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Background: The study was designed to explore the enhanced impact of nano-hydroxyapatite and emdogain on the survival and osteogenic/odontogenic differentiation of human stem cells isolated from the apical papilla (hSCAPs)., Materials and Methods: In this in vitro trial, hSCAPS obtained from intact impacted immature third molars were confirmed to have characteristic cell surface markers, then exposed to nanohydroxyapatite, emdogain, and nanohydroxyapatite coated with emdogain for durations of 1-3 days. The survival of apical papilla stem cells was measured using a methyl thiazolyl tetrazolium assay. The quantitative reverse transcription polymerase chain reaction, alkaline phosphatase activity (ALP) and Alizarin red staining were used to evaluate osteogenic-odontogenic differentiation. Analysis of the data was done using one-way ANOVA, t-test, and Mann-Whitney test (α = 0.05)., Results: At 1-3 days, emdogain exhibited no significant impact on the survival of human stem cells from the apical papilla. In contrast, nanohydroxyapatite (α > 0.05) and nanohydroxyapatite coated with emdogain demonstrated a notable reduction in cell survival compared to the control group (α < 0.05). The expression of dentin sialophosphoprotein, dentin matrix protein 1, and bone sialoprotein genes demonstrated a notable increase in the group treated with nanohydroxyapatite coated with emdogain compared to the other groups (α < 0.05), and furthermore, this group exhibited more pronounced mineralized nodules than the other experimental groups., Conclusion: In contrast to nanohydroxyapatite, Emdogain did not demonstrate a detrimental effect on the survival of hSCAPs. Nanohydroxyapatite, emdogain, and nanohydroxyapatite coated with emdogain increased osteogenic/odontogenic differentiation of hSCAPs., Competing Interests: Declarations. Competing interest: The authors do not have any relevant financial or non-financial interests to declare. The authors do not hold any financial or proprietary interests in the material discussed within this article. Ethical approval and consent to participate: The study was conducted according to the ISSCR Guidelines for Stem Cell Research and Clinical Translation and was given ethical clearance by the ethics committee of Hamadan University of Medical Sciences (IR.UMSHA.REC.1398.853). The procedure of the study was carried out in accordance with the ethical standards set out in the 1964 Helsinki Declaration and its subsequent amendments. Consent for publication: Not applicable. Informed consent: The trial participant received a comprehensive description of the procedures entailed. Prior to the tooth extraction procedure at the Department of Oral and Maxillofacial Surgery, Hamadan University of Medical Sciences, the participant provided informed consent by signing a form specifically for the use of the extracted teeth in research., (© 2025. The Author(s), under exclusive licence to Springer Nature B.V.)

Objective: The study evaluates three biologically active substances with known bone-inductive potential on previously decontaminated titanium (Ti) discs., Material and Methods: Rough and smooth Ti surfaces were contaminated with a multispecies biofilm and cleaned with a chitosan brush. Discs were treated either with injectable-platelet-rich fibrin (i-PRF), advanced platelet-rich fibrin (A-PRF+), or enamel matrix derivatives (EMDs) before osteoblast seeding., Results: Biocompatibility, adhesion, migration, and gene expression of runt-related transcription factor 2 (RUNX2), collagen Type I Alpha 2 (COL1a2), alkaline phosphatase (ALP), osteocalcin (OC), and osteonectin (ON) were performed. All the tested biologic agents similarly increased cell viability. Specifically, osteoblasts seeded over i-PRF and EMD-treated surfaces showed improvement in adhesion and migration and significantly increased ALP, OC, ON, RUNX-2, and COL1a2 mRNA levels up to 2.8 fold (p < 0.05) with no differences between Ti surfaces., Conclusions: i-PRF and EMD possess beneficial bioactive properties that enhance tissue healing and promote regeneration on thoroughly sterilized surfaces. Biologically active materials may hold the potential to influence the process of implant re-osseointegration, which warrants more research since sterilization of the affected surfaces under clinical conditions is still not reliably possible and remains one of the greatest challenges., (© 2024 The Author(s). Clinical Implant Dentistry and Related Research published by Wiley Periodicals LLC.)

Purpose: To evaluate the clinical and radiographic results of simultaneous implant placement using transcrestal sinus floor elevation (TSFE) with and without enamel matrix derivative (EMD) application., Materials and Methods: Twenty-four patients were randomly assigned into two groups: The EMD&#43;TSFE group (n &#61; 13 patients, 20 implants) received TSFE with EMD application, and the TSFE group (n &#61; 11 patients, 20 implants) received TSFE without EMD application. The patients were recalled at 3 (T3) and 12 (T12) months postsurgery. The residual bone height (RBH), implant protrusion length (IPL), peri-implant sinus bone level (SBL), endo-sinus bone gain (ESBG), and implant stability (ISQ) were measured. Multivariate regressions were performed for the groups., Results: At T3, the ESBG was 3.72 ± 0.85 mm in the EMD&#43;TSFE group and 3.10 ± 0.05 mm in the TSFE group, and there were statistically significant differences (P < .05). However, there were no statistically significant differences in ESBG at T12 between the groups (P > .05). ISQ values did not show a statistical difference between the groups at T1 and T3, but at T3 in the TSFE&#43;EMD group, there was a statistical increase in the intragroup evaluation compared to the TSFE group., Conclusions: The use of EMD in TSFE procedures is effective in new bone formation at the apical part of the implant during the early healing period, but in the long term, no significant difference was shown between cases in which EMD was or was not used in terms of new bone formation and primary and secondary stabilization.

Aim: This study compared the quality and quantity of newly formed bone in rabbits' critical-sized calvarial defects filled with enamel matrix derivative (EMD) combined with freeze-dried bone allograft (FDBA) vs FDBA alone., Materials and Methods: A total of 24 adult male white New Zealand rabbits were included. In each rabbit, three bone defects with a diameter of 8 mm were created on the calvarium bone; the first defect was left untreated, while the second was filled with FDBA, and the third was filled with EMD + FDBA. Twelve rabbits were randomly euthanized after a month, and the remaining 2 month postsurgery. Bone sections were histologically evaluated by hematoxylin and eosin and vascular endothelial growth factor (VEGF), alkaline phosphatase (ALP), osteoprotegerin (OPG), and receptor activator of NF-kappaB (RANK) immune-histochemical staining., Results: An improvement in the newly formed bone percentage was found in the defects filled with EMD + FDBA in comparison with FDBA and control defects at 1 month and 2 months postsurgery. Additionally, the expression of VEGF, ALP, OPG, and RANK showed highly significant differences in the defects filled with EMD + FDBA compared to the FDBA and control ones at 1 month postsurgery ( p = 0.001). Meanwhile, VEGF and ALP expression showed a significant decrease in defects filled with EMD + FDBA compared to the FDBA and control ones ( p = 0.001), while OPG and RANK expression showed non-significant differences between treated groups at 2 months postsurgery., Conclusion: Enamel matrix derivative combined with FDBA has a synergistic effect on bone formation and graft substitution. This combination accelerates the expression of VEGF, ALP, OPG, and RANK., Clinical Significance: The combination of EMD and FDBA accelerates and ameliorates the quality of newly formed bone, aiding in maxillofacial reconstruction. How to cite this article: Zakri RN, Grawish ME, Mowafey B, et al. Impact of Freeze-dried Corticocancellous Bone Allograft Combined with Enamel Matrix Derivative in the Treatment of Critical-sized Calvarial Bone Defects: An Animal Study. J Contemp Dent Pract 2024;25(5):424-431.

Objectives: To investigate the function of enamel matrix derivative (EMD)-liquid compared to EMD-gel (original Emdogain® with polyglycolic acid-carrier) in inducing soft tissue regeneration using a rat dorsal model., Material and Methods: Four subcutaneous pouches were created through dorsal skin incisions in 18 female Wistar rats and randomly allocated to the following groups: (1) sterile saline + non-crosslinked collagen matrix (CM), (2) EMD-gel + CM, and (3) EMD-liquid + CM. After 2 and 4 weeks of healing, the specimens were harvested and stained with Goldner's trichrome, hematoxylin and eosin, and were immunohistochemically stained with an anti-CD31 antibody., Results: The EMD-liquid group showed the thickest connective tissue compared to the other groups, with statistical significance both at 2 (p < 0.001) and 4 weeks (p = 0.011 and 0.023, respectively). The number of multinucleated giant cells was not significantly different among the groups for both periods. Moreover, there was a tendency to have more blood vessels over a longer period, and the highest number of blood vessels was observed in the EMD-liquid group at 4 weeks (p = 0.009 and 0036, respectively)., Conclusion: EMD-liquid-treated CM is advantageous compared to using CM alone or EMD-gel-treated CM, owing to the histomorphometric results that show significantly increased soft tissue thickness and number of blood vessels when EMD-liquid was pre-primed to CM., Clinical Relevance: EMD with a liquid carrier may be an appropriate biologic supplement to provide cell-inducing properties to the CM scaffold and is clinically more beneficial for phenotype modification therapy than CM only and EMD-gel-treated CM., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

The aim of this in vivo study was to investigate the effect of occlusal hypofunction on alveolar bone healing in the absence or presence of an enamel matrix derivative (EMD). A standardized fenestration defect over the root of the mandibular first molar in 15 Wistar rats was created. Occlusal hypofunction was induced by extraction of the antagonist. Regenerative therapy was performed by applying EMD to the fenestration defect. The following three groups were established: (a) normal occlusion without EMD treatment, (b) occlusal hypofunction without EMD treatment, and (c) occlusal hypofunction with EMD treatment. After four weeks, all animals were sacrificed, and histological (hematoxylin and eosin, tartrate-resistant acid phosphatase) as well as immunohistochemical analyses (periostin, osteopontin, osteocalcin) were performed. The occlusal hypofunction group showed delayed bone regeneration compared to the group with normal occlusion. The application of EMD could partially, but not completely, compensate for the inhibitory effects of occlusal hypofunction on bone healing, as evidenced by hematoxylin and eosin and immunohistochemistry for the aforementioned molecules. Our results suggest that normal occlusal loading, but not occlusal hypofunction, is beneficial to alveolar bone healing. Adequate occlusal loading appears to be as advantageous for alveolar bone healing as the regenerative potential of EMD.

Objectives: The present study was performed to compare the effectiveness of Ankaferd Blood Stopper ® (ABS) with enamel matrix derivatives (EMD) for treating fenestration defects in rats., Materials and Methods: Forty-eight male Wistar rats were randomly divided into six groups (each n = 8). Fenestration defects were created in all rats, to which ABS, EMD, or saline (S) was then applied. The rats were grouped and sacrificed at one of two different time points, as follows: ABS-10-group, ABS-treatment/sacrifice on day 10; EMD-10-group, EMD-treatment/sacrifice on day 10; S-10-group, S-treatment/sacrifice on day 10; ABS-38-group, ABS-treatment/sacrifice on day 38; EMD-38-group, EMD-treatment/sacrifice on day 38; and S-38-group, S-treatment/sacrifice on day 38. Then, histomorphometric analysis including measurements of new bone area (NBA) and new bone ratio (NBR), and immunohistochemical analysis including the determination of osteopontin (OPN) and type-III-collagen (C-III) expression were performed., Results: The NBA and NBR were significantly higher in the ABS-10-group and EMD-10-group compared to the S-10-group (p < .05), and in the EMD-38-group compared to the S-38-group (p < .05). The levels of C-III and OPN immunoreactivity were significantly higher in the ABS-10-group compared to the S-10-group (p < .017)., Conclusions: The results of this study suggested that ABS can promote early periodontal regeneration, although its efficacy seems to decrease over time., (© 2021 Wiley Periodicals LLC.)

Enamel matrix derivative (EMD) prepared from extracted porcine fetal tooth material can support the regrow of periodontal tissues. Previous findings suggest that EMD has anti-inflammatory properties and TGF-β activity in vitro. However, the anti-inflammatory activity of EMD is mediated via TGF-β has not been considered. To this aim, we first established a bioassay to confirm the anti-inflammatory activity of EMD. The bioassay was based on the RAW 264.7 macrophage cell line and proven with primary macrophages where EMD significantly reduced the forced expression of IL-6. We then confirmed the presence of TGF-β1 in EMD by immunoassay and by provoking the Smad2/3 nuclear translocation in RAW 264.7 macrophages. Next, we took advantage of the TGF-β receptor type I kinase-inhibitor SB431542 to block the respective signalling pathway. SB431542 reversed the anti-inflammatory activity of EMD and TGF-β in a bioassay when IL-6 and CXCL2 expression was driven by the LPS stimulation of RAW 264.7 macrophages. This central observation was supported by showing that SB431542 reversed the anti-inflammatory activity of EMD using IL-1β and TNF-α-stimulated ST2 bone marrow stromal cells. Together, these findings implicate that the TGF-β activity mediates at least part of the anti-inflammatory activity of EMD in vitro.

Background: Diabetes involves metabolic disorders in various tissues via hyperglycemia-induced oxidative stress. This study aimed to investigate the antioxidative effect of enamel matrix derivative (EMD) on periodontal regeneration in diabetes., Methods: Twenty-two rats were equally divided into streptozotocin (STZ)-induced diabetes or control group. Two months after induction of hyperglycemia, systemic oxidative stress was measured using urinary 8-hydroxy-2'-deoxyguanosine. EMD or saline was applied into the intrabony defects created in the bilateral maxillary molar. mRNA expressions of inflammatory and oxidative stress markers were quantified (n = 6). Histometric analyses and immunohistochemistry of superoxide dismutase-1 (SOD-1) were performed 7 days postoperatively (n = 5). For in vitro experiments, the bone marrow-derived mesenchymal stem cells were isolated from rat femur and cultured in a high glucose (HG) or control medium. Reactive oxygen species (ROS) measurement and alizarin red staining were performed with/without EMD., Results: Systemic oxidative stress was significantly higher in the diabetic group. The connective tissue attachment and cementum formation were significantly increased at EMD-treated sites in both diabetic and non-diabetic groups. The expression of nicotinamide adenine dinucleotide phosphate oxidase two and four was significantly lower at EMD-treated sites than at EMD-untreated sites in both diabetic and non-diabetic rats. Immunohistochemistry showed significantly higher SOD-1 expression at the EMD-treated site. In vitro, HG culture had significantly higher ROS production compared with control, which was downregulated by EMD. EMD treatment significantly recovered the impaired calcification in HG., Conclusion: EMD promoted early-phase wound healing and periodontal tissue regeneration in the surgically created bony defect of STZ-induced diabetic rat by suppressing hyperglycemia-induced oxidative stress., (© 2021 American Academy of Periodontology.)

Background : Pyroptosis is a caspase-dependent catabolic process relevant to periodontal disorders for which inflammation is central to the pathophysiology of the disease. Although enamel matrix derivative (EMD) has been applied to support periodontal regeneration, its capacity to modulate the expression of pyroptosis-related genes remains unknown. Considering EMD has anti-inflammatory properties and pyroptosis is linked to the activation of the inflammasome in chronic periodontitis, the question arises whether EMD could reduce pyroptosis signalling. Methods : To answer this question, primary macrophages obtained from murine bone marrow and RAW 264.7 macrophages were primed with EMD before being challenged by lipopolysaccharide (LPS). Cells were then analysed for pyroptosis-signalling components by gene expression analyses, interleukin-1β (IL-1β) immunoassay, and the detection of caspase-1 (CAS1). The release of mitochondrial reactive oxygen species (ROS) was also detected. Results : We report here that EMD, like the inflammasome (NLRP3) and CAS1 specific inhibitors-MCC950 and Ac-YVAD-cmk, respectively-lowered the LPS-induced expression of NLRP3 in primary macrophages (EMD: p = 0.0232; MCC950: p = 0.0426; Ac-YVAD-cmk: p = 0.0317). EMD further reduced the LPS-induced expression of NLRP3 in RAW 264.7 cells ( p = 0.0043). There was also a reduction in CAS1 and IL-1β in RAW 264.7 macrophages on the transcriptional level ( p = 0.0598; p = 0.0283; respectively), in IL-1β protein release ( p = 0.0313), and CAS1 activity. Consistently, EMD, like MCC950 and Ac-YVAD-cmk, diminished the ROS release in activated RAW 264.7 cells. In ST2 murine mesenchymal cells, EMD could not be tested because LPS, saliva, and IL-1β + TNF-α failed to provoke pyroptosis signalling. Conclusion : These findings suggest that EMD is capable of dampening the expression of pyroptosis-related genes in macrophages.

Objectives: This parallel, randomized controlled clinical trial evaluated the influence of bone substitutes (BS) on the efficacy of the non-incised papillae surgical approach (NIPSA) with enamel matrix derivate (EMD) in resolving deep, isolated, combined non-contained intrabony and supra-alveolar periodontal defects, preserving the soft tissue., Material and Methods: Twenty-four patients were randomized to treatment with NIPSA and EMD or NIPSA plus EMD and BS. Bleeding on probing (BoP), interproximal clinical attachment level (CAL), interproximal probing depth (PD), recession (REC), location of the tip of the papilla (TP), and width of the keratinized tissue (KT) were evaluated before surgery and at 1 year post-surgery (primary outcomes). Wound closure was assessed at 1 week post-surgery, and supra-alveolar attachment gain (SUPRA-AG) was recorded at 1 year post-surgery., Results: At 1 week, 87.5% of cases registered complete wound closure and there were no cases of necrosis, without differences between groups (p > .05). At 1 year, all cases showed negative BoP. A significant PD reduction (NIPSA + EMD 8.25 ± 2.70 mm vs. NIPSA + EMD + BS 6.83 ± 0.81 mm) and CAL gain (NIPSA + EMD 8.33 ± 2.74 mm vs. NIPSA + EMD + BS 7.08 ± 2.68 mm) were observed (p < .001) in both groups, without significant between-group differences (p > .05). The residual PD was < 5 mm in all defects (NIPSA + EMD 2.50 ± 0.67 mm vs. NIPSA + EMD + BS 2.67 ± 0.78 mm). Soft tissues were preserved without significant between-group differences (REC: NIPSA + EMD 0.25 ± 0.45 mm vs. NIPSA + EMD + BS 0.17 ± 0.58 mm, p > .05; KT: 0.00 ± 0.43 mm vs. 0.08 ± 0.67 mm, p > .05). There were improvements in the papilla in both groups (TP: NIPSA + EMD 0.33 ± 0.49 mm vs. NIPSA + EMD + BS 0.45 ± 0.52 mm, p > .05), which was only significant in the NIPSA EMD + BS group (0.45 ± 0.52 mm; p < .05). In both groups, CAL gain was recorded in the supra-alveolar component, showing full resolution of the intrabony component of the defect in all cases (SUPRA-AG: NIPSA + EMD 1.83 ± 1.11 mm vs. NIPSA + EMD + BS 2.00 ± 1.76 mm, p > .05)., Conclusions: NIPSA and EMD with or without BS seem to be a valid surgical approach in the treatment of isolated, deep non-contained periodontal defects. In our study, both treatments resulted in significant PD reduction and CAL gain, that extended in the supra-alveolar component, without differences with the use of BS. Both treatments resulted in soft tissue preservation. However, the addition of BS may improve interdental papillary tissue., Clinical Relevance: NIPSA, with or without bone substitutes, resulted in significant periodontal improvement, with soft tissue preservation in isolated, deep non-contained periodontal defects. The application of bone substitutes may provide interproximal soft tissue gain., Clinical Trial Registration: Clinicaltrials.gov: NCT04712630., (© 2021. The Author(s).)