws, divided into three age groups, were histomorphometrically investigated under light microscopy. The extension of the inflammatory infiltration correlated with age, the frequency of dental plaque and calculus deposition and the frequency of ulceration of the pocket epithelium (chi-square test p<0.01). Although the pathobiological width showed no significant differences among the different stages of inflammatory extension, the biological width was shorter in the most severe stage, in which the inflammatory infiltration reached the alveolar bone marrow, than in the other stages.
It was suggested that the deepening of the pocket, the resorption of the alveolar bone crest and the destruction of the periodontal ligament progressed migration of the junctional epithelium in the three earliest stages of inflammatory extension. In the most severe stage with inflammatory infiltration of the alveolar bone marrow, the deepening of the pocket and the destruction of the periodontal ligament progressed more rapidly than the resorption of the alveolar bone crest and the apical migration of the junctional epithelium.

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Some researchers postulated that only periodontal ligament (PDL) cells have the potential to establish new connective tissue attachment including the formation of new cementum. This concept, however, has recently been controversial. In order to clarify the origin of cells that can regenerate the periodontal tissues following flap surgery, the author examined cell proliferation and migration in explanted tissues which consisted of tooth, PDL, alveolar bone, and gingival connective tissue. Five adult monkeys with healthy gingiva were used. After gingival epithelium was excluded from gingiva and mucoperiosteal flaps were elevated on the buccal aspects of molars, a section of approximately 3 mm of crestal bone was removed. Subsequently, small tissue fragments of 2mm×2mm×4mm containing tooth, PDL, alveolar bone, and gingival connective tissue were obtained, and then cultured for 1, 3, 7, 10, and 14 days. The outgrowth of cells from the explants was observed every day under a phase-contrast microscope. Cells were labeled with bromo-deoxyuridine (BrdU) at each time period to evaluate DNA synthesis; the localization of labeled cells (Lc), namely, progenitor cells was immunostained with the anti-BrdU antibody. PDL was divided into 3 zones buccolingually to evaluate the Lc density at each compartment. In addition, specimens for transmission electron microscopy (TEM) were prepared. The Lc proliferated in the endosteal spaces and appeared to migrate from their openings into PDL. In addition, the Lc proliferation was seen around blood vessels. The Lc density in the bone side and the middle of PDL was higher than that in the cementum side. TEM findings showed that cells outgrew from the crestal bone and then migrated to the root surface. Thus cells likely migrated from crestal bone, openings of endosteal spaces into PDL, and paravascular regions in PDL after flap surgery, which suggests that they might be involved in the formation of new cementum after migrating to the root surface.

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This study was performed to know the best way of some periodontal treatments on the diseased root cementum by attachment and growth of cultured human gingival fibroblasts.
Four modalities of root treatments were performed as experimental groups: 1) ultrasonic scaling, Sc. 2) polishing by rubber cup and paste followed by scaling, Sc + Po. 3) shaved 20μm surface layer of cementum by curette scaler followed by scaling, Sc + 20μRp. 4) removed all cementum was by curette type scaler followed by scaling (throughly root planing), Sc + Rp. Three groups were made as controls: 1) no treatment of diseased cementum 2) throughly root planing of healthy teeth 3) no treartment of healthy teeth.
Root fragments were prepared from surgically extracted human healthy and periodontally involved teeth and each tublet had 3 mm diameter and 1 mm thickness. Each group had 10 tublets. The treated root fragments were placed in petridishes and seeded 1 x 10⁵/m/l fibroblasts in E-MEM containing 200 units/ml penicillin and 200 μm/m/l streptomycin followed by incubation for 72 h. After the tublets were fixed and stained, the number of attachments and growth cellls were counted by light microscopy and examined the morphologies by S. E. M.
The resutls were as follows:
1) the number of fibroblasts on the Sc-tublets was a few and their shapes were not enough to spread. 2) the number of cells attached on the Sc + Po-tublets was a few, but more than Sc group. 3) the number of cells attached on Sc + 20μRp and Sc + Rp groups was increased to the control level (C and C-Rp groups). Their cells were shaped flat and spreading, and they had many projections connecting to other cells.
These observations suggested that the treatments of shaving 20μm on the root surface and removing all cementum (throughly root planing) gave a good condition for attachment and growth of fibroblasts.

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The purpose of this study is to elucidate osteoblastic characteristics of two different cell populations derived from human alveolar bone. The cells from collagenase-treated bone fragments were termed E-AB, whereas the cells from untreated bone fragments were termed N-AB. Both cell populations were subcultured up to four generations, and used for this study. To determine phosphatase activity per 10⁴ cells, enzyme assay was carried out directly using the confluent monolayer in each well of a 96-well microplate. Protein synthesis was determined by 24 hr ¹⁴C-proline labeling. After the labeling, the cultured medium, cells and matrix layer were separately collected to determine ¹⁴Cproline incorporation. The radiolabeled proteins were separated by SDS-PAGE and determined by fluorography. The characteristics of both cell populations were as follows. The E-AB showed about 7-fold higher alkaline phosphatase activity than the N-AB's. On the other hand, no differences were observed in total acid phosphatase and tartrate -resistant acid phosphatase activities, which were less than 50% of the N-AB's alkaline phosphatase activity and about 25% of the total acid phosphatase activity, respectively. According to the results of ¹⁴C-proline labeling, both cell populations produced and secreted large quantities of Mrs of over 200 kDa proteins, procollagens and collagens into the culture media. However, the 28 K and 34 K proteins were much more evident in the E-AB's cultured medium. In both matrix layers, Mrs of 40-50 K proteins were incorporated, in addition of the large amounts of protein in the culture medium.

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The purpose of this study was to observe bacterial attachment and plaque formation on different kinds of suture materials in vitro and in vivo. Following three kinds of suture materials, braided silk suture, braided polyester suture, and monofilament nylon suture were used throughout this study. In vitro experiment, Actinomyces viscosus RF 7, Streptococcus sanguis 10556 and Streptococcus mutans PS 14 were incubated with each suture material in phosphate buffered saline for 5 hours, in brain heart infusion broth for 24 hours. Aritificial plaque formation on each suture material was also induced using S. mutans in 5% sucrose added BHI. In vivo experiment, at the time of periodontal flap surgery, the flap was positioned so as to obtain primary closure with each suture material. One week after flap surgery, the sutures were collected as the experimental specimens. After in vitro and in vivo experiment, all suture materials were processed for SEM and observed.
The following conclusions were obtained from this study.
1) The number of attached A. viscosus, S. sanguis and S. mutans to braided silk and polyester sutures were more pronounced than that to monofilament nylon suture.
2) The amount of artificial plaque on two kinds of braided sutures were more pronounced than that on monofilament suture.
3) On two kinds of braided sutures, bacterial movement due to wicking motion were confirmed.
4) In vivo experiment, the amount of plaque formation were more pronounced on two kinds of braided sutures than that on monofilament suture. Also, the bacterial colonies were formed on the part of braided sutures which penetrated into gingival tissues.

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We have previously reported that the deeper cementum in human periodontally involved teeth enhanced the formation of a new connective tissue attachment. The present study was undertaken to evaluate the air-powder abrasive system as an adjunct to curet the superficial cementum completely. Two patients, 38-and 66-year old males were selected ; their right maxillary second incisor, the pocket depth was more than 7 mm, was used. The labial root surface was sprayed with air-powder abrasive system for 10 seconds after 20 instrument strokes with a curet scaler. The interface of the root surface and regenerating connective tissue was observed 2 and 3 weeks after surgery by light and electron microscopy. In addition, histological measurement of the new connective tissue attachment was performed. At 2 week time point, the deeper cementum remained from 70 to 120μm in width on the root. The average measurement of length of the connective tissue attacment was 1.66 mm. At 3 week time point, the deeper cementum remained from 90 to 180μm in width. The average measurement of the length of the connective tissue attachment was 1.60 mm. Fibroblasts with well-developed rough endoplasmic reticulum and numerous ribosomes synthesized new fibrillar materials and collagen fibrils on the curetted cementum. The results suggested that the air-powder abrasive system after instrumentation with a curet scaler would be effective for the complete removal of bacteria in resorption lacunae and the remained superficial cementum.

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The purpose of this study was to evaluate the effects of clinical instrumentation with ultrasonic scaler (ENAC-3: SC-1, SC-5, SC-6, Diamond file) and a hand scaler (Gracey: # 11-12, 13-14) on the multirooted teeth which had been sectioned faciolingually in order to score residual calculus. After instrumentation with ENAC-3 and Gracey scalers, the same area was examined microscopically at a constant magnification in order to assess the percentage of residual calculus compared with the initial and final scoring of the calculus. Hopeless teeth, which had been designated for extraction for periodontal reasons, were also root planed with the same type of instrumentation and investigated in order to score the residual calculus on each of the root surfaces. In addition, root surface topography after instrumentation was examined by SEM and light microscopy.
The following results were obtained;
1) According to the microscopic evaluation, SC -1 and SC-5 produced the same smooth surface which could be obtained with a Gracey scaler.
2) At a low magnification of the light microscope and SEM, the topography results from SC-1, SC-5 and Gracey appeared smooth, but flat surfaces with occasional deep gouges were recognized at the more apical areas of surfaces planed with SC-1 and SC-5 scalers.

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The aim of the present study was to evaluate the response of deep angular bony defects to guided tissue regeneration (GTR). The patient sample included 16 subjects, 23-54 years of age, who presented with periodontal lesions which had a probing pocket depth (PD) that was ≥6mm after reevaluation. Baseline information, including PD and clinical attachment level (CAL) measurements, was recorded after reevaluation. Eight experimental and 8 control angular bony defects, randomly assigned, were included. Following flap elevation, scaling, root planing and removal of granulation tissue, on the experimental teeth, Gore-Tex periodontal material was placed and sutured, using a suspensory suture. The flaps were sutured tightly, assuring complete coverage of the material. After 4 to 6 weeks, the Gore-Tex membranes were removed. Clinical measurements were repeated at 3, 6 and 12 months following surgery. Changes from baseline in PD and CAL were calculated for each case. The results indicated that PD measurements were reduced by both procedures, but the reduction was greater for GTR at 3, 6 and 12 months. At 12 months the test sites showed 5.1.±1.6mm pocket reduction, while the control sites showed an average 2.4±1.5mm reduction in pocket depth. CAL recordings were improved by both treatments, but were better for GTR at 3, 6 and 12 months, with an average gain in CAL of 2.8±1.6 mm for the GTR and0.3±0.8mm for the controls. This suggests that GTR can improve the response to therapy for deep angular bony defects.

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A comprehensive examination was performed on twenty-nine (18 males and 11 females) circulatory disease patients treated with the drugs. The periodontal examination included number of teeth, probing depth (PD), gingival bleeding index (GBI), plaque control record (PCR), gingival index (GI), tooth mobility (TM) and bone index (BI). The patients were divided into the following groups: Ca group (taking calcium antagonists) ; (non-Ca group taking other drugs) ; Hyp group (evidence of gingival hyperplasia) and non-Hyp group (no evidence of gingival hyperplasia).
The mean age of all patients was 55. 44±8.39 range (39-79) years and sixteen patients received calcium antagonists. Nine patients (56.3%) presented gingival hyperplasia and belonged to the Ca group. Furthermore, except one patient, they had all been treated Nifedipine.
The Ca group had significantly deeper PD and less BI than the non-Ca group. The Hyp group had significantly less TM and BI than the non-Hyp group.
The hyperplastic gingiva showed fibrotic and nodular changes, some clefst of the marginal gingiva and various degrees of inflammation.
The gingival biopsies showed slight hyperkeratosis, elongation of the rete pegs, chronic inflammatory infiltration and dense collagen bundles in subepithelial connective tissue.

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Various laboratory techniques have been applied to the microbiological diagnosis of periodontal disease. However, certain disadvantages persist. In this study we examined the specific peptidase activity in periodontal pockets by using the chair-side test system, SK-013 as a microbiological diagnostics technique.
SK-013 can rapidly (within 15 min) evaluate the peptidase activities derived from periodontopathic bacteria such as Treponema denticola and Bacteroides species. We applied SK-013 to patients with periodontitis, gingivitis, dental caries, other types of periodontal infectious disease and periodontally healthy controls and then evaluated the clinical specificity of SK-013, assessing its cut-off level.
SK-013 activity was determined both spectrophotometrically and by using the color standard. Phase contrast microscopy was also utilized for determination of subgingival microorganisms. SK -013 activity was specifically detected in the periodontitis group. Setting its cut-off level at 0.2 Try U/ml based on the number of spirochetes in the subgingival pocket, we got good sensitivity, specificity, predictive value and efficacy in terms of the diagnostic value of SK-013. Using the same cut -off level, a good coincidence was also shown in both spectrophotometrically and by using the color standard.
These results indicate that SK-013 is a useful chair-side diagnostic method for periodontal disease.

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The purpose of the present study was to examine the diagnostic value of SK-013 (Sunstar, Osaka) in evaluating the efficacy of periodontal pocket curettage. SK-013 is a highly sensitive reagent which is used to measure the specific peptidase activities produced by Treponema denticola, Bacteroides (Porphyromonas) gingivalis and Bacteroides forsythus. Sixty-four periodontitis patients were selected for the study. Of the 64 patients, 33 were treated by plaque control instruction and periodontal pocket curettage (T group), and the other 31 patients received no treatment for periodontal disease (NT group). One site was monitored in each patient. The clinical status of each site was assessed using clinical indices such as the gingival index (GI), plaque index (PH), probing depth (PD), attachment level (AL), bleeding on probing (BOP) and tooth mobility. In the T group, the clinical and enzymatic examinations were made at five separate appointments as follows: (1) at baseline prior to plaque control instruction; (2) 4 weeks after plaque control instruction; and (3) 2, 4, and 8 weeks after periodontal pocket curettage. The same examinations were performed but, only at baseline and after 12 weeks, in the NT group. As a result, periodontal pockets were reduced in depth following plaque control instruction, while the enzymatic activity showed no significant change before and after plaque control. Two weeks after periodontal pocket curettage, both clinical indices (GI, PD, AL, BOP) and SK-013 activity were significantly reduced and these improvements were maintained for 4 weeks. No significant discrepancy was observed in the NT group between clinical assessment and enzymatic activity during the experimental period. This finding suggested that SK-013 is a clinically useful diagnostic method for assessing the effect of initial periodontal therapy.

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Periodontal tissues from patients with gingival hyperplasia caused by nifedipine were clinically and histopathologically examined and the microorganisms in the periodontal pockets were detected by phase-contrast microscopy.
The results obtained were asfollows:
1. No correlation was found between the amount of gingival hyperplasia and the plaque index.
2. No correlation between the amount of gingival hyperplasia and the total number of bacteria or proportional distribution of spirochetes and motile rods were found in the periodontal pockets by phase-contrast microscopy.
3. The microscopic sections showed epithelial hyperplasia with mild hyperparakeratosis, elongation of rete pegs, dense collagen bundles, engorged blood vessels, and moderate inflammatory cell infiltration predominantly with plasma cells.
4. Ultrastructurally, fibroblasts containing well -developed rough endoplasmic reticulum or phagocytosed collagen fibrils, and granular substances between the collagen fibrils were observed.

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This study was designed to make a multi-sided comparison between unexposed (intact) and esposed cementum.
The results were as follows:
1. A slight difference between groups could be found in Ca, P and Ca/P by X-ray microanalyzer.
2. The scanning electron microscopical observation showed that the cementicle in unexposed cementum and exposed cementum was observable as hypercalcified areas.
3. No statistical difference between groups could be found in the relative concentrations of the target elements Ca, P, F, 0 and Ca/P, but a statistical difference between groups could be found in N by X -ray photoelectron spectroscopy .
4. The contact angle in exposed cementum was demonstrated be more hydrophilic than in unexposed cementum.

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Care of defects in the oral mucosa, have been extensively discussed especially the donor site for the free gingival graft technique, which most frequently consists of palatal mucosa. The purpose of this study was to evaluate the protection and regeneration of palatal mucosa using dressing materials. The palatal areas of 2 healthy monkeys were designed as the experimental sites, from which split thickness gingival flaps of relatively uniform thickness were removed . They received a chitin membrane or lyophilized dura mater as a dressing. Healing responses were analysed quantitatively and observed histologically, in terms of the amounts of type I and III collagen- and keratin-proteins. Types and amounts of collagen were investigated by SDS-PAGE and keratin proteins were determined by ELISA and Western blotting. The results demonstrated that type I collagen production in both the chitin and dura mater experimental groups had increased more rapidly than in the group without dressing material. The detected increase in Type III collagen was markedly greater in the chitin group at 3 weeks after surgery. A great deal of keratin protein had been produced at 5 weeks with use of the chitin dressing. It was concluded that the chitin might protect the denuded palatal sites, promoting keratin production, and thereby facilitating rapid andeffective regeneration of the oral mucosa.

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Poly (L-lactic acid) microspheres containing tetracycline (TC-PLA-MS) were prepared using hydrophilic petrolatum or atelocollagen solution as the base. Clinical effects of the preparation were determined in 14 patients with periodontal disease after single administration to 57 sites involving periodontal pockets of more than 4 mm in depth. The results showed that bleeding on probing, total counts of anaerobic bacteria and the proportion of black pigmented bacteroides decreased significantly up to day 14 after administration in the TC-PLA -MS group, while, no significant difference was found in the probing depth, sulcus bleeding index and the proportion of spirochetes throught the study period. There was also no significant difference in the density of microorganisms in pockets between 200 1991.3. the TC-PLA-MS group and the controls after 7 days. No definite difference was noted in therapeutic effects between the different bases.

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