Stem cells from the apical papilla (SCAP) are essential for regenerative endodontic treatment. Although mineral trioxide aggregate (MTA) and Biodentine are widely used in regenerative endodontic treatment procedures, their effects on SCAP remain unclear. This study investigated the impact of ProRoot MTA and Biodentine on SCAP viability and mineralization in the presence of Enterococcus faecalis under aerobic and anaerobic environments. Stem cells from the apical papilla were isolated from three healthy donors and exposed to three different surface area-to-volume (SA:V) ratio extracts of ProRoot MTA and Biodentine for 21 days in aerobic or anaerobic conditions. Cell viability was assessed using a neutral red cytotoxicity assay, and mineralization was evaluated by measuring alkaline phosphatase (ALP) activity. No significant differences between ProRoot MTA and Biodentine regarding SCAP viability were detected; however, increased cytotoxicity was found (for both ProRoot MTA and Biodentine) at the highest SA:V ratio of extract used. Oxygen availability, as well as variability in responses of SCAP from the different donors, resulted in greater variation of ALP levels than did type of material. Both ProRoot MTA and Biodentine showed comparable effects on SCAP viability and mineralization, with high SA:V ratios of extracts resulting in increased cytotoxicity. Mineralization in SCAP is influenced by oxygen conditions and the presence of E. faecalis, elucidating the need for further in vivo studies to optimize regenerative endodontic treatment outcomes.

Oral and maxillofacial (OMF) tissue engineering has always been the subject of longstanding professional and academic debates. Despite rapidly evolving therapeutic approaches including reconstructive surgeries, tissue grafts, platelet-rich therapies, and mesenchymal stem cell-based treatments, there are remaining concerns regarding the therapeutic efficacy, safety profile, immunological compatibility, target-specificity, and ethical issues of such therapies. In addition to the multifactorial nature of OMF diseases, complex maxillofacial anatomy and neurovasculature further pinpoint the urgent need for breakthroughs in the era of OMF treatments. In recent years, exosomes have garnered immense popularity as the frontiers of regenerative medicine. Exosomes, natural nanovesicles secreted by a wide range of cells, act as nano messengers that harbor the rich molecular cargo received by their original cells and demonstrate important roles for diverse physiological and pathological intercellular communications. Among the different sources of exosomes, the oral cavity hosts a wide variety of mesenchymal stem cell-derived and salivary exosomes. Owing to the neural crest origin and multipotent differentiation, oral stem cell-derived exosomes hold great promise for OMF tissue engineering. Moreover, exosomes derived from pathological oral cavity cells and saliva samples provide diagnostic and prognostic signatures for different OMF diseases. This review highlights the cutting-edge diagnostic and therapeutic applications of oral cavity-derived exosomes in the field of OMF regeneration. Furthermore, we emphasize the existing challenges and constraints in exosome-based diagnostics and therapy, thus offering important perspectives for the future clinical application of exosomes derived from the oral cavity.

The tooth and its supporting periodontium are essential structures of the oral cavity, frequently compromised by conditions such as dental defects, aries, and periodontal diseases, which, if poorly treated, often lead to tooth loss. These conditions, affecting billions of people worldwide, remain significant healthcare and socio-economic challenges. Regenerative dentistry has emerged as a possible therapeutic option, leveraging advances in tissue engineering (TE), stem cell biology, and biophysical stimulation. Oral tissue-derived mesenchymal stem/stromal cells (OMSCs) hold great potential for dental and periodontal regeneration. Electrical stimulation (ES), a biophysical cue known to regulate key cellular behaviors such as migration, proliferation, and differentiation, has gained increasing attention for enhancing the therapeutic capacities of OMSCs. This review explores the biological properties of OMSCs under ES, its role in regenerative dentistry, and recent breakthroughs in ES-based dental and periodontal TE strategies. Furthermore, the current challenges and future directions for translating these innovative approaches into clinical practice are discussed.

Dental stem cells are widely viewed as good options for bone regeneration because of their ease of acquisition, innate ability to renew themselves, and ability to differentiate into different types of cells. However, the process of osteogenic differentiation of dental stem cells is orchestrated by an intricate system of regulatory mechanisms. Recent studies have demonstrated the critical impacts of circular RNAs (circRNAs) on osteogenic differentiation of dental stem cells. Exploring the roles and regulatory pathways of circRNAs in dental stem cells could identify novel targets and approaches for utilizing dental stem cell therapy in clinical settings. This review provides a comprehensive overview of the functions and mechanisms of circRNAs, with a particular focus on their expression patterns and regulatory roles in osteogenic differentiation of various dental stem cell types. Furthermore, this review discusses current research challenges in this field and proposes future directions for advancing our understanding of circRNA-mediated regulation in dental stem cell biology.

The aim was to evaluate the effect of single and double doses of low-level laser irradiation on proliferation of human dental pulp stem cells (DPSC) and expression of vascular endothelial growth factor (VEGF) and dentine sialoprotein (DSP).

In this experimental in vitro study, after confirming the stemness of DPSCs, the cells were cultured in Dulbecco's Modified Eagle Medium (DMEM) for MTT assay and VEGF-ELISA and osteogenic medium for DSP-ELISA. The wells containing DPSCs were divided into three main groups and 9 subgroups (n = 7). In groups with single low-level laser, 660-nm diode laser was irradiated at 100 mW and 3 J/cm2 energy density for 15 s. In groups with double doses of low-level laser the second identical irradiation was after 48 h. The MTT-assay and ELISA for DSP/VEGF (dentinogenic/angiogenic differentiation) were performed at 1, 7 and 14 days post irradiation. Using the SPSS software 20 (SPSS, Chicago, Ill, USA) with 95% confidence interval (P = 0.05), a two-way ANOVA test with Tukey's post hoc test was used for the effect of LLLI on VEGF and DSP. The One-Way ANOVA was used for of cell proliferation.

Higher proliferation rate in both single and double low-level laser was reported. The difference was statistically significant for double doses of low-level laser (P = 0.001, P = 0.020 and P = 0.000 for 1, 7 and 14 days, respectively). Also after one, 7 and 14 days, cells in significant increase in DSP (P > 0.05) and VEGF (P > 0.05) was observed that was significantly higher for double doses of low-level laser.

Low level laser enhanced the mitochondrial activity and proliferation of DPSCs. Increased production of DSP/VEGF indicates dentinogenic/angiogenic activity.

Low level laser increases the proliferation of DPSCs, elevates the production of VEGF (which means better angiogenesis in regenerative treatments) and increases the production of DSP (which means better dentinogenesis in vital pulp treatments).

Regenerative endodontic procedures (REPs) offer an alternative to apexification in necrotic immature permanent teeth, promoting continued root development and dentinal wall thickening. Success in REPs requires effective disinfection and the survival of dental-derived mesenchymal stem cells (DMSCs), such as dental pulp stem cells (DPSCs), stem cells from the apical papilla (SCAPs), and periodontal ligament stem cells (PDLSCs). This review investigates the biocompatibility of irrigation solutions, including sodium hypochlorite (NaOCl), ethylenediaminetetraacetic acid (EDTA), and chlorhexidine (CHX), on DMSCs. Following PRISMA guidelines, a comprehensive search was conducted in PubMed, Scopus, Web of Science, Cochrane, and SciELO, with the last update on March 4, 2024. Studies from January 2008 to April 2024 assessing viability, proliferation, migration, differentiation, and mineralization of DMSCs treated with NaOCl, EDTA, and CHX were included. The papers were selected using PICOS criteria and quality was assessed using the PRILE checklist and risk of bias with the Quality Assessment Tool for In Vitro Studies. Of 738 studies identified, 15 met inclusion criteria. The findings suggest that NaOCl and CHX exhibit lower biocompatibility towards DMSCs compared to EDTA. NaOCl and CHX are cytotoxic to DMSCs, while EDTA demonstrates favorable biocompatibility, promoting osteogenic differentiation and mineralization. This highlights potential implications for irrigant selection in regenerative procedures, as appropriate irrigants may enhance cellular survival and improve clinical outcomes.

The role of human Stem Cells from the Apical Papilla (SCAP) in tissue regeneration has been described, but their impact on modulating the apical inflammatory process by other surrounding cell populations, such as periodontal ligament fibroblasts (PLFs), is unclear. Therefore, we investigated the role of SCAP in the activation of PLFs in vitro.

Primary SCAP culture was used to obtain conditioned media (CM). A primary human PLF culture was established and stimulated with increasing concentrations of Escherichia coli lipopolysaccharide (LPS) (0.01, 0.1, and 1 µg/mL). At the 24 h time-point, an MTT viability assay was performed, and interleukin (IL)-6 and chemokine (CC-motif) ligand 2 (CCL2) levels were quantified by enzyme-linked immunosorbent assay. Then, PLFs were stimulated with LPS in the presence of SCAP-CM (1:5 dilution) for cell viability assessment and cytokine detection. The following groups were tested: PLF activated with LPS at concentrations of 0.01 and 1 µg/mL with or without SCAP-CM; a group with PLF stimulated by SCAP-CM alone; and a control group (proliferation medium only). The experiments were conducted in triplicate and sextuplicate. Statistical analyses were performed using analysis of variance followed by Tukey's post-hoc test, with statistical significance established at 5% (p=0.05).

The MTT assay showed no cytotoxicity of LPS or SCAP-CM on PLFs (p>0.05). The production of CCL2 and IL-6 significantly increased in the presence of SCAP-CM regardless of the presence of LPS (p<0.0001).

SCAP-CM significantly enhanced the release of proinflammatory cytokines by PLFs in vitro.

Based on the potential of DPSCs as the most promising candidates for bone tissue engineering, we comprehensively investigated the time-dependent cellular and molecular changes that occur during their osteodifferentiation. To analyze this area in-depth, we used both cellular and molecular approaches. Morphological changes were monitored using bright-field microscopy, while the production of mineral deposits was quantified spectrophotometrically. The expression of a key mesenchymal stem cell marker, CD90, was assessed via flow cytometry. Finally, protein-level changes in whole cells were examined by fluorescence microscopy. Our results show successful long-term osteodifferentiation of the patient's DPSCs within 25 days. In differentiated cells, mineralized extracellular matrix production gradually increased; in contrast, the expression of the specific stem cell marker CD90 significantly decreased. We observed dynamic changes in intracellular and extracellular proteins when collagen1 A1 and osteopontin appeared as earlier markers of osteogenesis, while apolipoprotein A2, bone morphogenetic protein 9, dentin sialophosphoprotein, and matrix metalloproteinase 8 were produced mainly in the late stages of this process. A decrease in actin microfilament expression indicated a reduction in cell proliferation, which could be used as another marker of osteogenic initiation. Our results suggest a coordinated process in vitro in which cells synthesize the necessary proteins and matrix components to regulate the growth of hydroxyapatite crystals and form the bone matrix.

The aim of this study was to investigate changes in the expression of members of the matrix metalloproteinases (MMPs) family in response to lipopolysaccharide (LPS) stimulation and to investigate the regulatory effects of BMP9 on MMPs.

The extracted human stem cells from the apical papilla (hSCAPs) were identified by flow cytometry, Alizarin Red staining, Oil Red O staining, and alkaline phosphatase staining. The appropriate LPS concentration for inducing inflammation in hSCAPs was determined using real-time quantitative PCR (RT-qPCR) and Cell Counting Kit-8 (CCK-8) assays. MMP expression in LPS-stimulated hSCAPs was evaluated by RT-qPCR. BMP9 was overexpressed in hSCAPs via recombinant adenovirus, and its effects on MMP regulation were assessed using RT-qPCR, Western blotting, and ELISA. All experiments were conducted in vitro. Data were analyzed by one-way ANOVA followed by Tukey's post-hoc comparison, with p < 0.05 considered significant.

The results showed that on the 3rd and 5th day after LPS stimulation, the expression of MMP-1, MMP-2, MMP-3, MMP-8, MMP-9, MMP-10, MMP-12, and MMP-13 in hSCAPs was significantly upregulated. On the 7th day after LPS induction, the expression of MMP-3, MMP-8, MMP-9 and MMP-13 in hSCAPs was significantly increased. When BMP9 was overexpressed in hSCAPs, the elevated MMPs were inhibited to varying degrees.

In the LPS-induced inflammatory environment, certain MMPs are elevated in hSCAP, with MMP-13 being the most significant. Overexpression of BMP9 can significantly inhibit elevated MMPs, suggesting that BMP9 may provide new insights and targets for the treatment of periapical periodontitis.

This study examines the effects of pulsed wave photobiomodulation (pwPBM) on the osteogenic differentiation of stem cells from the apical papilla (SCAP). Using 810 nm near-infrared (NIR) light with 300 Hz pulses and a 30% duty cycle, pwPBM was applied at a total energy density of 750 mJ/cm2. Osteogenesis was evaluated through both in vitro and in vivo analyses. In vitro experiments demonstrated significant enhancement of alkaline phosphatase (ALP) activity, along with upregulation of key osteogenesis-related genes and proteins, as confirmed by real-time polymerase chain reaction (PCR) and Western blot analyses. In vivo, histological assessments following SCAP transplantation revealed increased bone tissue formation, further corroborated by osteocalcin staining. These findings underscore the potential of pwPBM as an innovative and effective tool for dental tissue regeneration and engineering.

Periodontal bony defects pose a significant challenge in periodontology, necessitating advanced regenerative approaches to restore the lost structures. Stem cell-based therapies have emerged as a promising solution due to their ability to differentiate into various cells, modulating the regenerative microenvironment. This narrative review explores the potential of stem cells derived from multiple sources in treating periodontal bony defects. Additionally, we examine evidence from both animal and human studies, highlighting advancements, clinical outcomes, and limitations. By investigating these findings, this article provides a comprehensive overview of the advantages of stem cell-based therapies compared to other regenerative techniques in addressing periodontal bony defects and discusses the limitations of their translation into routine clinical practice.

This study aimed to establish and validate machine learning (ML) models to predict the prognosis of regenerative endodontic procedures (REPs) clinically, assisting clinicians in decision-making and avoiding treatment failure.

A total of 198 patients with 268 teeth were included for radiographic examination and measurement. Five Machine Learning (ML) models, including Random forest (RF), gradient boosting machine (GBM), extreme gradient boosting (XGB), Logistic regression (logR) and support vector machine (SVM) are implemented for the prediction on two datasets of follow-up periods of 1-year and 2-year, respectively. Using a stratified five folds of cross-validation method, each dataset is randomly divided into a training set and test set in a ratio of 8 : 2. Correlation analysis and importance ranking were performed for feature extraction. Seven performance metrics including area under curve (AUC), accuracy, F1-score, sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) were calculated to compare the predictive performance.

The RF (Accuracy = 0.91, AUC = 0.94; Accuracy = 0.84, AUC = 0.86) and GBM (Accuracy = 0.91, AUC = 0.93; Accuracy = 0.84, AUC = 0.85) had the best and similar performance simultaneously in the prediction of 1-year follow-up period and 2-year follow-up period, respectively. The variables applied to predict the primary outcome in REPs were ranked accordingly to their values of feature importance, including age, sex, etiology, the number of root canals, trauma type, swelling or sinus tract, periapical lesion size, root development stage, pre-operative root resorption, medicaments, scaffold, second REPs, previous root canal filling.

RF and GBM models outperformed XGB, logR, SVM models on the overall performance on our datasets, exhibiting the potential capability to predict the prognosis of REPs. The ranking of feature importance contributes to establishing the scoring system for prognosis prediction in REPs, assisting clinicians in decision-making.

The management of an endodontic failure in a traumatized tooth with an open apex presents a major dilemma. The arrest of root growth after traumatic injury is a substantial consequence of pulpal necrosis. Conventional endodontic treatment procedures will be challenging by the resulting thin, fragile dentinal walls, which will impede the appropriate debridement and optimal apical sealing. Apexification is a well-established procedure that is designed to treat or preserve a tooth with an incomplete root apex and necrotic pulpal tissue by promoting the formation of a calcified apical barrier through the application of a bioactive substance at the root apex. The present case report demonstrates a successful apexification procedure for an endodontically failed permanent central incisor with an open apex with a two-year follow-up time. The clinical and radiographical results revealed the absence of signs and symptoms and the formation of hard tissue at the root apex. The tooth was extracted for prosthodontic reasons and processed for histologic examination. The histologic evaluation revealed dentin-like and cementum-like tissues formed at the apical barrier.

Human stem cells derived from the apical papilla (SCAPs) are recognized for their multilineage differentiation potential and their capacity for functional tooth root regeneration. However, the molecular mechanisms underlying odonto/osteogenic differentiation remain largely unexplored. In this study, we utilized single-cell RNA sequencing (scRNA-seq) to conduct an in-depth analysis of the transcriptional changes associated with chemically induced osteogenesis in SCAPs.

scRNA-seq identified SCAPs as distinct subpopulations. Differentially expressed genes (DEGs) and Gene Ontology (GO) analyses were conducted to evaluate the potential function of each cluster. Pseudotime trajectory analysis was employed to elucidate the potential differentiation processes of the identified SCAP populations. To investigate the osteo/odontogenic potential of Deiodinase Iodothyronine Type 2 (DIO2) on SCAPs, we performed alkaline phosphatase staining, western blot analysis, Alizarin Red S staining and immunofluorescence staining. Additionally, SCAP components were transplanted into mouse calvarial defects to evaluate osteogenesis in vivo.

The analysis of cell clusters derived from our scRNA-seq data revealed a significant shift in cellular composition when cells were cultured in a mineralization induction medium compared to those cultured in a complete medium. Both groups exhibited heterogeneity, with some cells intrinsically predisposed to osteogenesis and others appearing to be primed for proliferative functions. Notably, we identified a subpopulation characterized by high expression of DIO2, which exhibited pronounced osteogenic activity during differentiation.

Our study is the first to reveal a shift in the cellular composition of SCAPs when cultured in a mineralization induction medium compared to a complete medium. Following both in vitro and in vivo validation, the DIO2+ subpopulation exhibited the highest transcriptional similarity to osteogenic function, suggesting its potential utility in tissue regeneration applications.

Cell homing, a process that leverages the body's natural ability to recruit cells and repair damaged tissues, presents a promising alternative to cell transplantation methods. Central to this approach is the recruitment of endogenous stem/progenitor cells-such as those from the apical papilla, bone marrow, and periapical tissues-facilitated by chemotactic biological cues. Moreover, biomaterial scaffolds embedded with signaling molecules create supportive environments, promoting cell migration, adhesion, and differentiation for the regeneration of the pulp-dentin complex. By analyzing in vivo animal studies using cell homing strategies, this review explores how biomolecules and scaffold materials enhance the recruitment of endogenous stem cells to the site of damaged dental pulp tissue, thereby promoting repair and regeneration. It also examines the key principles, recent advancements, and current limitations linked to cell homing-based regenerative endodontic therapy, highlighting the interplay of biomaterials, signaling molecules, and their broader clinical implications.

To assess the therapeutic effects of mesenchymal stem cell (MSC)-derived exosome therapy on periodontal regeneration and identify treatment factors associated with enhanced periodontal regeneration in recent preclinical studies.

Searches were conducted in PubMed, Cochrane Library, EMBASE, and Web of Science databases until October 10, 2024. A risk of bias (ROB) assessment was performed using the SYRCLE tool. Osteogenic-related parameters were used as the primary outcome measures.

In total, 1360 articles were identified, of which 17 preclinical studies were based on MSC-derived exosome therapy, and they demonstrated a beneficial effect on BV/TV (SMD = 13.99; 95% Cl = 10.50, 17.48; p < 0.00001), CEJ-ABC (SMD = -0.22; 95% Cl = -0.31, -0.13; p < 0.00001), BMD (SMD = 0.29; 95% Cl = 0.14, 0.45; p = 0.0002), and Tp.Sp (SMD = -0.08; 95% Cl= -0.15, -0.02; p = 0.02) compared with the control group. However, no significant differences were observed in Tp.Th (SMD = 0.03; 95% CI = 0.00, 0.07; p = 0.09) between the exosome-treated group and control group. Additionally, subgroup analysis indicated that preconditioned exosomes (p = 0.03) significantly improved BV/TV. In contrast, there were no significant differences in the enhancement of BV/TV with respect to the application method (p = 0.29), application frequency (p = 0.10), treatment duration (p = 0.15), or source of MSCs (p = 0.31).

MSC-derived exosomes show great promise for enhancing the quality of periodontal regeneration. However, more standardized and robust trials are needed to reduce heterogeneity and bias across studies and to confirm the therapeutic parameters associated with the enhancement of periodontal regeneration by MSC-derived exosomes.

CRD42024546236.

Revascularization procedures are used over apexification to treat teeth with necrotic pulp tissues and incomplete root formation. Clinically, inducing proliferation, migration, matrix deposition, and differentiation of stem cells from apical papilla (SCAPs) are critical for pulp regeneration. The study aimed to elucidate the impact of bone morphogenetic protein-4 (BMP-4) on plasminogen activation molecules and the osteogenic/odontogenic differentiation of SCAPs, as well as understand the related signaling mechanisms.

SCAPs were exposed to BMP-4 with or without signal transduction inhibitors. Cell viability was assessed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. mRNA levels were quantified using real-time PCR. Protein expression in SCAPs was analyzed through immunofluorescent staining or western blotting. Cellular protein production was measured with enzyme-linked immunosorbent assay.

BMP-4 induced suppressor of mother against decapentaplegic (Smad)1/5/8 and Smad2/3 phosphorylation and activation. It also promoted higher expression of osteogenic and odontogenic markers, including Osterix, N-cadherin, and secreted protein acidic and rich in cysteine (SPARC), in SCAPs. Additionally, BMP-4 stimulated connective tissue growth factor (CTGF), plasminogen activator inhibitor-1 (PAI-1), and urokinase plasminogen activator receptor (uPAR) expression, but inhibited uPA expression and production in SCAPs, indicating its role in matrix remodeling and cell migration. Inhibition of Smad2/3 with SB431542 and Smad1/5/8 with LDN193189 attenuated the BMP-4-induced expression Osx, N-cadherin, CTGF, SPARC, uPAR and PAI-1.

These results indicate that BMP-4 stimulates the osteogenic and odontogenic differentiation of SCAPs by regulating matrix turnover and mineralization-related proteins. Furthermore, these processes are associated with the induction of Smad2/3 and Smad1/5/8 of SCAPs by BMP-4.

Heat stress is essential for improving the efficacy of mesenchymal stem cell (MSC)-based regeneration medicine. However, it is still unclear whether and how heat stress influences the differentiation of stem cells from apical papilla (SCAPs). This research aimed to explore the potential mechanism of glucose-regulated protein 78 (GRP78) in regulating differentiation under heat stress in SCAPs.

The proliferation ability was assessed using the 5-Ethynyl-2'- deoxyuridine (EdU) assay, cell counting kit assay (CCK-8), and flow cytometry (FCM). The osteogenic and odontogenic differentiation capacities were investigated through Western blot, quantitative reverse transcription polymerase chain reaction (qRT-PCR), alkaline phosphatase (ALP) staining and activity assay, alizarin red S (ARS) staining, as well as immunofluorescence staining. Western blot and transmission electron microscopy (TEM) were used to detect autophagy.

Heat stress enhanced the osteogenic and odontogenic differentiation of SCAPs, but it did not significantly affect proliferation. Besides, GRP78 has been confirmed to modulate the differentiation induced by heat stress. Autophagy triggered by GRP78 enhanced osteogenic and odontogenic differentiation of SCAPs, while the knockdown of GRP78 or the inhibitor of autophagy suppressed the differentiation.

Heat stress induces osteogenic and odontogenic differentiation of SCAPs through GRP78-mediated autophagy.

Dental trauma can cause damage to the pulp tissue in immature teeth. Revascularization therapy is a possible option in the treatment of non-vital, immature permanent teeth with a history of trauma. The aim of this prospective study was to evaluate the radiographic and clinical results of immature teeth with a history of trauma treated by regenerative endodontic procedures and mineral trioxide aggregate apexification techniques.

Forty-one patients aged between 7 and 12 years with traumatized immature permanent maxillary incisors were included in the study. These patients were divided into two groups: those who had previously received endodontic treatment and those who had not. Twenty-four patients who applied directly to the university clinic and had not received endodontic treatment before were included in the regenerative endodontic protocol group (Group 1). Seventeen patients who had previously undergone endodontic intervention on their relevant teeth were included in the mineral trioxide aggregate apexification group (Group 2). The patients were followed for a period of 24 months. Clinical success rates were evaluated, and pre-treatment and control radiographs were analyzed to calculate the percentage increase in root dentin width and root length.

After 24 months of follow-up, positive periapical healing was detected in the radiographic findings in the majority of cases. In Group 1, a limited increase in root length and root dentin width was observed, while a narrowing in the apical opening was evident. The radiographic evaluation of two of the cases, which could be accessed 11 years later, emphasized the importance of long-term follow-up in assessing the effectiveness of the chosen methods.

The revascularization method is a treatment option that has positive results in terms of root development in teeth with necrotic pulp as a result of trauma.

The development of fluorescence-based methods for bioassays and medical diagnostics requires the design and synthesis of specific markers to target biological microobjects. However, biomolecular recognition in real cellular systems is not always as selective as desired. A new concept for creating fluorescent biomolecular probes, utilizing a fluorogenic dye and biodegradable, biocompatible nanomaterials, is demonstrated. The synthesis of a new dicationic asymmetric monomethine cyanine dye with benzo[d]thiazolium-N-propionamide and chloroquinoline end groups is presented. The photophysical properties of the newly synthesized dye were examined through the combined application of spectroscopic and theoretical methods. The applicability of the dye as a fluorogenic nucleic acid probe was proven by UV-VIS spectroscopy and fluorescence titration. The dye-nucleic acid interaction mode was investigated by UV-Vis and CD spectroscopy. The newly synthesized dicationic dye, like other similar fluorogenic structures, limited permeability, which restricts its use as a probe for RNA and DNA. To enhance cellular delivery, we utilized a patented technology that employs solid, insoluble lipid nanoparticles. This method ensures the complete introduction of the dye into cells while minimizing activity outside the cells. In our study involving two human cell lines, we observed improved penetration through the cell membrane and distinctive selectivity in visualizing nucleic acids within the cytoplasm and nucleus.

Dimethyl sulfoxide (DMSO) is widely used as an adjuvant in dissolving insoluble compounds in an aqueous medium; however, it can induce significant molecular changes in cells. The possible damages may occur obeying a tissue-specific profile, and the effect on human apical papilla cells (hAPC) remains unknown. Therefore, this study aimed to evaluate DMSO effects on the viability and mineralization activity in hAPC cultures in vitro and to establish standards of maximum concentrations for its use in laboratory routines. hAPCs were cultured, plated, and maintained in media containing increasing concentrations of Dimethyl sulfoxide (0.1%, 0.5%, 1%, 5%, and 10%) for 24 h, 48 h, 72 h, and 7 days. At each time point, the cells were subjected to the MTT assay. The Alizarin red S staining assay was performed to evaluate the osteo/odontogenic mineralization potential of hAPC DMSO-exposed (0.1%, 0.5%, and 1%) in the 21-day time-point. Statistical analysis was performed using one-way analysis of variance followed by Tukey's post hoc test (p<0.05). In general, the 5% and 10% DMSO concentrations were shown to be cytotoxic for hAPC at all analyzed time points, and the hAPC DMSO-stimulated presented higher osteo/odontogenic mineralization potential. Therefore, the 5% and 10% DMSO concentrations should be avoided, and the mineralization activity assay should be carefully designed in order to avoid biases at in vitro assays using hAPC cultures.

Stem cells offer great promise in dentistry by regenerating critical oral tissues. Derived from sources such as dental pulp, periodontal ligament, and dental follicle, these cells can be pluripotent or multipotent and are capable of differentiating into various cell types, including odontoblasts and osteoblasts. This regenerative potential could transform treatments for dental conditions like periodontal disease and pulpitis. Advancing this field requires understanding stem cell differentiation and exploring tissue engineering and biomaterial scaffolds. Despite the potential, evolving legal and ethical issues shape the integration of these innovations. Research in dental stem cells represents a significant advance in regenerative medicine, promising to enhance dental treatments and patient care.

Guided tissue regeneration (GTR) has been widely used in the periodontal treatment of intrabony and furcation defects for nearly four decades. The treatment outcomes have shown effectiveness in reducing pocket depth, improving attachment gain and bone filling in periodontal tissue. Although applying GTR could reconstruct the periodontal tissue, the surgical indications are relatively narrow, and some complications and race ethic problems bring new challenges. Therefore, it is challenging to achieve a consensus concerning the clinical benefits of GTR. With the appearance of stem cell-based regenerative medicine, mesenchymal stem/stromal cells (MSCs) have been considered a promising cell resource for periodontal regeneration. In this review, we highlight preclinical and clinical periodontal regeneration using MSCs derived from distinct origins, including non-odontogenic and odontogenic tissues and induced pluripotent stem cells, and discuss the transplantation procedures, therapeutic mechanisms, and concerns to evaluate the effectiveness of MSCs.

Ulcerative colitis is an inflammatory bowel disease (IBD) that involves inflammation of the colon lining and rectum. Although a definitive cure for IBD has not been identified, various therapeutic approaches have been proposed to mitigate the symptomatic presentation of this disease, primarily focusing on reducing inflammation. The aim of the present study was to evaluate the therapeutic potential of combining dental pulp stem cells (DPSCs) with sulfasalazine in an acetic acid-induced ulcerative colitis rat model and to assess the impact of this combination on the suppression of inflammatory cytokines and the regulation of oxidative stress in vivo.

Ulcerative colitis was induced in rats through transrectal administration of 3% acetic acid. The therapeutic effect of combining DPSCs and sulfasalazine on UC was evaluated by measuring the colonic weight/length ratio and edema markers; performing histopathological investigations of colon tissue; performing immunohistochemical staining for NF-κB-P65 and IL-1β; and evaluating oxidative stress and antioxidant indices via ELISA. Moreover, the proinflammatory markers NF-κB-P65, TNF-α and TLR-4 were assessed in colon tissue via ELISA. Furthermore, qRT‒PCR was used to estimate the expression levels of the TLR-4, NF-κB-P65, and MYD88 genes in colon tissue.

The investigated macroscopic and microscopic signs of inflammation were markedly improved after the combined administration of sulfasalazine and DPSCs, where a noticeable improvement in histological structure, with an intact mucosal epithelium and mild inflammatory infiltration in the mucosa and submucosa, with slight hemorrhage. The administration of either DPSCs or sulfasalazine, either individually or in combination, significantly reduced ROS levels and significantly increased XOD activity. The immunohistochemical results demonstrated that the combined administration of DPSCs and sulfasalazine attenuated NFκB-p65 and IL-1β expression. Finally, the combined administration of DPSCs and sulfasalazine significantly downregulated MyD88, NF-κB and TLR4 gene expression.

Cotreatment with DPSCs and sulfasalazine had synergistic effects on ulcerative colitis, and these effects were relieved.

The aim of this systematic review was to demonstrate the efficacy of topical application of corticosteroids in remineralization of dental pulp tissues to preserve their vitality and function.

An electronic search was performed using MEDLINE by PubMed, EMBASE, Web of Science (WOS), and Scopus databases. The inclusion criteria were in vitro studies that employed dental pulp tissue obtained from extracted healthy permanent human teeth and were subjected to topical administration of corticosteroids and evaluated tissue remineralization by performing any mineralization assay. A total of 11 studies were selected for inclusion. PRISMA guidelines were followed, and the methodological quality and risk of bias of the included studies were evaluated using the RoBDEMAT guidelines. Also, tables were designed for data extraction, including tissue mineralization and osteogenic differentiation as primary and secondary outcomes, respectively.

Alizarin Red S (ARS) has been able to demonstrate a possible mineralizing power of corticosteroids, applied at an adequate dose. The up-regulation of Alkaline phosphatase (ALP), osteocalcin (OCN), osteopontin (OSP), sialophosphoprotein (DSPP), runt-related transcription factor 2 (RUNX2), collagen type 1 alpha 1(COL1α1) and dentin matrix protein 1 (DMP-1) induced the osteogenic/odontogenic differentiation of dental pulp stem cells (DPSCs).

Deep carious lesions treatment is still challenging in restorative dentistry. Some treatments have been focused on dental pulp tissue remineralization to maintain the function and vitality. After corticosteroids topical application, mineral deposition and osteogenic differentiation have been detected.

The dental pulp's environment is essential for the regulation of mesenchymal stem cells' homeostasis and thus, it is of great importance to evaluate the materials used in regenerative procedures.

To assess in vitro (i) the effect of chitosan nanoparticles, 0.2% chitosan irrigation solution, Dual Rinse®, 17% EDTA, 10% citric acid and 2.5% NaOCl on DSCS viability; (ii) the effect of different concentrations of TGF-β1 on DCSC proliferation; and (iii) whether treatment with TGF-β1 following exposure to the different irrigation solutions could compensate for their negative effects.

(i) DSCS were treated with three dilutions (1:10, 1:100 and 1:1000) of the six irrigation solutions prepared in DMEM for 10 and 60 min to assess the effect on viability. (ii) The effect of different concentrations (0, 1, 5 and 10 ng/mL) of TGF-β1 on DCSC proliferation was assessed at 1, 3 and 7 days. (iii) The proliferative effect of TGF-β1 following 10-min exposure to 1:10 dilution of each irrigation solution was also tested. We used MTT assay to assess viability and proliferation. We performed statistical analysis using Prism software.

(i) The different endodontic irrigation solutions tested showed a significant effect on cell viability (p ≤ .0001). Significant interactions between the endodontic irrigation solutions and their dilutions were also found for all parameters (p ≤ .0001). Chitosan nanoparticles and 0.2% chitosan irrigation solution were the least cytotoxic to DSCS whilst 2.5% NaOCl was the most cytotoxic followed by 17% EDTA. (ii) TGF-β1 at concentrations of 1 and 5 ng/mL resulted in significantly higher proliferation compared to the control group. (iii) Exposure to 17% EDTA or 2.5% NaOCl for 10 min was sufficient to make DSCS cells refractory to the proliferative effects of TGF-β1. DSCS groups treated with TGF-β1 following exposure to chitosan nanoparticles, 0.2% chitosan irrigation solution, Dual Rinse® and 10% CA demonstrated significantly higher proliferation compared to non-TGF-β1-treated groups (p ≤ .0001, p ≤ .0001, p ≤ .0001 and p = .01 respectively).

The current study offers data that can be implemented to improve the outcome of regenerative endodontic procedures by using less toxic irrigation solutions and adding TGF-β1 to the treatment protocol.

Oral cavity stem cells (OCSCs) have been the focus of intense scientific efforts due to their accessibility and stem cell properties. The present work aims to compare the different characteristics of 6 types of dental stem cells derived from the oral cavity: dental pulp stem cells (DPSC), stem cells from human exfoliated deciduous teeth (SHED), periodontal ligament stem cells (PDLSC), stem cells from the apical papilla (SCAP), bone marrow mesenchymal stem cells (BMSC), and gingival mesenchymal stem cells (GMSC). Using immunofluorescence and real-time polymerase chain reaction techniques, we analysed the cells for stem cell, differentiation, adhesion, and extracellular matrix markers; the ability to proliferate in vitro; and multilineage differentiation potential. Markers such as vimentin, CD44, alkaline phosphatase, CD146, CD271, CD49f, Oct 3/4, Sox 9, FGF7, nestin, and BMP4 showed significant differences in expression levels, highlighting the heterogeneity and unique characteristics of each cell type. At the same time, we confirmed that all cell types successfully differentiated into osteogenic, chondrogenic, or adipose lineages, with different readiness. In conclusion, our study reveals the distinct properties and potential applications of various dental-derived stem cells. These findings contribute to a deeper understanding of OCSCs and their significance in future clinical applications.

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.

Calcium silicate-based bioceramics have been applied in endodontics as advantageous materials for years, many chemical components and new synthesizing methods were used to improve the base formulation of the materials for positively affecting the sealers properties. Recently, a novel biomaterial formulation, grounded in strontium silicate, has been introduced to the market, offering potential advancements in the field.

To comparatively analyze the cytotoxicity and cell migration effects of a novel strontium silicate-based bioceramic material (CRoot SP) and those of calcium silicate-based (iRoot SP) and epoxide amine resin (AH Plus) sealers on stem cells derived from rat apical papilla(rSCAPs).

rSCAPs were isolated and characterized in vitro and subsequently cultured in the presence of various concentrations of CRoot SP, iRoot SP and AH Plus extracts. Cytotoxicity was assessed by CCK-8 assay, and cell-migration capacity was assessed by using wound healing assays .

No significant differences in cell viability were observed in the 0.02 mg/mL and 0.2 mg/mL sealer groups. The cell viability of CRoot SP was consistently greater than that of iRoot SP at concentrations of 5 mg/mL and 10 mg/mL across all time points. Maximum cytotoxic effect was noted on day 5 with 10 mg/mL AH Plus.The scratch was partly healed by cell migration in all groups at 24 h, and the 0.02 mg/mL, and 0.2 mg/mL CRoot SP exerted beneficial effects on rSCAPs migration.

CRoot SP exhibited less cytotoxic than the iRoot SP and AH Plus extracts after setting. A lower concentration of CRoot SP thus promotes the cell migration capacity of rSCAPs, and it may achieve better tissue repair during root canal treatment.

Microorganisms, physical factors such as temperature or mechanical injury, and chemical factors such as free monomers from composite resin are the main causes of dental pulp diseases. Current clinical treatment methods for pulp diseases include the root canal therapy, vital pulp therapy and regenerative endodontic therapy. Regenerative endodontic therapy serves the purpose of inducing the regeneration of new functional pulp tissues through autologous revascularization or pulp tissue engineering. This article first discusses the current clinical methods and reviews strategies as well as the research outcomes regarding the pulp regeneration. Then the in vivo models, the prospects and challenges for regenerative endodontic therapy were further discussed.

DNA methylation as intensively studied epigenetic regulatory mechanism exerts pleiotropic effects on dental-derived mesenchymal stem cells (DMSCs). DMSCs have self-renewal and multidifferentiation potential. Here, this review aims at summarizing the research status about application of DMSCs in tissue engineering and clarifying the roles of DNA methylation in influencing the functions of DMSCs, with expectation of paving the way for its in-depth exploration in tissue engineering.

The current research status about influence of DNA methylation in DMSCs was acquired by MEDLINE (through PubMed) and Web of Science using the keywords 'DNA methylation', 'dental-derived mesenchymal stem cells', 'dental pulp stem cells', 'periodontal ligament stem cells', 'dental follicle stem cells', 'stem cells from the apical papilla', 'stem cells from human exfoliated deciduous teeth', and 'gingival-derived mesenchymal stem cells'.

This review indicates DNA methylation affects DMSCs' differentiation and function through inhibiting or enhancing the expression of specific gene resulted by DNA methylation-related genes or relevant inhibitors.

DNA methylation can influence DMSCs in aspects of osteogenesis, adipogenesis, immunomodulatory function, and so on. Yet, the present studies about DNA methylation in DMSCs commonly focus on dental pulp stem cells (DPSCs) and periodontal ligament stem cells (PDLSCs). Little has been reported for other DMSCs.

Circular RNA (circRNA) is a key player in regulating the multidirectional differentiation of stem cells. Previous research by our group found that the blue light-emitting diode (LED) had a promoting effect on the osteogenic/odontogenic differentiation of human stem cells from apical papilla (SCAPs). This research aimed to investigate the differential expression of circRNAs during the osteogenic/odontogenic differentiation of SCAPs regulated by blue LED.

SCAPs were divided into the irradiation group (4 J/cm2) and the control group (0 J/cm2), and cultivated in an osteogenic/odontogenic environment. The differentially expressed circRNAs during osteogenic/odontogenic differentiation of SCAPs promoted by blue LED were detected by high-throughput sequencing, and preliminarily verified by qRT-PCR. Functional prediction of these circRNAs was performed using Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) and the circRNA-miRNA-mRNA networks were also constructed.

It showed 301 circRNAs were differentially expressed. GO and KEGG analyses suggested that these circRNAs were associated with some signaling pathways related to osteogenic/odontogenic differentiation. And the circRNA-miRNA-mRNA networks were also successfully constructed.

CircRNAs were involved in the osteogenic/odontogenic differentiation of SCAPs promoted by blue LED. In this biological process, circRNA-miRNA-mRNA networks served an important purpose, and circRNAs regulated this process through certain signaling pathways.

Pregnancy induces significant changes in oral health because of hormonal fluctuations, making it a crucial period for preventive measures. Dental stem cells (DSCs), particularly those derived from the dental pulp and periodontal ligaments, offer promising avenues for regenerative therapies and, possibly, preventive interventions. While the use of DSCs already includes various applications in regenerative dentistry in the general population, their use during pregnancy requires careful consideration. This review explores recent advancements, challenges, and prospects in using DSCs to address oral health issues, possibly during pregnancy. Critical aspects of the responsible use of DSCs in pregnant women are discussed, including safety, ethical issues, regulatory frameworks, and the need for interdisciplinary collaborations. We aimed to provide a comprehensive understanding of leveraging DSCs to improve maternal oral health.

 The aim of this study was to evaluate the osteogenic differentiation ability and proliferation of apical papilla stem cells (SCAPs) using chitosan-coated nanohydroxyapatite and bioactive glass nanoparticles.

 Hydroxyapatite, chitosan-coated nanohydroxyapatite, and bioactive glass 45S5 nanoparticles were prepared and characterized using a transmission electron microscope and X-ray diffraction. SCAPs were harvested from freshly extracted impacted wisdom teeth, cultured, and characterized using flow cytometric analysis. Tested nanomaterials were mixed and samples were classified into five equal groups as follows: negative control group: SCAP with Dulbecco's modified eagle's medium, positive control group: SCAP with inductive media, first experimental group: nanohydroxyapatite with SCAP, second experimental group: chitosan-coated nanohydroxyapatite with SCAP, third experimental group: bioactive glass nanoparticles with SCAP. Osteoblastic differentiation was assessed using an alkaline phosphatase (ALP) assay. Receptor activator of nuclear factor kappa beta ligand (RANKL) expression was evaluated using specific polyclonal antibodies by fluorescence microscope. The proliferation of SCAP was assessed using cell count and viability of trypan blue in addition to an 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay.

 Isolated SCAP showed a nonhematopoietic origin. Chitosan-coated nanohydroxyapatite showed the highest ALP concentration followed by nanobioactive glass, nanohydroxyapatite, and negative control. Chitosan-coated nanohydroxyapatite showed the highest H score followed by nanobioactive glass, nanohydroxyapatite, and negative control in RANKL expression. Chitosan-coated nanohydroxyapatite showed the highest viable cell count.

 SCAP isolation is achievable from extracted fully impacted immature third molars. All tested biomaterials have the ability to induce osteogenic differentiation and proliferation of SCAP. Composite nanoparticle materials show better osteogenic differentiation and proliferation of SCAP than single nanoparticles.

Stem cells (SCs) undergo asymmetric division, producing transit-amplifying cells (TACs) with increased proliferative potential that move into tissues and ultimately differentiate into a specialized cell type. Thus, TACs represent an intermediary state between stem cells and differentiated cells. In the cornea, a population of stem cells resides in the limbal region, named the limbal epithelial stem cells (LESCs). As LESCs proliferate, they generate TACs that move centripetally into the cornea and differentiate into corneal epithelial cells. Upon limbal injury, research suggests a population of progenitor-like cells that exists within the cornea can move centrifugally into the limbus, where they dedifferentiate into LESCs. Herein, we summarize recent advances made in understanding the mechanism that governs the differentiation of LESCs into TACs, and thereafter, into corneal epithelial cells. We also outline the evidence in support of the existence of progenitor-like cells in the cornea and whether TACs could represent a population of cells with progenitor-like capabilities within the cornea. Furthermore, to gain further insights into the dynamics of TACs in the cornea, we outline the most recent findings in other organ systems that support the hypothesis that TACs can dedifferentiate into SCs.

Circular RNAs (circRNAs) are a novel class of endogenous non-coding RNAs (ncRNAs) which unlike linear RNAs, have a covalently closed continuous loop structure. circRNAs are found abundantly in human cells and their biology is complex. They feature unique expression to different types of cells, tissues, and developmental stages. To the present, the functional roles of circular RNAs are not fully understood. They reportedly act as microRNA (miRNA) sponges, therefore having key regulatory functions in diverse physiological and pathological processes. As for dentistry field, lines of evidence indicate that circRNAs play vital roles in the odontogenic and osteogenic differentiation of dental pulp stem cells (DPSCs) and periodontal ligament stem cells (PDLSCs). Abnormal expression of circRNAs have been found in other areas of pathology frequently reflected also in the oral environment, such as inflammation or bone and soft tissue loss. Therefore, circRNAs could be of significant importance in various fields in dentistry, especially in bone and soft tissue engineering and regeneration. Understanding the molecular mechanisms occurring during the regulation of oral biological and tissue remodeling processes could augment the discovery of novel diagnostic biomarkers and therapeutic strategies that will improve orthodontic and other oral therapeutic protocols.

To evaluate the osteogenic potential of platelet-rich fibrin (PRF) and low-level laser therapy (LLLT) on human stem cells from the apical papilla (SCAP) we isolated, characterized, and then cultured in an osteogenic medium cells with PRF and/or LLLT (660 nm, 6 J/m2-irradiation). Osteogenic differentiation was assessed by bone nodule formation and expression of bone morphogenetic proteins (BMP-2 and BMP-4), whereas the molecular mechanisms were achieved by qRT-PCR and RNA-seq analysis. Statistical analysis was performed by ANOVA and Tukey's post hoc tests (p < 0.05* and p < 0.01**). Although PRF and LLLT increased bone nodule formation after 7 days and peaked at 21 days, the combination of PRF + LLLT led to the uppermost nodule formation. This was supported by increased levels of BMP-2 and -4 osteogenic proteins (p < 0.005). Furthermore, the PRF + LLLT relative expression of specific genes involved in osteogenesis, such as osteocalcin, was 2.4- (p = 0.03) and 28.3- (p = 0.001) fold higher compared to the PRF and LLLT groups, and osteopontin was 22.9- and 1.23-fold higher, respectively (p < 0.05), after 7 days of interaction. The transcriptomic profile revealed that the combination of PRF + LLLT induces MSX1, TGFB1, and SMAD1 expression, after 21 days of osteogenic differentiation conditions exposition. More studies are required to understand the complete cellular and molecular mechanisms of PRF plus LLLT on stem cells. Overall, we demonstrated for the first time that the combination of PRF and LLLT would be an excellent therapeutic tool that can be employed for dental, oral, and craniofacial repair and other tissue engineering applications.

Notum is a secreted deacylase, which is crucial for tooth dentin development in mice. This study aimed to investigate the effect of NOTUM on the odontoblastic differentiation of human stem cells from the apical papilla (hSCAPs), to reveal the potential value of NOTUM in pulp-dentin complex regeneration.

The expression pattern of NOTUM in human tooth germs and during in vitro odontoblastic differentiation of hSCAPs was evaluated by immunohistochemical staining, and quantitative polymerase chain reaction, respectively. To manipulate the extracellular NOTUM level, ABC99 or small interfering RNA was used to down-regulate it, while recombinant NOTUM protein was added to up-regulate it. The effects of changing NOTUM level on the odontoblastic differentiation of hSCAPs and its interaction with the WNT/β-catenin signaling pathway were studied using alkaline phosphatase staining, alizarin red staining, quantitative polymerase chain reaction, and western blot.

NOTUM was observed in the apical papilla of human tooth germs. During in vitro odontoblastic differentiation of hSCAPs, NOTUM expression initially increased, while the WNT/β-catenin pathway was activated. Downregulation of NOTUM hindered odontoblastic differentiation. Recombinant NOTUM protein had varying effects on odontoblastic differentiation depending on exposure duration. Continuous addition of the protein inhibited both odontoblastic differentiation and the WNT/β-catenin pathway. However, applying the protein solely in the first 3 days enhanced odontoblastic differentiation and up-regulated the WNT/β-catenin pathway.

NOTUM demonstrated a bidirectional impact on in vitro odontoblastic differentiation of hSCAPs, potentially mediated by the WNT/β-catenin pathway. These findings suggest its promising potential for pulp-dentin complex regeneration.

To evaluate the cytocompatibility, bioactivity, and anti-inflammatory potential of the new pre-mixed calcium silicate cement-based sealers Bioroot Flow (BrF) and AH Plus Bioceramic Sealer (AHPbcs) on human periodontal ligament stem cells (hPDLSCs) compared to the epoxy resin-based sealer AH Plus (AHP).

Standardized discs and 1:1, 1:2, and 1:4 eluates of BrF, AHPbcs and AHP after setting were prepared. The following assays were performed: cell attachment and morphology via SEM, cell viability via a MTT assay, cell migration/proliferation via a wound-healing assay, cytoskeleton organization via immunofluorescence staining; cytokine release via ELISA; osteo/cemento/odontogenic marker expression via RT-qPCR, and cell mineralized nodule formation via Alizarin Red S staining. HPDLSCs were isolated from extracted third molars from healthy patients. Comparisons were made with hPDLSCs cultured in unconditioned (negative control) or osteogenic (positive control) culture media. Statistical significance was established at p < 0.05.

Both BrF and AHPbcs showed significantly positive results in the cytocompatibility assays (cell metabolic activity, migration, attachment, morphology, and cytoskeleton organization) compared with a negative control group, while AHP showed significant negative results. BrF exhibited an upregulation of at least one osteo/cementogenic marker compared to the negative and positive control groups. BrF showed a significantly higher calcified nodule formation than AHPbcs, the negative and positive control groups, while AHPbcs was higher than the negative control group. Both were also significantly higher than AHP group.

BrF and AHPbcs exhibit adequate and comparable cytocompatibility on hPDLSCs. BrF also promoted the osteo/cementogenic differentiation of hPDLSCs. Both calcium silicate-based sealers favored the downregulation of the inflammatory cytokine IL-6 and the calcified nodule formation from hPDLSCs. BrF exerted a significantly higher influence on cell mineralization than AHPbcs.

This is the first study to elucidate the biological properties and immunomodulatory potential of Bioroot Flow and AH Plus Bioceramic Sealer. The results act as supporting evidence for their use in root canal treatment.

The aim of this study was to analyze the effect of lipopolysaccharides (LPS) on the biological properties of stem cells from the apical papilla (SCAPs), such as viability, adhesion to dentin, odontoblast-like differentiation, mineralization, and release of immunomodulatory cytokines.

SCAPs were isolated from immature teeth of three donors (10 to 15 years old) and cultured in mineralizing media with or without 1 μg/mL lipopolysaccharide (LPS). Cells were seeded and cultured under standardized conditions; viability was assessed by MTT assay on days 1, 3, 5, and 7; adhesion to dentin was analyzed using an environmental scanning electron microscope after 2 days; the expression of odontogenic and mineralization genes (DSPP, DMP-1, OCN, Col1A1) was evaluated through qPCR after 14 days, mineralization was evaluated with alizarin red staining after 21 days; and the release of immunomodulatory cytokines (IL-6 and IL-10) was measured by ELISA after 1 and 7 days. The Kruskal-Wallis test was performed to detect the effect of LPS on SCAPs, followed by the Dunn-Sidak test.

LPS presence in the culture media affected SCAPs viability on day 5 and increased IL-6 secretion by day 7, however, SCAPs retained the adhesion to dentin and mineralization capacities, as well as the differentiation capacity into a mineralizing phenotype.

In conclusion, within the limitations of this in vitro study, and under the inflammatory microenvironment simulated in this study, stem cells from the apical papilla were found with retained adhesion capacity to dentin, differentiation into a mineralizing phenotype, mineralization, and release of IL-10.

The aim of the study was to investigate the morphometric parameters of dental pulp in open apices immature teeth in a sheep model after mechanical pulp exposure and restoration with reinforced zinc oxide-eugenol.

In this experimental study, a total of 12 immature mandibular central incisors from six adult male sheep, weighing 30-40 kg and with the age of 1 year old with Merino race were examined. After anesthesia, the pulps of the teeth in the case group were mechanically exposed and then were restored with reinforced zinc oxide-eugenol and amalgam. In the control group, the teeth remained intact. The animals were sacrificed at intervals of 2, 4, 6, and 8 weeks (E2, E4, E6, and E8) in the case and 2 and 8 weeks (C2 and C8) in the control groups. Then, their teeth were removed with the surrounding supporting tissues and alveolar bones. Tissue processing and staining were done, and the sections were examined under a light microscope. The Kruskal-Wallis and Mann-Whitney U tests were used to analyze the data and compare the changes between the two groups. P < 0.05 was considered statistically significant.

In response to mechanical exposure, reparative or tertiary dentin was formed, and its thickness increased during the time of the study. The thickness of the odontoblastic layer in the E4 group was the highest amount. The pulp chamber diameter in the C2 group was significantly larger than the other groups, and the diameter of the apical foramen in the E8 was decreased significantly compared to the controls (P < 0.05).

In response to mechanical exposure and restoration with reinforced zinc oxide-eugenol, some morphometric parameters of the dental pulp changed significantly in the sheep model compared to the controls.