Recombinant human enamelin produced in Escherichia coli promotes mineralization in vitro.

BACKGROUND: Enamelin is an enamel matrix protein that plays an essential role in the formation of enamel, the most mineralized tissue in the human body. Previous studies using animal models and proteins from natural sources point to a key role of enamelin in promoting mineralization events during enamel formation. However, natural sources of enamelin are scarce and with the current study we therefore aimed to establish a simple microbial production method for recombinant human enamelin to support its use as a mineralization agent. RESULTS: In the study the 32 kDa fragment of human enamelin was successfully expressed in Escherichia coli and could be obtained using immobilized metal ion affinity chromatography purification (IMAC), dialysis, and lyophilization. This workflow resulted in a yield of approximately 10 mg enamelin per liter culture. Optimal conditions for IMAC purification were obtained using Ni2+ as the metal ion, and when including 30 mM imidazole during binding and washing steps. Furthermore, in vitro mineralization assays demonstrated that the recombinant enamelin could promote calcium phosphate mineralization at a concentration of 0.5 mg/ml. CONCLUSIONS: These findings address the scarcity of enamelin by facilitating its accessibility for further investigations into the mechanism of enamel formation and open new avenues for developing enamel-inspired mineralized biomaterials.

Treatment of intrabony periodontal defects in controlled diabetic patients with an enamel matrix derivative: a split-mouth randomized clinical trial.

OBJECTIVE: This split-mouth randomized controlled trial aimed to evaluate the effect of enamel matrix derivative (EMD) associated with a simplified papilla preservation flap (SPPF) compared to SPPF alone in the surgical treatment of intrabony defects (ID) in type 2 diabetic mellitus (T2DM) patients. MATERIAL AND METHODS: Thirteen patients with controlled T2DM presenting with ID in at least two quadrants were included. In each patient, the test site (TS) was treated with SPPF plus EMD, whereas the control site (CS) was treated only with SPPF. Prior to surgery and at 6 months after intervention, the following parameters were evaluated: clinical attachment level (CAL), probing pocket depth (PPD), and gingival recession (GR). RESULTS: The TS and CS demonstrated a mean CAL gain of 3.31 ± 0.96 mm and 1.61 ± 1.12 mm, and a PPD reduction from 8.15 ± 0.98 to 3.00 ± 0.57 mm and 7.53 ± 0.96 to 4.69 ± 0.63 mm after 6 months, respectively. In both sites, the mean CAL gain and PPD reduction improved significantly after 6 months compared to baseline; however, the improvement was higher in the TS (p < 0.001). CONCLUSIONS: Both surgical procedures presented with clinical improvements in controlled T2DM patients. However, the additional use of EMD showed enhanced clinical results after 6 months with regard to CAL gain and PPD reduction. CLINICAL RELEVANCE: This study showed a better PPD reduction and CAL gain when an EMD was applied in addition to SPPF. Therefore, EMD may be used to enhance clinical outcomes in periodontal ID of controlled T2DM patients.

Molecular Cloning of Mouse Homologue of Enamel Protein C4orf26 and Its Phosphorylation by FAM20C.

It is widely accepted that cellular processes are controlled by protein phosphorylation and has become increasingly clear that protein degradation, localization and conformation as well as protein-protein interaction are the examples of subsequent cellular events modulated by protein phosphorylation. Enamel matrix proteins belong to members of the secretory calcium binding phosphoprotein (SCPP) family clustered on chromosome 4q21, and most of the SCPP phosphoproteins have at least one S-X-E motifs (S; serine, X; any amino acid, E; glutamic acid). It has been reported that mutations in C4orf26 gene, located on chromosome 4q21, are associated with autosomal recessive type of Amelogenesis Imperfecta (AI), a hereditary condition that affects enamel formation/mineralization. The enamel phenotype observed in patients with C4orf26 mutations is hypomineralized and partially hypoplastic, indicating that C4orf26 protein may function at both secretory and maturation stages of amelogenesis. The previous in vitro study showed that the synthetic phosphorylated peptide based on C4orf26 protein sequence accelerates hydroxyapatite nucleation. Here we show the molecular cloning of Gm1045, mouse homologue of C4orf26, which has 2 splicing isoforms. Immunohistochemical analysis demonstrated that the immunolocalization of Gm1045 is mainly observed in enamel matrix in vivo. Our report is the first to show that FAM20C, the Golgi casein kinase, phosphorylates C4orf26 and Gm1045 in cell cultures. The extracellular localization of C4orf26/Gm1045 was regulated by FAM20C kinase activity. Thus, our data point out the biological importance of enamel matrix-kinase control of SCPP phosphoproteins and may have a broad impact on the regulation of amelogenesis and AI.

The enamel protein ODAM promotes mineralization in a collagen matrix.

Purpose/aim of the study: Odontogenic ameloblast-associated protein (ODAM) is predominantly expressed during the maturation stage of enamel formation and interacts strongly with amelotin (AMTN). AMTN is involved in enamel mineralization, but the effect of ODAM on mineralization has not been investigated. This study determined whether ODAM was able to induce hydroxyapatite (HA) mineralization in modified simulated body fluid (SBF) and in a collagen matrix in vitro. MATERIALS AND METHODS: To monitor the kinetics of calcium phosphate mineralization, recombinant human (rh) ODAM protein in SBF buffer was incubated at 37°C and a light-scattering assay was conducted at intervals. To investigate the nucleation of ODAM in collagen matrix, the ODAM-impregnated collagen hydrogel was incubated in SBF buffer for 24 hours. Bovine serum albumin (BSA) was used as negative control. Mineral deposits were visualized using electron microscopy. RESULTS: The presence of rh-ODAM protein in SBF resulted in higher light-scattering values after 18-24 hours. Calcium phosphate precipitates were observed on the surface of the ODAM-treated, but not BSA-treated collagen hydrogel after 24 hours in SBF. TEM and SAED analyses showed that these crystals consisted of needle-like HA. CONCLUSION: Similar to AMTN, ODAM is able to promote HA nucleation in a dose-dependent manner in SBF, and even outside of its biological context in vitro.

Using Enamel Matrix Derivative to Improve Treatment Efficacy in Periodontal Furcation Defects.

PURPOSE: Furcations are complicated periodontal defects. Untreated furcations lead to loss of the involved teeth and supporting tissues. It has been demonstrated that regenerative biomaterials are beneficial in reconstruction of the bone surrounding furcation-affected teeth. These biomaterials range from bone grafts and nonresorbable/resorbable barrier membranes to biologics that are able to trigger inactive regenerative processes in periodontal tissues. Selection of appropriate material(s) to treat furcations is challenging. The aim of this article is to provide a comparative outlook on different biomaterials applicable in regeneration of furcations with a focus on enamel matrix derivative (EMD). METHODS: Scientific databases including PubMed/MEDLINE, ScienceDirect, and EMBASE were searched, and 28 articles were found primarily for this specific study. Full texts were studied to identify relevant studies; 17 studies were excluded because of irrelevancy, while 11 main studies were ultimately selected. Other references have been used for general statements. RESULTS: EMD is a protein complex widely used in the regeneration of different periodontal defects. To assess the effects of EMD for treatment of root furcations, clinical studies involving EMD with and without barrier membranes and bone grafts were selected and compared. Briefly, this study reveals that when EMD is combined with open flap debridement (OFD), guided tissue regeneration (GTR), or bone grafting (BG), the amount of class II furcations converted to class I increases significantly. EMD also reduces tissue swelling and patient discomfort after treatment. CONCLUSIONS: This study provides evidence to find the best combination of biomaterials to treat furcation defects. The best results are obtained if EMD is combined with ß-TCP/HA alloplastic bone grafts.

Long-term results of periodontal regenerative therapy: A retrospective practice-based cohort study.

AIM: Evaluation of the long-term effectiveness of regenerative treatment of intra-bony defects in periodontal practice. MATERIAL AND METHODS: A total of 1,008 intra-bony defects in 176 patients were analysed after using collagen-added deproteinized bovine bone mineral (DBBMc) with or without collagen membrane (CM) or enamel matrix derivative (EMD). Defects were classified as one- and two-wall and as shallow (≤6 mm), moderate (>6 and <11 mm) and deep (≥11 mm). Radiographic bone level changes were evaluated after 1 year, 2 to 4 years and 5 to 10 years. RESULTS: Mean radiographic defect fill was 3.8 mm after 1 year and remained stable up to 10 years. Deep and moderate defects showed a higher degree of fill than shallow defects (53.3%, 49.2%, 42.9%). Tooth loss amounted to 2.6%, was dependent on initial defect size (1.2% shallow, 1.4% moderate, 5.7% deep defects) and occurred mainly due to endodontic reasons. CONCLUSIONS: Within the limits of the retrospective study design, the findings indicate that periodontal treatment using DBBMc with or without CM or EMD can lead to long-term defect reduction and tooth survival for up to 10 years in the setting of a periodontal practice.

Enamel matrix protein derivative and/or synthetic bone substitute for the treatment of mandibular class II buccal furcation defects. A 12-month randomized clinical trial.

OBJECTIVE: This study aims to clinically evaluate the treatment of mandibular class II furcation defects with enamel matrix derivative (EMD) and/or a bone substitute graft made of ß-tricalcium phosphate/hydroxyapatite (ßTCP/HA). MATERIALS AND METHODS: Forty-one patients, presenting a mandibular class II buccal furcation defect, probing pocket depth (PPD) ≥4 mm and bleeding on probing, were included. They were randomly assigned to the groups: 1-EMD (n = 13); 2-ßTCP/HA (n = 14); 3-EMD + ßTCP/HA (n = 14). Plaque index (PI), gingival index (GI), relative gingival margin position (RGMP), relative vertical and horizontal attachment level (RVCAL and RHCAL), and PPD were evaluated at baseline and 6 and 12 months. The mean horizontal clinical attachment level gain was considered the primary outcome variable. RESULTS: No significant intragroup differences were observed for RGMP, but significant changes were observed for RVCAL, RHCAL, and PPD for all groups (p < 0.05). After 12 months, the mean horizontal clinical attachment level gain was 2.77 ± 0.93 mm for EMD, 2.64 ± 0.93 mm for ßTCP/HA, and 2.93 ± 0.83 mm for EMD + ßTCP/HA, with no significant differences among the groups. At the end of the study, 85.3 % of the sites were partially closed; however, no complete closure was observed. CONCLUSION: EMD + ßTCP/HA does not provide a significant advantage when compared to the isolated approaches. All three tested treatments promote significant improvements and partial closure of class II buccal furcation defects. Based on its potential to induce periodontal regeneration, EMD may be considered an attractive option for this type of defect, but complete closure remains an unrealistic goal. CLINICAL RELEVANCE: The partial closure of buccal furcation defects can be achieved after the three tested approaches. However, the combined treatment does not provide a significant benefit when compared to the isolated approaches.

Five-year clinical results for treatment of intrabony defects with EMD, guided tissue regeneration and open-flap debridement: a case series.

BACKGROUND AND OBJECTIVE: Although regenerative periodontal surgery with EMD or guided tissue regeneration (GTR) has been shown to enhance periodontal regeneration, there are limited data on the long-term results following these treatment modalities. The purpose of the present study was to investigate the long-term clinical outcomes in intrabony defects following regenerative periodontal surgery with EMD or GTR compared with open-flap debridement (OFD). MATERIAL AND METHODS: Data from 40 subjects (44 teeth), with no history of smoking or systemic diseases that could interfere with periodontal disease and who received one of three surgical procedures (EMD, GTR or OFD) for two- or three-wall intrabony defects, were analyzed. Postoperative reduction in probing pocket depth, gain in clinical attachment level, gingival recession and percentage bone fill were compared at 1, 3 and 5 years. RESULTS: Reduction in probing pocket depth after GTR was significantly higher than after OFD at 1 and 3 years postoperatively, but there was no difference between the groups at 5 years. The gains in clinical attachment level for EMD (at 3 and 5 years) and for GTR (at 1, 3 and 5 years) were significantly greater than for OFD. Gingival recession after treatment with EMD and GTR showed a tendency toward positive results, whereas no such tendency was observed for OFD. Postoperative percentage bone fill for EMD and GTR was significantly greater than for OFD at 3 and 5 years. CONCLUSIONS: This is a retrospective study and an exploratory report with a high risk of bias. Within the limits of the current study, it may be concluded that superior gains in clinical attachment level and improved percentage bone fill can be obtained with EMD and GTR when compared with OFD, and these can be maintained over a period of 5 years.

Regulation of calcium phosphate formation by native amelogenins in vitro.

Our previous in vitro studies have shown that recombinant full-length porcine amelogenin rP172 can transiently stabilize amorphous calcium phosphate (ACP) and uniquely guide the formation of well-aligned bundles of hydroxyapatite (HA) crystals, as seen in the secretory stage of amelogenesis. This functional capacity is dependent on the hydrophilic C-terminal domain of full-length amelogenin. However, we have also found that native phosphorylated (single S-16 site) forms of full-length (P173) and C-terminal cleaved (P148) amelogenins can stabilize ACP for > 2 d and prevent HA formation. The present study was carried out to test the hypothesis that, at reduced concentrations, native full-length P173 also has the capacity to guide ordered HA formation. The effect of P148 and P173 concentrations (0.2-2.0 mg/ml) on the rate of spontaneous calcium phosphate precipitation was monitored via changes in solution pH, while mineral phases formed were assessed using TEM. At higher P173 concentrations (1.0-2.0 mg/ml), limited mineral formation occurred and only ACP nanoparticles were observed during a 48 h period. However, at 0.4 mg/ml P173, a predominance of organized bundles of linear, needle-like HA crystals were observed. At 0.2 mg/ml of P173, limited quantities of less organized HA crystals were found. Although P148 similarly stabilized ACP, it did not guide ordered HA formation, like P173. Hence, the establishment of the hierarchical enamel structure during secretory stage amelogenesis may be regulated by the partial removal of full-length amelogenin via MMP20 proteolysis, while predominant amelogenin degradation products, like P148, serve to prevent uncontrolled mineral formation.

Periodontal healing after application of enamel matrix derivative in surgical supra/infrabony periodontal defects in rats with streptozotocin-induced diabetes.

BACKGROUND AND OBJECTIVE: Epidemiologic and clinical studies have indicated that diabetes is a risk factor for periodontal disease progression and healing. The aim of the present study was to evaluate short-term healing after enamel matrix derivative (EMD) application in combined supra/infrabony periodontal defects in diabetic rats. MATERIAL AND METHODS: Thirty male Wistar rats were initially divided into two groups, one with streptozotocin-induced diabetes and another one with healthy (non-diabetic) animals. Bony defects were surgically created on the mesial root of the first maxillary molars. After root surface planing and EDTA conditioning, EMD was applied to the roots at one side of the maxillae, while those on the contralateral sides were left untreated. Animals were killed 3 wk after surgery, and block sections were prepared for histologic and histomorphometric analysis. RESULTS: There was statistically significant more gingival recession in diabetic animals than in non-diabetic animals. The length of the junctional epithelium was significantly shorter in the EMD-treated sites in both diabetic and normoglycemic rats. Sulcus depth and length of supracrestal soft connective tissue showed no statistically significant differences between groups. In all animals, new bone formation was observed. Although new bone occurred more frequently in healthy animals, the extent of new bone was not significantly different between groups. In none of the teeth, a layer of new cementum was detectable. EMD had no influence on bone or cementum regeneration. Adverse reactions such as excessive inflammation due to bacterial root colonization, ankylosis and bone fractures were exclusively observed in diabetic animals, irrespective of EMD treatment. CONCLUSION: Within the limits of the present study, it can be concluded that periodontal healing was impaired in streptozotocin-induced diabetic rats. EMD had no beneficial effects on new bone and cementum formation during short-term healing in this defect model and could not ameliorate the adverse effects in the systemically compromised animals.

Long-term clinical outcomes of periodontal regeneration with enamel matrix derivative: A retrospective cohort study with a mean follow-up of 10 years.

BACKGROUND: Despite the large body of evidence on the efficacy of enamel matrix derivative (EMD) in the treatment of periodontal intrabony defects, few studies reported long-term data (≥10-year). METHODS: Periodontal patients treated with regenerative surgery with EMD between 1999 and 2012 were invited to participate in a clinical examination. The following clinical parameters were recorded and compared at baseline (T0), 6 months after surgery (T1) and after at least 8 years of follow-up (T2): probing depth (PD), gingival recession (GR), clinical attachment level (CAL), plaque and bleeding scores. The primary outcome variable was CAL change. RESULTS: Forty-one patients with 75 treated teeth were available for analysis. Out of these, 68 (tooth survival rate: 90.7%) reached the latest follow-up with a mean observation period of 10.3 years (range: 8.0 to 21.3). The most frequent reason for tooth loss was recurrence of periodontal disease. Tooth survival curves showed a statistically significant difference between smokers and non-smokers (P = 0.028). Mean CAL changed from 8.43 ± 1.86 (T0) to 6.47 ± 1.70 (T1) (P < 0.001) and to 5.91 ± 1.83 (T2) (P < 0.001). At T1, a CAL gain of ≥3 mm was measured in 35% of the defects whereas at T2 it was detected in 51% of cases. CONCLUSIONS: Within their limitations, the present results have shown that in intrabony defects, the clinical improvements obtained following regenerative surgery with EMD can be maintained on a mean period of 10 years. Smoking status and maxillary molars were correlated with an increased risk for tooth and CAL loss, respectively.

Periodontal regenerative therapy in patients with type 2 diabetes using minimally invasive surgical technique with enamel matrix derivative under 3-year observation: A prospective cohort study.

BACKGROUND: Information regarding periodontal regenerative therapy in patients with diabetes mellitus (DM) is limited. This pilot study compared the regenerative outcomes of minimally invasive periodontal surgery using enamel matrix derivative (EMD) between DM and non-DM patients. METHODS: This prospective study included deep intrabony defects in patients with or without type 2 DM. Minimally invasive surgical technique (MIST) or modified MIST (M-MIST) using EMD, without bone graft materials, was performed. Periodontal examination and intraoral radiography were performed at baseline, 6 months, and 1 and 3 years after surgery. RESULTS: Ten sites of 10 subjects in the DM group, and 20 sites of 18 subjects in non-DM group were evaluated (mean age; 67.5 ± 7.6 and 63.1 ± 9.7, respectively). Probing depth significantly decreased from 7.1 ± 1.6 and 7.0 ± 1.3 mm to 2.2 ± 0.9 and 2.3 ± 1.1 mm at the 1-year examination in the DM and non-DM groups, respectively. Clinical attachment level (CAL) gain and radiographical defect fill at the 3-year examination were 3.8 ± 1.1 mm and 58.3% ± 10.4%, respectively, in the DM group, and 4.1 ± 1.1 mm and 65.5% ± 18.8%, respectively, in the non-DM group, showing no significant differences between the groups. Multiple regression analysis showed no significant association of CAL gain with DM or age after adjustments for relevant confounders. CONCLUSIONS: This is the first documented study of successful periodontal tissue regeneration in patients with DM. Minimally invasive surgery combined with EMD yielded significant clinical attachment gain and bone fill in the DM and non-DM groups at comparable levels.

Comparative evaluation of fluoride varnishes, self-assembling peptide-based remineralization agent, and enamel matrix protein derivative on artificial enamel remineralization in vitro.

BACKGROUND: One of the most unfavorable side effects of fixed orthodontic treatment is white spot lesions (WSLs). Although the most important approach is prevention of WSLs, it is also essential to evaluate the efficacy of the remineralization agents. However, there is no concurrence in the literature with respect to the remineralization process of these agents. The objective of the present study was to evaluate the effects of different fluoride varnishes, enamel matrix protein, and self-assembling peptide derivatives with varying chemical compositions on remineralization of artificially created WSLs in vitro using quantitative light-induced fluorescence (QLF). METHODS: Artificial WSLs were created on bovine enamel samples using acidic buffer solution (pH 5, 10 days). Specimens were randomly allocated to six groups (n = 10/group): (1) Emdogain (Straumann, Basel, Switzerland), (2) Curodont Repair (Credentis AG, Switzerland), (3) Duraphat (Colgate-Palmolive, New York, NY), (4) Clinpro XT (3 M ESPE, Pymble, New South Wales, Australia), (5) Enamel Pro Varnish (Premier Dental Products, PA, USA), and (6) control (untreated). The agents were applied to the WSLs according to the manufacturers' instructions. Fluorescence loss (ΔF), lesion area (area), and impact (ΔQ) values of enamel surfaces were quantified by QLF-D BiluminatorTM (Inspektor-Pro, Amsterdam, The Netherlands) at baseline and after 7, 14, and 21 days of application of the respective materials. RESULTS: ΔF value presented a significantly decreasing trend throughout the 21 days for all groups except the Duraphat and Enamel Pro varnishes. The changes between 14th and 21st days of the Clinpro XT varnish application were significantly higher than Emdogain, Curodont, and Enamel Pro. The Curodont group showed higher lesion area changes between the first and second week in comparison to the Emdogain, Clinpro XT, and Enamel Pro groups, whereas Clinpro XT assured the highest reduction from the second to the third week of the observation period. CONCLUSIONS: The fluorescence loss was significantly reduced with enamel matrix protein, self-assembling peptide, and light-curable fluoride varnishes in the analysis for 21 days. Curodont and Clinpro XT were effective in diminishing the fluorescence loss and lesion area compared to the Duraphat, Enamel Pro fluoride varnishes, and Emdogain in different time points.

Purification of Developing Enamel Matrix Proteins Using Preparative SDS-PAGE.

In this chapter we discuss the potential of preparative SDS-PAGE for use in purifying native developing enamel matrix proteins. We believe that the methodology has the potential to provide the relatively large-scale single-step purification of any enamel protein that can be resolved as a single band during analytical SDS-PAGE. Of course, a single band on analytical SDS-PAGE does not guarantee absolute purity as the band may be comprised of two or more proteins migrating at the same apparent molecular weight on the gel. Where absolute purity is required, the methodology can be used in conjunction with other techniques such as ion-exchange chromatography or reverse-phase chromatography. We do not see preparative SDS-PAGE replacing chromatographic methodologies but believe that it can provide another powerful tool to add to the battery of purification techniques already available to researchers in the field.

Loss of transforming growth factor-ß1 in epithelium cells affects enamel formation in mice.

OBJECTIVES: In order to understand the specific in vivo function of transforming growth factor-beta1 (TGF-ß1), we successfully established aTGF-ß1 deficient mouse model using a conditional knockout method. In the present study, we aimed to further understand the potential role of TGF-ß1 in enamel formation. DESIGN: Transgenic mice withoutTGF-ß1 in epithelial cells were generated. Scanning electron microscopy and micro-computed tomography analysis were used to detect the dental appearance, enamel microstructure and tooth density. Histological analysis was used to examine the residual organic matrix of enamel. Quantitative real-time polymerase chain reaction was used to analyze the expressions of enamel matrix proteins at the mRNA level. RESULTS: The enamel of mandibular molars and incisors inTGF-ß1 conditional knockout mice displayed severe attrition and lower density compared with the wild-type littermates. A slender microstructure of enamel rod was observed, and enamel matrix proteins were retained in the enamel space at the maturation stage in conditional knockout mice. Moreover, the expressions of enamel matrix protein-encoding genes, such as amelogenin (Amelx), ameloblastin (Ambn), Enamelin (Enam) and matrix metalloproteinase-20 (Mmp-20), were increased in enamel organs of conditional knockout mice. On the other hand, the expressions of Amelotin (Amtn), kallikrein-related peptidase-4 (Klk4), C4orf26 and WD repeat-containing protein 72 (Wdr72) were dramatically decreased at the transition and maturation stages. CONCLUSIONS: TGF-ß1 played an important role in enamel mineralization through decreasing synthesis ofAmelx, Ambn and Enam and increasing synthesis of Klk4, Amtn, Corf26 and Wdr72.

Comparative expression of the four enamel matrix protein genes, amelogenin, ameloblastin, enamelin and amelotin during amelogenesis in the lizard Anolis carolinensis.

BACKGROUND: In a recent study, we have demonstrated that amelotin (AMTN) gene structure and its expression during amelogenesis have changed during tetrapod evolution. Indeed, this gene is expressed throughout enamel matrix deposition and maturation in non-mammalian tetrapods, while in mammals its expression is restricted to the transition and maturation stages of amelogenesis. Previous studies of amelogenin (AMEL) gene expression in a lizard and a salamander have shown similar expression pattern to that in mammals, but to our knowledge there are no data regarding ameloblastin (AMBN) and enamelin (ENAM) expression in non-mammalian tetrapods. The present study aims to look at, and compare, the structure and expression of four enamel matrix protein genes, AMEL, AMBN, ENAM and AMTN during amelogenesis in the lizard Anolis carolinensis. RESULTS: We provide the full-length cDNA sequence of A. carolinensis AMEL and AMBN, and show for the first time the expression of ENAM and AMBN in a non-mammalian species. During amelogenesis in A. carolinensis, AMEL, AMBN and ENAM expression in ameloblasts is similar to that described in mammals. It is noteworthy that AMEL and AMBN expression is also found in odontoblasts. CONCLUSIONS: Our findings indicate that AMTN is the only enamel matrix protein gene that is differentially expressed in ameloblasts between mammals and sauropsids. Changes in AMTN structure and expression could be the key to explain the structural differences between mammalian and reptilian enamel, i.e. prismatic versus non-prismatic.

Periodontal treatment in a generalized severe chronic periodontitis patient: A case report with 7-year follow-up.

The aim of the periodontal treatment is to provide healthy and functional dentition all through a lifetime. In this report, periodontal treatment of a 42-year-old male patient with generalized severe chronic periodontitis is presented. He received initial periodontal treatment together with adjunctive antimicrobials. The devital teeth were endodontically treated, and free gingival grafts were placed at the inadequate keratinized tissue zones before regenerative surgery. Following the surgical treatment using enamel matrix derivatives and xenogenic bone graft combination, the patient was put on a strict recall program. After 12 months, favorable clinical and radiographical improvements were obtained. The 7-year maintenance of the present case with several initially hopeless teeth has been shown and discussed in this report. It can be concluded that optimum oral hygiene level as well as the positive cooperation of the patient enhanced the success of periodontal treatment results even in extremely severe periodontal destruction.

Novel apexification method in a non-vital tooth with an open apex: a case report.

Many materials have been introduced for apexification each having their own advantages and disadvantages. This case report aims to present a new method of apexification using a combination of deproteinized bovine bone mineral (DBBM) and enamel matrix derivative (EMD). After irrigating the canal of the maxillary right canine with 2.5 % sodium hypochlorite, a mixture of Bio-Oss and EMD was packed into the apical region for formation of an apical barrier and the canal was obturated by thermoplastic gutta percha technique with AH26 sealer; coronal seal was achieved by resin bonded composite. The size of the periapical lesion decreased significantly after 3, 6, 12 and 18-months. The patient had no radiographic signs or clinical symptoms at 24-month follow up and complete maturation of the apex and healing of the periapical bone were achieved.

Efficacy of enamel matrix derivative with freeze-dried bone allograft or demineralized freeze-dried bone allograft in intrabony defects: a randomized trial.

BACKGROUND: Promising clinical outcomes have been reported with the combination of enamel matrix derivative (EMD) and allograft materials. Direct comparison between EMD with a freeze-dried bone allograft (FDBA) and a demineralized FDBA (DFDBA) was evaluated in one case series study. To date, no randomized controlled trial has been reported. Therefore, a well-controlled randomized clinical trial was conducted to determine the relative efficacy of EMD/FDBA versus EMD/DFDBA when managing intrabony defects. METHODS: A randomized parallel trial was conducted in a private practice from April 2004 to October 2011. Sixty-nine patients were randomly assigned to one of three groups: EMD/FDBA (EF) intervention group (n = 23), EMD/DFDBA (ED) intervention group (n = 23), and EMD alone without graft material (E) as a negative control group (n = 23). All of the grafting material had minocycline added. Each patient had an intrabony defect. The primary outcomes were the absolute change in probing depth (PD) reduction and clinical attachment level (CAL) gain from baseline to 1- and 3-year follow-up. Intrabony defects were surgically treated with EMD/FDBA, EMD/DFDBA, or EMD alone. RESULTS: Sixty-seven patients (EF, n = 21: ED, n = 23; E, n = 23) were analyzed. All groups demonstrated significant improvement in PD reduction and CAL gain from baseline. The changes for PD were as follows (mm, 95% confidence interval [CI]): at 1 year: EF (4.4 mm, 4.0 to 4.7), ED (3.7 mm, 3.4 to 4.0), and E (control) (3.3 mm, 3.0 to 3.6); at 3 years: EF (4.4 mm, 4.1 to 4.8), ED (3.7 mm, 3.4 to 4.0), and E (3.1 mm, 2.8 to 3.4). The changes for CAL were as follows (mm, 95% CI): at 1 year: EF (4.1 mm, 3.8 to 4.5), ED (3.5 mm, 3.0 to 4.0), and E (3.0 mm, 2.5 to 3.6); at 3 years: EF (4.2 mm, 3.7 to 4.7), ED (3.6 mm, 3.1 to 4.1), and E (3.0 mm, 2.5 to 3.5). The intervention groups (EF and ED) showed better treatment outcomes than the control group at 1 and 3 years. Statistically, the two bone-graft groups were not significantly different from each other at 1 and 3 years. CONCLUSIONS: Both EMD/FDBA and EMD/DFDBA interventions resulted in greater soft tissue improvement at 1 and 3 years of follow-up compared to EMD alone. Both graft materials worked well in managing deep intrabony defects when combined with EMD.

Effect of enamel matrix derivative on bone formation around intraosseous titanium implant: An experimental study in canine model.

BACKGROUND: The purpose of this study was to perform a histological, histomorphometrical, and immunohistochemical evaluation of the effect of Enamel matrix derivative (EMD) on bone formation around titanium dental implant. MATERIALS AND METHODS: In this animal study, 12 implants (10 × 3.8 mm) were inserted in the tibia bone of three dogs of Iranian breed. Two implants were placed in each tibia with EMD only on the left side. The dogs were sacrificed 2, 4, and 6 weeks after implantation. Following decalcification of the implants' surrounding tissue and preparation of 4 µm thick sections, they were stained with hematoxylin and eosin (H and E) and immunohistochemical (IHC) stain for osteopontin (OPN) marker. Histomorphometric evaluation was performed via measurement of the percentage of the woven, lamellar, and total generated bone. Light microscopy osteoblastic intensity of OPN in osteoblasts and bone matrix was also evaluated Data were analyzed by Wilcoxon signed Ranks, and Mc Nemar tests. RESULTS: In both control and EMD-applied groups, bone formation was recognized around the implants at the 4(th) week postimplantation. The percentage of total generated bone in the test group was higher than the control group, although being not statistically significant (P value = 0.917). Osteoclasts exhibited significantly higher proliferation activity compared the control group when stimulated by EMD (P value = 0.027). On average, the staining intensity in osteoblasts and extracellular matrix of bone, in EMD-applied subjects was higher than those of the controls (P value = 0.167 and P value = 0.414, respectively). CONCLUSION: EMD enhanced bone formation around dental implants, but this increase was not significant.