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Background Objective Methods Results Conclusion Dental pulp tissue engineering is expected to become an ideal treatment for irreversible pulpitis and apical periodontitis. However, angiogenesis and neurogenesis for functional pulp regeneration have not yet met the standard for large‐scale clinical application, and need further research.This review focused on the potential mechanisms of angiogenesis and neurogenesis in pulp regeneration, including stem cell types, upstream and downstream regulatory molecules and cascade signalling pathways, thereby providing a theoretical basis and inspiring new ideas to improve the effectiveness of dental pulp tissue engineering.An electronic literature search was carried out using the keywords of ‘pulp regeneration’, ‘stem cell transplantation’, ‘dental pulp stem cells’, ‘angiogenesis’ and ‘neurogenesis’. The resulting literature was screened and reviewed.Stem cells used in dental pulp tissue engineering can be classified as dental‐derived and non‐dental‐derived stem cells, amongst which dental pulp stem cells (DPSC) have achieved promising results in animal experiments and clinical trials. Multiple molecules and signalling pathways are involved in the process of DPSC‐mediated angiogenic and neurogenetic regeneration. In order to promote angiogenesis and neurogenesis in pulp regeneration, feasible measures include the addition of growth factors, the modulation of transcription factors and signalling pathways, the use of extracellular vesicles and the modification of bioscaffold materials.Dental pulp tissue engineering has had breakthroughs in preclinical and clinical studies in vivo. Overcoming difficulties in pulpal angiogenesis and neurogenesis, and achieving functional pulp regeneration will lead to a significant impact in endodontics. [ABSTRACT FROM AUTHOR]

The dehydrated human amnion–chorion membranes (dHACMs) derived from the human placenta have emerged as a promising biomaterial for dental pulp regeneration owing to their unique biological and structural properties. The purpose of this review is to explore the potentials of dHACMs in dental pulp tissue engineering, focusing on their ability to promote cellular proliferation, differentiation, angiogenesis, and neurogenesis. dHACMs are rich in extracellular matrix proteins and growth factors such as TGF-β1, FGF2, and VEGF. They also exhibit significant anti-inflammatory and antimicrobial properties, creating an optimal environment for dental pulp regeneration. The applications of dHACMs in regenerative endodontic procedures are discussed, highlighting their ability to support the formation of dentin and well-vascularized pulp-like tissue. This review demonstrates that dHACMs hold significant potential for enhancing the success of pulp regeneration and offer a biologically based approach to preserve tooth vitality and improve tooth survival. Future research is expected to focus on conducting long-term clinical studies to establish their efficacy and safety. [ABSTRACT FROM AUTHOR]

Objective: To develop a detergent-enzymatic method and evaluate the quality of a decellularized pulp scaffold for regenerative endodontics.Materials and methods: Biomaterial and mesenchymal stem cells (MSCs) were derived from dental pulp that was obtained following third molar extraction indicated for orthodontic reasons in patients aged 14-18 years. The detergent-enzymatic method enabled to obtain a decellularized scaffold from pulp samples. The proliferative activity and viability of dental pulp-derived MSCs were assessed using trypan blue staining and XTT assay. To assess tissue response, Wistar rats underwent subcutaneous implantation of native and decellularized dental pulp. Explanted samples were stained with hematoxylin-eosin on days 7 and 14.Results: The detergent-enzymatic treatment of the dental pulp demonstrated the absence of nuclear material, whereas the histoarchitecture of the dental pulp was disturbed. The DNA content in the sample of the decellularized scaffold was 22.79 ± 2.1 ng/mg of tissue; the amount of DNA in the native sample was 78.5 ± 5.4 ng/mg of tissue. According to XTT assay results, no cytotoxicity of the decellularized scaffold against MSCs was found. Biopsy specimens of the rats with implanted decellularized dental pulp were characterized by no signs of inflammation.Conclusions: The study results will enable to create a biomaterial that can be the base of a tissue-engineered structure of the dental pulp and be used for the regeneration of the pulp-dentin complex.

Successful dental pulp regeneration is closely associated with rapid revascularization and angiogenesis, processes driven by the Jagged1(JAG1)/Notch signaling pathway. However, soluble Notch ligands have proven ineffective in activating this pathway. To overcome this limitation, a Notch signaling hydrogel is developed by indirectly immobilizing JAG1, aimed at precisely directing the regeneration of vascularized pulp tissue. This hydrogel displays favorable mechanical properties and biocompatibility. Cultivating dental pulp stem cells (DPSCs) and endothelial cells (ECs) on this hydrogel significantly upregulate Notch target genes and key proangiogenic markers expression. Three‐dimensional (3D) culture assays demonstrate Notch signaling hydrogels improve effectiveness by facilitating encapsulated cell differentiation, enhancing their paracrine functions, and promoting capillary lumen formation. Furthermore, it effectively communicates with the Wnt signaling pathway, creating an odontoinductive microenvironment for pulp‐dentin complex formation. In vivo studies show that short‐term transplantation of the Notch signaling hydrogel accelerates angiogenesis, stabilizes capillary‐like structures, and improves cell survival. Long‐term transplantation further confirms its capability to promote the formation of pulp‐like tissues rich in blood vessels and peripheral nerve‐like structures. In conclusion, this study introduces a feasible and effective hydrogel tailored to specifically regulate the JAG1/Notch signaling pathway, showing potential in advancing regenerative strategies for dental pulp tissue. [ABSTRACT FROM AUTHOR]

Background: Dental pulp regeneration aims to restore the function and vitality of the dental pulp, which is the soft tissue inside the tooth. Research in this field is effective in trying to improve clinical practices and procedures. This study aimed to analyze the literature related to dental pulp regeneration and to create a documented research perspective for this field. Materials and Methods: This bibliometric study analyzes the research outputs of the subject area of dental pulp regeneration indexed in the Web of Science database between 2013 and 2023. SciMAT software was used to visualize and predict the trends in research on the topic. Results: In general, it has been observed that the stem cell cluster consistently had the highest number of articles across all periods.As we progressed through time, the significance of this cluster continued to grow, eventually becoming a crucial component in the motor theme during the last period. In addition, a significant portion of the studies conducted during different periods focused on identifying suitable materials for scaffold formation. Various materials, including polymers and bioactive glasses, were proposed as viable options for scaffold formation in different periods. Conclusion: Dental pulp stem cells (DPSCs), scaffold networks, growth factors, and regulatory factors are the three main factors that influence dental pulp regeneration. By analyzing maps and thematic clusters in dental pulp regeneration research, as well as considering indicators such as repetition frequency, centrality, and citation of these clusters, researchers can identify the strengths, weaknesses, and gaps in current research. [ABSTRACT FROM AUTHOR]

Background and Objectives: The present systematic review aims to assess the success rate of the pulp regeneration treatment, according to the American Association of Endodontists (AAE) criteria, using different bioactive materials in permanent teeth of pediatric subjects (6-17 years of age). Materials and Methods: The study protocol was registered on PROSPERO and adhered to the Preferred Reporting Items for Systematic Reviews and Meta-analyses statement. The question formulation was accomplished using the PICO model, and an electronic search was carried out on Scopus, MEDLINE/PubMed, Web of Science, and Cochrane databases till April 1, 2023. A total of 30 studies were established to fulfill the inclusion criteria of this systematic review. Results: A total of 273 teeth have been treated with pulp regeneration treatment. By comparing different biomaterials and the success criteria defined by the AAE, the material associated with a higher success rate was found to be the white mineral trioxide aggregate. However, the overall success rate of pulp regeneration treatment was reported for 248 out of 273 teeth (91.20%). Conclusions: Data obtained support the potential that regenerative endodontics aids in continuing root development in permanent immature teeth. Further studies are needed for a more extensive evaluation of the use of different biomaterials and the success rate in regenerative endodontics. [ABSTRACT FROM AUTHOR]

Abstract Dental pulp regeneration is a promising strategy for addressing tooth disorders. Incorporating this strategy involves the fundamental challenge of establishing functional vascular networks using dental pulp stem cells (DPSCs) to support tissue regeneration. Current therapeutic approaches lack efficient and stable methods for activating DPSCs. In the study, we used a chemically modified microRNA (miRNA)-loaded tetrahedral-framework nucleic acid nanostructure to promote DPSC-mediated angiogenesis and dental pulp regeneration. Incorporating chemically modified miR-126-3p into tetrahedral DNA nanostructures (miR@TDNs) represents a notable advancement in the stability and efficacy of miRNA delivery into DPSCs. These nanostructures enhanced DPSC proliferation, migration, and upregulated angiogenesis-related genes, enhancing their paracrine signaling effects on endothelial cells. This enhanced effect was substantiated by improvements in endothelial cell tube formation, migration, and gene expression. Moreover, in vivo investigations employing matrigel plug assays and ectopic dental pulp transplantation confirmed the potential of miR@TDNs in promoting angiogenesis and facilitating dental pulp regeneration. Our findings demonstrated the potential of chemically modified miRNA-loaded nucleic acid nanostructures in enhancing DPSC-mediated angiogenesis and supporting dental pulp regeneration. These results highlighted the promising role of chemically modified nucleic acid-based delivery systems as therapeutic agents in regenerative dentistry and tissue engineering. Graphical abstract

Dental pulp regeneration is challenging in endodontics. Cellular therapy is an alternative approach to induce dental pulp regeneration. Mesenchymal stromal cells (MSCs) have the capacity to induce dental pulp-like tissue formation. In this study, we evaluated the capacity of allogeneic bone marrow MSCs (BM-MSCs) to regenerate pulp following necrosis and apical periodontitis in children's permanent immature apex teeth. Patients aged 8 to 12 years with pulp necrosis and apical periodontitis were evaluated. The study included 15 teeth (13 incisors and 2 molars) from 14 patients (8 boys and 6 girls). Radiographic evaluation showed periapical radiolucency and immature apex teeth. There was no response to cold or electric pulp testing. The root canal of each tooth was cleaned, shaped, and Ca(OH) 2 used as an interappointment medication. Cryopreserved allogeneic BM-MSCs were thawed, expanded, incorporated into preclotted platelet-rich plasma, and implanted into the tooth's pulp cavity. They were sealed with bioceramic cement and composite. Sensibility, apical foramen, calcium deposits within the root canal, and resolution of periapical lesions were evaluated in each tooth over the following 12 months. Based on 9 variables established for dental pulp-like tissue regeneration, all MSC-treated teeth showed evidence of successful regeneration. Clinical and radiographic evaluation of the treated teeth showed periapical lesion healing, sensitivity to cold and electricity, decreased width of the apical foramen, and mineralization within the canal space. Transplantation of allogeneic MSCs induces the formation of dental pulp-like tissue in permanent immature apex teeth with pulp necrosis and apical periodontitis. Implant of MSCs constitutes a potential therapy in regenerative endodontics in pediatric dentistry. Future studies incorporating a larger sample size may confirm these results. [ABSTRACT FROM AUTHOR]

Objectives: Migration and differentiation of human dental pulp stem cells (hDPSCs) is a vital and key factor in the success of reparative dentin formation for maintenance of pulp vitality. Pulp capping materials are used to stimulate DPSCs to induce new dentin formation. Thus, the aim of the present study was to compare the response of DPSCs to four commercially available pulp capping materials: a bioactive bioceramic (Material 1), a nonresinous ready‐to‐use bioceramic cement (Material 2), a bioactive composite (Material 3), and a biocompatible, dual‐cured, resin‐modified calcium silicate (Material 4). Materials and Methods: hDPSCs were isolated and cultured from freshly extracted teeth and were then characterized by flow cytometry and multilineage differentiation. Discs prepared from pulp capping materials were tested with hDPSCs and MTT (3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide) assay, cell migration assay and odontogenic differentiation assay was performed. Expression of osteogenic markers (osteopontin, RUNX family transcription factor 2, osteocalcin) and the odontogenic marker (dentin sialophosphoprotein) was detected using reverse transcription‐polymerase chain reaction. Results: Materials 1, 2, and 3 generated more cell viability than Material 4. Furthermore, Material 4 showed the least wound exposure percentage, while Material 3 showed the highest percentage. Enhanced mineralization was found in hDSCPs cultured with Material 3, followed by Material 1, and then Material 2, while Material 4 revealed the least calcified mineralization. Conclusions: The results of this study were inconclusive regards contemporary bioceramic materials designed for vital pulp therapy as they have different effects on hDPSC. Further testing for cytotoxicity using live‐dead staining, animal experiments, clinical trials, and independent analyses of these biomaterials is necessary for clinicians to make an informed decision for their use. [ABSTRACT FROM AUTHOR]

Abstract Objectives Migration and differentiation of human dental pulp stem cells (hDPSCs) is a vital and key factor in the success of reparative dentin formation for maintenance of pulp vitality. Pulp capping materials are used to stimulate DPSCs to induce new dentin formation. Thus, the aim of the present study was to compare the response of DPSCs to four commercially available pulp capping materials: a bioactive bioceramic (Material 1), a nonresinous ready‐to‐use bioceramic cement (Material 2), a bioactive composite (Material 3), and a biocompatible, dual‐cured, resin‐modified calcium silicate (Material 4). Materials and Methods hDPSCs were isolated and cultured from freshly extracted teeth and were then characterized by flow cytometry and multilineage differentiation. Discs prepared from pulp capping materials were tested with hDPSCs and MTT (3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide) assay, cell migration assay and odontogenic differentiation assay was performed. Expression of osteogenic markers (osteopontin, RUNX family transcription factor 2, osteocalcin) and the odontogenic marker (dentin sialophosphoprotein) was detected using reverse transcription‐polymerase chain reaction. Results Materials 1, 2, and 3 generated more cell viability than Material 4. Furthermore, Material 4 showed the least wound exposure percentage, while Material 3 showed the highest percentage. Enhanced mineralization was found in hDSCPs cultured with Material 3, followed by Material 1, and then Material 2, while Material 4 revealed the least calcified mineralization. Conclusions The results of this study were inconclusive regards contemporary bioceramic materials designed for vital pulp therapy as they have different effects on hDPSC. Further testing for cytotoxicity using live‐dead staining, animal experiments, clinical trials, and independent analyses of these biomaterials is necessary for clinicians to make an informed decision for their use.

Background: Regenerative endodontic procedures (REPs) have emerged as a transformative approach to treating immature permanent teeth with necrotic pulp tissue. Materials and Methods: A prospective study was conducted, enrolling 100 patients with immature permanent teeth requiring REPs. All procedures were performed by a single experienced endodontist following established protocols. Patients were followed up for a minimum of 5 years' post-treatment. Clinical examinations, radiographic assessments, and patient-reported outcomes were recorded at regular intervals. Data were analyzed using statistical methods to determine the success rates, complications, and factors influencing long-term outcomes. Results: The results of this original research reveal a significant and sustained success rate for REPs. After a minimum follow-up period of 5 years, an arbitrary value of 92% for tooth survival was achieved. Radiographic assessments demonstrated consistent healing of apical lesions, and continued root development was observed in the majority of cases. Patient-reported outcomes indicated a high level of satisfaction with the procedure. Complications such as crown discoloration and tooth fracture occurred in a minority of cases but were effectively managed without compromising the overall success of REPs. Conclusion: This original research provides strong evidence for the long-term success of REPs in the treatment of immature permanent teeth with necrotic pulp tissue. The high tooth survival rate, continued root development, and patient satisfaction support the efficacy of REPs as a reliable treatment option.

The present challenge in dental pulp tissue engineering scaffold materials lies in the development of tissue-specific scaffolds that are conducive to an optimal regenerative microenvironment and capable of accommodating intricate root canal systems. This study utilized porcine dental pulp to derive the decellularized extracellular matrix (dECM) via appropriate decellularization protocols. The resultant dECM was dissolved in an acid pepsin solution to form dECM hydrogels. The analysis encompassed evaluating the microstructure and rheological properties of dECM hydrogels and evaluated their biological properties, including in vitro cell viability, proliferation, migration, tube formation, odontogenic, and neurogenic differentiation. Gelatin methacrylate (GelMA) hydrogel served as the control. Subsequently, hydrogels were injected into treated dentin matrix tubes and transplanted subcutaneously into nude mice to regenerate dental pulp tissue in vivo. The results showed that dECM hydrogels exhibited exceptional injectability and responsiveness to physiological temperature. It supported the survival, odontogenic, and neurogenic differentiation of dental pulp stem cells in a 3D culture setting. Moreover, it exhibited a superior ability to promote cell migration and angiogenesis compared to GelMA hydrogel in vitro. Additionally, the dECM hydrogel demonstrated the capability to regenerate pulp-like tissue with abundant blood vessels and a fully formed odontoblast-like cell layer in vivo. These findings highlight the potential of porcine dental pulp dECM hydrogel as a specialized scaffold material for dental pulp regeneration. [ABSTRACT FROM AUTHOR]

Background: Dental pulp regeneration aims to restore the function and vitality of the dental pulp, which is the soft tissue inside the tooth. Research in this field is effective in trying to improve clinical practices and procedures. This study aimed to analyze the literature related to dental pulp regeneration and to create a documented research perspective for this field. Materials and Methods: This bibliometric study analyzes the research outputs of the subject area of dental pulp regeneration indexed in the Web of Science database between 2013 and 2023. SciMAT software was used to visualize and predict the trends in research on the topic. Results: In general, it has been observed that the stem cell cluster consistently had the highest number of articles across all periods. As we progressed through time, the significance of this cluster continued to grow, eventually becoming a crucial component in the motor theme during the last period. In addition, a significant portion of the studies conducted during different periods focused on identifying suitable materials for scaffold formation. Various materials, including polymers and bioactive glasses, were proposed as viable options for scaffold formation in different periods. Conclusion: Dental pulp stem cells (DPSCs), scaffold networks, growth factors, and regulatory factors are the three main factors that influence dental pulp regeneration. By analyzing maps and thematic clusters in dental pulp regeneration research, as well as considering indicators such as repetition frequency, centrality, and citation of these clusters, researchers can identify the strengths, weaknesses, and gaps in current research.

Background: Regenerative endodontic procedures (REPs) have emerged as a transformative approach to treating immature permanent teeth with necrotic pulp tissue. Materials and Methods: A prospective study was conducted, enrolling 100 patients with immature permanent teeth requiring REPs. All procedures were performed by a single experienced endodontist following established protocols. Patients were followed up for a minimum of 5 years' post-treatment. Clinical examinations, radiographic assessments, and patient-reported outcomes were recorded at regular intervals. Data were analyzed using statistical methods to determine the success rates, complications, and factors influencing long-term outcomes. Results: The results of this original research reveal a significant and sustained success rate for REPs. After a minimum follow-up period of 5 years, an arbitrary value of 92% for tooth survival was achieved. Radiographic assessments demonstrated consistent healing of apical lesions, and continued root development was observed in the majority of cases. Patient-reported outcomes indicated a high level of satisfaction with the procedure. Complications such as crown discoloration and tooth fracture occurred in a minority of cases but were effectively managed without compromising the overall success of REPs. Conclusion: This original research provides strong evidence for the long-term success of REPs in the treatment of immature permanent teeth with necrotic pulp tissue. The high tooth survival rate, continued root development, and patient satisfaction support the efficacy of REPs as a reliable treatment option. [ABSTRACT FROM AUTHOR]

BACKGROUND: Revascularization plays an important role in tissue engineering. Dental pulp chamber has its unique anatomy structure, which is surrounded by hard dentin. The blood supply of dental pulp only comes from the narrow apical opening, which makes revascularization more difficult in dental pulp regeneration. OBJECTIVE: To conclude the research advances focusing on the designs of signal molecules, scaffolds and cells in promoting vascularization in dental pulp regeneration. METHODS: We searched the articles on PubMed and CNKI databases with the keywords of “dental pulp regeneration, revascularization, growth factor, material, scaffold, dental pulp stem cell, prevascularization, coculture” in Chinese and English, respectively. Finally, 69 articles met the criteria for review. RESULTS AND CONCLUSION: (1) Vascular endothelial growth factor, platelet-derived growth factor and fibroblast growth factor are the most studied proangiogenic factors, which promote angiogenesis by promoting the proliferation and migration of endothelial and perivascular cells, the recruitment of stem cells and their differentiation to endothelial cells. (2) To maintain the appropriate concentration of pro-angiogenic factors in the microenvironment, the addition of heparin, laponite and cellulose nanocrystals to the scaffolds can solubilize, encapsulate, adsorb, and covalently cross-link signal molecules, which mimic in vivo storage and release mechanisms. Natural materials including decellularized dental pulp matrix, platelet-rich plasma, and platelet-rich fibrin are rich in bioactive molecules and their structures can protect and control the release of these signal molecules. (3) Different cell sub-populations of dental pulp stem cells have different angiogenic potentials. Moreover, stem cells can be transfected by biological and chemical vectors to improve the cells’ ability to secrete signal molecules and to promote the transformation of stem cells to endothelial cells. Therefore, the angiogenic capacity of transfected cells is significantly enhanced. Co-implantation of stem cells and endothelial cells can also enhance angiogenic capacity. (4) Prevascularization may be the most powerful means to promote rapid vascularization of dental pulp regeneration. In vitro co-culture of endothelial cells and stem cells with a certain ratio can form microtissues containing vascular structures. Microvascular fragments can be obtained by enzymatic digestion of adipose tissue, in which tubular structures and functional cells can promote the development of new blood vessels and anastomosis with host vessels. (5) Current clinical studies on revascularization of dental pulp are limited to dental pulprevascularization, where signal molecules are added while stimulating the formation of blood clots in the root canal and the effects are observed. While other research directions mainly involve in vivo trials in small and large animals, and further clinical trials are still needed to observe regenerative effects, mitigate side effects, and optimize the preparation process. [ABSTRACT FROM AUTHOR]

OBJECTIVE: To summarize research progress on forming dental pulp-like tissue from dental pulp stem cells via a co-culture system, as well as provide ideas for stem cell therapy for dental pulp tissue regeneration. METHODS: The first author searched PubMed, CNKI, and Wanfang databases for relevant literature published before May 2022. The English search terms were “dental pulp regeneration, dental pulp stem cell, co-culture, tissue engineering, signaling pathway”, and the Chinese search terms were “dental pulp regeneration, dental pulp stem cells, co-culture, tissue engineering, scaffold, signaling pathway”. Totally 67 articles meeting the criteria were summarized and described. RESULTS AND CONCLUSION: (1) Different combinations of cells are used in co-culture systems to form the main body of the cell and the combined body. (2) Direct co-culture and indirect co-culture are the two types of combination modes at present. Through the intervention of biological scaffold materials and a change in culture conditions, the co-culture outcome can be divided into two-dimensional regenerated tissues and three-dimensional regenerated tissues. (3) Direct co-culture creates an extracellular matrix and microenvironment that facilitates stem cell proliferation and differentiation through intercellular contact. (4) Indirect co-culture addresses the problem of insufficient sources of combined cells and immune rejection between individuals. The biological factors produced by the combined cells can be more easily made into clinical drugs and biological scaffold materials. (5) Though the co-culture system has superior performance in angiogenesis, there are few reports on other structures, and the regeneration of tissue from the co-culture outcome is uncertain. Furthermore, there are limitations to applying newly generated tissue generated by the co-culture system to clinical practice due to tooth anatomical constraints and rejection mechanisms. [ABSTRACT FROM AUTHOR]

One of the difficulties of pulp regeneration is the rapid vascularization of transplanted engineered tissue, which is crucial for the initial survival of the graft and subsequent pulp regeneration. At present, prevascularization techniques, as emerging techniques in the field of pulp regeneration, has been proposed to solve this challenge and have broad application prospects. In these techniques, endothelial cells and pericytes are cocultured to induce intercellular communication, and the cell coculture is then introduced into the customized artificial vascular bed or induced to self-assembly to simulate the interaction between cells and extracellular matrix, which would result in construction of a prevascularization system, preformation of a functional capillary network, and rapid reconstruction of a sufficient blood supply in engineered tissue after transplantation. However, prevascularization techniques for pulp regeneration remain in their infancy, and there remain unresolved problems regarding cell sources, intercellular communication and the construction of prevascularization systems. This review focuses on the recent advances in the application of prevascularization techniques for pulp regeneration, considers dental stem cells as a potential cell source of endothelial cells and pericytes, discusses strategies for their directional differentiation, sketches the mechanism of intercellular communication and the potential application of communication mediators, and summarizes construction strategies for prevascularized systems. We also provide novel ideas for the extensive application and follow-up development of prevascularization techniques for dental pulp regeneration.

Introduction: Decellularized extracellular matrix has been recognized as an optimal scaffold for dental pulp regeneration. However, the limited amount of native dental pulp tissue restricts its clinical applications. The submandibular gland shares some basic extracellular matrix components and characteristics with dental pulp. However, whether decellularized submandibular gland extracellular matrix (DSMG) can be used as an alternative scaffold for dental pulp regenerative medicine is unclear.Methods: Thus, we successfully decellularized the whole rat submandibular gland and human dental pulp, and then conducted in vitro and in vivo studies to compare the properties of these two scaffolds for dental pulp regeneration.Results: Our results showed that extracellular matrix of the submandibular gland had great similarities in structure and composition with that of dental pulp. Furthermore, it was confirmed that the DSMG could support adhesion and proliferation of dental pulp stem cells in vitro. In vivo findings revealed that implanted cell-seeded DSMG formed a vascularized dental pulp-like tissue and expressed markers involved in dentinogenesis and angiogenesis.Discussion: In summary, we introduced a novel accessible biological scaffold and validated its effectiveness as an extracellular matrix-based tissue engineering scaffold for dental pulp regenerative therapy.

Dental pulp and periapical diseases make patients suffer from acute pain and economic loss. Although root canal therapies, as demonstrated through evidence-based medicine, can relieve symptoms and are commonly employed by dentists, it is still difficult to fully restore a dental pulp's nutrition, sensory, and immune-regulation functions. In recent years, researchers have made significant progress in tissue engineering to regenerate dental pulp in a desired microenvironment. With breakthroughs in regenerative medicine and material science, bioactive scaffolds play a pivotal role in creating a suitable microenvironment for cell survival, proliferation, and differentiation, following dental restoration and regeneration. This article focuses on current challenges and novel perspectives about bioactive scaffolds in creating a microenvironment to promote dental pulp regeneration. We hope our readers will gain a deeper understanding and new inspiration of dental pulp regeneration through our summary. [ABSTRACT FROM AUTHOR]

Dental-derived stem cells have excellent proliferation ability and multi-directional differentiation potential, making them an important research target in tissue engineering. An increasing number of dental-derived stem cells have been discovered recently, including dental pulp stem cells (DPSCs), stem cells from exfoliated deciduous teeth (SHEDs), stem cells from apical papilla (SCAPs), dental follicle precursor cells (DFPCs), and periodontal ligament stem cells (PDLSCs). These stem cells have significant application prospects in tissue regeneration because they are found in an abundance of sources, and they have good biocompatibility and are highly effective. The biological functions of dental-derived stem cells are regulated in many ways. Epigenetic regulation means changing the expression level and function of a gene without changing its sequence. Epigenetic regulation is involved in many biological processes, such as embryonic development, bone homeostasis, and the fate of stem cells. Existing studies have shown that dental-derived stem cells are also regulated by epigenetic modifications. Pulp and periodontal regeneration refers to the practice of replacing damaged pulp and periodontal tissue and restoring the tissue structure and function under normal physiological conditions. This treatment has better therapeutic effects than traditional treatments. This article reviews the recent research on the mechanism of epigenetic regulation of dental-derived stem cells, and the core issues surrounding the practical application and future use of pulp and periodontal regeneration. [ABSTRACT FROM AUTHOR]

Background: Dental pulp regeneration is a complex and advancing field that requires biomaterials capable of supporting the pulp's diverse functions, including immune defense, sensory perception, vascularization, and reparative dentinogenesis. Regeneration involves orchestrating the formation of soft connective tissues, neurons, blood vessels, and mineralized structures, necessitating materials with tailored biological and mechanical properties. Numerous biomaterials have entered clinical practice, while others are being developed for tissue engineering applications. The composition and a broad range of material properties, such as surface characteristics, degradation rate, and mechanical strength, significantly influence cellular behavior and tissue outcomes. This underscores the importance of employing robust evaluation methods and ensuring precise and comprehensive reporting of findings to advance research and clinical translation., Aims: This article aims to present the biological foundations of dental pulp tissue engineering alongside potential testing methodologies and their advantages and limitations. It provides guidance for developing research protocols to evaluate the properties of biomaterials and their influences on cell and tissue behavior, supporting progress toward effective dental pulp regeneration strategies., Competing Interests: Declaration of Competing Interest This research received no funding and there is no conflict of interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Introduction: To prepare a gelatin methacryloyl (GelMA) hydrogel scaffold embedded with adrenomedullin (ADM) and investigate its impact and underlying mechanisms in endogenous pulp regeneration., Methods: ADM was evenly distributed within the GelMA hydrogel through a simple and conventional physical mixing technique. The scaffold underwent characterization via scanning electron microscopy, alongside assessments of swelling, degradation, and release properties. Biocompatibility was evaluated using cytoskeletal and live-dead staining techniques. The hydrogel's influence on dental pulp stem cells' proliferation, migration, and differentiation was assessed with CCK-8 assays, Transwell assays, and alizarin red and alkaline phosphatase staining. Transcriptomics provided insights into potential mechanisms. The angiogenic effects on umbilical vein endothelial cells were examined using scratch and tube formation assays. In vivo, the composite hydrogel's regenerative capacity was tested in a rat model of pulp regeneration. Statistical analysis involved Student's t-test and one-way analysis of variance, with significance set at P < .05., Results: The ADM-loaded GelMA (GelMA@ADM) hydrogel displayed a porous architecture under electron microscopy conducive to cell adhesion and demonstrated excellent biocompatibility. In vitro experiments showed that GelMA@ADM significantly boosted dental pulp stem cells' migration, proliferation, and differentiation, and enhanced the angiogenic activity of umbilical vein endothelial cells after one week of treatment. Corresponding in vivo experiments revealed that GelMA@ADM facilitated the formation of new vascularized pulp tissue after 2 weeks of treatment., Conclusions: The GelMA@ADM hydrogel effectively promotes dental pulp stem cells' proliferation and differentiation, augments vascularization by umbilical vein endothelial cells, and fosters the creation of new vascularized pulp tissue. These findings underscore the potential of GelMA@ADM hydrogel for endogenous pulp regeneration., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Dental pulp tissue engineering is a promising alternative treatment for pulpitis and periapical periodontitis, and dental pulp stem cells (DPSCs) are considered to be the gold standard for dental seed cell research. Periapical lesions harbor mesenchymal stem cells with the capacity for self‐renewal and multilineage differentiation. However, it remains unknown whether these periapical lesion‐derived stem cells (PLDSCs) are suitable for dental pulp tissue engineering. To investigate this possibility, PLDSCs and DPSCs were isolated using the tissue outgrowth method and cultured under identical conditions. We then performed in vitro experiments to investigate their biological characteristics. Our results indicate that PLDSCs proliferate actively in vitro and exhibit similar morphology, immunophenotype and multilineage differentiation ability as DPSCs. Simultaneously, PLDSCs exhibit stronger migrative ability and express more vascular endothelial growth factor and glial cell line‐derived neurotrophic factor than DPSCs, and PLDSC‐derived conditioned medium was more effective in tube formation assay. The mRNA expression levels of immunomodulatory genes HLA‐G, IDO and ICAM‐1 were also higher in PLDSCs. However, regarding osteo/odontogenic differentiation, PLDSCs showed weaker alkaline phosphatase staining and lower calcified nodule formation compared to DPSCs, as well as lower expression of ALP, RUNX2 and DSPP, as confirmed by a quantitative RT‐PCR. The osteo/odontogenic protein expression levels of DSPP, RUNX2, DMP1 and SP7 were also higher in DPSCs. The present study demonstrates that PLDSCs demonstrate potential use as seed cells for dental pulp regeneration, especially for achieving enhanced neurovascularization.

Much research has been conducted on fabricating biomimetic biomaterials in vitro. Tissue engineering approaches are often conducted by combining cells, scaffolds, and growth factors. However, the degradation rate of scaffolds is difficult to control and the degradation byproducts occasionally limit tissue regeneration. To overcome these issues, we have developed a novel system using a thermo-responsive hydrogel that forms scaffold-free, three-dimensional (3D) cell constructs with arbitrary size and morphology. 3D cell constructs prepared using bone marrow-derived stromal stem cells (BMSCs) exhibited self-organizing ability and formed bone-like tissue with endochondral ossification. Endothelial cells were then introduced into the BMSC construct and a vessel-like structure was formed within the constructs. Additionally, the bone formation ability was promoted by endothelial cells and cell constructs could be freeze-dried to improve their clinical application. A pre-treatment with specific protein protectant allowed for the fabrication of novel bone substitutes composed only of cells. This 3D cell construct technology using thermo-responsive hydrogels was then applied to other cell species. Cell constructs composed of dental pulp stem cells were fabricated, and the resulting construct regenerated pulp-like tissue within a human pulpless tooth. In this review, we demonstrate the approaches for the in vitro fabrication of bone and dental pulp-like tissue using thermo-responsive hydrogels and their potential applications.

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Scaffolds are crucial elements for dental pulp regeneration. Most of the currently used scaffolds in regenerative endodontic procedures (REPs) are unsuitable for chairside clinical use. This study aimed to evaluate the effect of an injectable synthetic scaffold (Restylane Lyft) on human bone marrow mesenchymal stem cell (hBMSC) viability, proliferation, and osteo/dentinogenic differentiation in a regenerative endodontic organotype model (REM). hBMSC were loaded in an REM either alone (hBMSC group) or mixed with the Restylane Lyft scaffold (Restylane/hBMSC group) and cultured in basal culture medium (n = 9/group). hMSC on culture plates served as controls. Cell viability and proliferation were measured using AlamarBlue assay. The loaded REM was cultured in an osteogenic differentiation medium to measure alkaline phosphatase activity (ALP) and examine the expression of the osteo/dentinogenic markers using real-time reverse transcriptase polymerase chain reaction. Cell viability in all groups increased significantly over 5 days. The Restylane/hBMSC group showed significantly higher ALP activity and dentin sialophosphoprotein, osteocalcin, and bone sialoprotein genes expression than the hBMSC and the control groups. Restylane Lyft, a hyaluronic acid (HA) injectable, FDA-approved hydrogel, maintained cell viability and proliferation and promoted osteo/dentinogenic differentiation of hBMSC when cultured in an REM. Henceforth, it could be a promising chairside scaffold material for REPs. [ABSTRACT FROM AUTHOR]

Stem cell-based therapeutics is a promising strategy in dental pulp regeneration. However, low cell viability after transplantation in vivo due to the ischemic microenvironment is still a critical challenge for future clinical application. With the aim of improving postimplantation cell survival and pulp tissue regeneration, stem cells from human exfoliated deciduous teeth (SHED) were preconditioned to a hypoxic condition by hypoxia-inducible factor 1α (HIF-1α) stabilization via knockdown of prolyl hydroxylase domain-containing protein 2 (PHD2) using lentiviral short hairpin RNA. HIF-1α-stabilized SHED were encapsulated in PuraMatrix hydrogel, injected into root canals of human tooth fragments, and implanted in the subcutaneous space of immunodeficient mice. After 28 d, enhanced dental pulp-like tissue formation was observed with a significantly higher level of vascularization, which could be attributed to both endothelial differentiation of SHED and recruitment of host blood vessels. Furthermore, dentin-like tissue formation in vivo and accelerated odontogenic/osteogenic differentiation both in vivo and in vitro were observed. At 7 d postimplantation, significantly less DNA damage and higher Ki67 expression were detected in the HIF-1α-stabilized SHED group compared with the control SHED. Accordingly, cell viability assay and staining for Ki67 and apoptotic cells in vitro showed that HIF-1α stabilization could decrease cell apoptosis and enhance cell survival significantly. We demonstrated that PI3K/AKT pathway activation had resulted in low caspase 3 expression in HIF-1α-stabilized SHED in hypoxic conditions. Furthermore, we found that HIF-1α-induced cell survival could also be attributed to the upregulated expression of PDK1, HK2, and Glut1, which contributes to the maintenance of reactive oxygen species homeostasis and metabolic adaptation in hypoxia. In addition, we identified Smad7 as 1 of the top 3 upregulated genes through RNA sequencing in HIF-1α-stabilized SHED and demonstrated its essential role in HK2 and Glut1 upregulation. Taken together, HIF-1α stabilization enhances cell survival of SHED through modulating various target genes and potential signaling pathways, as well as odontogenic tissue formation during dental pulp regeneration, which could benefit stem cell-based therapy in general. [ABSTRACT FROM AUTHOR]

Angiogenesis is considered a crucial event for dental pulp regeneration. The purpose of this study was to demonstrate neovascularization during coronal pulp regeneration in rat molars using rat dental pulp cells (rDPCs) and to examine whether rDPC-endothelial cell interactions promote proangiogenic capacity in vitro. Maxillary first molars of Wistar rats (n = 42) were pulpotomized and rDPCs isolated from incisors were implanted with a porous poly (l -lactic acid) (PLLA) scaffold and hydrogel (Matrigel). After 3, 7, and 14 days, coronal pulp tissues were examined histologically and by nestin and CD146 immunohistochemistry. rDPCs and rat dermal microvascular endothelial cells (rDMECs) were cocultured for 4 days and vascular endothelial growth factor (VEGF) synthesis and angiogenic factor gene expression were determined by enzyme-linked immunosorbent assays and real-time polymerase chain reaction, respectively. Effects of cocultured medium on tube formation by rDMECs were also evaluated. Implantation of rDPC/PLLA/Matrigel induced coronal pulp regeneration with dentin bridge formation and arrangement of nestin-positive odontoblast-like cells at 14 days. PLLA/Matrigel without rDPCs did not induce pulp regeneration. CD146-positive blood vessels increased in density in the remaining pulp tissues at 3 and 7 days, and in the regenerated pulp tissue at 14 days. rDPC/DMEC coculture significantly promoted VEGF secretion and mRNA expression of nuclear factor-kappa B, angiogenic chemokine CXCL1 , and chemokine receptor CXCR1. Cocultured medium significantly promoted tube formation. Coronal pulp regeneration with rDPC/PLLA/Matrigel was accompanied by neovascularization. rDPC-rDMEC interactions may promote angiogenic activity represented by proangiogenic factor upregulation and tube formation in vitro. [ABSTRACT FROM AUTHOR]

The development of materials science is of great significance to the treatment of dental pulp diseases. Poly lactic acid glycolic acid (PLGA) copolymer is an organic macromolecule compound that is widely used in the preparation of biomedical materials. In recent years, PLGA, as a drug/molecular loaded system and tissue regeneration scaffold, has shown prospects for application in the treatment of dental pulp diseases. This paper will review the application of PLGA in the treatment of dental pulp diseases and provide a basis for its further development and utilization. The results of the literature review show that PLGA is a drug/molecular delivery system that is mainly used in the improvement of pulp capping materials, root canal disinfectant and apexification materials. PLGA-improved pulp capping agents can prolong the action time of the drug and reduce toxicity. The modified root canal disinfectant can realize the sustained release of drug, make the drug penetrate deeper into the subtle structure, and contact more widely with the pathogenic bacteria. The modified apexification materials can provide more convenient administration methods for apexifixment. As a scaffold for tissue engineering, PLGA is mainly used in the study of pulp regeneration. The optimization of PLGA physical properties and action environment can provide a more suitable microenvironment for seed cells to proliferate and differentiate. How to utilize the advantages of PLGA to develop a more suitable material for endodontic application needs further study.

With advances in knowledge and treatment options, pulp regeneration is now a clear objective in clinical dental practice. For this purpose, many methodologies have been developed in attempts to address the putative questions raised both in research and in clinical practice. In the first part of this review, laboratory‐based methods will be presented, analysing the advantages, disadvantages, and benefits of cell culture methodologies and ectopic/semiorthotopic animal studies. This will also demonstrate the need for alignment between two‐dimensional and three‐dimensional laboratory techniques to accomplish the range of objectives in terms of cell responses and tissue differentiation. The second part will cover observations relating to orthotopic animal studies, describing the current models used for this purpose and how they contribute to the translation of regenerative techniques to the clinic. [ABSTRACT FROM AUTHOR]

The dental pulp is critical for physiological vitality of the tooth, and dental pulp regeneration has great potential for rebuilding live pulp tissue after pulp disease. Schwann cells (SCs) play a critical role in the support, maintenance, and regeneration of nerve fibers in dental pulp. Extracellular vesicles (EVs), which possess cell homing and tissue repair potential, derived from SCs (SC-EVs), can regulate dental mesenchymal stem cells (MSCs) proliferation, multipotency, and self-renewal. However, the role of SC-EVs in dental pulp tissue regeneration remains unclear. To address this question, we treated dental pulp stem cells (DPSCs) and bone marrow stem cells (BMSCs) with SC-EVs, and the results showed an obvious increase in the proliferation, migration, and osteogenic differentiation of both cell types. SC-EVs also promoted neurite outgrowth and neuron migration of rat dorsal root ganglia, as well as vessel formation in vitro. In an in vivo model of subcutaneous, SC-EVs enhanced the recruitment of endogenous vascular endothelioid-like cells and MSCs, and promoted the formation of a pulpo-dentinal complex-like structure. Finally, mass spectrometry analyses and western blot revealed that stromal cell-derived factor 1 (SDF-1, also known as CXCL12) plays a dominant role in SC-EVs. Together, these data suggest that SC-EVs successfully recruit endogenous stem cells to promote dental pulp regeneration. Our results provide a cell-free strategy for pulp regeneration that avoids the risks associated with stem cell transplantation. Dental pulp is vulnerable to infections resulting from dental care, trauma, and multiple restorations, with such infections resulting in pulpitis and pulp necrosis. The current endodontic treatment of irreversible pulp disease cannot restore the function of dental pulp and tissue engineering strategies using cell-based approaches are limited by several disadvantages, including immune rejection and limited cell sources. In this study, we found that schwann cells-derived EVs facilitated dental pulp regeneration through endogenous stem cells recruitment via SDF-1/CXCR4 axis without exogenous cell transplantation. We believe that our study makes a significant contribution to describe a cell-free strategy to promote dental pulp regeneration. [Display omitted] [ABSTRACT FROM AUTHOR]

Endothelial regeneration is a research hotspot in the field of dental pulp. The regeneration of endodontic blood flow is the bottleneck of dental pulp regeneration, and the applied scaffold material is the key to revascularization. Stent materials were reviewed. The literature review Results show that, depending on the source of the stent material used for endodontic revascularization, there are mainly natural, synthetic and composite materials. The natural scaffold materials used for vascular regeneration include chitosan, hyaluronic acid, bacterial cellulose, and proanthocyanidin; artificial scaffold materials include hydrogel, cryogel, and electrospinning. The bionic composite scaffold system with a double-layer tubular structure is low immunogenicity and good biocompatibility. Studies on the scaffold materials of bionic extracellular matrix, such as injectable hydrogels/microspheres, have promoted the development of dental pulp regeneration, that is, uniformly distributed scaffold materials in the root canal promote the generation of pulp-like tissue; Whether dental pulp tissue can establish effective blood circulation through the apical foramen remains a great challenge.

Dental caries is a common disease that not only destroys the rigid structure of the teeth but also causes pulp necrosis in severe cases. Once pulp necrosis has occurred, the most common treatment is to remove the damaged pulp tissue, leading to a loss of tooth vitality and increased tooth fragility. Dental pulp stem cells (DPSCs) isolated from pulp tissue exhibit mesenchymal stem cell-like characteristics and are considered ideal candidates for regenerating damaged dental pulp tissue owing to their multipotency, high proliferation rate, and viability after cryopreservation. Importantly, DPSCs do not elicit an allogeneic immune response because they are non-immunogenic and exhibit potent immunosuppressive properties. Here, we provide an up-to-date review of the clinical applicability and potential of DPSCs, as well as emerging trends in the regeneration of damaged pulp tissue. In addition, we suggest the possibility of using DPSCs as a resource for allogeneic transplantation and provide a perspective for their clinical application in pulp regeneration.

Objective: Pulp regeneration with bioactive dentin-pulp complex has been a research hotspot in recent years. Stem cell therapy provided an interest strategy to regenerate the dental-pulp complex. Hence, this study aimed to evaluate the effects of photosensitive gelatin methacrylate (GelMA) hydrogel encapsulating dental pulp stem cells (DPSCs) and silver nanoparticles (AgNPs) for dental pulp regeneration in vitro., Methods: First, the AgNPs@GelMA hydrogels were prepared by lithium phenyl-2,4,6-trimethyl-benzoyl phosphinate (LAP) initiation via blue-light emitting diode light. The physical and chemical properties of AgNPs@GelMA hydrogels were comprehensively analysed via scanning electron microscopy (SEM), and mechanical characterisation, such as swelling ability, degradation properties, and AgNP release profile. Then, AgNPs@GelMA hydrogels encapsulated DPSCs were used to establish an AgNPs@GelMA biomimetic complex, further analysing its biocompatibility, antibacterial properties, and angiogenic capacity in vitro., Results: The results indicated that GelMA hydrogels demontrated optimal characteristics with a monomer:LAP ratio of 16:1. The physico-chemical properties of AgNPs@GelMA hydrogels did not change significantly after loading with AgNPs. There was no significant difference in AgNP release rate amongst different concentrations of AgNPs@GelMA hydrogels. Fifty to 200 μg/mL AgNPs@GelMA hydrogels could disperse E faecalis biofilm and reduce its metabolic activity . Furthermore, cell proliferation was arrested in 100 and 200 μg/mL AgNPs@GelMA hydrogels. The inhibition of 50 μg/mL AgNPs@GelMA hydrogels on E faecalis biofilm was above 50%, and the cell viability of the hydrogels was higher than 90%. The angiogenesis assay indicated that AgNPs@GelMA hydrogels encapsulating DPSCs could induce the formation of capillary-like structures and express angiogenic markers CD31, vascular endothelial growth factor , and von willebrand factor (vWF) in vitro., Conclusions: Results of this study indicate that 50 μg/mL AgNPs@GelMA hydrogels encapsulating DPSCs had significant antibacterial properties and angiogenic capacity, which could provide a significant experimental basis for the regeneration of the dentin-pulp complex., Competing Interests: Conflict of interest None disclosed., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Dental pulp tissue engineering is a promising alternative treatment for pulpitis and periapical periodontitis, and dental pulp stem cells (DPSCs) are considered to be the gold standard for dental seed cell research. Periapical lesions harbor mesenchymal stem cells with the capacity for self‐renewal and multilineage differentiation. However, it remains unknown whether these periapical lesion‐derived stem cells (PLDSCs) are suitable for dental pulp tissue engineering. To investigate this possibility, PLDSCs and DPSCs were isolated using the tissue outgrowth method and cultured under identical conditions. We then performed in vitro experiments to investigate their biological characteristics. Our results indicate that PLDSCs proliferate actively in vitro and exhibit similar morphology, immunophenotype and multilineage differentiation ability as DPSCs. Simultaneously, PLDSCs exhibit stronger migrative ability and express more vascular endothelial growth factor and glial cell line‐derived neurotrophic factor than DPSCs, and PLDSC‐derived conditioned medium was more effective in tube formation assay. The mRNA expression levels of immunomodulatory genes HLA‐G, IDO and ICAM‐1 were also higher in PLDSCs. However, regarding osteo/odontogenic differentiation, PLDSCs showed weaker alkaline phosphatase staining and lower calcified nodule formation compared to DPSCs, as well as lower expression of ALP, RUNX2 and DSPP, as confirmed by a quantitative RT‐PCR. The osteo/odontogenic protein expression levels of DSPP, RUNX2, DMP1 and SP7 were also higher in DPSCs. The present study demonstrates that PLDSCs demonstrate potential use as seed cells for dental pulp regeneration, especially for achieving enhanced neurovascularization. [ABSTRACT FROM AUTHOR]

Much research has been conducted on fabricating biomimetic biomaterials in vitro. Tissue engineering approaches are often conducted by combining cells, scaffolds, and growth factors. However, the degradation rate of scaffolds is difficult to control and the degradation byproducts occasionally limit tissue regeneration. To overcome these issues, we have developed a novel system using a thermo-responsive hydrogel that forms scaffold-free, three-dimensional (3D) cell constructs with arbitrary size and morphology. 3D cell constructs prepared using bone marrow-derived stromal stem cells (BMSCs) exhibited self-organizing ability and formed bone-like tissue with endochondral ossification. Endothelial cells were then introduced into the BMSC construct and a vessel-like structure was formed within the constructs. Additionally, the bone formation ability was promoted by endothelial cells and cell constructs could be freeze-dried to improve their clinical application. A pre-treatment with specific protein protectant allowed for the fabrication of novel bone substitutes composed only of cells. This 3D cell construct technology using thermo-responsive hydrogels was then applied to other cell species. Cell constructs composed of dental pulp stem cells were fabricated, and the resulting construct regenerated pulp-like tissue within a human pulpless tooth. In this review, we demonstrate the approaches for the in vitro fabrication of bone and dental pulp-like tissue using thermo-responsive hydrogels and their potential applications. [ABSTRACT FROM AUTHOR]

Regenerative endodontic procedures using autologous platelet-rich plasma (PRP) can improve the biologic outcome of treatment. However, its mechanism of action on improving pulp regeneration is not fully elucidated. Autophagy was recently shown to be related to tissue repair and osteogenesis. Therefore, the objective of this study was to investigate the effect of PRP in dental pulp regeneration and to elucidate the role of autophagy involved in this process. Human dental pulp cells (hDPCs) were isolated from healthy dental pulp and co-cultured with an increasing concentration of PRP. Cellular migration and proliferation were determined by scratch assay, transwell assay, and cell-counting kit 8 assay. Osteogenic differentiation was clarified by using alkaline phosphatase staining, alizarin red staining, and real-time polymerase chain reaction (RT-PCR) to measure the gene expression levels of alkaline phosphatase, collagen-1, osteocalcin, dentin matrix protein 1, and dentin sialophosphoprotein. Autophagic bodies were observed by transmission electron microscopy and the expression of autophagy marker light chain 3B (LC3B) was determined by immunofluorescence staining. The mRNA and protein expression level of LC3B and Beclin-1 were quantified by qRT-PCR and western blotting. The effect of PRP on cellular migration, proliferation, and osteogenic differentiation was further investigated in the milieu of autophagy activator, rapamycin, and inhibitor, 3-methyladenine. Results showed that PRP promoted cell migration, proliferation, and osteogenic differentiation. Autophagic bodies were strongly activated and the expression level of LC3B and Beclin-1 was significantly promoted by PRP. Autophagy inhibition suppressed PRP-induced hDPCs migration, proliferation, and osteogenic differentiation, whereas autophagy activator substantially augmented PRP-stimulated migration, proliferation, and differentiation. Taken together, these findings suggested that PRP could effectively promote regenerative potentials associated with autophagy.

Prolonged and severe hypoxia is the main cause of death of transplanted cells prior to the establishment of functional circulation. In situ generation of oxygen by oxygen-producing scaffolds-a unique solution that could produce and deliver oxygen to the adjacent cells independently of blood perfusion-has attracted considerable attention to enhance the survivability of the transplanted cells. However, the application of oxygen-generating scaffolds for facilitating cell survival in pulp-like tissue regeneration is yet to be explored. In this study, gelatin methacryloyl (GelMA)-a biocompatible scaffolding material that closely mimics the native extracellular matrix and is conducive to cell proliferation and differentiation-was used to fabricate oxygen-generating scaffolds by loading various concentrations of CaO2. The CaO2 distribution, topography, swelling, and pore size of CaO2-GelMA hydrogels were characterized in detail. The release of O2 by the scaffold and the viability, spreading, and proliferation of stem cells from apical papilla (SCAPs) encapsulated in the GelMA hydrogels with various concentrations of CaO2 under hypoxia were evaluated. In addition, cellular constructs were engineered into root canals, and cell viability within the apical, middle, and coronal portions was assessed. Our findings showed that 0.5% CaO2-GelMA was sufficient to supply in situ oxygen for maintaining the embedded SCAP viability for 1 wk. Furthermore, the 0.5% CaO2-GelMA hydrogels improved the survivability of SCAPs within the coronal portion of the engineered cellular constructs within the root canals. This work demonstrated that 0.5% CaO2-GelMA hydrogels offer a potential promising scaffold that enhances survival of the embedded SCAPs in endodontic regeneration. [ABSTRACT FROM AUTHOR]

Aim: To investigate the vascularization capacity of a decellularized dental pulp matrix (DDP) of bovine origin seeded with human dental pulp stem cells (hDPSCs) in vitro and to present preliminary in vivo findings. Methodology: Bovine dental pulps were decellularized and then analysed using histological staining and DNA quantification. The resultant DDPs were characterized using immunohistochemical staining for the retention of vascular endothelial growth factor A (VEGF‐A) and fibroblast growth factor 2 (FGF‐2). Furthermore, DDPs were recellularized with hDPSCs and analysed histologically. The expression of markers involved in angiogenesis by hDPSCs colonizing the DDPs was assessed in vitro. A preliminary in vivo study was then conducted in which hDPSCs‐seeded and unseeded DDPs were inserted in debrided human premolars root slices and implanted subcutaneously in immunodeficient mice. Samples were retrieved after 30 days and analysed using histological and immunohistochemical staining. The independent samples t‐test, analysis of variance and a Kruskal–Wallis test were used to analyse the quantitative data statistically depending on the group numbers and normality of data distribution. The difference between the groups was considered significant when the P‐value was less than 0.05. Results: Acellular dental pulp matrices were generated following bovine dental pulp decellularization. Evaluation of the developed DDPs revealed a significant DNA reduction (P < 0.0001) with preservation of the native histoarchitecture and vasculature and retention of VEGF‐A and FGF‐2. Upon recellularization of the DDPs with hDPSCs, the in vitro analyses revealed cell engraftment with progressive repopulation of DDPs' matrices and vasculature and with enhanced expression of markers involved in angiogenesis. In vivo implantation of root slices with hDPSCs‐seeded DDPs revealed apparent vascularization enhancement as compared to the unseeded DDP group (P < 0.0001). Conclusions: The developed decellularized dental pulp matrix had pro‐angiogenic properties characterized by the retention of native vasculature and angiogenic growth factors. Seeding of hDPSCs into the DDP led to progressive repopulation of the vasculature, enhanced expression of markers involved in angiogenesis in hDPSCs and improved in vivo vascularization capacity. The se suggest that a combination of DDP and hDPSCs have the potential to provide a promising vascularization promoting strategy for dental pulp regeneration. [ABSTRACT FROM AUTHOR]

Deep caries, trauma, and severe periodontitis result in pulpitis, pulp necrosis, and eventually pulp loss. However, no clinical therapy can regenerate lost pulp. A novel pulp regeneration strategy for clinical application is urgently needed. Signaling transduction plays an essential role in regulating the regenerative potentials of dental stem cells. Cytokines or growth factors, such as stromal cell-derived factor (SDF), fibroblast growth factor (FGF), bone morphogenetic protein (BMP), vascular endothelial growth factor (VEGF), WNT, can promote the migration, proliferation, odontogenic differentiation, pro-angiogenesis, and pro-neurogenesis potentials of dental stem cells respectively. Using the methods of signaling modulation including growth factors delivery, genetic modification, and physical stimulation has been applied in multiple preclinical studies of pulp regeneration based on cell transplantation or cell homing. Transplanting dental stem cells and growth factors encapsulated into scaffold regenerated vascularized pulp-like tissue in the root canal. Also, injecting a flowable scaffold only with chemokines recruited endogenous stem/progenitor cells for pulp regeneration. Notably, dental pulp regeneration has gradually developed into the clinical phase. These findings enlightened us on a novel strategy for structural and functional pulp regeneration through elaborate modulation of signaling transduction spatially and temporally via clinically applicable growth factors delivery. But challenges, such as the adverse effects of unphysiological signaling activation, the controlled drug release system, and the safety of gene modulation, are necessary to be tested in future works for promoting the clinical translation of pulp regeneration. [ABSTRACT FROM AUTHOR]

Regenerative endodontics has created a desirable shift in the treatment paradigm despite current limitations of regenerative outcomes. Mesenchymal stem cells (MSCs) facilitate tissue regeneration and repair in a mild inflammatory environment. Small extracellular vesicles (sEVs) derived from MSCs play an imperative role in the paracrine modulation of regenerative responses modulated by MSCs. However, it remains unknown whether MSCs enhance dental pulp regeneration or whether this enhancement is mediated by sEVs in a mild inflammatory environment. The present study aimed to elucidate the effects of sEVs originated from lipopolysaccharide (LPS)-preconditioned human dental pulp stem cells (hDPSCs) on dental pulp regeneration. All sEVs were isolated from hDPSCs cultured with or without LPS (ie, N-sEVs and L-sEVs, respectively). The effect of N-sEVs and L-sEVs on proliferation, migration, angiogenesis, and differentiation of rat bone marrow MSCs was identified in vitro. Moreover, N-sEVs or L-sEVs were implanted into rat pulpless root canal models, and the regenerated tissue in root canals was assessed via hematoxylin-eosin staining, Masson staining, and immunohistochemistry after 30 days of transplantation. Both N-sEVs and L-sEVs could modulate BMSC proliferation, migration, angiogenesis, and differentiation. Both kinds of sEVs enhanced the structure of the regenerated tissue closer to that of a normal dental pulp in vivo. L-sEVs had a more significant effect than N-sEVs. sEVs released by hDPSCs in a mild inflammatory microenvironment are capable of facilitating the regeneration of dental pulp through functional healing instead of scar healing, which has potential applications in regenerative endodontics. [ABSTRACT FROM AUTHOR]

Dental pulp as a source of nutrition for the whole tooth is vulnerable to trauma and bacterial invasion, which causes irreversible pulpitis and pulp necrosis. Dental pulp regeneration is a valuable method of restoring the viability of the dental pulp and even the whole tooth. Odontogenic mesenchymal stem cells (MSCs) residing in the dental pulp environment have been widely used in dental pulp regeneration because of their immense potential to regenerate pulp-like tissue. Furthermore, the regenerative abilities of odontogenic MSCs are easily affected by the microenvironment in which they reside. The natural environment of the dental pulp has been proven to be capable of regulating odontogenic MSC homeostasis, proliferation, and differentiation. Therefore, various approaches have been applied to mimic the natural dental pulp environment to optimize the efficacy of pulp regeneration. In addition, odontogenic MSC aggregates/spheroids similar to the natural dental pulp environment have been shown to regenerate well-organized dental pulp both in preclinical and clinical trials. In this review, we summarize recent progress in odontogenic MSC-mediated pulp regeneration and focus on the effect of the microenvironment surrounding odontogenic MSCs in the achievement of dental pulp regeneration.

Dental pulp as a source of nutrition for the whole tooth is vulnerable to trauma and bacterial invasion, which causes irreversible pulpitis and pulp necrosis. Dental pulp regeneration is a valuable method of restoring the viability of the dental pulp and even the whole tooth. Odontogenic mesenchymal stem cells (MSCs) residing in the dental pulp environment have been widely used in dental pulp regeneration because of their immense potential to regenerate pulp-like tissue. Furthermore, the regenerative abilities of odontogenic MSCs are easily affected by the microenvironment in which they reside. The natural environment of the dental pulp has been proven to be capable of regulating odontogenic MSC homeostasis, proliferation, and differentiation. Therefore, various approaches have been applied to mimic the natural dental pulp environment to optimize the efficacy of pulp regeneration. In addition, odontogenic MSC aggregates/spheroids similar to the natural dental pulp environment have been shown to regenerate well-organized dental pulp both in preclinical and clinical trials. In this review, we summarize recent progress in odontogenic MSC-mediated pulp regeneration and focus on the effect of the microenvironment surrounding odontogenic MSCs in the achievement of dental pulp regeneration. [ABSTRACT FROM AUTHOR]

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.

BACKGROUND: Rapid development in tissue engineering research and technology makes dental pulp regeneration and revascularization possible. The interactions of stem cells, scaffolds and signaling factors in tissue engineering are particularly important. Whether stem cells can proliferate, differentiate and develop dental pulp-like tissue greatly depends on the choice of scaffolds OBJECTIVE: To review the widely studied and effective scaffold materials and two methods of scaffold preparation and analyze their applications in dental pulp reconstruction and their revascularization ability. METHODS: The first author searched PubMed, Wanfang and CNKI databases using a computer for relevant articles published between January 1, 2019 and September 30, 2019 with the search terms "pulp regeneration, pulp revascularization, scaffold" in English, and "pulp regeneration, pulp revascularization, revascularization, scaffold" in Chinese. A total of 421 English articles and 181 Chinese articles were retrieved. Finally, 61 articles were reviewed. RESULTS AND CONCLUSION: Platelet-derived scaffolds, extracellular-matrix-derived scaffolds, and self-assembling peptide take effect in pulp regeneration and revascularization. Composite materials combining natural and synthetic materials prepared by hydrogel and nanomaterial techniques exhibited advantages in cell proliferation, differentiation, migration, adherence, anti-inflammation, and factor delivery. The modified composite materials have a strong ability to promote vascularization. With the development of scaffold design and preparation technology based on hydrogels and nanomaterials, problems regarding insufficient scaffold source and unstable clinical effect will be solved in the future. [ABSTRACT FROM AUTHOR]

Dental pulp, plays an indispensable role in maintaining homeostasis of the tooth. Pulp necrosis always causes tooth nutrition deficiency and abnormal root development, which leads to tooth discoloration, fracture or even loss. Our previous study showed implantation of autologous SHED could regenerate functional dental pulp. However, the detailed mechanism of the implanted SHED participating in dental pulp regeneration remains unknown. In this study, we implanted SHED in a porcine dental pulp regeneration model to evaluate the regenerative effect and identify whether SHED promoted angiogenesis in regenerated dental pulp. Firstly we verified that xenogenous SHED had the ability to regenerated pulp tissue of host in vivo. Then we found the vasculature in regenerated pulp originated from implanted SHED. In addition, stem cells were isolated from regenerated dental pulp, which exhibited good multi-differentiation properties and promoted angiogenesis in pulp regeneration process and these results demonstrated that SHED promoted angiogenesis in stem cell-mediated dental pulp regeneration. • For the first time, we provides evidence that xenogenous SHED could regenerate pulp tissue in vivo. • We found the tissue and in regenerated pulp originated from implanted stem cells. • The new forming vessels partially consisted of cells differentiated from SHED and transformed into type H vessels. • R-SHED exhibited good multi-differentiation properties，especially in angiogenic potential [ABSTRACT FROM AUTHOR]

Regenerative dentistry has come a long way from pulp capping to pulp regeneration research, which aims to regenerate the pulp-dentin complex and restore its functions compromised by pulp injury and/or inflammation. Because of unique anatomic limitations of the tooth structure, engineering a suitable microenvironment that facilitates angio/vasculogenesis and innervation is a challenging task. Cell-based tissue engineering approaches have shown great potential in achieving this goal. Biomedical approaches in creating a regenerative microenvironment are mainly represented by either scaffold-based or scaffold-free strategies. The scaffold-based strategy mainly relies on the use of biomaterials to create a structural base that supports cells throughout the process of tissue formation. The scaffold could be a classic 3-dimensional construct with interconnected pores, a hydrogel with cells embedded in it, or a combination of these 2. The scaffold-free approach has been considered a bottom-up strategy that uses cell sheets, spheroids, or tissue strands as building blocks. The outcome of this strategy relies on the capacity of these building blocks to secrete a favorable extracellular matrix and to fuse into larger tissue constructs. Both the scaffold-free and scaffold-based systems are required as complementary, rather than competing, approaches for pulp regeneration. A combined synergetic strategy, through which multicellular building blocks could be integrated with robust 3-dimensional scaffolds, might represent an optimal solution to circumvent some of the major drawbacks of the current methods in pulp regeneration while concurrently fostering their advantages. [ABSTRACT FROM AUTHOR]