Receptor activator of NF-kappaB ligand (RANKL) and CD 31 expressions in chronic periodontitis patients before and after surgery.

AIM OF THE STUDY: The present study investigated the hypothesis that upregulation of receptor activator of NF-kappaB ligand (RANKL) expression may be associated with upregulation of endothelial cell activitiy, which is common for periods of periodontal bone loss in chronic periodontitis. MATERIAL AND METHODS: RANKL expression of activated cells in soft tissue biopsies with CD 31 activity and the presence of RANKL and osteoprotegerin (OPG) in gingival crevicular fluid (GCF) were assessed in chronic periodontitis patients. Biopsies from 17 patients and 10 healthy subjects were immunohistochemically analyzed. Clinical measurements [plaque index (PI), the gingival index (GI), probing pocket depth (PPD), clinical attachment level (CAL) and gingival bleeding index (GBI)] and GCF samples were obtained before and after periodontal therapy. RESULTS: CD31 staining did not support the assumption that endothelium-like cells were predominantly associated with RANKL expression. CONCLUSIONS: RANKL-positive cells were widely distributed in periodontitis patients giving only partial support to the hypothesis that RANKL expression is restricted to T- and B-cell activation.

Are putative periodontal pathogens reliable diagnostic markers?

Periodontitis is one of the most common chronic inflammatory diseases. A number of putative bacterial pathogens have been associated with the disease and are used as diagnostic markers. In the present study, we compared the prevalence of oral bacterial species in the subgingival biofilm of generalized aggressive periodontitis (GAP) (n = 44) and chronic periodontitis (CP) (n = 46) patients with that of a periodontitis-resistant control group (PR) (n = 21). The control group consisted of subjects at least 65 years of age with only minimal or no periodontitis and no history of periodontal treatment. A total of 555 samples from 111 subjects were included in this study. The samples were analyzed by PCR of 16S rRNA gene fragments and subsequent dot blot hybridization using oligonucleotide probes specific for Aggregatibacter (Actinobacillus) actinomycetemcomitans, Porphyromonas gingivalis, Prevotella intermedia, Tannerella forsythia, a Treponema denticola-like phylogroup (Treponema phylogroup II), Treponema lecithinolyticum, Campylobacter rectus, Fusobacterium spp., and Fusobacterium nucleatum, as well as Capnocytophaga ochracea. Our data confirm a high prevalence of the putative periodontal pathogens P. gingivalis, P. intermedia, and T. forsythia in the periodontitis groups. However, these species were also frequently detected in the PR group. For most of the species tested, the prevalence was more associated with increased probing depth than with the subject group. T. lecithinolyticum was the only periodontopathogenic species showing significant differences both between GAP and CP patients and between GAP patients and PR subjects. C. ochracea was associated with the PR subjects, regardless of the probing depth. These results indicate that T. lecithinolyticum may be a diagnostic marker for GAP and C. ochracea for periodontal health. They also suggest that current presumptions of the association of specific bacteria with periodontal health and disease require further evaluation.

Ridge augmentation and maxillary sinus grafting with a biphasic calcium phosphate: histologic and histomorphometric observations.

OBJECTIVES: This retrospective study reports on histologic and histomorphometric observations performed on human biopsies harvested from sites augmented exclusively by biphasic calcium phosphate [BCP: hydroxyapatite (HA)/ tricalcium phosphate (TCP) 60/40] and healed for a minimum of 6 months. MATERIALS AND METHODS: Five patients benefited from three augmentation regimens (i.e.: one-stage lateral augmentation; two-stage lateral augmentation; and two-stage sinus grafting). In all patients, a degradable collagen membrane served as a cell-occlusive barrier. Core biopsies were obtained from lateral as from crestal aspects 6-10 months after augmentation surgeries. For histologic and histomorphometric evaluations, the non-decalcified tissue processing was performed. RESULTS: The histological examination of 11 biopsies showed graft particles frequently being bridged by the new bone, and a close contact between the graft particles and newly formed bone was seen in all samples. The mean percentages of newly formed bone, soft tissue compartment, and graft material were 38.8% (+/-5.89%), 41.75% (+/-6.08%), and 19.63% (+/-4.85%), respectively. Regarding bone-to-graft contact values, the percentage of bone coverage of graft particles for all biopsies ranged from 27.83% to 80.17%. The mean percentage of bone coverage was 55.39% (+/-13.03%). CONCLUSIONS: Data from the present study demonstrated osteoconductivity scores for the BCP material (HA/TCP 60/40) in patients resembling those previously shown for grafting materials of xenogenic and alloplastic origin.

Minimally invasive flap surgery and enamel matrix derivative in the treatment of localized aggressive periodontitis: case report.

Localized aggressive periodontitis is a distinct entity of periodontal disease and is characterized by deep vertical bony defects that typically affect the first molars and incisors of young patients. Therapy is usually aimed at reducing the pathogenic microflora through scaling and root planing and the administration of systemic antibiotics. However, conservative periodontal therapy may result in reparative wound healing with limited regeneration of the lost tissues. Periodontal surgery combined with enamel matrix derivative has been introduced as a method to promote regeneration of the lost periodontium and has been studied extensively in the treatment of chronic periodontitis. This case report describes the treatment of a 27-year-old patient displaying severe localized aggressive periodontitis with documented disease progression. After initial therapy consisting of scaling and root planing and systemic administration of amoxicillin and metronidazole, the vertical defects were treated by minimally invasive access flaps combined with application of enamel matrix derivative. Clinical, microbiologic, and radiographic findings are reported for up to 1.5 years after initial therapy, indicating good efficacy of the therapeutic strategy and stability of the treatment outcome.

Timing affects the clinical outcome of adjunctive systemic antibiotic therapy for generalized aggressive periodontitis.

BACKGROUND: Systemic antibiotics improve the outcome of scaling and root planing (SRP) in patients exhibiting severe periodontitis. This study evaluated the influence of timing of adjunctive systemic antibiotics in the sequence of periodontal therapy. METHODS: Two cohorts of patients with generalized aggressive periodontitis and treated by SRP, adjunctive antibiotics, and supportive periodontal therapy (SPT) were analyzed retrospectively. Cohort A (17 patients; 36 +/- 5 years of age) received systemic amoxicillin/metronidazole immediately after SRP ("immediate"); cohort B (17 patients; 36 +/- 4 years of age) received the same regimen 3 months after SRP, following SPT, including subgingival reinstrumentation ("late"). Clinical parameters, including probing depth (PD), relative attachment level (RAL), bleeding on probing (BOP), and suppuration, were recorded with a pressure-sensitive electronic probe at baseline and 3 and 6 months after SRP. RESULTS: Significant time*group interactions were found for all clinical parameters except BOP, i.e., timing of antibiotic therapy affected the course of clinical changes over time. Immediate antibiotic therapy produced significantly higher initial changes (0 to 3 months) in PD and RAL. Late antibiotic therapy at 3 months resulted in additional significant improvements in all clinical parameters between 3 and 6 months. In initially deep sites (baseline PD >6 mm), improvements in PD and RAL over 6 months were significantly higher with immediate antibiotic therapy compared to late antibiotic therapy. CONCLUSION: Within the limits of a retrospective analysis, these findings indicate that administration of amoxicillin/metronidazole immediately after initial SRP provides more PD reduction and RAL "gain" in initially deep sites than late administration at SPT with reinstrumentation after 3 months.

Influence of periodontal therapy on the regulation of soluble cell adhesion molecule expression in aggressive periodontitis patients.

BACKGROUND: Inflammatory periodontal disease is associated with an increased risk of cardiovascular disease. Circulating cell adhesion molecules (CAM) (intercellular adhesion molecule-1 [ICAM-1], vascular cell adhesion molecule-1 [VCAM-1], and E-selectin) have been suggested as potential candidate markers of endothelial dysfunction, which contribute to the pathogenesis of cardiovascular diseases. The regulation of CAM in subjects with severe periodontitis and the influence of periodontal intervention on systemic CAM levels are not clear. The aim of this study was to determine whether intensive periodontal therapy reduces serum levels of CAM in patients with generalized aggressive periodontitis. METHODS: Blood samples were collected at six treatment time points from 21 patients with previously untreated generalized aggressive periodontitis (mean age: 34.6 +/- 4.3 years). Patients received subgingival scaling and root planing and antibiotic therapy and were monitored over a 6-month recall period. Serum levels of soluble ICAM-1 (sICAM-1), VCAM-1 (sVCAM-1), and E-selectin (sE-selectin) were measured by enzyme-linked immunosorbent assay. RESULTS: sE-selectin plasma levels decreased significantly (P <0.01) during periodontal therapy. Mean plasma levels were 65.95 ng/ml before treatment and 44.71 ng/ml 6 months after antibiotic therapy. sICAM-1 and sVCAM-2 serum levels were unaffected by therapeutic intervention. CONCLUSIONS: Periodontal therapy reduces plasma sE-selectin levels. Whether this leads to a reduction in risk of future cardiovascular events in patients with aggressive periodontal disease warrants further studies.

Enamel matrix derivative induces connective tissue growth factor expression in human osteoblastic cells.

BACKGROUND: Enamel matrix derivative (EMD) stimulates the production of transforming growth factor-beta (TGF-beta), which has been suggested to play a role in mediating the effects of EMD in periodontal tissue regeneration. Connective tissue growth factor (CTGF) is a mediator of TGF-beta and promotes cell development. The interaction between EMD and CTGF is unknown. This study explored the effects of EMD on CTGF expression in human osteoblastic cells and whether the interaction is modulated by the TGF-beta signaling pathway. Also, the roles of CTGF in cell proliferation, cell cycle progression, and mineralized nodule formation of EMD-induced osteoblastic cultures were examined. METHODS: Human osteoblastic cells (Saos-2) were treated with 25 to 100 microg/ml EMD with or without the addition of TGF-beta inhibitor. CTGF mRNA expression was detected by reverse transcription-polymerase chain reaction (RT-PCR), and CTGF protein levels were assayed by Western blot analysis. In addition, cell cycle progression and DNA synthesis were determined by flow cytometry and 5-bromo-2'-deoxyuridine (BrdU) incorporation following EMD treatment with or without CTGF antibody. Mineralization was examined by alizarin red staining and quantified by elution with cetylpyridinium chloride. RESULTS: Western blot and RT-PCR analysis demonstrated a dose-dependent increase of CTGF expression by EMD. EMD-induced CTGF expression was reduced significantly in the presence of TGF-beta inhibitor. Cell cycle and BrdU analysis revealed an increase in cell proliferation following EMD treatment, which was due to an increase in the percentage of cells in the G2/M phase of the cell cycle. No significant effect was found when anti-CTGF antibody was added. Conversely, mineralization was inhibited significantly in EMD-treated cultures in the presence of anti-CTGF antibody. CONCLUSIONS: EMD stimulates CTGF expression, and the interaction is modulated via TGF-beta in osteoblastic cells. Also, CTGF affects EMD-induced osteoblastic mineralization but not cell proliferation. To our knowledge, these results provide novel insight into EMD-CTGF interaction, two biomodifiers that have therapeutic relevance to tissue engineering and regeneration.

In vitro attachment of osteoblasts on contaminated rough titanium surfaces treated by Er:YAG laser.

Microbial contamination of implant surfaces inhibits formation of new osseous tissues. Biocompatibility of sandblasted large grid (SLA) surface, after previous in vitro cocultivation with Porphyromonas gingivalis and concomitant Er:YAG laser irradiation of microorganisms, was tested by attachment of newly cultured osteoblasts. A total of 36 customized titanium cubes with SLA surface were placed into human osteoblast culture for 14 days. After removal of 1 control cube, 35 other cubes were contaminated with precultured P. gingivalis (ATCC33277) and incubated in broth medium for 1 week. Ablation was carried out on 32 cubes. Each side was treated for 23.5 s with a pulsed, water-cooled laser beam. After irradiation, cubes were again placed into fresh osteoblast culture for 2 weeks. One randomly selected single side per cube was analyzed by scanning electron microscope in 22 cubes. On other 10 cubes, vitality of attached cells was tested with ethidiumbromide staining by fluorescence microscopy. Three negative controls revealed constantly adherent P. gingivalis, and no osteoblasts were detectable after P. gingivalis contamination on the surfaces. Laser-treated specimens showed newly attached osteoblasts, extending over 50-80% of the surface. Positive control cube (without bacterial contamination) showed over 80% cell coverage of the surface. Vitality of widely stretched osteoblasts was confirmed by FITC staining. Our results indicate that Er:YAG laser was effective in removing P. gingivalis and cell compounds, offering an acceptable surface for new osteoblast attachment.

Moderate effect of enamel matrix derivative (Emdogain Gel) on Porphyromonas gingivalis growth in vitro.

OBJECTIVE: The aim of this study was to analyse the antibacterial effects of Emdogain Gel or its constituents on the growth of the suspected periodontopathogen Porphyromonas gingivalis. STUDY DESIGN: The effects of the proteins of enamel matrix derivative (EMD), the commercial product Emdogain Gel or its vehicle propylene glycol alginate (PGA) (Straumann, Switzerland) on P. gingivalis growth were determined by two methods: broth dilution assay (BDA) and agar diffusion assay (ADA). RESULTS: BDA-Emdogain Gel inhibited moderately the growth of P. gingivalis, whereas EMD showed no effect. The PGA vehicle inhibited the growth completely. ADA-Emdogain Gel resulted in some inhibition in growth but was not significantly different from control. EMD revealed no zone of inhibition. PGA demonstrated statistically significant zones of inhibition. CONCLUSION: Emdogain Gel shows moderate antibacterial activities against P. gingivalis. These properties seem to be due to the PGA component of the gel preparation.

Simvastatin promotes cell metabolism, proliferation, and osteoblastic differentiation in human periodontal ligament cells.

BACKGROUND: Simvastatin is one of the cholesterol lowering drugs. Recent studies demonstrated that it has a bone stimulatory effect. Periodontal ligament (PDL) cells are believed to play an important role in periodontal regeneration; that is, they may differentiate into specific cells which make cementum, bone, and attachment apparatus. It would be of interest whether simvastatin has a positive effect on PDL cells. Therefore, effects of simvastatin on cell proliferation and osteoblastic differentiation in PDL cells were analyzed. METHODS: Human PDL cells were cultured in monolayer with simvastatin for 24 and 72 hours and cell metabolism and proliferation were determined. To analyze osteoblastic differentiation, human PDL cells were cultured in organoid culture for 7, 14, and 21 days and alkaline phosphatase (ALP) activity, osteopontin (OPN), bone morphogenetic protein (BMP) -2, osteocalcin (OCN), and calcium contents were measured. They were co-treated by simvastatin and mevalonate. RESULTS: Simvastatin enhanced cell proliferation and metabolism dose-dependently after 24 hours. Simvastatin also stimulated ALP activity of human PDL cells dose-dependently, and maximum effect was obtained at the concentration of 10(8) M. In time dependent analysis, 10(8) M simvastatin stimulated ALP activity and osteopontin content after 7 days and calcium contents after 21 days. BMP-2 and OCN contents were not detected. Moreover this statin-enhanced ALP activity was abolished by mevalonate. CONCLUSION: These results suggest that at low concentration, simvastatin exhibits positive effect on proliferation and osteoblastic differentiation of human PDL cells, and these effects may be caused by the inhibition of the mevalonate pathway.

Coverage of Miller class I and II recession defects using enamel matrix proteins versus coronally advanced flap technique: a 2-year report.

BACKGROUND: The aim of this study was to evaluate a comparison of the coronally advanced flap procedure with or without the use of enamel matrix proteins in the treatment of recession defects. METHODS: This 2-year study was conducted as a blinded, split-mouth, placebo-controlled, and randomized design. Thirty patients from two dental schools with two paired buccal recession defects were chosen. Surgical recession coverage was performed as the coronally advanced flap technique. One site was additionally treated with derivative (EMD) and the other site with a placebo (propylene glycol alginate [PGA]). A blinded examiner assessed pre- and post-surgical measurements. Measurements comprised the height and width of the gingival recession, height of keratinized tissue, probing attachment level, probing depth, and alveolar bone level. RESULTS: Twenty-four months after therapy, both treatment modalities showed significant root coverage and probing attachment gain. The mean gingival recession decreased from 3.6 to 0.8 mm for the EMD-treated sites and from 3.8 to 1.4 mm for the control sites. However, this difference was not statistically significant (P = 0.122). Similarly, all other clinical parameters did not differ significantly in the between-group comparison except for the recession width (P = 0.027) and probing depth (P = 0.046) exhibiting higher reductions in the EMD group. Complete root coverage could be maintained over 2 years in 53% of the EMD versus merely 23% in the control group. A total of 47% of the treated recessions in the control group deteriorated again in the second year after therapy compared to 22% in the EMD group. CONCLUSION: Enamel matrix derivative seems to provide better long-term results.

Immediate substitution of central incisors with an unusual enamel paraplasia by a newly developed titanium implant: a case report.

The replacement of incisors with an unfavorable hard and soft tissue environment in the maxilla can be a challenging procedure in terms of esthetic outcome. The present report describes the replacement of two hopeless central incisors by newly developed titanium implants (TE Implant, Straumann) for immediate placement. This type of implant was chosen to compensate for the natural esthetics, which became compromised because of periodontitis and an unusual root anatomy. Both incisors presented with an atypical enamel paraplasia characterized by a circumferential enamel projection in apical direction. The consequence was a markedly reduced surface for periodontal attachment. Two weeks after implant placement, two acrylic resin crowns were cemented onto the new temporary titanium abutments. Five months later, the definitive prosthetic restoration was processed by porcelain pressed onto galvanic-cap crowns, which were cemented to standard wide-neck abutments. Control radiographs showed uneventful healing.

[Critical assessment of microbiological diagnostics in periodontal diseases with special focus on Porphyromonas gingivalis]. / Kritische Beurteilung mikrobiologischer Diagnostik bei marginaler Parodontitis unter besonderer Berüchsichtigung von Porphyomonas gingivalis.

Periodontitis is caused by an opportunistic infection with pathogenic microorganisms of the oral biofilm. In this paper, we discuss the usefulness of microbial diagnostics with respect to the differential diagnosis or the treatment approaches of periodontal diseases. Several diagnostic techniques, based on morphological, enzymatic, cultural, genetic or antigenetic properties have been established to analyze the microbial flora. Among the bacterial species some virulent genotypes of P. gingivalis play an important role in the etiology of periodontitis. Expression of fimbriae or different proteases have been identified as potential virulence factors of this gram negative anaerobic rod. To date a characterization of virulence of specific strains or a correlation between expression of different virulence factors and distinct periodontal conditions, however, is missing. Therefore, the importance of a routine identification of P. gingivalis still needs further evaluation.

The effect of cigarette smoking on gingival bleeding.

BACKGROUND: The purpose of this study was to investigate the dose-dependent effect of cigarette smoking upon gingival bleeding on probing (BOP) in a large representative sample of the United States population (National Health and Nutrition Examination Survey III). METHODS: Weighted multiple logistic regression was used to model bleeding on probing of 141,967 mesio-buccal sites in 12,385 individuals with complete case records on all covariates. Adjustments were made for age, gender, race/ethnicity, number of missing teeth, tooth type/jaw, root caries, full crown coverage, socioeconomic status (poverty/income ratio), and survey characteristics. The model stratified by presence of calculus (CALC) and increased probing depth (PD > or = 4 mm). Generalized estimating equations were used to account for dependence of sites within subjects. RESULTS: Smoking had a strong suppressive effect on gingival bleeding. The effect was strongest in heavier smokers (> 10 cigarettes/day) and smallest in former smokers. In healthy sites (no CALC, PD < or = 3 mm), the odds ratio (OR) of bleeding for sites in heavier smokers compared to never-smokers was 0.56 (95% CI: 0.45-0.70). Sites with CALC and/or PD > or = 4 mm were more likely to bleed in never-smokers (OR: 5.7; 95% CI: 4.3-7.6). This relationship was less evident among heavier smokers (OR: 1.3; 95% CI: 0.8-1.9). The effect of smoking did not differ between maxillary and mandibular molars, premolars, or incisors. CONCLUSION: Smoking exerts a strong, chronic, and dose-dependent suppressive effect on gingival bleeding on probing.

Effects of lipopolysaccharide extracted from Prevotella intermedia on bone formation and on the release of osteolytic mediators by fetal mouse osteoblasts in vitro.

Prevotella intermedia, a Gram-negative obligate anaerobic black-pigmented oral bacterium, belongs to a small group of microorganisms that is closely associated with the initiation of periodontal diseases. Lipopolysaccharide (LPS), an outer membrane component, is one of the main virulence factors of this bacterium. The aim of this study was to examine the effects of Prev. intermedia lipopolysaccharide, extracted by the hot-phenol-water method, on differentiation (alkaline phosphatase activity) and mineralisation (calcium incorporation) of fetal mouse calvarial cells in vitro and to determine the release of the important osteolytic factors nitric oxide, interleukin-6 (IL-6) and matrix metalloproteinases by these cells after treatment with different concentrations of Prev. intermedia lipopolysaccharide (0.2-25 microg/ml). By gelatin zymography, we also characterized the matrix metalloproteinases released by these osteoblasts. Treatment with Prev. intermedia lipopolysaccharide dose-dependently inhibited bone formation by reducing alkaline phosphatase activity and calcium incorporation and induced the release of nitric oxide, IL-6 and the latent proforms of MMP-2 and MMP-9 by fetal mouse osteoblasts in organoid culture. These results indicate that the lipopolysaccharide from Prev. intermedia not only participates in periodontal tissue destruction and alveolar bone resorption, but also inhibits bone formation.

Effects of enamel matrix derivative on proliferation and differentiation of primary osteoblasts.

OBJECTIVE: The aim of this study was to investigate the effects of enamel matrix derivative (EMD) on proliferation, protein synthesis, and mineralization in primary mouse osteoblasts. STUDY DESIGN: Osteoblasts were obtained from mouse calvaria by enzymatic digestion and grown in monolayer together with EMD (2-100 microg/ml). Metabolic activity and cell proliferation were determined by tetrazolium salt assay (MTT) and by 5-bromo-2'-deoxyuridine (BrdU) incorporation. For differentiation studies, a 3-dimensional organoid culture system was used. Osteoblastic differentiation was estimated by alkaline phosphatase (ALP) activity and calcium content. Collagen synthesis was assessed by [(3)H]-proline incorporation. Morphologic observations were made by electron microscopy. RESULTS: EMD treatments increased metabolic cell activity and BrdU incorporation. In the organoid cultures, ALP activity and calcium accumulation were enhanced by EMD treatment, but [(3)H]-proline incorporation was not. Morphologically, an increased deposition of mineralized nodules was found. CONCLUSIONS: EMD treatment enhanced cellular activities of primary osteoblasts and might support the regeneration of periodontal bony defects.

Periapical and periodontal healing after osseous grafting and guided tissue regeneration treatment of apicomarginal defects in periradicular surgery: results after 12 months.

OBJECTIVE: The aim of the present study was to evaluate the periapical and periodontal healing of apicomarginal defects 12 months after periradicular surgery and guided tissue regeneration in a series of consecutively treated patients. STUDY DESIGN: Patients with apicomarginal defects who were referred for periradicular surgery were included. Apicomarginal defects were grafted with Bio-Oss bone mineral and covered with a Bio-Gide membrane. Periodontal probing depths (PPDs) and relative attachment levels were measured preoperatively and 12 months postoperatively with a manual force-controlled probe. Periapical healing was assessed clinically and radiographically. RESULTS: Of the 23 defects in 22 patients for whom follow-up data were available, 19 were considered clinically and radiographically successful, 2 were doubtful, and 2 were failures. Overall, the baseline median PPD decreased from 9.0 mm to 3.0 mm, corresponding to a median relative attachment level gain of 2.8 mm. In the case of periodontic-endodontic lesions, the median baseline PPD decreased from 9.8 mm to 4.0 mm, corresponding to a median relative attachment level gain of 4.2 mm. Defects that involved a proximal root surface had a significantly higher residual PPD than did defects not involving a proximal root surface. CONCLUSIONS: Guided tissue regeneration treatment of apicomarginal defects yields good results in terms of periapical and periodontal healing after 12 months and should be considered as an adjunct to periradicular surgery in such cases.

Apicomarginal defects in periradicular surgery: classification and diagnostic aspects.

The prognosis of periradicular surgery is affected by the amount and location of bone loss. Apicomarginal defects are localized bony defects encompassing the total root length, and periradicular surgery on these teeth is associated with a lower success rate. This paper reviews the etiology, pathogenesis, and morphology of apicomarginal defects as encountered in periradicular surgery on the basis of a series of 24 consecutively treated patients. Periodontal data were recorded before surgery in all patients, and apicomarginal defects were diagnosed after flap reflection or, if applicable, apicoectomy. On the basis of the findings in these cases and on theoretic considerations, a classification system for apicomarginal defects with potential therapeutic and prognostic implications is presented and several criteria for differential diagnosis are discussed.

Stimulation of osteoblasts with Emdogain increases the expression of specific mineralization markers.

OBJECTIVE: The purpose of this study was to determine the effects of enamel matrix derivative on mRNA expression of markers related to periodontal healing. STUDY DESIGN: Murine osteoprogenitor cells (MC3T3-E1) were grown for 12 and 16 days in mineralization media and stimulated with 100 microg/mL Emdogain (EMD). Cell cultures treated with 2% and 10% fetal calf serum (FCS) served as control. The mRNA expression of bone sialoprotein (BSP), osteopontin (OPN), and runt-related protein 2 (Runx2) was analyzed by real-time polymerase chain reaction. One-way analysis of variance was used for statistical analysis. RESULTS: Stimulation with EMD significantly (P < .01) enhanced mRNA expression of BSP up to 13.9-fold and of OPN up to 3.2-fold at day 16 compared with the 2% FCS control. The expression of mRNA for transcription factor Runx2 was not significantly changed. CONCLUSION: The beneficial effects seen in periodontal regeneration after treatment with EMD may be related to an increase of the mineralization markers BSP and OPN at mRNA level.

Cytobiocompatibility of collagen and ePTFE membranes on osteoblast-like cells in vitro.

In guided bone regeneration (GBR), a semipermeable membrane is placed over an osseous defect to create a secluded environment in which bone formation can proceed without ingrowth of connective tissue cells from the overlaying soft tissue. Although the cell-occlusive property of GBR membranes appear to be essential to new bone formation, the role of transmembrane tissue fluid diffusion is not known. The objective of this study was to evaluate the degree to which diffusion across commonly used GBR membranes can support functional properties of osteoblast-like cells in vitro. Cells from an established osteoblast-like line (SAOS-2) were cultured on membranes of cross-linked collagen, noncross linked collagen, and ePTFE. The membranes rested on metal grids which allowed the membranes to lightly contact the surface of the culture medium. As a control, cells were directly plated and cultured in control wells. At days 7 and 21, cells were harvested by scraping the membranes or culture wells and analyzed for expression of alkaline phosphatase (ALP), core binding factor 1 (cbfa-1), bone sialoprotein-2 (BSP-2), and osteocalcin (OC). Expression was determined by quantitative real-time PCR. Glucose-6-phosphate dehydrogenase (G6PD) served as a reference gene. The membranes were examined by transmission light microscopy. RT-PCR revealed up-regulation of ALP of up to 60-fold and of cbfa-1 and BSP of up to threefold relative to G6PDH. Expression of OC was less then onefold. The expression profile for each of the four genes tested demonstrated small variations among cells grown on different membranes. Microscopic observations revealed remnants of undisrupted osteoblast-like cells attached to both collagen membranes. Cell morphology and spatial arrangement indicated that vitality was maintained. Diffusion through the three membranes evaluated in this study was sufficient to support osteoblast-like cell differentiation.