A Comparison Between Three Types of Scaffolds for Pulp Regeneration: A Histological Study on Dogs.

OBJECTIVES: This study aims to compare the application of three types of normal scaffolds-native chitosan, enzymatically modified chitosan, and blood clot (BC)-on pulp regeneration in the teeth of experimental dogs through histological examination, to determine the quantity and type of new tissues formed within the root canal. MATERIALS AND METHODS: The research sample consisted of 32 root canals from 20 premolars of two male local experimental dogs. The sample was randomly divided into a control group, in which no intervention was performed on the teeth, and three experimental groups based on the type of scaffold used: the BC group, the native chitosan combined with BC (NCS + BC) group, and the enzymatically modified chitosan combined with BC (EMCS + BC) group. Mechanical and chemical cleaning of the canals was performed, followed by the application of the studied scaffolds within the root canals. After 3 months, the teeth were extracted and prepared for histological study, where two variables were studied: the percentage of total vital tissue (soft and hard; VT%) and the percentage of soft vital tissue only (ST%). A one-way ANOVA and Bonferroni tests were used to determine significant differences between the groups at a 95% confidence level. RESULTS: The VT% values were significantly higher in the EMCS + BC group compared to both the NCS + BC and BC groups. The ST% values were also significantly higher in the EMCS + BC group compared to the BC group. However, no significant differences in ST% values were observed between the NCS + BC group and either the BC or EMCS + BC groups. CONCLUSIONS: Within the limitations of this study, we conclude that the application of enzymatically modified chitosan scaffolds combined with BC yields superior results in pulp regeneration, which contributes to the formation of pulp-like tissue and cells resembling odontoblasts, as well as apex closure with tissue resembling bone tissue.

Bovine pulp extracellular matrix hydrogel for regenerative endodontic applications: in vitro characterization and in vivo analysis in a necrotic tooth model.

BACKGROUND: Regenerative endodontic procedures (REPs) offer the promise of restoring vitality and function to a previously necrotic and infected tooth. However, the nature of regenerated tissues following REPs remains unpredictable and uncontrollable. Decellularized extracellular matrix scaffolds have gained recent attention as scaffolds for regenerative endodontics. OBJECTIVES: Preparation and characterization of a bovine dental pulp-derived extracellular matrix (P-ECM) hydrogel for regenerative endodontic applications. Biocompatibility and regenerative capacity of the prepared scaffold were evaluated in vivo in a canine animal model. METHODS: Fifteen freshly extracted bovine molar teeth were used to prepare P-ECM hydrogels following approval of the institutional review board of the faculty of dentistry, Alexandria University. Decellularization and lyophilization of the extracted pulp tissues, DNA quantification and histological examination of decellularized P-ECM were done. P-ECM hydrogel was prepared by digestion of decellularized pulps. Prepared scaffolds were evaluated for protein content and release as well as release of VEGF, bFGF, TGF-ß1 and BMP2 using ELISA. Rabbit dental pulp stem cells' (rDPSCs) viability in response to P-ECM hydrogels was performed. Finally, proof-of-concept of the regenerative capacity of P-ECM scaffolds was assessed in an infected mature canine tooth model following REPs versus blood clot (BC), injectable platelet-rich fibrin (i-PRF) or hyaluronic acid (HA). Statistical analysis was done using independent t test, the Friedman test and chi-square tests (p value ≤ 0.05). RESULTS: DNA was found to be below the cut-off point (50 ng/mg tissue). Histological evaluation revealed absence of nuclei, retention of glycosaminoglycans (GAGs) and collagen content, respectively. P-ECM hydrogel had a total protein content of (493.12 µg/µl) and protein release was detected up to 14 days. P-ECM hydrogel also retained VEGF, bFGF, TGF-ß1 and BMP2. P-ECM hydrogel maintained the viability of rDPSCs as compared to cells cultured under control conditions. P-ECM hydrogel triggered more organized tissues compared to BC, i-PRF and HA when used in REPs for necrotic mature teeth in dogs. Periapical inflammation was significantly less in HA and P-ECM groups compared to blood-derived scaffolds. CONCLUSION: Bovine dental pulp-derived extracellular matrix (P-ECM) hydrogel scaffold retained its bioactive properties and demonstrated a promising potential in regenerative endodontic procedures compared to conventional blood-derived scaffolds.

Preparation and characterization of bovine dental pulp-derived extracellular matrix hydrogel for regenerative endodontic applications: an in vitro study.

BACKGROUND: The use of biological scaffolds in regenerative endodontics has gained much attention in recent years. The search for a new biomimetic scaffold that contains tissue-specific cell homing factors could lead to more predictable tissue regeneration. The aim of this study was to prepare and characterize decellularized bovine dental pulp-derived extracellular matrix (P-ECM) hydrogels for regenerative endodontic applications. METHODS: Freshly extracted bovine molar teeth were collected. Bovine dental pulp tissues were harvested, and stored at -40º C. For decellularization, a 5-day protocol was implemented incorporating trypsin/EDTA, deionized water and DNase treatment. Decellularization was evaluated by DNA quantification and histological examination to assess collagen and glycosaminoglycans (GAGs) content. This was followed by the preparation of P-ECM hydrogel alone or combined with hyaluronic acid gel (P-ECM + HA). The fabricated scaffolds were then characterized using protein quantification, hydrogel topology and porosity, biodegradability, and growth factor content using Enzyme-linked immunosorbent assay (ELISA): transforming growth factor beta-1(TGF-ß1), basic fibroblast growth factor (bFGF), bone morphogenetic protein 2 (BMP-2) and vascular endothelial growth factor (VEGF). RESULTS: Decellularization was histologically confirmed, and DNA content was below (50 ng/mg tissue). P-ECM hydrogel was prepared with a final ECM concentration of 3.00 mg/ml while P-ECM + HA hydrogel was prepared with a final ECM concentration of 1.5 mg/ml. Total protein content in P-ECM hydrogel was found to be (439.0 ± 123.4 µg/µl). P-ECM + HA showed sustained protein release while the P-ECM group showed gradual decreasing release. Degradation was higher in P-ECM + HA which had a significantly larger fiber diameter, while P-ECM had a larger pore area percentage. ELISA confirmed the retention and release of growth factors where P-ECM hydrogel had higher BMP-2 release, while P-ECM + HA had higher release of TGF-ß1, bFGF, and VEGF. CONCLUSIONS: Both P-ECM and P-ECM + HA retained their bioactive properties demonstrating a potential role as functionalized scaffolds for regenerative endodontic procedures.

Flavonoids modulate regenerative-related cellular events in LPS-challenged dental pulp cells.

OBJECTIVE: To investigate the effects of quercetin (QU), hesperetin (HT), and taxifolin (TX) on human dental pulp cells (hDPCs) chronically exposed to lipopolysaccharide (LPS). METHODS: First, the cytotoxicity (alamarBlue) and bioactivity (biomineralization, Alizarin Red) of QU, HT, and TX concentrations were evaluated on healthy hDPCs. Then, the effects of non-cytotoxic and bioactive concentrations were investigated on hDPCs after previous stimulation with E. coli LPS (10 µg/mL) for 7 days. Cell culture media with and without LPS were used as positive and negative controls, respectively. Cell viability (alamarBlue), NF-κB activation (immunofluorescence), reactive oxygen species production (ROS, H2DCFDA probe), cell migration (Transwell), inflammation-related gene expression (RT-qPCR), and odontogenic differentiation (RT-qPCR and alizarin red) were evaluated (n = 8). Data were analyzed using confidence intervals and ANOVA (α = 5 %). RESULTS: The concentrations of 20 µM QU, 20 µM HT, and 200 µM TX reduced cell viability by more than 30 %. The 5 µM QU, 10 µM HT, and 100 µM TX concentrations were cytocompatible and stimulated biomineralization by healthy hDPCs. These concentrations were tested under the LPS challenge, and cell viability and odontogenic differentiation were significantly increased, while ROS production and inflammatory response were significantly decreased. In addition, the flavonoids significantly stimulated cell migration, reduced NF-κB activation, and increased biomineralization by LPS-challenged hDPCs compared to cells exposed to LPS alone and without any other treatment. CONCLUSION: Flavonoids can modulate the metabolism of hDPCs chronically exposed to LPS in vitro, stimulating cellular events compatible with stem cell-based regenerative processes. CLINICAL SIGNIFICANCE: Flavonoids may be explored as adjuvant therapeutic agents during pulp capping to counteract chronic inflammatory conditions and stimulate regeneration of the dentin-pulp complex in caries-affected teeth, thereby preserving tooth vitality.

Management Strategies for Immature Teeth with Pulp Necrosis: An Umbrella Review of Systematic Reviews.

Introduction: This review evaluates the effectiveness of treatment modalities for immature teeth with pulp necrosis, focusing on calcium hydroxide (CH) and mineral trioxide aggregate (MTA) apexification, as well as regenerative endodontic treatments (RETs). Recent advancements and clinical outcomes are highlighted. Materials and Methods: A comprehensive search of MEDLINE (PubMed), Embase, Cochrane Library, Scopus, and grey literature was conducted from inception to July 2024. Systematic reviews and meta-analyses (SR/MAs) assessing apexification and RET outcomes in immature teeth with pulp necrosis were included. Studies were selected based on predefined criteria, and data on study design, interventions, and outcomes were extracted. Methodological quality was evaluated using the AMSTAR-2 tool. Results: 31 SR/MAs were included. The quality ranged from critically low to low, except one rated as high. MTA apexification was more effective than CH for faster apical barrier formation, though overall success rates were similar. MTA is preferred for its efficiency, but standardized protocols are needed, and tooth discoloration was noted as a potential complication. RET generally outperforms apexification in root maturation, with platelet concentrates like platelet-rich plasma (PRP) and platelet-rich fibrin (PRF) showing promising results; PRP was associated with greater root length, while PRF showed superior apical healing. Variability in RET outcomes was noted due to the lack of standardized protocols. Comparative studies of RET versus apexification showed no significant differences in survival or overall success rates. RET often provides better apical closure and root development, though results vary. Both approaches are viable, but more research with standardized protocols and larger samples is needed to establish definitive clinical advantages. Conclusions: MTA apexification and RET are viable alternatives to CH apexification, with RET showing greater potential for root development and apical healing. Future research should focus on developing standardized protocols and uniform RET guidelines, and evaluating long-term outcomes to establish efficacy and safety.

Time-Dependent Morphological Changes in Traumatic Immature Teeth With Necrotic Pulps Following Regenerative Endodontic Treatment: A Retrospective Study.

BACKGROUND/AIM: Regenerative endodontic treatment is a promising approach for healing periapical lesions and continuous root maturation. Although previous studies have reported its outcomes, the dynamics of morphological changes over time remain unclear. Therefore, this study aimed to evaluate changes in the periapical status and root dimensions over a 60-month follow-up period. MATERIALS AND METHODS: The follow-up duration, periapical status changes, calcific barrier formation, degree of apical closure and radiographic root area changes were compared with those of the last follow-up in this retrospective study. Radiographic root area changes were calculated as the difference between the total root and total canal areas. RESULTS: Fifty-eight patients (81 teeth) underwent regenerative endodontic treatment during the study period, of whom 32 patients (36 teeth, 62%) were included. The survival and success rates of the treated teeth were 100% and 94.4%, respectively. All teeth developed a calcific bridge in the cervical third of the root canal, indicating the presence of vital tissue. Apical narrowing (partial or total) was observed in 75% of the cases. The root maturation stage affected the percentage increase in the radiographic root area. Teeth in Cvek stages II-III showed a higher radiographic root area increase than more mature teeth. All tooth radiographic root areas increased significantly in the initial 20 months of the treatment and moderately thereafter. CONCLUSIONS: Regenerative endodontic treatment is a safe approach for traumatised immature teeth. The presence of a radiographic calcified bridge may be an early indication of treatment success. The main complete tooth morphological changes occur after approximately 20 months posttreatment. These findings may help clinicians better understand the time-dependent changes in the root morphology after treatment, improve the follow-up schedule and predict the progress of healing during follow-up visits.

What is the impact of Endodontic Irrigant Solutions on the Viability of Stem Cells from Apical Papilla in an in-vitro setting: A Systematic Review.

Background: The success of regenerative endodontic procedures (REPs) is significantly influenced by the choice of endodontic irrigant solution. However, the impact of these solutions on the viability of stem cells from the apical papilla (SCAP), a critical component of the REP, remains a subject of ongoing debate. Objective: This study aimed to investigate the effects of various endodontic irrigant solutions on the viability of stem cells from the apical papilla in an in vitro setting. Methods: A systematic literature search was conducted using databases such as PubMed/Medline, Scopus, the Cochrane Library, Web of Science, Embase, gray literature, and reference lists up to August 2023. The search was limited to in vitro studies investigating the impact of endodontic irrigant solutions on SCAP viability. The risk of bias in these studies was evaluated using the Joanna Briggs Institute's checklist. Results: Of the 131 articles retrieved, 14 were selected for review. The effects of eighteen different root canal irrigants, such as ethylenediaminetetraacetic acid, sodium hypochlorite, chlorhexidine, and citric acid, on the viability of SCAPs were evaluated. The risk-of-bias analysis showed a high risk in sample randomization and size justification but a low risk in other areas. Discussion: The effects of endodontic irrigant solutions on the viability of SCAPs are concentration dependent. Concentrations higher than 1.5% sodium hypochlorite, 2 % chlorhexidine, 10 % citric acid, and 2.5 % EDTA significantly reduced cell viability. However, additional research is necessary to determine the effect of these irrigants on tissue regeneration.

Enriched advanced platelet-rich fibrin plus gold nanoparticles against Enterococcus faecalis for its potential use in revascularization for necrotic immature permanent teeth.

Background: Regenerative endodontic procedures allow reinforcement of root canal wall and continuation of root development, opening new therapeutic possibilities. The root canal system of infected teeth is colonized by a variety of microorganisms, which hinder the regenerative process, leading to treatment failure if not adequately addressed, thereby requiring careful attention to microbial control. Aim and Objective: The aim of the study was to assess the antimicrobial activity of advanced platelet-rich fibrin (A-PRF) and gold nanoparticles (AuNps) against Enterococcus faecalis. Materials and Methods: Intravenous blood (5-6 ml) was drawn from four healthy individuals, and A-PRF was prepared through centrifugation at 1500 revolutions per minute (rpm) for 14 min. A-PRF was doped with 3 µl of AuNps and centrifuged at 1000 rpm for 1 min. Antimicrobial activity was assessed using disk diffusion; inhibition zones were measured. For minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC), A-PRF + AuNps were added to the microbial broth at varying concentrations to determine growth inhibition and microbial death. Results: Disk diffusion assays revealed significant antibacterial effects against E. faecalis. Norfloxacin displayed the highest mean zone of inhibition (20.33 ± 1.53 mm), followed by the Test group (A-PRF + AuNPs) (19.33 ± 0.58 mm). Multiple comparisons indicated significant differences (P < 0.001). MIC of A-PRF + AuNPs against E. faecalis was 0.031 mg/ml, with MBC at 0.015 mg/ml. Conclusion: The addition of AuNPs to A-PRF offers the potential for sustained growth factor release while maintaining the sterility of the canal, leading to successful revitalization and regeneration. The combined use of A-PRF + AuNps shows promise for enhancing revascularization in necrotic immature permanent teeth.

The Differentiation Potential of Apical Papilla Cells in Relation to Tenascin-C and Syndecan-1 Expression and Their Potential Role in Regeneration.

Introduction: This study investigated the distribution pattern of tenascin-C and syndecan-1 in the dental mesenchyme during root development of immature swine teeth in order to define the differentiation dynamics of both pulp tissue progenitors and apical papilla cells, as well as to assess the adequacy criticize of the apical papilla to induce dentin-pulp regeneration. Methods: Three 7-month-old miniature swine were used in this study. A total of 12 teeth, including two immature permanent incisors and two premolar teeth of each case, were extracted and processed for histological and immunohistochemical analysis. Different populations of mesenchymal cells located at the root apex were morphologically evaluated in hematoxylin-eosin serial sections. Additionally, the distribution patterns of tenascin-C and syndecan-1 were assessed immunohistochemically. Results: Syndecan-1 was strongly expressed in the dental pulp, particularly along the odontoblasts of the root and the newly deposited predentin layer. Tenascin-C was intensely expressed in the dental pulp. The apical papilla and dental follicle showed no expression of either molecule. Conclusions: Cell differentiation potential in the developing swine apex is progressively restricted to the newly formed dental pulp, whereas phenotypic expression of apical papilla cells remains undetermined unless the new microenvironment triggers cell differentiation towards the odontoblastic lineage.

Triple Antibiotic Paste: A Game Changer in Endodontics.

Triple antibiotic paste has emerged as a game changer in the field of endodontics, offering a novel approach to the management of refractory endodontic infections and regenerative endodontics. This review article provides an overview of the composition, mechanism of action, clinical applications, advantages, and potential limitations of triple antibiotic paste in endodontic therapy. Additionally, recent advancements and future directions for the use of triple antibiotic paste in endodontics are also discussed.

The in vitro efficacy of advanced platelet-rich fibrin plus versus injectable platelet-rich fibrin on the proliferation, migration, and differentiation of stem cells from apical papilla.

Background/purpose: Platelet-rich fibrin (PRF) is a promising host-derived scaffold for regenerative endodontic treatment. This study investigated the effects of advanced PRF plus (A-PRF+) and injectable PRF (i-PRF) on the proliferation, migration, and differentiation of stem cells from apical papilla (SCAPs). Materials and methods: A-PRF+ and i-PRF were prepared using a DUO Quattro centrifuge following a standard protocol. A-PRF+ and i-PRF extract were diluted in Dulbecco's modified Eagle's medium and Ham's F-12 medium (DMEM/F12) to produce the experimental culture medium. DMEM/F12 and DMEM/F12 supplemented with 10% foetal bovine serum (FBS) were used as the negative control (NC) and positive control (PC) media, respectively. The proliferative ability of SCAPs was assessed using a counting method (haemocytometer). The migration ability was examined using a scratch-wound assay. Alkaline phosphatase, bone sialoprotein, dentin matrix protein 1, and dentin sialophosphoprotein expression were measured to determine the differentiation ability. Results: The proliferation, migration, and differentiation of SCAPs in the A-PRF+ group were similar to those of the PC group. In the i-PRF group, the cell number was significantly (p < 0.01) lower than that of the A-PRF+ group on days 8 and 10; the percentage of the scratched area on days 1 and 2 was significantly higher than in the A-PRF+ group (p < 0.05). The mRNA expression levels of biomarkers in the i-PRF group were similar to those in the A-PRF+ group. Conclusion: Both A-PRF+ and i-PRF induce SCAPs proliferation, migration, and differentiation. However, A-PRF+ was superior in supporting the proliferation and migration of SCAPs.

Sphingosine-1-phosphate receptor 1-mediated odontogenic differentiation of mouse apical papilla-derived stem cells.

Background/purpose: Sphingosine-1-phosphate (S1P) exhibits receptor-mediated physiological effects by facilitating the differentiation of mesenchymal stem cells toward the osteoblast lineage. This study aimed to determine the effect of S1P on odontogenic differentiation of mouse immortalized stem cells of dental apical papilla (iSCAP) and assess the distribution of the S1P receptor 1 (S1PR1) in the apical papilla and the root canal wall of immature rat molars. Materials and methods: Immunostaining for S1PR1 was conducted at the apex of the rat mandibular first molar and within the root canal wall. The iSCAP was treated with S1P and bone morphogenetic protein (BMP)-9 (for comparison), and the expression levels of the odontogenic differentiation marker were evaluated via real-time reverse-transcriptase quantitative polymerase chain reaction and enzyme-linked immunosorbent assay. Mineralization and lipid droplet formation were evaluated via Alizarin red and Oil red O staining. Results: S1PR1-positive cells were expressed in areas of both apical papilla and dentin-pulp interface of root canal wall. During the odontogenic differentiation of iSCAP, S1P and BMP-9 increased the expression of the differentiation marker mRNA and secreted proteins including dentin sialophosphoprotein, dentin matrix phosphoprotein 1, and matrix extracellular phosphoglycoprotein. The S1PR1 signaling pathway is involved in the action of S1P, but not that of BMP-9. S1PR1 signaling also facilitated mineralization in iSCAP and suppressed the differentiation of these cells into adipocytes. Conclusion: S1P induced odontogenic differentiation of iSCAP through S1PR1. Furthermore, S1PR1-positive cells were expressed in the apical papilla of immature rat molars and in the dentin-pulp interface where odontoblast-like cells exist.

Research progress of dental pulp regeneration treatment. / å†ç”Ÿæ€§ç‰™é«“æ²»ç–—çš„ç ”ç©¶è¿›å±•.

The dental pulp is the only soft tissue structure within the tooth, serving functions such as sensation and nutrition. However, the dental pulp is highly susceptible to necrosis due to external factors. Currently, root canal therapy is the most commonly used treatment for pulp necrosis. Nevertheless, teeth treated with root canal therapy are prone to secondary infections and adverse outcomes like vertical root fractures. Regenerative endodontic therapy has emerged as a solution, aiming to replace damaged tooth structures, including dentin, root structure, and the pulp-dentin complex cells. This approach demonstrates significant advantages in addressing clinical symptoms and achieving regeneration of the root and even the pulp. Since the discovery of dental pulp stem cells, regenerative endodontic therapy has gained new momentum. Advances in cell transplantation and cell homing techniques have rapidly developed, showing promising potential for clinical applications.

Biocompatibility and antimicrobial effect of demineralised dentin matrix hydrogel for dental pulp preservation.

Regeneration of dentin and preserving pulp vitality are essential targets for vital pulp therapy. Our study aimed to evaluate a novel biomimetic pulp capping agent with increased dentin regenerative activities. To produce demineralised dentin matrix (DDM) particles, human extracted teeth were ground and treated with ethylene diamine tetra-acetic acid solution. DDM particles were added to sodium alginate and this combination was dripped into a 5% calcium chloride to obtain DDM hydrogel (DDMH). The eluants of both DDMH and mineral trioxide aggregate (MTA) were tested using an MTT assay to detect their cytotoxic effect on dental pulp stem cells (DPSC). Collagen-I (COL-I) gene expression was analysed on DPSC exposed to different dilutions of pulp capping material eluants by real-time quantitative polymerase chain reaction. Acridine orange staining was used to monitor the cell growth over the tested materials. Agar diffusion assay was utilised to test the antibacterial effect of DDMH and MTA compared to controls. MTT assay revealed that neat eluates of DDMH promoted DPSC viability. However, neat eluates of MTA were cytotoxic on DPSC after 72 h of culture. Moreover, DPSC were capable of growth and attached to the surface of DDMH, while they showed a marked reduction in their number when cultured on the MTA surface for one week, as shown by the acridine orange stain. In DPSC cultured with DDMH eluates, the COL-I gene was overexpressed compared to those cultured with MTA eluants. DDMH had significant antimicrobial activity in comparison to MTA after 24 h incubation. This in vitro study showed that DDMH could be an alternative pulp capping agent for regenerative endodontics.

Arrest and repair of inflammatory root resorption after an endodontic regeneration procedure - a hypothesis and case report.

INTRODUCTION: External Inflammatory Root Resorption (EIRR) is a significant complication that can occur following traumatic dental injuries, with a prevalence of approximately 18%. Most cases occur during the early stage of the mixed dentition. Specifically, EIRR occurs in approximately 5-8% of luxation injuries, 30% of replanted teeth following avulsion, and 38% of intruded teeth. Conventional methods for addressing EIRR in immature teeth pose several challenges. This often requires numerous dental visits where Ledermix® and calcium hydroxide are used, which may significantly prolong the treatment. Additionally, the effect of prolonged use of calcium hydroxide medication in the root canal is debatable. Recent publications have highlighted the ability of regenerative endodontic treatment (RET) to effectively stop and repair external inflammatory root resorption (EIRR) in a relatively brief time, yielding impressive results. Nevertheless, the underlying mechanism responsible for this effect remains unclear. METHODS: A hypothesis is proposed and drawn from existing data, explaining the mechanism by which RET triggers alterations in the root dimensions of necrotic immature teeth, facilitating continuous root maturation. The hypothesis suggests that bioactive molecules, including growth factors, might be able to penetrate the denuded dentin, reach the resorbed area, and attract stem cells from the surrounding periodontal ligament (PDL) and adjacent bone, leading to the arrest of the resorption process. RESULTS: This recruitment may trigger repair mechanisms, ultimately resulting in the coverage of the denuded dentin with a new layer of PDL, cementoid, and cementum. CONCLUSIONS: A hypothesis of the potential mechanism in which RET may arrest EIRR is presented along with a case report.

Antimicrobial Silk Fibroin Methacrylated Scaffolds for Regenerative Endodontics.

INTRODUCTION: Recognizing the necessity of novel disinfection strategies for improved bacterial control to ultimately favor tissue regeneration, this study developed and characterized antibiotics-laden silk fibroin methacrylated (SilkMA) scaffolds for regenerative endodontics. METHODS: SilkMA-based solutions (10% w/v) containing Clindamycin (CLI) or Tinidazole (TIN) (0 - control; 5, 10, or 15% w/w) or the combination of both drugs (BiMix CLI/TIN 10%) were electrospun and photocrosslinked. Morphology and composition were assessed using scanning electron microscopy and Fourier-transform infrared spectroscopy. Additionally, swelling and degradation profiles were also determined. Cytotoxicity was evaluated in stem cells from apical papilla. Antibacterial efficacy was tested using direct and indirect contact assays against Aggregatibacter actinomycetemcomitans/Aa, Actinomyces naeslundii/An, Enterococcus faecalis/Ef, and Fusobacterium nucleatum/Fn. E. faecalis biofilm inhibition on dentin discs was specifically evaluated for BiMix-laden scaffolds. Data were statistically analyzed with a significance level of 5%. RESULTS: Scanning electron microscopy revealed that all scaffolds had similar characteristics, including fiber morphology and bead absence. Fourier-transform infrared spectroscopy showed the incorporation of CLI and TIN into the fibers and in BiMix scaffolds. Antibiotic-laden scaffolds exhibited lower swelling capacity than the control and were degraded entirely after 45 days. Scaffolds laden with CLI, TIN, or BiMix throughout all time points did not reduce stem cells from apical papilla's viability. CLI-laden scaffolds inhibited the growth of Aa, An, and Ef, while TIN-laden scaffolds inhibited Fn growth. BiMix-laden scaffolds significantly inhibited Aa, An, Ef, and Fn in direct contact, and their aliquots inhibited An and Fn through indirect contact, with additional biofilm inhibition against Ef. CONCLUSIONS: BiMix-laden SilkMA scaffolds are cytocompatible and exhibit antimicrobial effects against endodontic pathogens, indicating their therapeutic potential as a drug delivery system for regenerative endodontics.

Anti-inflammatory potential of casein enzymatic hydrolysate/gelatin methacryloyl scaffolds for vital pulp therapy.

OBJECTIVES: To synthesize casein enzymatic hydrolysate (CEH)-laden gelatin methacryloyl (GelMA) fibrous scaffolds and evaluate the cytocompatibility and anti-inflammatory effects on dental pulp stem cells (DPSCs). MATERIALS AND METHODS: GelMA fibrous scaffolds with 10%, 20%, and 30% CEH (w/w) and without CEH (control) were obtained via electrospinning. Chemo-morphological, degradation, and mechanical analyses were conducted to evaluate the morphology and composition of the fibers, mass loss, and mechanical properties, respectively. Adhesion/spreading and viability of DPSCs seeded on the scaffolds were also assessed. The anti-inflammatory potential on DPSCs was tested after the chronic challenge of cells with lipopolysaccharides (LPS), followed by treatment with extracts obtained after immersing the scaffolds in α-MEM. The synthesis of the pro-inflammatory cytokines IL-6, IL-1α, and TNF-α was measured by ELISA. Data were analyzed by ANOVA/post-hoc tests (α = 5%). RESULTS: CEH-laden electrospun fibers had a larger diameter than pure GelMA (p ≤ 0.036). GelMA scaffolds laden with 20% and 30% CEH had a greater mass loss. Tensile strength was reduced for the 10% CEH fibers (p = 0.0052), whereas no difference was observed for the 20% and 30% fibers (p ≥ 0.6736) compared to the control. Young's modulus decreased with CEH (p < 0.0001). Elongation at break increased for the 20% and 30% CEH scaffolds (p ≤ 0.0038). Over time, DPSCs viability increased across all groups, indicating cytocompatibility, with CEH-laden scaffolds exhibiting greater cell viability after seven days (p ≤ 0.0166). Also, 10% CEH-GelMA scaffolds decreased the IL-6, IL-1α, and TNF-α synthesis (p ≤ 0.035). CONCLUSION: CEH-laden GelMA scaffolds facilitated both adhesion and proliferation of DPSCs, and 10% CEH provided anti-inflammatory potential after chronic LPS challenge. CLINICAL RELEVANCE: CEH incorporated in GelMA fibrous scaffolds demonstrated the potential to be used as a cytocompatible and anti-inflammatory biomaterial for vital pulp therapy.

Mahidol Study 2: Treatment Outcomes and Prognostic Factors of Regenerative Endodontic Procedures in Immature Permanent Teeth.

INTRODUCTION: Studies on the clinical outcomes and prognostic factors of regenerative endodontic procedures (REPs) in a large population and long-term recall period have been limited. Therefore, the aims of this study were to evaluate treatment outcomes and prognostic factors of REPs. METHODS: Immature permanent teeth treated with REPs with a minimum one-year follow-up period were included. Treatment outcomes (functional retention, healed rate, root development, and sensibility test response) and any prognostic factors were analyzed with multivariable Cox regression, linear regression, and modified Poisson regression. RESULTS: One-hundred-twenty REPs teeth with a mean 41.7-month recall period were included with a functional retention rate of 97.5%. The healed, healing, and diseased rates of REPs were 80%, 9.2%, and 10.8%, respectively. Significant prognostic factors for healed were age (<12 years old) and root development stage (stages 4 and 5). Changes in the apical diameter, root length, root width, and radiographic root area after REPs were 56.8%, 8.3%, 23.2%, and 21.7%, respectively. Significant prognostic factors for continued root development were age and etiology of pulpal disease (from caries or dental anomalies). The sensibility test response rate was 41.7% with significant positive factors of ethylenediaminetetraacetic acid irrigation and capping material level above the cemento-enamel junction. CONCLUSION: REPs demonstrated high functional retention and healed rates. Patients <12 years old presented a higher healed rate and continued root development (excluding root length). Dental caries or anomalies had higher continued root development compared with dental trauma. The sensibility test response was related to ethylenediaminetetraacetic acid irrigation and level capping material.

Top 100 most-cited scientific articles in regenerative endodontics 2019-2023: A bibliometric analysis.

BACKGROUND: Bibliometric analysis is a critical indicator of the influence and relevance of scientific papers, whilst also highlighting key contributors and gaps in knowledge in a scientific field. OBJECTIVES: To update and analyse the 100 most-cited papers in regenerative endodontics from 2019 to 2023. METHODS: A search of the most-cited recent papers focusing on regenerative endodontics using journals included in the category, 'Dentistry, Oral Surgery & Medicine', in the Clarivate Web of Science database from 2019 to 2023 was performed. Three researchers conducted the study selection and data extraction. Data extraction included publication title and year, authors, number and mean number of citations, institution, country and continent, study design, journal title, keywords and research topic. Citation counts were also collected in Google Scholar and Scopus databases. Graphical bibliometric networks were created using VOSviewer software. RESULTS: The number of citations of the 100 most-cited articles ranged from 6 to 85. Most were published in 2020 (n = 48), principally in the Journal of Endodontics (47%), followed by International Endodontic Journal (13%), Journal of Dental Research (6%) and Dental Materials (6%). Laboratory study was the most common study design amongst the included papers (n = 47), followed by narrative reviews (n = 17) and observational studies (n = 16). The most frequent first author on the top three most-cited papers was Hacer Aksel, whilst Adham A. Azim (n = 6; 89 citations) contributed most to the top 100 articles. The institution from which most articles originated was the University of Hong Kong (China) (n = 5; 81 citations), whereas the corresponding authors were predominantly from the United States of America (USA) (n = 31; 560 citations). The VOSviewer map of co-authorship demonstrated research collaborative clusters. 'Regenerative endodontics' and 'stem-cells' were the most employed keywords (37 and 36 occurrences respectively). DISCUSSION: The current study was designed not only to showcase the most influential papers in regenerative endodontics since 2019 but also to provide a better understanding of global research in this area over the last five years. CONCLUSIONS: This bibliometric analysis highlighted papers, authors, institutions and keywords in regenerative endodontics. The 100 most-cited papers primarily consisted of laboratory studies published in the USA, focusing on evaluating biomaterials and scaffold design strategies in contact with stem cells. Clinical studies and systematic reviews representing higher levels of scientific evidence are currently not the most influential in the regenerative endodontic field.

Advancements and challenges in stem/progenitor cell transplantation for dentin-pulp regeneration: a systematic review of animal studies (part I).

Dentin-pulp regeneration through stem/progenitor cell transplantation represents a promising frontier in regenerative endodontics. This systematic review meticulously evaluates animal studies to investigate the efficacy of stem cell therapy in repairing/regenerating the dentine-pulp complex in mature/immature animal teeth. Employing a comprehensive electronic search of PubMed and Scopus databases up to October 2023, relevant English studies were identified/assessed. Evaluation parameters encompassed radiographic and histological assessments of dentin-pulp complex formation. Outcome measures included pulp-like and dentin-like tissues regeneration, apical healing, dentin thickening, apical closure, and dentinal bridge formation. The risk-of-bias assessment adhered to the Systematic Review Centre for Laboratory Animal Experimentation (SYRCLE) guidelines. Out of 3250 identified articles, 23 animal experiments were included, categorized into regenerative procedures in mature teeth (n=11), regenerative procedures in immature teeth (n=4), and vital pulp therapy (n=8). Despite the promising potential, the bias in the included studies was high. Notably, Various scaffolds, and growth factors were employed, highlighting the heterogeneity across the studies. Dental pulp stem cells (DPSCs) and bone marrow stem cells, especially specific subfractions, demonstrated notable regenerative potential: hypoxic conditions and extracellular vesicles from preconditioned DPSCs enhanced regeneration, with considerations of cell fate. Donor age impacted regeneration, and challenges persisted in pulpotomy and direct pulp capping. Scaffold and growth factor choices influenced outcomes, underscoring the need for standardized strategies. Despite the promise, clinical viability faces hurdles, necessitating further investigation into adverse effects, optimized scaffolds, and regulatory considerations. This systematic review illuminates the potential of stem cell transplantation for dentin-pulp complex regeneration. The overall evidence quality, influenced by study heterogeneity and biases, underscores the need for cautious interpretation of findings. Future studies should refine methodologies and establish reliable histological parameters for meaningful advancements in dentin-pulp regeneration.