Cellular senescence in dental pulp stem cells

Platelet-rich plasma (PRP) is an emerging therapeutic application because PRP contains various growth factors that have beneficial effects on tissue regeneration and engineering. Mesenchymal stem cells and PRP derived from peripheral blood have been well studied. In this study, we investigated the effects of PRP derived from human umbilical cord blood (UCB-PRP) on proliferation, alkaline phosphatase (ALP) activity, and osteogenic differentiation of stem cells from human exfoliated deciduous teeth (SHEDs), dental pulp stem cells (DPSCs), and periodontal ligament stem cells (PDLSCs). Three types of dental stem cells were primarily isolated and characterized by flow cytometric analysis. Dental stem cells were exposed to various concentrations of UCB-PRP, which resulted in the proliferation of dental stem cells. Treatment with 2% UCB-PRP resulted in the highest level of proliferation. The ALP activity of DPSCs and PDLSCs increased following treatment with UCB-PRP in a dose-dependent manner up to a concentration of 2%. ALP activity decreased with higher concentration of UCB-PRP. The effects of UCB-PRP on calcium deposition were similar to those on proliferation and ALP activity. Treatment with 2% UCB-PRP resulted in the highest calcium depositions in DPSCs and PDLSCs; however, treatment with 1% UCB-PRP resulted in the highest calcium deposition in SHEDs. The concentrations of platelet-derived growth factor-AB and transforming growth factor-β1 in UCB-PRP were investigated and found to be comparable to the amounts in peripheral blood. Overall, UCB-PRP had beneficial effects on the proliferation and osteogenic differentiation of dental stem cells. Determination of the optimal concentration of UCB-PRP requires further investigation for clinical applications.

Stem cell migration is a critical step during the repair of damaged tissues. In order to achieve appropriate cell-based therapies for tooth and periodontal ligament repair it is necessary first to understand the dynamics of tissue-specific stem cell populations such as dental pulp stem cells (DPSC) and dental follicle stem cells (DFSC). Using time-lapse imaging, we analysed migratory and proliferative capabilities of these two human stem cell lines in vitro. When cultured alone, both DPSC and DFSC exhibited low and irregular migration profiles. In co-cultures, DFSC, but not DPSC, spectacularly increased their migration activity and velocity. DFSC rapidly surrounded the DPSC, thus resembling the in vivo developmental process, where follicle cells encircle both dental epithelium and pulp. Cell morphology was dependent on the culture conditions (mono-culture or co-culture) and changed over time. Regulatory genes involved in dental cell migration and differentiation such as TWIST1, MSX1, RUNX2, SFRP1 and ADAM28, were also evaluated in co-cultures. MSX1 up-regulation indicates that DPSC and DFSC retain their odontogenic potential. However, DPSC lose their capacity to differentiate into odontoblasts in the presence of DFSC, as suggested by RUNX2 up-regulation and TWIST1 down-regulation. In contrast, the unchanged levels of SFRP1 expression suggest that DFSC retain their potential to form periodontal tissues even in the presence of DPSC. These findings demonstrate that stem cells behave differently according to their environment, retain their genetic memory, and compete with each other to acquire the appropriate territory. Understanding the mechanisms involved in stem cell migration may lead to new therapeutic approaches for tooth repair.

Dental pulp stem cells (DPSCs) are a population of adult stem cells with self-renewal capacity and multilineage differentiation potential, widely utilized in tissue engineering and regenerative medicine. However, with increasing donor age or prolonged in vitro expansion, DPSCs gradually exhibit senescence-associated phenotypes, including reduced proliferative capacity and impaired differentiation potential. This review comprehensively summarizes current advances in the study of DPSCs senescence. First, it explores how the aging dental pulp microenvironment influences the biological characteristics of DPSCs. It then focuses on the interplay between DPSCs senescence and mitochondrial dysfunction, epigenetic alterations, as well as key signaling pathways such as p53/p21/p16. On this basis, recent molecular strategies for delaying DPSCs senescence are discussed, such as melatonin can alleviate DPSCs senescence by inhibiting the expression of matrix metalloproteinase, and metformin could interfere DPSCs senescence by AMPK/mTOR signaling pathway, etc. Lastly, a comparative analysis of senescence characteristics among DPSCs, bone marrow and adipose derived mesenchymal stem cells, is conducted to contextualize the unique and shared features of these cell types. Collectively, this review could provide a theoretical foundation and practical insights for advancing anti-senescence strategies in DPSC-based regenerative applications.

Objective. The adult spinal cord of mammals contains a certain amount of neural precursor cells, but these endogenous cells have a limited capacity for replacement of lost cells after spinal cord injury. The exogenous stem cells transplantation has become a therapeutic strategy for spinal cord repairing because of their immunomodulatory and differentiation capacity. In addition, dental stem cells originating from the cranial neural crest might be candidate cell sources for neural engineering. Approach. Human dental follicle stem cells (DFSCs), stem cells from apical papilla (SCAPs) and dental pulp stem cells (DPSCs) were isolated and identified in vitro, then green GFP-labeled stem cells with pellets were transplanted into completely transected spinal cord. The functional recovery of rats and multiple neuro-regenerative mechanisms were explored. Main results. The dental stem cells, especially DFSCs, demonstrated the potential in repairing the completely transected spinal cord and promote functional recovery after injury. The major involved mechanisms were speculated below: First, dental stem cells inhibited the expression of interleukin-1β to reduce the inflammatory response; second, they inhibited the expression of ras homolog gene family member A (RhoA) to promote neurite regeneration; third, they inhibited the sulfonylurea receptor1 (SUR-1) expression to reduce progressive hemorrhagic necrosis; lastly, parts of the transplanted cells survived and differentiated into mature neurons and oligodendrocytes but not astrocyte, which is beneficial for promoting axons growth. Significance. Dental stem cells presented remarkable tissue regenerative capability after spinal cord injury through immunomodulatory, differentiation and protection capacity.

Characterization of the Cellular Responses of Dental Mesenchymal Stem Cells to the Immune System

Aim: This study aims to review the unexplored area in regenerative prosthodontics using induced pluripotent stem (IPS) cells. Materials and Methods: A computer-based search was done from the year 1981 to 2020, but more specifically for IPS cells from 2006 and, the data were collected from various sources such as the National Center for Biotechnology Information (via PubMed), Medline semantic scholar, BioMed Central, Directory of Open Access Journals, Science Open, and Bielefeld Academic Search Engine using the keywords stem cells in dentistry, IPS cells, embryonic stem cells (ESCs), prosthodontic regeneration using induced pluripotent cells, stem cells from the periodontal ligament, dental pulp, deciduous tooth, and dental follicle. We have also systematically reviewed the method to generate IPS cells and their research in obtaining various organs that is of concern in dentistry and especially prosthodontics. Results: Based on the keywords and cross-referencing, 69 articles were valid and selected for review. The chosen articles are organized according to the subheadings in the order of their publication date. This literature is based on studies conducted on animals, and it does not include the transplantation of IPS cells in humans. In 2006, when Yamanaka and Takahashi discovered IPS cells, there was a spike rise in the study of these cells. The number of clinical trials for IPS cells was around 75%, whereas for ESCs was just 25%. Most of the clinical trials for IPS cells have been done in the USA (37%) whereas India accounts for only 1% of the study. Conclusion: This review article suggests the future of IPS cells in regenerative dentistry and prosthodontics. It gives information on the types of stem cells, why IPS cells have been chosen for the review, the source of IPS cells, the method of induction of IPS cells, and their use in prosthodontics and implantology.

To compare the proliferation and capacity of differentiation to vascular endothelial cells and angiogenesis induction among stem cells from human exfoliated deciduous teeth (SHED), dental pulp stem cells (DPSC) and human bone marrow mesenchymal stem cells (BMSC) from orofacial bone. SHED and DPSC were isolated from pulp tissue of the patients. BMSC were isolated from orthognathic or alveolar surgical sites. The surface markers of the cells were detected by flowcytometry. Cell counting kit-8 (CCK-8) assays were conducted to detect the proliferation ability of the cells. The cells were induced into endothelial cells with conditional medium and then the induced cells were cultured in Matrigel medium. The expression of angiogenesis-related genes such as platelet endothelial cell adhesion molecule-1 (PECAM-1/CD31), vascular endothelial growth factor (VEGF), vascular endothelial growth factor receptor 1 (VEGFR1), vascular endothelial growth factor receptor 2 (VEGFR2) and von Willebrand Factor (vWF) were quantified by real-time PCR. The cells were cultured in chick embryo chorioallantoic membrane (CAM) and the vessels were counted after 5 days. The cell surface markers CD73, CD90, CD105 and CD146 of all the stem cells were positive, CD34 and CD45 were negative. The CD146 positive rate of SHED and DPSC was higher than that of BMSC. SHED had a higher proliferation rate than DPSC and BMSC. After angiogenic induction for 14 d, 3 kinds of cells emanated pseudopodia formed grid structure long vasculature in Matrigel media. The total length of tube formation of induced BMSC (7 759.7 μm) and SHED (7 734.3 μm) was higher than DPSC (5 541.0 μm). The meshes number of induced SHED (70.7) was higher than DPSC (60) and BMSC (53.7) in Matrigel medium. The expression of CD31, VEGFR2 and vWF genes of SHED were higher than those of BMSC and DPSC. VEGFR1 gene expression of BMSC was higher than that of the other groups, and SHED was higher than DPSC. The expression of VEGF showed no difference among the cells. No deference was showed between the effect of the stem cells and negative control on new formed vessels in CAM. The total length of vessels of SHED (30.4 mm) was higher than that of the negative control (20.9 mm) and BMSC (28.0 mm). SHED, DPSC and BMSC can differentiate into vascular endothelial cells. SHED showed a stronger angiogenesis differentiation and proliferation potential compared with DPSC and BMSC.

The pluripotent nature of embryonic stem cells (ESCs) makes them amenable for regenerative therapies because they can differentiate into cells that form all tissue types within the body (Zandstra and Nagy, 2001). The potential drawbacks to the use of ESCs for cellular therapies include the obvious ethical dilemmas of obtaining ESCs, the potential of cancer or tumor formation and the risk of immunogenic rejection (Wobus and Boheler, 2005). Therefore adult stem cell sources with multipotent and pluripotent potential have been sought as an alternative for ESCs including mesenchymal stem cell (MSCs) and tissuederived specific stem cells. Interestingly, the isolation of a population of dental stem cells derived ectodermally from the neural crest (NC) have been shown to be multipotent and give rise to multifarious cell types that result in the development of many of the body’s organs or tissues (Huang et al., 2009a; Huang et al., 2009b). Stem cells extracted from dental tissues including dental pulp, periodontal ligament, apical papilla and dental follicle precursor cells have an expansive differentiation potential with respect to mesodermal and ectodermal lineages. Currently there are six types of dental stem cells that are well characterized and described both in vitro and in vivo (Gronthos et al., 2000; Huang et al., 2009a; Karaoz et al., 2010; Miura et al., 2003; Morsczeck et al., 2005; Seo et al., 2004; Sonoyama et al., 2006). Some dental stem cells lines have been shown to express ESC markers Oct4, Nanog, Sox2 and Klf4 and NC markers p75, Sox10, Slug and Nestin suggesting that dental stem cells may be able to become many of the same tissues as ESCs (Huang et al., 2009a). Further, dental stem cells have been shown to differentiate into neurogenic, adipogenic, cardiomyogenic, chondrogenic, myogenic and osteogenic lineages (Huang et al., 2009a; Karaoz et al., 2010; Miura et al., 2003; Seo et al., 2004; Sonoyama et al., 2006; Zhang et al., 2006). Since dental stem cells have been shown to differentiate into a multitude of cell types, their potential for use in tissue regeneration may be boundless. We are currently using dental stem cells to investigate the mechanisms of mechanotransduction elicited during dynamic cyclic compression for chondrogenesis. Our long term goal is to develop technology and protocols utilizing dental stem cells and biomechanical force for reparative medicine and tissue regeneration of cartilage. This review

Dental stem cells (DSCs) holds the ability to differentiate into numerous cell types. This property makes these cells particularly appropriate for therapeutic use in regenerative medicine. We report evidence that when DSCs undergo osteogenic differentiation, the osteoblast-like cells can be reverted back to a stem-like state and then further differentiated toward the osteogenic phenotype again, without gene manipulation. We have investigated two different MSCs types, both from dental tissues: dental follicle progenitor stem cells (DFPCs) and dental pulp stem cells (DPSCs). After osteogenic differentiation, both DFPCs and DPSCs can be reverted to a naïve stem cell-like status; importantly, dedifferentiated DSCs showed a greater potential to further differentiate toward the osteogenic phenotype. Our report aims to demonstrate for the first time that it is possible, under physiological conditions, to control the dedifferentiation of DSCs and that the rerouting of cell fate could potentially be used to enhance their osteogenic therapeutic potential. Significantly, this study first validates the use of dedifferentiated DSCs as an alternative source for bone tissue engineering.

Stem Cells are characterized by their properties of self renewal and multipotency.These stem cells are localized inside our body at specialized niches, which are known to have low oxygen tension.Dental pulp tissue found in the middle of the tooth is one such niche which experiences hypoxic conditions due to its anatomical location.However under in-vitro conditions, stem cells are cultured under normoxia condition (21% O 2 concentration).It has been known since decades that low oxygen tension in tissue cultures has positive effect on the cell growth.In this study, we tested this hypothesis on in vitro isolated, expanded, Dental Pulp Stem Cells (DPSCs) grown under low oxygen.The comparative evaluation was done between DPSCs grown in normoxia (21% O 2 ) and Hypoxia (3% O 2 ) on the basis of proliferation, aging, maintenance of stemness and differentiation potential.Our study indicates that DPSCs when grown in hypoxia exhibited better renewal capacity, lesser population time and hence decreased differentiation.Thus, this study emphasizes that hypoxic culture conditions is better in vitro culture condition for expansion of DPSCs in large scale and maintenance of their stemness for therapeutic purpose.

Effects of hydrogen peroxide, doxorubicin and ultraviolet irradiation on senescence of human dental pulp stem cells

BackgroundSeveral controversies exist about the methods of harvesting and eventual utilization of stem cells in Medicine and Dentistry. The objective of the study was to investigate the awareness, attitude and knowledge of the use of stem cells in Dentistry among Nigerian Dentists.MethodsThis descriptive, cross-sectional study was conducted among dentists selected from both private and public health sectors, in some of the major cities in Nigeria.ResultsThe majority of the participants were ≤35 years in age, male, Pentecostal Christians, possessed a postgraduate qualification, had practiced for ≤5 years and were specialists or specializing. In this study, 153(81.0%) of the participants reported awareness about the use of stem cells in dentistry which was significantly associated with qualification and type of practice. Most of the respondents 114 (60.3%) had a poor knowledge of the use of stem cells in Dentistry. This was significantly associated with type of practice and awareness about stem cell use in dentistry but binary logistic regression showed awareness as the only determinant of knowledge. About three-quarters 142 (75.1%) of the participants exhibited positive attitude towards stem cell use. This had a positive non-significant association with knowledge and reported awareness.ConclusionData from this study revealed a high level of awareness, positive attitude to and poor knowledge of the use of stem cells in Dentistry among a cross section of Nigerian Dentists.

Bone regeneration and bone fixation strategies in dentistry utilize scaffolds containing regenerating-competent cells as a replacement of the missing bone portions and gradually replaced by autologous tissues. Mesenchymal stem cells represent an ideal cell population for scaffold-based tissue engineering. Among them, dental pulp stem cells (DPSCs) and periosteal stem cells (PeSCs) have the potential to differentiate into a variety of cell types including osteocytes, suggesting that they can be used with this purpose. However, data on bone regeneration properties of these types of cells in scaffold-based tissue engineering are yet insufficient.In this study, we evaluated temporal dynamic bone regeneration (measured as a percentage of bone volume on the total area of the defect) induced by DPSCs or PeSCs when seeded with different scaffolds to fill critical calvarial defects in SCID Beige nude mice. Two commercially available scaffolds (granular deproteinized bovine bone with 10% porcine collagen and granular β;-tricalcium phosphate) and one not yet introduced on the market (a sponge of agarose and nanohydroxyapatite) were used. The results showed that tissue-engineered constructs did not significantly improve bone-induced regeneration process when compared with the effect of scaffolds alone. In addition, the data also showed that the regeneration induced by β;-tricalcium phosphate alone was higher after 8 weeks than that of scaffold seeded with the 2 stem cell lines. Altogether these findings suggest that further studies are needed to evaluate the potential of DPSCs and PeSCs in tissue construct and identify the appropriate conditions to generate bone tissue in critical-size defects.

For several years, scientists have focused on studying mesenchymal stem cells because of their ability for self-regeneration, their potential for differentiation into multiple lineages (e.g., osteogenic, chondrogenic, and adipogenic cells), and their low immunogenicity and remarkable capacity to modulate the immune system. The importance of these cells ranges from preserving tissue health and repairing injured tissues in their nearby areas, to their use in scientific investigation for the treatment of neurodegenerative, autoimmune, and cardiovascular diseases. Despite the availability of various tissue sources, such as bone marrow and adipose tissue, their collection and handling may not always be easily achievable. However, dental tissues, such as the pulp and periodontal ligament, are relatively accessible supplies that do not require complex or stressful interventions for the patient. Here, we provide a detailed description of each step involved in the isolation and characterization of mesenchymal stem cells from the pulp and periodontal ligament using monoclonal antibodies ensuring a high level of effectiveness. In addition, we also describe a technique to generate the cellular presenescence state. © 2026 The Author(s). Current Protocols published by Wiley Periodicals LLC. Basic Protocol: Isolation of human periodontal ligament and dental pulp stem cells Support Protocol 1: Characterization of human periodontal ligament and dental pulp stem cells Support Protocol 2: Induction of presenescence of human periodontal ligament and dental pulp stem cells.

Teeth are a kind of masticatory organs of special histological origin, unique to vertebrates, playing an important role in chewing, esthetics, and auxiliary pronunciation. In the past decades, with the development of tissue engineering and regenerative medicine, the studies of mesenchymal stem cells (MSCs) gradually attracted the interest of researchers. Accordingly, several types of MSCs have been successively isolated in teeth or teeth-related tissues, including dental pulp stem cells, periodontal ligament stem cells, stem cells from human exfoliated deciduous teeth, dental follicle stem cells, stem cells from apical papilla and gingival mesenchymal stem cells. These dental stem cells (DSCs) are easily accessible, possess excellent stem cell characteristics, such as high proliferation rates and profound immunomodulatory properties. Small-molecule drugs are widely used and show great advantages in clinical practice. As research progressed, small-molecule drugs are found to have various complex effects on the characteristics of DSCs, especially the enhancement of biological characteristics of DSCs, which has gradually become a hot issue in the field of DSCs research. This review summarizes the background, current status, existing problems, future research directions, and prospects of the combination of DSCs with three common small-molecule drugs: aspirin, metformin, and berberine.