Effects of EDTA and saline as the final irrigation in regenerative endodontic procedures on the migration, proliferation, and differentiation of human stem cells from the apical papilla.

OBJECTIVES: To evaluate the effect of EDTA and saline as the final irrigation in regenerative endodontic procedures (REPS) on the attachment, proliferation, migration, and differentiation of stem cells from the apical papilla (SCAPs). MATERIALS AND METHODS: Dentin specimens from 140 human third molars were irrigated with various protocols-group 1: normal sterile saline (NSS), group 2: EDTA, group 3: EDTA then 5 mL NSS, or group 4: EDTA then 20 mL NSS. The specimens were used in cell assays. For cell proliferation, SCAPs were seeded on dentin, and the cell viability on days 1, 3, and 7 was determined using an MTT assay. At day 3, the attached cells' morphology was observed using SEM, and cell migration was investigated using a transwell migration assay. The ALP activity and odonto/osteogenic differentiation gene expression were evaluated at days 7, 14, and 21 using an ALP activity assay and RT-qPCR. RESULTS: On days 3 and 7, group 4 demonstrated more viable cells than group 1 (p < 0.01). The amount of migrated cells in groups 2, 3, and 4 was greater compared with group 1 (p < 0.05). Moreover, SCAP differentiation was similar between groups. CONCLUSIONS: Irrigating dentin with EDTA alone or with EDTA then NSS promoted SCAP migration. However, a final irrigation with 20 mL NSS after EDTA promoted SCAP proliferation without affecting their differentiation. CLINICAL RELEVANCE: When using a blood clot as a scaffold, a final flushing with 20 mL NSS after EDTA could be beneficial for clinical REP protocols.

Web-Based Survey on Regenerative Endodontic Practices among Members of the American Association of Endodontists.

INTRODUCTION: The American Association of Endodontics (AAE) Regenerative Endodontics Committee reports here the web-based survey data of regenerative endodontic procedures performed by the AAE members from 2008 to 2019. METHODS: The web-based survey consisted of 2 questionnaires, a revascularization and a follow-up, including clinical and radiographic data at 3, 6, 12 or >12 months after treatment. Data from demographics, etiology, clinical protocols, radiographic, and clinical outcome as perceived by the clinicians were recorded. From 927 entries, 184 full cases were submitted and 126 were suitable for radiographic analysis. The data were divided into cases with 6-12 months and cases with >12 months recall time. Descriptive statistics and univariate analyses were performed. RESULTS: Predominantly patients were male (63%), average 10 years-old with anterior teeth (77.2%) due to trauma (69.6%). A wide variation in regenerative endodontic procedures protocols was reported by the AAE members. The most common clinical protocols used minimal instrumentation (75.5%), 2.5% or higher concentrations of sodium hypochlorite (83.7%), and antibiotic paste as intracanal medication (58.7%). The majority reported blood clot formation (56%) and most common coronal capping material was white mineral trioxide aggregates (50.5%). Increases in radiographic root length, and radiographic root area were proportional to the time lapsed after treatment. Overall, case outcomes were categorized by clinicians as successful (54.3%), uncertain (23.9%), and unsuccessful (3.8%). CONCLUSIONS: This web-based survey provides a valuable perspective on case selection, clinical protocols, and perceived outcomes for regenerative endodontic procedures and supports the need for future higher level evidence studies.

Intrafibrillar silicification of collagen scaffolds for sustained release of stem cell homing chemokine in hard tissue regeneration.

Traditional bone regeneration strategies relied on supplementation of biomaterials constructs with stem or progenitor cells or growth factors. By contrast, cell homing strategies employ chemokines to mobilize stem or progenitor cells from host bone marrow and tissue niches to injured sites. Although silica-based biomaterials exhibit osteogenic and angiogenic potentials, they lack cell homing capability. Stromal cell-derived factor-1 (SDF-1) plays a pivotal role in mobilization and homing of stem cells to injured tissues. In this work, we demonstrated that 3-dimensional collagen scaffolds infiltrated with intrafibrillar silica are biodegradable and highly biocompatible. They exhibit improved compressive stress-strain responses and toughness over nonsilicified collagen scaffolds. They are osteoconductive and up-regulate expressions of osteogenesis- and angiogenesis-related genes more significantly than nonsilicified collagen scaffolds. In addition, these scaffolds reversibly bind SDF-1α for sustained release of this chemokine, which exhibits in vitro cell homing characteristics. When implanted subcutaneously in an in vivo mouse model, SDF-1α-loaded silicified collagen scaffolds stimulate the formation of ectopic bone and blood capillaries within the scaffold and abrogate the need for cell seeding or supplementation of osteogenic and angiogenic growth factors. Intrafibrillar-silicified collagen scaffolds with sustained SDF-1α release represent a less costly and complex alternative to contemporary cell seeding approaches and provide new therapeutic options for in situ hard tissue regeneration.

Biologically based treatment of immature permanent teeth with pulpal necrosis: a case series.

This case series reports the outcomes of 8 patients (ages 9-4 years) who presented with 9 immature permanent teeth with pulpal necrosis and apical periodontitis. During treatment, 5 of the teeth were found to have at least some residual vital tissue remaining in the root canal systems. After NaOCI irrigation and medication with ciprofloxacin, metronidazole, and minocycline, these teeth were sealed with mineral trioxide aggregate and restored. The other group of 4 teeth had no evidence of any residual vital pulp tissue. This second group of teeth was treated with NaOCl irrigation and medicated with ciprofloxacin, metronidazole, and minocycline followed by a revascularization procedure adopted from the trauma literature (bleeding evoked to form an intracanal blood clot). In both groups of patients, there was evidence of satisfactory postoperative clinical outcomes (1-5 years); the patients were asymptomatic, no sinus tracts were evident, apical periodontitis was resolved, and there was radiographic evidence of continuing thickness of dentinal walls, apical closure, or increased root length.

Effect of Sodium Hypochlorite in Dental Unit Waterline on Aerosolized Bacteria Generated from Endodontic Procedures.

INTRODUCTION: Aerosol generation in a dental setting is a critical concern, and approaches that aim at decreasing bacterial load in aerosols are of high priority for dental professionals. The objectives of this study were to evaluate the relative effect of various endodontic procedures on the generation and dissemination of aerosols and the effect of 0.1% sodium hypochlorite (NaOCl) in dental unit waterlines (DUWLs) on the bacterial load in the generated aerosols in a clinical setting. METHODS: The study was completed in 2 phases. The classic passive sampling technique using brain-heart infusion agar plates was used. Agar plates were strategically placed throughout the operatory at predefined locations. In phase 1, to evaluate the effect of different endodontic procedures on the generation and dissemination of aerosols, we collected a total of 38 samples. After baseline collection, test samples were collected during vital pulp therapy (VPT) full pulpotomy (n = 10), nonsurgical root canal therapy (NSRCT, n = 10), surgical root canal therapy (SRCT, n = 10), and incision and drainage (n = 8) procedures. Bacterial growth was expressed as colony-forming units at 48 hours after sample collection. Data were analyzed using 1-way analysis of variance with the Tukey multiple comparison post hoc test. In phase 2, to evaluate the effect of 0.1% NaOCl in the DUWL on the bacterial load in the generated aerosols, a total of 30 samples were collected. All procedures including VPT (n = 10), NSRCT (n = 10), and SRCT (n = 10) were performed with 0.1% NaOCl in the DUWL. Bacterial growth was expressed as colony-forming units at 48 hours after sample collection. Data were analyzed using 2-way analysis of variance with the Tukey multiple comparison post hoc test. RESULTS: All endodontic procedures generated aerosols at all tested locations, except incision and drainage. Aerosols were disseminated as far as 3 m from the patient's head with no significant difference between various locations (P > .05). VPT procedures generated the maximum number of aerosols compared with NSRCT and SRCT. Adding 0.1% NaOCl to DUWLs significantly reduced the bacterial load in the generated aerosols in all treatment groups compared with groups treated with untreated waterlines (P < .05). No significant difference was noted in the bacterial load between all groups with treated waterlines (P > .05). CONCLUSIONS: All tested endodontic procedures led to the generation and dissemination of contaminated aerosols, and the addition of 0.1% NaOCl as a biocide to the DUWL led to a statistically significant reduction in the bacterial load.

Microbiome Changes during Regenerative Endodontic Treatment Using Different Methods of Disinfection.

INTRODUCTION: The purpose of this study was to characterize qualitatively and quantitatively the changes in the endodontic microbiome, in teeth with necrotic pulp, open apexes, and apical periodontitis, with 3 antimicrobial protocols, undertaken in a multicenter clinical trial. METHODS: Microbiological samples were collected from 116 regenerative endodontic teeth, and 97 qualified for inclusion. The teeth were randomly divided into 3 treatment groups: apexification (APEX), regeneration (REGEN), and revascularization (REVASC), all in 2 appointments. The group variables in the first appointment irrigants, and second appointment irrigants and medicaments were as follows: APEX: 5.25%-6% NaOCl, 5.25%-6% NaOCl + 17% EDTA and calcium hydroxide; REGEN: 1.25% NaOCl, 17% EDTA, and 0.1 mg/mL triple antibiotic paste (TAP); and REVASC 5.25% NaOCl, saline, and 1 g/mL TAP, respectively. Sampling was done upon access (S0), after irrigation in the first appointment (S1), and after using medication and irrigation in the second appointment (S2). RESULTS: Quantitative polymerase chain reaction analysis of the 16S ribosomal RNA gene showed significant reduction in bacterial load from S0 to S2 in all groups; however, the APEX and REVASC groups had significantly less residual DNA than the REGEN group (P = .0045). The relative abundance of Bacteroidetes, Fusobacteria, Spirochaetes, and Synergistetes were reduced with the treatment rendered. However, relative abundance of Firmicutes and Actinobacteria was not changed, and that of Proteobacteria increased. LEfSe analysis showed that reduction in bacterial taxa was more in REVASC than APEX, which in turn was more than in REGEN. CONCLUSION: Enhanced antimicrobial protocols lead to better reduction in quantitative and qualitative parameters of the endodontic microflora.

Coronavirus Disease 19 (COVID-19): Implications for Clinical Dental Care.

The recent spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and its associated coronavirus disease has gripped the entire international community and caused widespread public health concerns. Despite global efforts to contain the disease spread, the outbreak is still on a rise because of the community spread pattern of this infection. This is a zoonotic infection, similar to other coronavirus infections, that is believed to have originated in bats and pangolins and later transmitted to humans. Once in the human body, this coronavirus (SARS-CoV-2) is abundantly present in nasopharyngeal and salivary secretions of affected patients, and its spread is predominantly thought to be respiratory droplet/contact in nature. Dental professionals, including endodontists, may encounter patients with suspected or confirmed SARS-CoV-2 infection and will have to act diligently not only to provide care but at the same time prevent nosocomial spread of infection. Thus, the aim of this article is to provide a brief overview of the epidemiology, symptoms, and routes of transmission of this novel infection. In addition, specific recommendations for dental practice are suggested for patient screening, infection control strategies, and patient management protocol.

Clinical Outcomes of Immature Teeth Treated with Regenerative Endodontic Procedures-A San Antonio Study.

INTRODUCTION: Factors that influence clinical outcomes for regenerative endodontic procedures (REPs) are unknown. This retrospective study aimed to assess tooth healing, root development, pulp vitality, and esthetics post-REPs and categorize them into clinician- and patient-centered outcomes. Furthermore, this study identified significant predictors affecting such outcomes. METHODS: Immature permanent teeth diagnosed with pulp necrosis treated with REPs between 2008 and 2018 with a minimum of 1-year follow-up were included. Outcomes included success and survival rates, changes in root development using 2-dimensional radiographic root area (RRA) and 3-dimensional measurements, pulp vitality, and tooth discoloration. Predictor variables of success included age, sex, etiology of pulp necrosis (PN), type of medicament, sodium hypochlorite (NaOCl) concentration, type of biomaterial used over the blood clot, and preoperative apical diagnosis. Statistical analyses included Cox proportional hazard analyses and generalized regression models. RESULTS: Fifty-one teeth with an average of 2.1 years of follow-up satisfied the criteria. The survival rate was 92%. The success rate was 84.3% with age, etiology of PN, type of medicament, and NaOCl concentration being significant predictors of failure. Root development occurred in 91.4% of cases with age, sex, etiology of PN, type of medicament, NaOCl concentration, and apical diagnosis being significant predictors for RRA change. Positive pulp sensibility responses were associated with greater RRA change, and, finally, the type of biomaterial was a significant predictor for tooth discoloration after treatment. CONCLUSIONS: REPs provide a high survival rate. Patient and clinical factors may affect outcomes, and this knowledge may help to define the criteria for optimal treatment planning of REPs.

Regeneration potential of the young permanent tooth: what does the future hold?

During the last 10-15 years, there has been a tremendous increase in our clinical "tools" (ie, materials, instruments, and medications) and knowledge from the trauma and tissue engineering fields that can be applied to regeneration of a functional pulp-dentin complex. In addition, recent case reports indicate that biologically based endodontic therapies can result in continued root development, increased dentinal wall thickness, and apical closure when treating cases of necrotic immature permanent teeth. The purpose of this review was to summarize these findings and illustrate a path forward for the development and evaluation of regenerative endodontic therapies.

Effect of Bioceramic Materials on Proliferation and Odontoblast Differentiation of Human Stem Cells from the Apical Papilla.

INTRODUCTION: In regenerative endodontic treatment (RET), practitioners favor the placement of bioceramics as sealing materials over blood clots. It is important to understand the interaction between sealing material and cells in the root canal. The purpose of this study was to compare the effectiveness of various bioceramic materials (ProRoot MTA [Dentsply, Tulsa, OK], Biodentine [Septodont, Saint-Maur-des-Fossés, France], and RetroMTA [BioMTA, Seoul, Korea]) as sealing materials in RET for the proliferation and differentiation of stem cells from the apical papilla (SCAPs). METHODS: SCAPs were seeded at 20,000 cells/well and cultured with soluble agents of testing materials through a transwell culture plate. The proliferation of SCAPs was investigated using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay on days 1, 3, 7, and 14 of testing. Alizarin red staining and quantitative real-time polymerase chain reaction were used for SCAP differentiation at different time points (1, 7, 14, and 21 days). The odontoblast genes expressed are dentin matrix acidic phosphoprotein 1, dentin sialophosphoprotein, osteocalcin, and matrix extracellular phosphoglycoprotein, which were used in this study. The SCAPs were cultured in odonto/osteogenic induction medium and also contacted soluble agents from the testing materials. RESULTS: All 3 tested biomaterials induced SCAP proliferation. The Biodentine, ProRootMTA, and RetroMTA groups showed significant SCAP proliferation on days 7 and 14 compared with the control. In regard to odontoblastic differentiation, only Biodentine showed positive alizarin red staining. The highest expressions of dentin matrix acidic phosphoprotein 1, dentin sialophosphoprotein, and matrix extracellular phosphoglycoprotein were found on day 21 in the Biodentine group. The expression of osteocalcin was found to be significant on day 7. CONCLUSIONS: Biodentine, ProRootMTA, and RetroMTA can induce SCAP proliferation. Biodentine induced significant SCAP differentiation among the 3 materials.

A Comparison of 3 Quantitative Radiographic Measurement Methods for Root Development Measurement in Regenerative Endodontic Procedures.

INTRODUCTION: The outcomes of an immature tooth with necrotic pulp treated with regenerative endodontic procedures (REPs) were assessed clinically and radiographically. Root maturation is an important outcome of REPs, and several radiographic measurement methods have been used to measure this. The aim of this study was to compare radiographic measurement methods, measuring root maturation in immature teeth with necrotic pulp treated with REPs. METHODS: Seventy-one radiographic images of REP cases were measured and compared using 3 radiographic measurement methods described by Bose et al (2009), Alobaid et al (2014), and Flake et al (2014). The intraclass correlation coefficient values were evaluated using the intra- and interobserver reliability test and the effect of the stage of root development. RESULTS: The intra- and interobserver reliability for Alobaid et al's method and Flake et al's method were slightly higher than Bose et al's method as quantified by the intraclass correlation coefficient without a significant difference (P > .05). The stage of root development did not affect the reliability of the measurement methods. A high level of agreement was found among the 3 stages of root development for all 3 quantitative radiographic measurement methods. CONCLUSIONS: All 3 quantitative radiographic measurement methods exhibited high agreement regarding reliability. The stage of root development did not have an impact on the reliability of the measurement methods.