Efficacy of salivary versus subgingival bacterial sampling for the detection and quantification of periodontal pathogens

Purpose@#The aim of this study was to investigate the efficacy and validity of subgingival bacterial sampling using a retraction cord, and to evaluate how well this sampling method reflected changes in periodontal conditions after periodontal therapy. @*Methods@#Based on clinical examinations, 87 subjects were divided into a healthy group (n=40) and a periodontitis group (n=47). Clinical measurements were obtained from all subjects including periodontal probing depth (PD), bleeding on probing (BOP), clinical attachment loss (CAL), and the plaque index. Saliva and gingival crevicular fluid (GCF) as a subgingival bacterial sample were sampled before and 3 months after periodontal therapy. The salivary and subgingival bacterial samples were analyzed by reverse-transcription polymerase chain reaction to quantify the following 11 periodontal pathogens: Aggregatibacter actinomycetemcomitans (Aa), Porphyromonas gingivalis (Pg), Tannerella forsythus (Tf), Treponema denticola (Td), Prevotella intermedia (Pi), Fusobacterium nucleatum (Fn), Pavimonas micra (Pm), Campylobacter rectus (Cr), Prevotella nigrescens (Pn), Eikenella corrodens (Ec), and Eubacterium nodatum (En). @*Results@#Non-surgical periodontal therapy resulted in significant decreases in PD (P<0.01), CAL (P<0.01), and BOP (P<0.05) after 3 months. Four species (Pg, Tf, Pi, and Pm) were significantly more abundant in both types of samples in the periodontitis group than in the healthy group. After periodontal therapy, Cr was the only bacterium that showed a statistically significant decrease in saliva, whereas statistically significant decreases in Cr, Pg, and Pn were found in GCF. @*Conclusions@#Salivary and subgingival bacterial sampling with a gingival retraction cord were found to be equivalent in terms of their accuracy for differentiating periodontitis, but GCF reflected changes in bacterial abundance after periodontal therapy more sensitively than saliva.

Efficacy of salivary versus subgingival bacterial sampling for the detection and quantification of periodontal pathogens

Purpose@#The aim of this study was to investigate the efficacy and validity of subgingival bacterial sampling using a retraction cord, and to evaluate how well this sampling method reflected changes in periodontal conditions after periodontal therapy. @*Methods@#Based on clinical examinations, 87 subjects were divided into a healthy group (n=40) and a periodontitis group (n=47). Clinical measurements were obtained from all subjects including periodontal probing depth (PD), bleeding on probing (BOP), clinical attachment loss (CAL), and the plaque index. Saliva and gingival crevicular fluid (GCF) as a subgingival bacterial sample were sampled before and 3 months after periodontal therapy. The salivary and subgingival bacterial samples were analyzed by reverse-transcription polymerase chain reaction to quantify the following 11 periodontal pathogens: Aggregatibacter actinomycetemcomitans (Aa), Porphyromonas gingivalis (Pg), Tannerella forsythus (Tf), Treponema denticola (Td), Prevotella intermedia (Pi), Fusobacterium nucleatum (Fn), Pavimonas micra (Pm), Campylobacter rectus (Cr), Prevotella nigrescens (Pn), Eikenella corrodens (Ec), and Eubacterium nodatum (En). @*Results@#Non-surgical periodontal therapy resulted in significant decreases in PD (P<0.01), CAL (P<0.01), and BOP (P<0.05) after 3 months. Four species (Pg, Tf, Pi, and Pm) were significantly more abundant in both types of samples in the periodontitis group than in the healthy group. After periodontal therapy, Cr was the only bacterium that showed a statistically significant decrease in saliva, whereas statistically significant decreases in Cr, Pg, and Pn were found in GCF. @*Conclusions@#Salivary and subgingival bacterial sampling with a gingival retraction cord were found to be equivalent in terms of their accuracy for differentiating periodontitis, but GCF reflected changes in bacterial abundance after periodontal therapy more sensitively than saliva.

The bactericidal effect of an atmospheric-pressure plasma jet on Porphyromonas gingivalis biofilms on sandblasted and acid-etched titanium discs

PURPOSE: Direct application of atmospheric-pressure plasma jets (APPJs) has been established as an effective method of microbial decontamination. This study aimed to investigate the bactericidal effect of direct application of an APPJ using helium gas (He-APPJ) on Porphyromonas gingivalis biofilms on sandblasted and acid-etched (SLA) titanium discs. METHODS: On the SLA discs covered by P. gingivalis biofilms, an APPJ with helium (He) as a discharge gas was applied at 3 different time intervals (0, 3, and 5 minutes). To evaluate the effect of the plasma itself, the He gas–only group was used as the control group. The bactericidal effect of the He-APPJ was determined by the number of colony-forming units. Bacterial viability was observed by confocal laser scanning microscopy (CLSM), and bacterial morphology was examined by scanning electron microscopy (SEM). RESULTS: As the plasma treatment time increased, the amount of P. gingivalis decreased, and the difference was statistically significant. In the SEM images, compared to the control group, the bacterial biofilm structure on SLA discs treated by the He-APPJ for more than 3 minutes was destroyed. In addition, the CLSM images showed consistent results. Even in sites distant from the area of direct He-APPJ exposure, decontamination effects were observed in both SEM and CLSM images. CONCLUSIONS: He-APPJ application was effective in removing P. gingivalis biofilm on SLA titanium discs in an in vitro experiment.

A clinical pilot study of jawbone mineral density measured by the newly developed dual-energy cone-beam computed tomography method compared to calibrated multislice computed tomography

PURPOSE: This clinical pilot study was performed to determine the effectiveness of dual-energy cone-beam computed tomography (DE-CBCT) in measuring bone mineral density (BMD).MATERIALS AND METHODS: The BMD values obtained using DE-CBCT were compared to those obtained using calibrated multislice computed tomography (MSCT). After BMD calibration with specially designed phantoms, both DE-CBCT and MSCT scanning were performed in 15 adult dental patients. Three-dimensional (3D) Digital Imaging and Communications in Medicine data were imported into a dental software program, and the defined regions of interest (ROIs) on the 3-dimensional surface-rendered images were identified. The automatically-measured BMD values of the ROIs (g/cm³), the differences in the measured BMD values of the matched ROIs obtained by DE-CBCT and MSCT 3D images, and the correlation between the BMD values obtained by the 2 devices were statistically analyzed.RESULTS: The mean BMD values of the ROIs for the 15 patients as assessed using DE-CBCT and MSCT were 1.09±0.07 g/cm³ and 1.13±0.08 g/cm³, respectively. The mean of the differences between the BMD values of the matched ROIs as assessed using DE-CBCT and calibrated MSCT images was 0.04±0.02 g/cm³. The Pearson correlation coefficient between the BMD values of DE-CBCT and MSCT images was 0.982 (r=0.982, P<0.001).CONCLUSION: The newly developed DE-CBCT technique could be used to measure jaw BMD in dentistry and may soon replace MSCT, which is expensive and requires special facilities.

Periodontal pathogens and the association between periodontitis and rheumatoid arthritis in Korean adults

PURPOSE: Periodontitis and rheumatoid arthritis (RA) share a similar inflammatory pathogenesis. Porphyromonas gingivalis (Pg) can induce anticyclic-citrullinated peptide autoantibodies (anti-CCP antibodies), a key factor in the development of RA. This study aimed at evaluating the relationships between the 2 diseases and identifying the clinical implications thereof, with a focus on periodontal pathogens in Korean adults. METHODS: A total of 260 RA patients and 86 age- and sex-matched control patients without arthritis were enrolled in this prospective cross-sectional study. Periodontal indices and the prevalence and amount of periodontal pathogens were compared between the groups. Correlations between periodontal and RA indices were examined, as were correlations between 9 periodontal pathogens and RA indices. RESULTS: The RA group had significantly higher values than the control group for all investigated periodontal indices (P < 0.05) except the number of teeth. The gingival index (GI) was correlated with the disease activity score 28 (DAS28) (r = 0.125, P = 0.049), RA disease duration (r = 0.253, P < 0.001), erythrocyte sedimentation rate (ESR) (r = 0.162, P = 0.010), and anti-CCP antibody titer (r = 0.205, P = 0.004). Probing pocket depth (PPD) was correlated with ESR (r = 0.139, P = 0.027) and anti-Pg antibody titer (r = 0.203, P = 0.001). Bleeding on probing (BOP) was correlated with DAS28 (r = 0.137, P = 0.030), RA disease duration (r = 0.202, P = 0.001), ESR (r = 0.136, P = 0.030), anti-Pg antibody titer (r = 0.177, P = 0.005), and anti-CCP antibody titer (r = 0.188, P = 0.007). Clinical attachment level (CAL) and periodontitis severity were correlated with anti-Pg antibody titer (the former r = 0.201, P = 0.002; the latter r = 0.175, P = 0.006). The quantity of Pg was positively correlated with the serum anti-Pg antibody titer (r = 0.148, P = 0.020). CONCLUSIONS: The GI, BOP, and PPD showed positive relationships with several RA indices. The anti-Pg antibody titer had positive relationships with PPD, BOP, CAL, and periodontitis severity. Thus, increasing values of periodontal indices could be used as a risk indicator of disease development in RA patients, and an increasing anti-Pg antibody titer could be considered as a warning sign in RA patients suffering with periodontitis.

Magnoliae Cortex and maize modulate Porphyromonas gingivalis-induced inflammatory reactions

PURPOSE: The aim of this study was to evaluate the capacity of single and combined applications of the bark of the stems and roots of Magnolia officinalis Rehd. et Wils. (Magnoliae Cortex) and Zea mays L. (maize) to modulate inflammation in RAW 264.7 cells stimulated with Porphyromonas gingivalis. METHODS: RAW 264.7 cells were stimulated with P. gingivalis, and Magnoliae Cortex and/or maize was added. Cytotoxicity and the capacity to modulate inflammation were determined with a methylthiazol tetrazolium (MTT) assay, nitrite production, enzyme-linked immunosorbent assay (ELISA), and western blotting. RESULTS: Treatment with Magnoliae Cortex and/or maize inhibited nuclear transcription factor κB (NF-κB) pathway activation and nuclear p44/42 mitogen-activated protein kinase (MAPK) and inducible nitric oxide synthase (iNOS) protein expression in P. gingivalis-stimulated RAW 264.7 cells. Moreover, the treatments suppressed cytokines (prostaglandin E2 [PGE2], interleukin [IL]-1β, and IL-6) and nitrite production. CONCLUSIONS: Both Magnoliae Cortex and maize exerted an anti-inflammatory effect on P. gingivalis-stimulated RAW 264.7 cells, and this effect was more pronounced when the extracts were combined. These findings show that these extracts may be beneficial for slowing the progression of periodontal disease.

Comparison of marginal bone loss between internal- and external-connection dental implants in posterior areas without periodontal or peri-implant disease

PURPOSE: The purpose of this retrospective study with 4–12 years of follow-up was to compare the marginal bone loss (MBL) between external-connection (EC) and internal-connection (IC) dental implants in posterior areas without periodontal or peri-implant disease on the adjacent teeth or implants. Additional factors influencing MBL were also evaluated. METHODS: This retrospective study was performed using dental records and radiographic data obtained from patients who had undergone dental implant treatment in the posterior area from March 2006 to March 2007. All the implants that were included had follow-up periods of more than 4 years after loading and satisfied the implant success criteria, without any peri-implant or periodontal disease on the adjacent implants or teeth. They were divided into 2 groups: EC and IC. Subgroup comparisons were conducted according to splinting and the use of cement in the restorations. A statistical analysis was performed using the Mann-Whitney U test for comparisons between 2 groups and the Kruskal-Wallis test for comparisons among more than 2 groups. RESULTS: A total of 355 implants in 170 patients (206 EC and 149 IC) fulfilled the inclusion criteria and were analyzed in this study. The mean MBL was 0.47 mm and 0.15 mm in the EC and IC implants, respectively, which was a statistically significant difference (P < 0.001). Comparisons according to splinting (MBL of single implants: 0.34 mm, MBL of splinted implants: 0.31 mm, P=0.676) and cement use (MBL of cemented implants: 0.27 mm, MBL of non-cemented implants: 0.35 mm, P=0.178) showed no statistically significant differences in MBL, regardless of the implant connection type. CONCLUSIONS: IC implants showed a more favorable bone response regarding MBL in posterior areas without peri-implantitis or periodontal disease.

A randomized controlled clinical study of periodontal tissue regeneration using an extracellular matrix-based resorbable membrane in combination with a collagenated bovine bone graft in intrabony defects

PURPOSE: The purpose of this study was to investigate the feasibility of regenerative therapy with a collagenated bone graft and resorbable membrane in intrabony defects, and to evaluate the effects of the novel extracellular matrix (ECM)-based membrane clinically and radiologically. METHODS: Periodontal tissue regeneration procedure was performed using an ECM-based resorbable membrane in combination with a collagenated bovine bone graft in intrabony defects around the teeth and implants. A novel extracellular matrix membrane (NEM) and a widely-used membrane (WEM) were randomly applied to the test group and the control group, respectively. Cone-beam computed tomography images were obtained on the day of surgery and 6 months after the procedure. Alginate impressions were taken and plaster models were made 1 week and 6 months postoperatively. RESULTS: The quantity of bone tissue, the dimensional changes of the surgically treated intrabony defects, and the changes in width and height below the grafted bone substitutes showed no significant difference between the test and control groups at the 6-month examination. CONCLUSIONS: The use of NEM for periodontal regeneration with a collagenated bovine bone graft showed similar clinical and radiologic results to those obtained using WEM.

Comparative, randomized, double-blind clinical study of alveolar ridge preservation using an extracellular matrix-based dental resorbable membrane in the extraction socket

PURPOSE: The aim of this study was to radiographically and clinically compare the effect of extracellular matrix (ECM) membranes on dimensional alterations following a ridge preservation procedure. METHODS: One of 2 different ECM membranes was applied during a ridge preservation procedure. A widely used ECM membrane (WEM; Bio-Gide, Geistlich Biomaterials, Wolhusen, Switzerland) was applied in the treatment group and a newly developed ECM membrane (NEM; Lyso-Gide, Oscotec Inc., Seongnam, Korea) was applied in the control group. Cone-beam computed tomography (CBCT) scans and alginate impressions were obtained 1 week and 6 months after the ridge preservation procedure. Results were analyzed using the independent t-test and the nonparametric Mann-Whitney U test. RESULTS: There were no significant differences between the ECM membranes in the changes in the dimension, width, and height of the extraction socket or the quantity of bone tissue. CONCLUSIONS: The NEM showed comparable clinical and radiographic results to the WEM following the ridge preservation procedure.

Rescue of a periodontally compromised tooth by non-surgical treatment: a case report

PURPOSE: This article describes a case of the successful non-surgical management of a periodontally compromised maxillary premolar. METHODS: A combination therapy, including root planing, occlusal adjustment, and tooth splinting, was applied. Clinical and radiographic examinations were performed during the 16-month follow-up period. RESULTS: All periodontal parameters were improved. There were dramatic decreases (3–6 mm) in the probing pocket depth, tooth mobility, and marginal bone loss. Interestingly, gradual resolution of the periapical radiolucency and alveolar bone regeneration were observed in the radiographs, and the periodontal condition was maintained during the follow-up period. CONCLUSIONS: Within the limits of this study, these results demonstrate the importance of natural tooth preservation through proper periodontal treatment and occlusal adjustment of the periodontally compromised tooth, which is typically targeted for tooth extraction and dental implantation.

New method of assessing the relationship between buccal bone thickness and gingival thickness

PURPOSE: The aim of this study was to determine the relationship between buccal bone thickness and gingival thickness by means of a noninvasive and relatively accurate digital registration method. METHODS: In 20 periodontally healthy subjects, cone-beam computed tomographic images and intraoral scanned files were obtained. Measurements of buccal bone thickness and gingival thickness at the central incisors, lateral incisors, and canines were performed at points 0–5 mm from the alveolar crest on the superimposed images. The Friedman test was used to compare buccal bone and gingival thickness for each depth between the 3 tooth types. Spearman's correlation coefficient was calculated to assess the correlation between buccal bone thickness and gingival thickness. RESULTS: Of the central incisors, 77% of all sites had a buccal thickness of 0.5–1.0 mm, and 23% had a thickness of 1.0–1.5 mm. Of the lateral incisors, 71% of sites demonstrated a buccal bone thickness <1.0 mm, as did 63% of the canine sites. For gingival thickness, the proportion of sites <1.0 mm was 88%, 82%, and 91% for the central incisors, lateral incisors, and canines, respectively. Significant differences were observed in gingival thickness at the alveolar crest level (G0) between the central incisors and canines (P=0.032) and between the central incisors and lateral incisors (P=0.013). At 1 mm inferior to the alveolar crest, a difference was found between the central incisors and canines (P=0.025). The lateral incisors and canines showed a significant difference for buccal bone thickness 5 mm under the alveolar crest (P=0.025). CONCLUSIONS: The gingiva and buccal bone of the anterior maxillary teeth were found to be relatively thin (<1 mm) overall. A tendency was found for gingival thickness to increase and bone thickness to decrease toward the root apex. Differences were found between teeth at some positions, although the correlation between buccal bone thickness and soft tissue thickness was generally not significant.

Efficacy of sonic-powered toothbrushes for plaque removal in patients with peri-implant mucositis

PURPOSE: The aim of this study was to evaluate the effectiveness of powered toothbrushes for plaque control in patients with peri-implant mucositis, in comparison with manual toothbrushes. METHODS: This randomized, prospective, controlled, clinical parallel study compared the efficacy of manual and powered toothbrushes for plaque control in implant restorations. Patients with bleeding on probing, no residual pocket depth (as indicated by a pocket probing depth > or = 5 mm), and no radiological peri-implant bone loss were eligible for this study. Patients were requested to complete a questionnaire describing their oral hygiene habits. The duration and frequency of tooth brushing were recorded by subjects in order to assess their compliance. Clinical parameters, including the modified plaque index (mPI), the modified sulcus bleeding index (mSBI), and clinical photographs (buccal and lingual views) were recorded at baseline and at one-month and two-month follow-up visits. RESULTS: Statistically significant differences between patients who used manual toothbrushes and those who used powered toothbrushes were found regarding the frequency of tooth brushing per day and the duration of brushing at one-month and two-month follow-up visits, while no statistically significant differences were found relating to other oral hygiene habits. A statistically significant difference in patient compliance for tooth brushing was found at one month, while no difference was found at two months. Statistically significant decreases in the mPI and the mSBI were observed in both groups from baseline to the one- and two-month follow-ups. The overall reduction of these parameters was not significantly different between the two groups, except for mPI reduction between baseline and one month of follow-up. CONCLUSIONS: Sonic-powered toothbrushes may be a useful device for plaque control in patients with peri-implant mucositis.

Evaluation of the correlation between insertion torque and primary stability of dental implants using a block bone test

PURPOSE: Implant stability at the time of surgery is crucial for the long-term success of dental implants. Primary stability is considered of paramount importance to achieve osseointegration. The purpose of the present study was to investigate the correlation between the insertion torque and primary stability of dental implants using artificial bone blocks with different bone densities and compositions to mimic different circumstances that are encountered in routine daily clinical settings. METHODS: In order to validate the objectives, various sized holes were made in bone blocks with different bone densities (#10, #20, #30, #40, and #50) using a surgical drill and insertion torque together with implant stability quotient (ISQ) values that were measured using the Osstell Mentor. The experimental groups under evaluation were subdivided into 5 subgroups according to the circumstances. RESULTS: In group 1, the mean insertion torque and ISQ values increased as the density of the bone blocks increased. For group 2, the mean insertion torque values decreased as the final drill size expanded, but this was not the case for the ISQ values. The mean insertion torque values in group 3 increased with the thickness of the cortical bone, and the same was true for the ISQ values. For group 4, the mean insertion torque values increased as the cancellous bone density increased, but the correlation with the ISQ values was weak. Finally, in group 5, the mean insertion torque decreased as the final drill size increased, but the correlation with the ISQ value was weak. CONCLUSIONS: Within the limitations of the study, it was concluded that primary stability does not simply depend on the insertion torque, but also on the bone quality.

Effects of immunosuppressants, FK506 and cyclosporin A, on the osteogenic differentiation of rat mesenchymal stem cells

PURPOSE: The purpose of this study was to investigate the effects of the immunosuppressants FK506 and cyclosporin A (CsA) on the osteogenic differentiation of rat mesenchymal stem cells (MSCs). METHODS: The effect of FK506 and CsA on rat MSCs was assessed in vitro. The MTT assay was used to determine the deleterious effect of immunosuppressants on stem cell proliferation at 1, 3, and 7 days. Alkaline phosphatase (ALP) activity was analyzed on days 3, 7, and 14. Alizarin red S staining was done on day 21 to check mineralization nodule formation. Real-time polymerase chain reaction (RT-PCR) was also performed to detect the expressions of bone tissue-specific genes on days 1 and 7. RESULTS: Cell proliferation was promoted more in the FK506 groups than the control or CsA groups on days 3 and 7. The FK506 groups showed increased ALP activity compared to the other groups during the experimental period. The ALP activity of the CsA groups did not differ from the control group in any of the assessments. Mineralization nodule formation was most prominent in the FK506 groups at 21 days. RT-PCR results of the FK506 groups showed that several bone-related genes-osteopontin, osteonectin, and type I collagen (Col-I)-were expressed more than the control in the beginning, but the intensity of expression decreased over time. Runx2 and Dlx5 gene expression were up-regulated on day 7. The effects of 50 nM CsA on osteonectin and Col-I were similar to those of the FK506 groups, but in the 500 nM CsA group, most of the genes were less expressed compared to the control. CONCLUSIONS: These results suggest that FK506 enhances the osteoblastic differentiation of rat MSCs. Therefore, FK506 might have a beneficial effect on bone regeneration when immunosuppressants are needed in xenogenic or allogenic stem cell transplantation to treat bone defects.

The biological effects of fibrin-binding synthetic oligopeptides derived from fibronectin on osteoblast-like cells

PURPOSE: The aim of this study was to investigate the effects of synthetic fibronectin (FN) fragments, including fibrin binding sites from amino-terminal FN fragments containing type I repeats 1 to 5, on osteoblast-like cell activity. METHODS: Oligopeptides ranging from 9 to 20 amino acids, designated FF1, FF3, and FF5, were synthesized by a solid-phase peptide synthesizing system, and we investigated the effects of these peptides on cell attachment and extent of mineralization using confocal microscopy, 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays, and Alizarin red S staining. RESULTS: FF3 and FF5 peptides increased the number of attached human osteoblastic cells, and FF3 administration led to prominent cell spreading. Mineralization was increased in FF3 and FF5 compared to FF1 and the untreated control. CONCLUSIONS: Taken together, it can be concluded that the fibrin-binding oligopeptides FF3 and FF5 enhanced cell attachment and mineralization on osteoblast-like cells. These results indicate that FF3 and FF5 have the potential to increase osteoblast-like cell activity. Performing an in vivo study may provide further possibilities for surface modification of biomimetic peptides to enhance osteogenesis, thus improving the regeneration of destroyed alveolar bone.

A Hyperactive Neutrophil Phenotype in Aggressive Periodontitis

Although neutrophils function in both defense and tissue destruction, their defensive roles have rarely been studied in association with periodontitis. We hypothesized that peripheral neutrophils are pre-activated in vivo in periodontitis and that hyperactive neutrophils would show enhanced phagocytic ability as well as an increased production of reactive oxygen species (ROS). Peripheral blood neutrophils from patients with aggressive periodontitis and age/gender-matched healthy subjects (10 pairs) were isolated. The levels of CD11b and CD64 expression on the neutrophils and the level of plasma endotoxin were determined by flow cytometry and a limulus amebocyte lysate test, respectively. In addition, neutrophils were subjected to a flow cytometric phagocytosis assay and luminol-enhanced chemiluminescence for non-opsonized Fusobacterium nucleatum in parallel. The neutrophilsfrom most patients expressed increased levels of both CD11b and CD64. In addition, the plasma from these patients tended to contain a higher level of endotoxin than the healthy controls. In contrast, no differences were found between the two groups with regard to phagocytosis or ROS generation by F. nucleatum. The ability to phagocytose F. nucleatum was found to positively correlate with the ability to produce ROS. In conclusion, peripheral neutrophils from patients with aggressive periodontitis are hyperactive but not hyperreactive to F. nucleatum.

Response of osteoblast-like cells cultured on zirconia to bone morphogenetic protein-2

PURPOSE: The aim of this study was to compare osteoblast behavior on zirconia and titanium under conditions cultured with bone morphogenetic protein-2. METHODS: MC3T3-E1 cells were cultured on sandblasted zirconia and sandblasted/etched titanium discs. At 24 hours after seeding MC3T3-E1, the demineralized bone matrix (DBM) gel alone and the DBM gel with bone morphogenetic protein-2 (BMP-2) were added to the culture medium. The surface topography was examined by confocal laser scanning microscopy. Cellular proliferation was measured at 1, 4, and 7 days after gel loading. Alkaline phosphatase activity was measured at 7 days after gel loading. The mRNA expression of ALPase, bone sialoprotein, type I collagen, runt-related transcription factor 2 (Runx-2), osteocalcin, and osterix were evaluated by real-time polymerase chain reaction at 4 days and 7 days. RESULTS: At 1, 4, and 7 days after loading the DBM gel alone and the DBM gel with BMP-2, cellular proliferation on the zirconia and titanium discs was similar and that of the groups cultured with the DBM gel alone and the DBM gel with BMP-2 was not significantly different, except for titanium with BMP-2 gel. ALPase activity was higher in the cells cultured with BMP-2 than in the other groups, but there was no difference between the zirconia and titanium. In ALPase, bone sialoprotein, osteocalcin, Runx-2 and osterix gene expression, that of cells on zirconia or titanium with BMP-2 gel was much more highly increased than titanium without gel at day 7. The gene expression level of cells cultured on zirconia with BMP-2 was higher than that on titanium with BMP-2 at day 7. CONCLUSIONS: The data in this study demonstrate that the osteoblastic cell attachment and proliferation of zirconia were comparable to those of titanium. With the stimulation of BMP-2, zirconia has a more pronounced effect on the proliferation and differentiation of the osteoblastic cells compared with titanium.

Initial adhesion of bone marrow stromal cells to various bone graft substitutes

PURPOSE: The aim of this study is to determine whether certain biomaterials have the potential to support cell attachment. After seeding bone marrow stromal cells onto the biomaterials, we investigated their responses to each material in vitro. METHODS: Rat bone marrow derived stromal cells were used. The biomaterials were deproteinized bovine bone mineral (DBBM), DBBM coated with fibronectin (FN), synthetic hydroxyapatite (HA), HA coated with FN, HA coated with beta-tricalcium phosphate (TCP), and pure beta-TCP. With confocal laser scanning microscopy, actin filaments and vinculin were observed after 6, 12, and 24 hours of cell seeding. The morphological features of cells on each biomaterial were observed using scanning electron microscopy at day 1 and 7. RESULTS: The cells on HA/FN and HA spread widely and showed better defined actin cytoskeletons than those on the other biomaterials. At the initial phase, FN seemed to have a favorable effect on cell adhesion. In DBBM, very few cells adhered to the surface. CONCLUSIONS: Within the limitations of this study, we can conclude that in contrast with DBBM not supporting cell attachment, HA provided a more favorable environment with respect to cell attachment.

Immunomodulatory effect of canine periodontal ligament stem cells on allogenic and xenogenic peripheral blood mononuclear cells

PURPOSE: The aim of this study was to investigate the immunomodulatory effects of canine periodontal ligament stem cells on allogenic and xenogenic immune cells in vitro. METHODS: Mixed cell cultures consisting of canine stem cells (periodontal ligament stem cells and bone marrow stem cells) and allogenic canine/xenogenic human peripheral blood mononuclear cells (PBMCs) were established following the addition of phytohemagglutinin. The proliferation of PBMCs was evaluated using the MTS assay. The cell division of PBMCs was analyzed using the CFSE assay. The apoptosis of PBMCs was assessed using the trypan blue uptake method. RESULTS: Periodontal ligament stem cells and bone marrow stem cells inhibited the proliferation of allogenic and xenogenic PBMCs. Both periodontal ligament stem cells and bone marrow stem cells suppressed the cell division of PBMCs despite the existence of a mitogen. No significant differences in the percentages of apoptotic PBMCs were found among the groups. CONCLUSIONS: Canine periodontal ligament stem cells have an immunomodulatory effect on allogenic and xenogenic PBMCs. This effect is not a product of apoptosis of PBMCs but is caused by the inhibition of cell division of PBMCs.

Biological effects of a porcine-derived collagen membrane on intrabony defects

PURPOSE: To prolong the degradation time of collagen membranes, various cross-linking techniques have been developed. For cross-linking, chemicals such as formaldehyde and glutaraldehyde are added to collagen membranes, but these chemicals could adversely affect surrounding tissues. The aim of this study is to evaluate the ability of porous non-chemical cross-linking porcine-derived collagen nanofibrous membrane to enhance bone and associated tissue regeneration in one-wall intrabony defects in beagle dogs. METHODS: The second and third mandibular premolars and the first molars of 2 adult beagles were extracted bilaterally and the extraction sites were allowed to heal for 10 weeks. One-wall intrabony defects were prepared bilaterally on the mesial and distal side of the fourth mandibular premolars. Among eight defects, four defects were not covered with membrane as controls and the other four defects were covered with membrane as the experimental group. The animals were sacrificed 10 weeks after surgery. RESULTS: Wound healing was generally uneventful. For all parameters evaluating bone regeneration, the experimental group showed significantly superior results compared to the control. In new bone height (NBh), the experimental group exhibited a greater mean value than the control (3.04 +/- 0.23 mm/1.57 +/- 0.59, P = 0.003). Also, in new bone area (NBa) and new bone volume (NBv), the experimental group showed superior results compared to the control (NBa, 34.48 +/- 10.21% vs. 5.09 +/- 5.76%, P = 0.014; and NBv, 28.04 +/- 12.96 vs. 1.55 +/- 0.57, P = 0.041). On the other hand, for parameters evaluating periodontal tissue regeneration, including junctional epithelium migration and new cementum height, there were no statistically significant differences between two groups. CONCLUSIONS: Within the limitations of this study, this collagen membrane enhanced bone regeneration at one-wall intrabony defects. On the other hand, no influence of this membrane on periodontal tissue regeneration could be ascertained in this study.