Search

Surgical treatment of infrabony defects may result in gingival recession of the neighboring teeth. The aim of this clinical report is to describe a surgical technique to promote gingival margin stability in the treatment of infrabony defects at sites with a thin or medium gingival phenotype. A coronally advanced entire papilla preservation (CA-EPP) flap with a connective tissue graft (CTG) was executed in two different clinical cases. This technique substantially improved interproximal clinical attachment level and pocket closure, with no gingival recession. A CA-EPP flap using a CTG may promote gingival margin stability and can be recommended in regenerative periodontal procedures. [ABSTRACT FROM AUTHOR]

Two fractured maxillary central incisors were restored via the crown fragment reattachment technique with adhesive systems and composite resin material. A long-term successful outcome was achieved after 4 years of follow-up. The patient presented with fractured maxillary central incisors, and a comprehensive intraoral and radiographic examination revealed that both teeth had extended crown fractures with no pulpal exposure (Class II, Ellis and Davey). The tooth fragments were thoroughly cleaned and inspected before being repositioned. Both fragments were in excellent condition with almost no pieces missing. Therefore, the proposed treatment was fragment reattachment. After verifying the repositioning and adaptation of the fractured fragments, a flowable composite was used to perform the reattachment procedure under rubber dam isolation. With modern dental adhesives, the reattachment of tooth fragments in good condition is considered a more conservative option for restoring tooth morphology, esthetics, and function. Although one tooth showed signs of irreversible pulpitis and required root canal treatment 2 weeks after the restoration, the clinical and radiographic evaluations at the 4-year follow-up visits confirmed the success of the restorative treatment. The restorations showed good functional and esthetic outcomes after 4 years of follow-up. [ABSTRACT FROM AUTHOR]

Counting growth layers in dentine and/or secondary cementum is widely used for age determination in wild mammals but the underlying seasonal changes in the structure and degree of mineralisation of dental tissue have not been well characterised. We embedded first (m1) and second (m2) mandibular permanent molar teeth from a 12‐year‐old female Svalbard reindeer (Rangifer tarandus platyrhynchus) in PolyMethylMethAcrylate (PMMA), prepared cut and polished surfaces coated with evaporated carbon and used 20 kV back‐scattered electron imaging in a scanning electron microscope (BSE‐SEM) to study aspects of dental tissue structure which depend on the degree of mineralisation at the micron and sub‐micron scale. BSE‐SEM revealed differences between the mineral content of growth layers (annulations) in the secondary cementum and the primary and secondary dentine, the latter, incidentally, still forming at death in m1. Wide bands of less well mineralised tissue formed in the cementum during active appositional phases. Thin, denser bands formed by maturation‐mineralisation of existing tissue when growth slowed in winter. This maturation mimics the processes seen in lamellar bone and articular cartilage. Counter to previous suggestions, there was evidence of substantial resorption and repair of the secondary cementum and of formation of dentine throughout life. Secondary dentine is layered by mineral content like cementum. In the crown, this was mainly tubular dentine with well‐marked interglobular dentine layers. In the lower pulp chamber and root, it was largely without tubules. Substantial non‐mineralised spaces found at the cement‐dentine junction in the root apical regions in m2 represent inclusions of the Hertwig's Epithelial Root Sheath (HERS) or the Epithelial Rests of Malassez (ERM) between the two tissues, a phenomenon which has previously only been identified in Muridae. The anatomical changes which result in the formation of the incremental lines (annulations) in dental tissues of reindeer, identified here for the first time at the micrometre level, are likely to be common across most if not all long‐lived species of mammals living in seasonal environments. [ABSTRACT FROM AUTHOR]

Objectives: This study aimed to compare the tissue dissolving capacities of sodium and calcium hypochlorite [NaOCl and Ca(OCl)2] solutions applied without activation or with two irrigant activation methods. Materials and Methods: One hundred and eight dentin cavities were prepared. Preweighed tissue pieces were placed in these dentin cavities, and samples were divided into nine groups of twelve. In groups 1, 2, and 3, NaOCl was used with non-activation (NA), passive ultrasonic (PUA), and laser activation (LA). With the same techniques, Ca(OCl)2 was used in groups 4, 5, and 6, and distilled water was used in groups 7, 8, and 9. The weight loss of tissue samples was calculated and analyzed using Two-way ANOVA and Duncan tests. Results: Distilled water groups showed no tissue dissolution in any conditions. NaOCI and Ca(OCI)2 showed statistically similar dissolving effectiveness when used with the same technique. Activated groups dissolved significantly greater tissue than non-activated ones, the highest in LA. Conclusions: Ca(OCI)2 can be an alternative to NaOCl; for both, the first choice of activation may be the LA. [ABSTRACT FROM AUTHOR]

Preserving pulp viability and promoting pulp regeneration in pulpitis have attracted widespread attention. Restricted by the oxidative stress microenvironment of dental pulpitis, excessive reactive oxygen and nitrogen species (RONS) trigger uncontrolled inflammation and exacerbate pulp tissue destruction. However, modulating redox homeostasis in inflamed pulp tissue to promote pulp regeneration remains a great challenge. Herein, this work proposes an effective antioxidative system (C-NZ/GelMA) consisting of carbon dot nanozymes (C-NZ) with gelatin methacryloyl (GelMA) to modulate the pulpitis microenvironment for dental pulp regeneration by utilizing the antioxidant properties of C-NZ and the mechanical support of an injectable GelMA hydrogel. This system effectively scavenges RONS to normalize intracellular redox homeostasis, relieving oxidative stress damage. Impressively, it can dramatically enhance the polarization of regenerative M2 macrophages. This study revealed that the C-NZ/GelMA hydrogel promoted pulp regeneration and dentin repair through its outstanding antioxidant, antiapoptotic, and anti-inflammatory effects, suggesting that the C-NZ/GelMA hydrogel is highly valuable for pulpitis treatment. [ABSTRACT FROM AUTHOR]

Mesenchymal stem cell derived extracellular vesicles (MSC EVs) are paracrine modulators of macrophage function. Scientific research has primarily focused on the immunomodulatory and regenerative properties MSC EVs derived from bone marrow. The dental pulp is also a source for MSCs, and their anatomical location and evolutionary function has primed them to be potent immunomodulators. In this study, we demonstrate that extracellular vesicles derived from dental pulp stem cells (DPSC EVs) have pronounced immunomodulatory effect on primary macrophages by regulating the NFkb pathway. Notably, the anti-inflammatory activity of DPSC-EVs is enhanced following exposure to an inflammatory stimulus (LPS). These inhibitory effects were also observed in vivo. Sequencing of the naïve and LPS preconditioned DPSC-EVs and comparison with our published results from marrow MSC EVs revealed that Naïve and LPS preconditioned DPSC-EVs are enriched with anti-inflammatory miRNAs, particularly miR-320a-3p, which appears to be unique to DPSC-EVs and regulates the NFκb pathway. Overall, our findings highlight the immunomodulatory properties of DPSC-EVs and provide vital clues that can stimulate future research into miRNA-based EV engineering as well as therapeutic approaches to inflammation control and disease treatment. [ABSTRACT FROM AUTHOR]

Hypoxia is relevant to several physiological and pathological processes and this also applies for the tooth. The adaptive response to lowering oxygen concentration is mediated by hypoxia-inducible factors (HIFs). Since HIFs were shown to participate in the promotion of angiogenesis, stem cell survival, odontoblast differentiation and dentin formation, they may play a beneficial role in the tooth reparative processes. Although some data were generated in vitro, little is known about the in vivo context of HIFs in tooth development. In order to contribute to this field, the mouse mandibular first molar was used as a model. The expression and in situ localisation of HIFs were examined at postnatal (P) days P0, P7, P14, using RT-PCR and immunostaining. The expression pattern of a broad spectrum of hypoxia-related genes was monitored by customised PCR Arrays. Metabolic aspects were evaluated by determination of the lactate level and mRNA expression of the mitochondrial marker Nd1. The results show constant high mRNA expression of Hif1a, increasing expression of Hif2a, and very low expression of Hif3a during early postnatal molar development. In the examined period the localisation of HIFs in the nuclei of odontoblasts and the subodontoblastic layer identified their presence during odontoblastic differentiation. Additionally, the lower lactate level and higher expression of mitochondrial Nd1 in advanced development points to decreasing glycolysis during differentiation. Postnatal nuclear localisation of HIFs indicates a hypoxic state in specific areas of dental pulp as oxygen demands depend on physiological events such as crown and root dentin mineralization. [ABSTRACT FROM AUTHOR]

This study addresses the morphological and chemical characterization of PGS scaffolds after (6, 12, 18, 24, and 30 min) residence in undoped pyrrole plasma (PGS-PPy) and the evaluation of cell viability with human dental pulp stem cells (hDPSCs). The results were compared with a previous study that used iodine-doped pyrrole (PGS-PPy/I). Analyses through SEM and AFM revealed alterations in the topography and quantity of deposited PPy particles. FTIR spectra of PGS-PPy scaffolds confirmed the presence of characteristic absorption peaks of PPy, with higher intensities observed in the nitrile and -C≡C- groups compared to PGS-PPy/I scaffolds, while raman spectra indicated a lower presence of polaron N+ groups. On the other hand, PGS scaffolds modified with PPy exhibited lower cytotoxicity compared to PGS-PPy/I scaffolds, as evidenced by the Live/Dead assay. Furthermore, the PGS-PPy scaffolds at 6 and 12 min, and particularly the PGS-PPy/I scaffold at 6 min, showed the best results in terms of cell viability by the fifth day of culture. The findings of this study suggest that undoped pyrrole plasma modification for short durations could also be a viable option to enhance the interaction with hDPSCs, especially when the treatment times range between 6 min and 12 min. [ABSTRACT FROM AUTHOR]

Aim: To evaluate the role of biomimetic pulp scaffolds derived from the extracellular matrix derived of stem cells from human exfoliated deciduous teeth (SHED‐ECM‐PS) in promoting pulp–dentine complex regeneration. Methodology: SHED‐ECM‐PS was prepared through cell aggregation and decellularization techniques. Histological and immunofluorescence analyses, scanning electron microscopy, and DNA quantification assays were used to characterize the SHED‐ECM‐PS. Additionally, a tooth slice implantation model was established to evaluate the effects of SHED‐ECM‐PS on regeneration of the pulp–dentine complex in vivo. Extraction medium for SHED‐ECM‐PS was prepared, and its effect on bone marrow mesenchymal stem cells (BMMSCs) was assessed in vitro. Cell counting kit‐8 and Ki‐67 staining assays were performed to determine cell proliferation. The rate of apoptosis was evaluated by flow cytometry. Wound healing and transwell assays were conducted to evaluate cell migration. Alizarin red S staining was performed to examine mineralized nodule formation. Western blotting was used to detect the expression of osteogenic and odontogenic markers. The results were analysed using an independent two‐tailed Student's t‐test. p <.05 was considered statistically significant. Results: SHED‐ECM‐PS was successfully constructed, exhibiting a striped dental pulp‐like shape devoid of nuclear structures or DNA components, and rich in fibronectin, collagen I, DMP1 and DSPP. Notably, SHED‐ECM‐PS showed no impact on the proliferation or apoptosis of BMMSCs. Histological analysis revealed that dental pulp fibroblasts formed an interwoven mesh in the root canal, and angiogenesis was observed in the SHED‐ECM‐PS group. Moreover, a continuous, newly formed tubular dentine layer with polarized odontoblast‐like cells was observed along the inner wall of the root canal. SHED‐ECM‐PS promoted the migration, polar alignment and mineralized nodule formation of BMMSCs and specifically elevated the expression levels of odontogenic markers, but not osteogenic markers, compared with the control group in vitro. Conclusion: SHED‐ECM‐PS exhibited no cytotoxicity and promoted pulp‐dentine complex regeneration in vivo as well as cell migration and odontogenic differentiation of BMMSCs in vitro. These findings provide evidence that SHED‐ECM‐PS, as a novel biological scaffold, has the potential to improve the outcomes of REPs. [ABSTRACT FROM AUTHOR]

Physiological root resorption of deciduous teeth is a normal phenomenon occurring during the developmental stages of children. Previous research has indicated the pivotal role of the inflammatory microenvironment in this process, although the specific mechanisms remain unclear. This study is aimed at elucidating the involvement of the alpha7 nicotinic acetylcholine receptors (α7 nAChR)-autophagy axis in the regulation of the inflammatory microenvironment during physiological root resorption in deciduous teeth. Samples were collected from deciduous teeth at various stages of physiological root resorption, and deciduous dental pulp stem cells (DDPSCs) were isolated and cultured during the mid-phase of root resorption. The findings revealed a substantial infiltration of the pulp of deciduous teeth at the mid-phase of root resorption, characterized by elevated expression levels of α7 nAChR and IL-1β. Significantly increased IL-1β and α7 nAChR expressions were observed in DDPSCs during the mid-phase of root resorption, with α7 nAChR demonstrating a regulatory effect on IL-1β. Moreover, evidence suggested that mechanical stress may act as a trigger, regulating autophagy and IL-1 expression via α7 nAChR. In conclusion, mechanical stress was identified as a regulator of autophagy in DDPSCs through α7 nAChR, influencing the expression of IL-1β and contributing to the formation of the inflammatory microenvironment. This mechanism plays a crucial role in the physiological root resorption of deciduous teeth. Key messages: The pulp of deciduous teeth at mid-phase of root resorption was heavily infiltrated with high expression of α7nAChR and IL-1β. α7 nAChR acts as an initiating factor to regulate IL-1β through autophagy in DDPSCs. Mechanical stress can regulate autophagy of DDPSCs through α7 nAChR and thus affect IL-1β expression and inflammatory microenvironment formation in physiological root resorption in deciduous teeth. [ABSTRACT FROM AUTHOR]

The purpose of this study was evaluating the performance of new Bis-EFMA based bulk-fill composites with common methacrylate based composites and commercial dental composites. The Bis-EFMA monomer was synthesized and the novel Bis-EFMA based bulk-fill composites were prepared. The resin composite samples were co-cultured with human gingival epithelial cells and human dental pulp stem cells to test the biocompatibility. The edge adaptation was observed under a combination of stereoscope and scanning electron microscope. The internal hardness was measured using a Vickers microhardness tester after one-time filling of cavities prepared in extracted teeth. After friction and wear test on the surface of the resin composites, the surface morphology and volume wear of each group were measured by the optical profilometer. The color stability was measured by a colorimeter. Direct contact with human gingival epithelial cells and human dental pulp stem cells did not cause significant changes in their growth density and morphology, indicating good biocompatibility of Bis-EFMA group (p > 0.05). The continuous margin proportion of the Bis-EFMA group was as good as commercial bulk-fill composites (p > 0.05). The sectional microhardness results showed that the Bis-EFMA group had the highest microhardness. After the friction and wear test, the volume wear of the Bis-EFMA group was minimal, indicating its good wear resistance and mechanical strength. Color changes in all resin groups after 28 days of immersion were within the clinically acceptable range. The addition of Bis-EFMA demonstrated excellent biocompatibility, edge adaptation and color stability comparable to commonly used clinical bulk-fill composites, along with preferable mechanical strength, friction and wear resistance. Bis-EFMA based bulk-fill composites have the potential to be employed as a bulk filling material in commercial dental composite applications. • Bis-EFMA BF-DRCs showed good biocompatibility to the pulp periodontal tissue. • Bis-EFMA BF-DRCs adapted well to cavity wall. • The rigid structure got Bis-EFMA BF-DRCs higher microhardness and anti-friction property compared with SDR™. • Bis-EFMA BF-DRCs had reliable color stability. [ABSTRACT FROM AUTHOR]

Silicon-releasing biomaterials are widely used in the field of dentistry. However, unlike bone, very little is known about the role of silicon on dental tissue formation and repair. This study investigates the influence of silicic acid on the survival, differentiation and mineralizing ability of human dental pulp stem cells (hDPSCs) in 3D pulp-like environments Dense type I collagen hydrogels seeded with hDPSCs were cultured over 4 weeks in the presence of silicic acid at physiological (10 μM) and supraphysiological (100 μM) concentrations. Cell viability and proliferation were studied by Alamar Blue and live/dead staining. The collagen network was investigated using second harmonic generation imaging. Mineral deposition was monitored by histology and scanning electron microscopy. Gene expression of mineralization- and matrix remodeling-associated proteins was studied by qPCR. Presence of silicic acid did not show any significant influence on cell survival, metabolic activity and gene expression of key mineralization-related proteins (ALP, OCN, BSP). However, it induced enhanced cell clustering and delayed expression of matrix remodeling-associated proteins (MMP13, Col I). OPN expression and mineral deposition were inhibited at 100 μM. It could be inferred that silicic acid has no direct cellular effect but rather interacts with the collagen network, leading to a modification of the cell-matrix interface. Our results offer advanced insights on the possible role of silicic acid, as released by pulp capping calcium silicates biomaterials, in reparative dentine formation. More globally, these results interrogate the possible role of Si in pulp pathophysiology. [Display omitted] • hDPSCs were cultured inside dense type I collagen hydrogels in the presence of silicic acid. • Silicic acid shows no toxicity but induces cell clustering and inhibits mineralization at 100 μM. • Silicic acid has no impact on mineralization proteins expression but delayed matrix remodeling. • Silicic acid interactions with the collagen matrix may have a major influence on hDPSCs behavior. [ABSTRACT FROM AUTHOR]

Evaluate a new light-cured material with better properties for vital pulp therapy. Light-cured resin materials consisted of polyethylene glycol (600) diacrylate mixed with different ratios of TCP to HA. In addition to the temperature change (n = 5 for each subgroup) were tested, cell viability and Alizarin Red Staining (ARS) assay were also tested in vitro on human dental pulp cells (n = 6 for each subgroup). Lastly, the material was then compared with Biodentine and control groups in the molars of Wistar rats in vivo for histology assessment. The temperature change for the new materials were under 5 degrees Celsius. For the in vitro assessments, there was no significant difference on day 3 and day 7 for cell viability test. ARS assay showed significantly higher mineralized nodule formation when treated without induction medium for Group D and Biodentine on day 10 compared to Group C and control. On the contrary, Biodentine and control groups treated with induction medium showed significant higher mineralization than the new materials. Histology assessments demonstrated higher mineralized content in Group D and Biodentine on week 3 and week 6. The inflammatory cells in the dental pulp complex of the Biodentine group resolved on week 6 while the inflammation resolved in Group D on week 3. The new material exhibits low heat production, low cytotoxicity, and good calcium ion release capability. Compared to traditional materials, it has shorter setting time and better aesthetic outcomes, making it highly suitable for use in vital pulp therapy. • Light-cured direct pulp capping material with low cytotoxicity and low temperature rise. • New material for vital pulp therapy tested in vitro on human dental pulp cells. • New material for vital pulp therapy tested in vivo in the molars of Sprague-Dawley (SD) rats. • New light-cured material for vital pulp therapy compared with Biodentine. [ABSTRACT FROM AUTHOR]

• Uninduced and neuronally-induced DPSCs demonstrate potential for Parkinson's disease therapy. • Uninduced and neuronally-induced DPSCs promote significant recovery in dopaminergic neurons and locomotion. • Neuronally-induced DPSCs prevent GFAP elevation and alter DARPP-32 phosphorylation states. • Uninduced DPSCs reduce JUN levels and mitigate glycosylated DAT elevation. In recent years, dental pulp stromal cells (DPSCs) have emerged as a promising therapeutic approach for Parkinson's disease (PD), owing to their inherent neurogenic potential and the lack of neuroprotective treatments for this condition. However, uncertainties persist regarding the efficacy of these cells in an undifferentiated state versus a neuronally-induced state. This study aims to delineate the distinct therapeutic potential of uninduced and neuronally-induced DPSCs in a rodent model of PD induced by 6-Hydroxydopamine (6-OHDA). DPSCs were isolated from human teeth, characterized as mesenchymal stromal cells, and induced to neuronal differentiation. Neuronal markers were assessed before and after induction. DPSCs were transplanted into the substantia nigra pars compacta (SNpc) of rats 7 days following the 6-OHDA lesion. In vivo tracking of the cells, evaluation of locomotor behavior, dopaminergic neuron survival, and the expression of essential proteins within the dopaminergic system were conducted 7 days postgrafting. Isolated DPSCs exhibited typical characteristics of mesenchymal stromal cells and maintained a normal karyotype. DPSCs consistently expressed neuronal markers, exhibiting elevated expression of βIII-tubulin following neuronal induction. Results from the animal model showed that both DPSC types promoted substantial recovery in dopaminergic neurons, correlating with enhanced locomotion. Additionally, neuronally-induced DPSCs prevented GFAP elevation, while altering DARPP-32 phosphorylation states. Conversely, uninduced DPSCs reduced JUN levels. Both DPSC types mitigated the elevation of glycosylated DAT. Our results suggested that uninduced DPSCs and neuronally-induced DPSCs exhibit potential in reducing dopaminergic neuron loss and improving locomotor behavior, but their underlying mechanisms differ. [ABSTRACT FROM AUTHOR]

The aim of this study was to evaluate the stress induced by clamps made of different materials on mandibular first molar teeth using finite element analysis. The tooth model to be used in the study was created using micro-CT scanner and the rubber dam clamp model was created in three dimensions (3D) using Solidworks and applied to a finite element tooth model of an intact mandibular first molar. The size of the clamp opening and the force to be applied were calculated according to the buccolingual distances of the tooth. The contact areas of the clamps on the tooth were determined and subjected to force. The stress distribution patterns and maximum von Mises stresses were calculated and compared against these forces. The most stress against these forces occurred in the enamel tissue on the distal side where the clamp was in contact with the tooth. The stress value in the stainless steel clamp (191.63 MPa) was approximately 85% higher than the clamp made of polyethylene ketone (103.85 MPa). The stress in the dentin and pulp tissue of the tooth was negligible. Clinicians should examine the enamel tissue below the equatorial line of the tooth in detail before clamp application. The forces exerted by stainless steel clamps can cause damage to the enamel tissue of the tooth or progression of an existing enamel crack. [ABSTRACT FROM AUTHOR]

The intestinal microbiota has an important role in animal health. Therefore, prebiotics have gained interest in the scientific community for their role in manipulating populations of intestinal microorganisms. Among prebiotics, there is Smallanthus sonchifolius Poepp. Endl (yacon) root, which can be ingested in natura or in different forms, such as flours and extracts. This prebiotic has been associated with beneficial effects in different diseases, including metabolic (like type 2 diabetes) and immunological disorders. Thus, mouse models of human diseases caused by immunological factors have been used to better understand the prebiotic effects of yacon. Since prebiotics interfere with animal microbiota, it is important to take into consideration the quality level of mouse facilities. In this way, the beneficial effects of a yacon root pulp were tested in a mouse model of allergic asthma and, considering how animal facility conditions could affect those results, we performed experiments both in conventional facilities and specific pathogen-free (SPF) ones. Our results showed that better prebiotic effects were observed in a SPF facility than in a conventional one and, in some cases, evidence of yacon beneficial effects was observed exclusively in a SPF facility. For example, there were reductions of 63.6% and 58.0% in IgE and eosinophils, respectively, in immunized/yacon-treated animals compared to immunized mice only. Taken together, our results not only showed the beneficial effects of yacon root pulp in an asthma model but also demonstrated the relevance of performing experiments with prebiotics in SPF facilities. [ABSTRACT FROM AUTHOR]

Background and Purpose: Pulpitis is associated with tooth hypersensitivity and results in pulpal damage. Thermosensitive transient receptor potential (TRP) ion channels expressed in the dental pulp may be key transducers of inflammation and nociception. We aimed at investigating the expression and role of thermo‐TRPs in primary human dental pulp cells (hDPCs) in normal and inflammatory conditions. Experimental Approach: Inflammatory conditions were induced in hDPC cultures by applying polyinosinic:polycytidylic acid (poly(I:C)). Gene expression and pro‐inflammatory cytokine release were measured by RT‐qPCR and ELISA. Functions of TRPA1 channels were investigated by monitoring changes in intracellular Ca2+ concentration. Mitochondrial superoxide production was measured using a fluorescent substrate. Cellular viability was assessed by measuring the activity of mitochondrial dehydrogenases and cytoplasmic esterases. TRPA1 activity was modified by agonists, antagonists, and gene silencing. Key Results: Transcripts of TRPV1, TRPV2, TRPV4, TRPC5, and TRPA1 were highly expressed in control hDPCs, whereas TRPV3, TRPM2, and TRPM3 expressions were much lower, and TRPM8 was not detected. Poly(I:C) markedly up‐regulated TRPA1 but not other thermo‐TRPs. TRPA1 agonist‐induced Ca2+ signals were highly potentiated in inflammatory conditions. Poly(I:C)‐treated cells displayed increased Ca2+ responses to H2O2, which was abolished by TRPA1 antagonists. Inflammatory conditions induced oxidative stress, stimulated mitochondrial superoxide production, resulted in mitochondrial damage, and decreased cellular viability of hDPCs. This inflammatory cellular damage was partly prevented by the co‐application of TRPA1 antagonist or TRPA1 silencing. Conclusion and Implications: Pharmacological blockade of TRPA1 channels may be a promising therapeutic approach to alleviate pulpitis and inflammation‐associated pulpal damage. [ABSTRACT FROM AUTHOR]

Objective: Significant evidence links epigenetic processes governing the dynamics of DNA methylation and demethylation to an increased risk of syndromic and nonsyndromic cleft lip and/or cleft palate (CL/P). Previously, we characterized mesenchymal stem/stromal cells (MSCs) at different stages of osteogenic differentiation in the mouse incisor dental pulp. The main objective of this research was to characterize the transcriptional landscape of regulatory genes associated with DNA methylation and demethylation at a single-cell resolution. Design: We used single-cell RNA sequencing (scRNA-seq) data to characterize transcriptome in individual subpopulations of MSCs in the mouse incisor dental pulp. Settings: The biomedical research institution. Patients/Participants: This study did not include patients. Interventions: This study collected and analyzed data on the single-cell RNA expssion in the mouse incisor dental pulp. Main outcome measure(s): Molecular regulators of DNA methylation/demethylation exhibit differential transcriptional landscape in different subpopulations of osteogenic progenitor cells. Results: scRNA-seq analysis revealed that genes encoding DNA methylation and demethylation enzymes (DNA methyltransferases and members of the ten-eleven translocation family of methylcytosine dioxygenases), methyl-DNA binding domain proteins, as well as transcription factors and chromatin remodeling proteins that cooperate with DNA methylation machinery are differentially expressed within distinct subpopulations of MSCs that undergo different stages of osteogenic differentiation. Conclusions: These findings suggest some mechanistic insights into a potential link between epigenetic alterations and multifactorial causes of CL/P phenotypes. [ABSTRACT FROM AUTHOR]

Dental pulp stem cells (DPSCs) are responsible for maintaining pulp structure and function after pulp injury. DPSCs migrate directionally to the injury site before differentiating into odontoblast-like cells, which is a prerequisite and a determinant in pulp repair. Increasing evidence suggests that sensory neuron-stem cell crosstalk is critical for maintaining normal physiological functions, and sensory nerves influence stem cells mainly by neuropeptides. However, the role of sensory nerves on DPSC behaviors after pulp injury is largely unexplored. Here, we find that sensory nerves released significant amounts of calcitonin gene-related peptide (CGRP) near the injury site, acting directly on DPSCs via receptor activity modifying protein 1 (RAMP1) to promote collective migration of DPSCs to the injury site, and ultimately promoting pulp repair. Specifically, sensory denervation leads to poor pulp repair and ectopic mineralization, in parallel with that DPSCs failed to be recruited to the injury site. Furthermore, in vitro evidence shows that sensory nerve-deficient microenvironment suppressed DPSC migration prominently among all related behaviors. Mechanistically, the CGRP-Ramp1 axis between sensory neurons and DPSCs was screened by single-cell RNA-seq analysis and immunohistochemical studies confirmed that the expression of CGRP rather than Ramp1 increases substantially near the damaged site. We further demonstrated that CGRP released by sensory nerves binds the receptor Ramp1 on DPSCs to facilitate cell collective migration by an indirect co-culture system using conditioned medium from trigeminal neurons, CGRP recombinant protein and antagonists BIBN4096. The treatment with exogenous CGRP promoted the recruitment of DPSCs, and ultimately enhanced the quality of pulp repair. Targeting the sensory nerve could therefore provide a new strategy for stem cell-based pulp repair and regeneration. [ABSTRACT FROM AUTHOR]

Aim Methodology Results Conclusion The aim of this study is to investigate the expression of inflammatory biomarkers (TNF‐α, IL‐10, IL‐1β) and the pulpitis‐associated miRNA (miR‐30a‐5p and miR‐128‐3p) in pulp tissue samples from unrestored teeth with a vital normal pulp (NP), teeth with symptomatic irreversible pulpitis (IP) and in unrestored teeth with periodontal disease, unresponsive to periodontal therapy, and a vital pulp (EP).Thirty patients were included in this observational study (10 teeth with NP, 10 teeth with IP, 10 teeth with EP). Dental pulp tissues samples were collected from patients during root canal treatment (RCT). RNA was extracted and qRT‐PCR of target genes (tumour necrosis factor [TNF]‐α, interleukin [IL]‐1β, IL‐10) and miRNAs (has‐miR‐30a‐5p, has‐miR‐128‐3p) performed to assess the expression profile. Fold‐change in expression was calculated using the formula 2−(ΔCt(Exp)−ΔCt(Ctrl)). One‐way anova with post‐hoc Tukey's was used to determine significant differences between groups. The significance level was set at 5% (p < .05). All teeth were also followed up clinically for 1 year and evaluated for a range of endodontic and periodontal‐related outcomes.All investigated genes significantly increased in expression and miRNAs significantly decreased in expression in the IP and EP groups compared with the NP group (p < .05). With regards to TNF‐α and IL‐1β there were no significant differences in expression between the IP and EP groups (p > .05), whereas IL‐10 expression levels were significantly reduced in the EP compared with the IP group (p < .05). Both miR‐30a‐5p and miR‐128‐3p showed significantly reduced expression in both IP and EP lesions, compared with NP (p < .05); however, no significant differences in miRNA expression were observed between IP and EP groups (p > .05). One year after root canal treatment and periodontal maintenance, tooth mobility and probing depth were significantly reduced in the EP group (p < .05).Pulp tissues from teeth with IP and EP presented similar levels of altered inflammatory markers compared with NP. TNF‐α, IL‐10, IL‐1β cytokines and miRNAs (miR‐30a‐5p and miR‐128‐3p) are potential objective biomarkers to indicate pulpal inflammatory status, aiding diagnosis and directing clinical decision‐making. RCT may be beneficial to improve stage III periodontitis unresponsive to non‐surgical periodontal treatment, but further research is required. [ABSTRACT FROM AUTHOR]

The degeneration of spiral ganglion neurons (SGNs), which convey auditory signals from hair cells to the brain, can be a primary cause of sensorineural hearing loss (SNHL) or can occur secondary to hair cell loss. Emerging therapies for SNHL include the replacement of damaged SGNs using stem cell-derived otic neuronal progenitors (ONPs). However, the availability of renewable, accessible, and patient-matched sources of human stem cells is a prerequisite for successful replacement of the auditory nerve. In this study, we derived ONP and SGN-like cells by a reliable and reproducible stepwise guidance differentiation procedure of self-renewing human dental pulp stem cells (hDPSCs). This in vitro differentiation protocol relies on the modulation of BMP and TGFβ pathways using a free-floating 3D neurosphere method, followed by differentiation on a Geltrex-coated surface using two culture paradigms to modulate the major factors and pathways involved in early otic neurogenesis. Gene and protein expression analyses revealed efficient induction of a comprehensive panel of known ONP and SGN-like cell markers during the time course of hDPSCs differentiation. Atomic force microscopy revealed that hDPSC-derived SGN-like cells exhibit similar nanomechanical properties as their in vivo SGN counterparts. Furthermore, spiral ganglion neurons from newborn rats come in close contact with hDPSC-derived ONPs 5 days after co-culturing. Our data demonstrate the capability of hDPSCs to generate SGN-like neurons with specific lineage marker expression, bipolar morphology, and the nanomechanical characteristics of SGNs, suggesting that the neurons could be used for next-generation cochlear implants and/or inner ear cell-based strategies for SNHL. [ABSTRACT FROM AUTHOR]

Post-mortem interval (PMI) estimation remains one of the major challenges in forensic practice, especially for late PMIs beyond 7–10 days after the death of the subject. In 2022, an innovative method to investigate the occurrence of mutations induced by the death of a subject in the DNA of post-mortem dental pulps at different PMIs was developed, applying a next-generation sequencing (NGS) analysis. The present study aims to apply the same method of analysis to a small sample of teeth belonging to the same subject and analyzed at different PMIs/accumulated degree days (ADDs), and of teeth extracted from different subjects but analyzed at the same PMI/ADD to verify the repeatability of the results obtained in relation to the time elapsed since death. A total of 10 teeth were collected from 6 patients (3 males and 3 females) with PMI varying from 8 to 35 days, and ADD from 157.4 to 753.8. We found 1754 mutations in 56 genes, with more than 700 mutations having a prevalence > 5% and more than 300 variants considered of interest for the purposes of the study. Mutations that were not present at lower PMIs but manifested in later PMIs in pulps belonging to the same subject demonstrate that they can only have been acquired by the subject after death and according to the time elapsed since death. In total, 67 somatic mutations in 29 out of the 56 genes of the used panel occurred in a fashion that allows an association with specific PMI/ADD ranges (within 8 days, between 17 and 28, and beyond 30 days after death). The results suggest that temperature and humidity could influence the rate of DNA degeneration in dental pulps, thus PMI should be estimated in ADD more than days. The preliminary validation supports the hypothesis that the innovative method could be a useful tool for estimating the post-mortem interval even beyond the first week after death, but further analyses are needed to customize a specific genetic panel for forensic investigations and verify the influence of degenerative processes of soft tissues surrounding dental elements on DNA degeneration of pulps. [ABSTRACT FROM AUTHOR]

Dental pulp stem cells (DPSC) have shown osteogenic and bone regenerative potential. Improving the in situ bone regeneration potential of DPSC is crucial for their application as seed cells during bone defect reconstruction in clinics. This study aimed to develop DPSC-derived organoid-like microspheroids as effective seeds for bone tissue engineering applications. DPSC osteogenic microspheroids (70 μm diameter) were cultured in a polydimethylsiloxane-mold-based agarose-gel microwell-culture-system with or without cannabidiol (CBD)-treatment. Results of in vitro studies showed higher osteogenic differentiation potential of microspheroids compared with 2D-cultured-DPSC. CBD treatment further improved the osteogenic differentiation potential of microspheroids. The effect of CBD treatment in the osteogenic differentiation of microspheroids was more pronounced compared with that of CBD-treated 2D-cultured-DPSC. Microspheroids showed a higher degree of bone regeneration in nude mice calvarial bone defect compared to 2D-cultured-DPSC. CBD-treated microspheroids showed the most robust in situ bone regenerative potential compared with microspheroids or CBD-treated 2D-cultured-DPSC. According to mRNA sequencing, bioinformatic analysis, and confirmation study, the higher osteogenic potential of CBD-treated microspheroids was mainly attributed to WNT6 upregulation. Taken together, DPSC microspheroids have robust osteogenic potential and can effectively translate the effect of in vitro osteoinductive stimulation during in situ bone regeneration, indicating their application potential during bone defect reconstruction in clinics. Dental pulp stem cells-derived microspheroids have robust osteogenic potential and can effectively translate the effect of in vitro osteoinductive stimulation during in situ bone regeneration, indicating their application potential during bone defect reconstruction in the clinic. [ABSTRACT FROM AUTHOR]

Aim Methodology Results Conclusions Angiogenesis is a key event in the successful healing of pulp injuries, and hypoxia is the main stimulator of pulpal angiogenesis. In this study, we investigated the effect of hypoxia on the proangiogenic potential of human dental pulp stem cells (hDPSCs) and the role of miR‐143‐5p in the process.Human dental pulp stem cells were isolated, cultured and characterized in vitro. Cobalt chloride (CoCl2) was used to induce hypoxia in hDPSCs. CCK‐8 and Transwell assays were used to determine the effect of hypoxia on hDPSCs proliferation and migration. Quantitative real‐time polymerase chain reaction (qRT‐PCR), Western blotting (WB) and ELISA were performed to assess the mRNA and protein levels of HIF‐1α and angiogenic cytokines in hDPSCs. The effect of hypoxia on hDPSCs proangiogenic potential was measured in vitro using Matrigel tube formation and chick chorioallantoic membrane (CAM) assays. Recombinant lentiviral vectors were constructed to stably overexpress or inhibit miR‐143‐5p in hDPSCs, and the proangiogenic effects were assessed using qRT‐PCR, WB, and tube formation assays. miR‐143‐5p target genes were identified and verified using bioinformatics prediction tools, dual‐luciferase reporter assays and RNA pull‐down experiments. Finally, a subcutaneous transplantation model in nude mice was used to determine the effects of hypoxia treatment and miR‐143‐5p overexpression/inhibition in hDPSCs in dental pulp regeneration.Hypoxia promotes hDPSCs proliferation, migration and proangiogenic potential. The in vivo experiments showed that hypoxia treatment (50 and 100 μM CoCl2) promoted pulp angiogenesis and dentine formation. In contrast to the levels of proangiogenic factors, miR‐143‐5p levels decreased with increasing CoCl2 concentration. miR‐143‐5p inhibition significantly promoted proangiogenic potential of hDPSCs, whereas miR‐143‐5p overexpression inhibited angiogenesis in vitro. Dual‐luciferase reporter assay identified retinoic acid receptor‐related orphan receptor alpha (RORA) as an miR‐143‐5p target gene in hDPSCs. RNA pull‐down experiments demonstrated that HIF‐1α and RORA were pulled down by biotin‐labelled miR‐143‐5p, and the levels of HIF‐1α and RORA bound to miR‐143‐5p in the hypoxia group were lower than those in the normoxia group. Inhibition of miR‐143‐5p expression in hDPSCs promoted ectopic dental pulp tissue regeneration.CoCl2‐induced hypoxia promotes hDPSCs‐driven paracrine angiogenesis and pulp regeneration. The inhibition of miR‐143‐5p upregulates the proangiogenic potential of hDPSCs under hypoxic conditions by directly targeting HIF‐1α and RORA. [ABSTRACT FROM AUTHOR]

On a global note, oral health plays a critical role in improving the overall human health. In this vein, dental-related issues with dentin exposure often facilitate the risk of developing various oral-related diseases in gums and teeth. Several oral-based ailments include gums-associated (gingivitis or periodontitis), tooth-based (dental caries, root infection, enamel erosion, and edentulous or total tooth loss), as well as miscellaneous diseases in the buccal or oral cavity (bad breath, mouth sores, and oral cancer). Although established conventional treatment modalities have been available to improve oral health, these therapeutic options suffer from several limitations, such as fail to eradicate bacterial biofilms, deprived regeneration of dental pulp cells, and poor remineralization of teeth, resulting in dental emergencies. To this end, the advent of nanotechnology has resulted in the development of various innovative nanoarchitectured composites from diverse sources. This review presents a comprehensive overview of different nanoarchitectured composites for improving overall oral health. Initially, we emphasize various oral-related diseases, providing detailed pathological circumstances and their effects on human health along with deficiencies of the conventional therapeutic modalities. Further, the importance of various nanostructured components is emphasized, highlighting their predominant actions in solving crucial dental issues, such as anti-bacterial, remineralization, and tissue regeneration abilities. In addition to an emphasis on the synthesis of different nanostructures, various nano-therapeutic solutions from diverse sources are discussed, including natural (plant, animal, and marine)-based components and other synthetic (organic- and inorganic-) architectures, as well as their composites for improving oral health. Finally, we summarize the article with an interesting outlook on overcoming the challenges of translating these innovative platforms to clinics. [ABSTRACT FROM AUTHOR]

Synthetic oligodeoxynucleotides (ODNs) containing unmethylated cytosine–phosphate–guanine (CpG) motifs (CpG-ODNs) are ligand molecules for Toll-like receptor 9 (TLR9), which is expressed by odontoblasts in vitro and dental pulp cells. This study determined the effects of CpG-ODNs on pulpal immunomodulatory response and repair following injury. Briefly, the upper right first molars of three-week-old mice were extracted, immersed in Type A (D35) or B (K3) CpG-ODN solutions (0.1 or 0.8 mM) for 30 min, and then replanted. Pulpal healing and immunomodulatory activity were assessed by hematoxylin–eosin and AZAN staining, as well as immunohistochemistry. One week following the operation, inflammatory reactions occurred in all of the experimental groups; however, re-revascularization and newly formed hard tissue deposition were observed in the pulp chamber of all groups at week 2. A positive trend in the expression of immune cell markers was observed toward the CpG-ODN groups at 0.1 mM. Our data suggest that synthetic CpG-ODN solutions at low concentrations may evoke a long-lasting macrophage–TLR9-mediated pro-inflammatory, rather than anti-inflammatory, response in the dental pulp to modulate the repair process and hard tissue formation. Further studies are needed to determine the effects of current immunomodulatory agents in vitro and in vivo and develop treatment strategies for dental tissue regeneration. [ABSTRACT FROM AUTHOR]

Simple Summary: When joints are harmed due to injury, inheritance, or aging, it is difficult for the body to heal damaged cartilage. This is a major cause of pain and disability worldwide. With increasing life expectancy, millions of adults are affected, and enormous costs are imposed on health care systems. Scientists are attempting to use stem cells to create new cartilage-like tissue as a potential solution. However, the most suitable stem cell type for cartilage repair has not been defined. This study compared the ability to generate cartilage-like tissue by using two stem cell types that were derived from human bone marrow and dental pulp. To produce cartilage, the tissue's environment should be simulated for stem cells in a laboratory. Therefore, the low-oxygen tension of joint cartilage was simulated using a simple and inexpensive chemical, cobalt chloride. The results showed that dental pulp stem cells could produce higher quality tissue that closely resembled human joint cartilage in terms of its structure and organization that could form the primary base of normal function. While further research is necessary, these initial findings represent a promising step toward using stem cells more beneficially to enhance a patient's quality of life. The limited self-repair capacity of articular cartilage is a challenge for healing injuries. While mesenchymal stem/stromal cells (MSCs) are a promising approach for tissue regeneration, the criteria for selecting a suitable cell source remain undefined. To propose a molecular criterion, dental pulp stem cells (DPSCs) with a Hox-negative expression pattern and bone marrow mesenchymal stromal cells (BMSCs), which actively express Hox genes, were differentiated towards chondrocytes in 3D pellets, employing a two-step protocol. The MSCs' response to preconditioning by cobalt chloride (CoCl2), a hypoxia-mimicking agent, was explored in an assessment of the chondrogenic differentiation's efficiency using morphological, histochemical, immunohistochemical, and biochemical experiments. The preconditioned DPSC pellets exhibited significantly elevated levels of collagen II and glycosaminoglycans (GAGs) and reduced levels of the hypertrophic marker collagen X. No significant effect on GAGs production was observed in the preconditioned BMSC pellets, but collagen II and collagen X levels were elevated. While preconditioning did not modify the ALP specific activity in either cell type, it was notably lower in the DPSCs differentiated pellets compared to their BMSCs counterparts. These results could be interpreted as demonstrating the higher plasticity of DPSCs compared to BMSCs, suggesting the contribution of their unique molecular characteristics, including their negative Hox expression pattern, to promote a chondrogenic differentiation potential. Consequently, DPSCs could be considered compelling candidates for future cartilage cell therapy. [ABSTRACT FROM AUTHOR]

Aims: Dental pulp stem cells (DPSCs) contain a population of stem cells with a broad range of differentiation potentials, as well as more lineage‐committed progenitors. Such heterogeneity is a significant obstacle to experimental and clinical applications. The aim of this study is to isolate and characterize a homogenous neuronal progenitor cell population from human DPSCs. Methodology: Polysialylated‐neural cell adhesion molecule (PSA‐NCAM+) neural progenitors were isolated from the dental pulp of three independent donors using magnetic‐activated cell sorting (MACS) technology. Immunofluorescent staining with a panel of neural and non‐neural markers was used to characterize the magnetically isolated PSA‐NCAM+ fraction. PSA‐NCAM+ cells were then cultured in Neurobasal A supplemented with neurotrophic factors: dibutyryl cyclic‐AMP, neurotrophin‐3, B27 and N2 supplements to induce neuronal differentiation. Both PSA‐NCAM+ and differentiated PSA‐NCAM+ cells were used in Ca2+ imaging studies to assess the functionality of P2X3 receptors as well as membrane depolarization. Results: PSA‐NCAM+ neural progenitors were isolated from a heterogeneous population of hDPSCs using magnetic‐activated cell sorting and anti‐PSA‐NCAM MicroBeads. Flow cytometry analysis demonstrated that immunomagnetic sorting significantly increased the purity of PSA‐NCAM+ cells. Immunofluorescent staining revealed expression of pan‐neuronal and mature neuronal markers, PGP9.5 and MAP2, respectively, as well as weak expression of the mature sensory markers, peripherin and islet1. ATP‐induced response was mediated predominately by P2X3 receptors in both undifferentiated and differentiated cells, with a greater magnitude observed in the latter. In addition, membrane depolarizations were also detected in cells before and after differentiation when loaded with fast‐voltage‐responding fluorescent molecule, FluoVolt™ in response to potassium chloride. Interestingly, only differentiated PSA‐NCAM+ cells were capable of spontaneous membrane oscillations. Conclusions: In summary, DPSCs contain a population of neuronal progenitors with enhanced neural differentiation and functional neural‐like properties that can be effectively isolated with magnetic‐activated cell sorting (MACS). [ABSTRACT FROM AUTHOR]

Background: The European Society of Endodontology published in 2023, the S3‐level clinical practice guidelines, which supersede the Quality Guidelines for Endodontic Treatment published in 2006. Objectives: This review aims to summarize and compare the above guidelines to support their dissemination. Method: A narrative synthesis of the main differences alongside tabulation according to the main themes. Results: Three tables were prepared according to the following themes: diagnosis of pulpal and apical condition; treatment of pulpitis; and treatment of nonvital pulp and apical periodontitis. Conclusions: A compared and simplified message regarding the most recent clinical practice guidelines has been prepared. Registration: Not applicable as a narrative review. [ABSTRACT FROM AUTHOR]

Objective: To investigate the incidence of accessory canals and the variation in root canal morphology of permanent mandibular incisors and canines in Pakistani population using Cone Beam Computed Tomography (CBCT). Methods: A cross-sectional study was conducted in University College of Dentistry, The University of Lahore, Pakistan after getting institutional ethical permission from January 2020 to September 2022. The data included records of 111 patients consisting of 444 permanent mandibular incisors and 222 permanent mandibular canines. Accessory root, root canals and Vertucci canal configuration for each tooth was recorded. Data analysis was done using SPSS v20. Descriptive statistics were calculated for each anatomical parameter. Chi-square test was applied to determine association of gender with the presence of accessory roots and root canals. Results: Among the 111 records evaluated, 48.6% were males and 51.4% were females. No accessory root was found in the central and lateral incisors. However, an accessory root was found in 4.9% of the canines. The incidence of accessory canals in the central incisors, lateral incisors and canines was 18.9%, 25.2% and 10.4% respectively. The most common canal configuration in teeth with accessory canals was Type-III, followed by Type-II and Type-V. Conclusion: None of the central or lateral incisor showed accessory root while it was detected in 4.9% canines. The frequency of accessory root canal was found to be 18.9%, 25.2% and 10.4% in central incisors, lateral incisors and canines respectively. The most common canal configuration was Type-I, followed by Type-III and Type-II. Type-V, VI and VII were less common. [ABSTRACT FROM AUTHOR]

The local charge density and distribution of extracellular membranes play a crucial role in the various cellular processes, such as regulation and localization of membrane proteins, electrophysiological signal transduction, transcriptional control, cell growth, and cell death. In this study, a novel scanning ion conductance microscopy‐based method is employed to extracellular membrane mapping. This method allows to not only visualize the dynamic topography and surface charge distribution around individual cells, but also distinguish the charge difference. To validate the accuracy and effectiveness of this method, the charge density on model sample surfaces are initially manipulated and the charge sensing mechanism using finite element modeling (FEM) is explored subsequently. By applying this method, both the extracellular charge distributions and topography structures of normal and senescent human dental pulp stem cells (hDPSCs) are able to monitor. Interestingly, it is observed that the surface charge became significantly more negative after cellular senescence. This innovative approach enables us to gain valuable insights into surface charge changes during cellular senescence, which can contribute to a better understanding of the underlying mechanisms and potential therapeutic strategies for age‐related diseases. [ABSTRACT FROM AUTHOR]

Medication-related osteonecrosis of the jaw is a serious disease occurring in patients with cancer and osteoporosis, who are undergoing treatment with antiresorptive agents (ARAs) such as bisphosphonate (BP) or denosumab, an antibody targeting receptor activator of NF-κB ligand. Recently, stem cell-based therapy has been shown to be effective in preventing the development of bisphosphonate-related osteonecrosis of the jaw. However, studies on denosumab-related osteonecrosis of the jaw (DRONJ) remain limited. Here, the efficacy of treatment with dental pulp stem cell conditioned media (DPSC-CM) in preventing DRONJ in a murine model was evaluated. Local administration of DPSC-CM into the extraction socket of a mouse with DRONJ decreased the number of empty osteocyte lacunae and the prevalence of ONJ. In tissues surrounding the extraction sockets in the DPSC-CM-treated group, the expression of inflammatory cytokines was attenuated and that of osteogenesis-related molecules was enhanced compared to that in the control group. Further, the expression of Wnt signaling molecules, which had been suppressed, was improved. These findings collectively suggest that DPSC-CM prevents ONJ development in a murine DRONJ model. [ABSTRACT FROM AUTHOR]

Objective(s): This study aimed to investigate the impact of bioactive plant extracts on the proliferation and migration of dental pulp stem cells (DPSCs) and their potential implications for dental care, focusing on the nurse-caring aspect. Materials and Methods: TDPSCs were cultured on gelatin polymer scaffolds mimicking the extracellular matrix (ECM) environment. Bioactive plant extracts with antibacterial, anti-inflammatory, and anti-oxidant properties were incorporated into the gelatin polymer at concentrations ranging from 0.1% to 2.0%. Proliferation and migration assays were performed, considering nurse-caring practices during the experiments. Results: Treatment with specific bioactive plant extracts significantly enhanced DPSC proliferation, showing a 2.5-fold increase compared to the control groups. The migration assay revealed a substantial increase in cell migration distance, with treated cells covering an average distance of 400-500 µm compared to 220-260 µm in the control group. Treated cells also exhibited improved viability and metabolic activity, with a 30% increase in cell viability and a 10-20% increase in metabolic activity compared to the control group. Conclusion: This study demonstrates that bioactive plant extracts have the potential to enhance DPSC proliferation, migration, viability, and metabolic activity. These findings support the use of these extracts in dental care, benefiting from the nurse-caring practices. [ABSTRACT FROM AUTHOR]

Teeth are vulnerable to structural compromise, primarily attributed to carious lesions, in which microorganisms originating from the oral cavity deteriorate the mineralized structures of enamel and dentin, subsequently infiltrating the underlying soft connective tissue, known as the dental pulp. Nonetheless, dental pulp possesses the necessary capabilities to detect and defend against bacteria and their by-products, using a variety of intricate defense mechanisms. The pulp houses specialized cells known as odontoblasts, which encounter harmful substances produced by oral bacteria. These cells identify pathogens at an early stage and commence the immune system response. As bacteria approach the pulp, various cell types within the pulp, such as different immune cells, stem cells, fibroblasts, as well as neuronal and vascular networks, contribute a range of defense mechanisms. Therefore, the immune system is present in the healthy pulp to restrain the initial spread of pathogens, and then in the inflamed pulp, it prepares the conditions for necrosis or regeneration, so inflammatory response mechanisms play a critical role in maintaining tissue homeostasis. This review aims to consolidate the existing literature on the immune system in dental pulp, encompassing current knowledge on this topic that explains the diverse mechanisms of recognition and defense against pathogens exhibited by dental pulp cells, elucidates the mechanisms of innate and adaptive immunity in inflamed pulp, and highlights the difference between inflamed and normal pulp tissue. [ABSTRACT FROM AUTHOR]

To investigate the transdentinal effects of surface reaction-type pre-reacted glass-ionomer (S-PRG) fillers on odontoblast-like cells. An eluate of S-PRG fillers was obtained by dissolving the particles in distilled water (1:1 m/v). Dentin discs with similar permeability were mounted into artificial pulp chambers and MDPC-23 cells were seeded on their pulpal surface. The occlusal surface was treated with (n = 10): ultrapure water (negative control – NC), hydrogen peroxide (positive control – PC), S-PRG eluate exposure for 1 min (S-PRG 1 min), or S-PRG filler eluate exposure for 30 min (S-PRG 30 min). After 24 h, cell viability (alamarBlue) and morphology (SEM) were evaluated. The extract obtained from transdentinal diffusion was applied to MDPC-23 pre-cultured in plates for another 24 h to evaluate viability (alamarBlue, 1, 3, and 7 days), gene expression of Col1a1 , Alpl , Dspp , and Dmp1 (RT-qPCR, 1 and 7 days), and mineralization (Alizarin Red, 7 days). Data were analyzed with ANOVA (α = 5 %). While S-PRG 1 min did not differ from NC, S-PRG 30 min reduced 17.9 % viability of cells from discs. S-PRG treatments resulted in low cell detaching from dentin, and the remaining cells exhibited typical morphology or minor cytoplasmic contraction. S-PRG 30 min slightly increased cell viability (6 %) 1 day after contact with the extract. S-PRG treatments upregulated the expression of the investigated genes, especially after 1 day. S-PRG 30 min stimulated mineralization activity by 39.7 %. S-PRG filler eluate does not cause transdentinal cytotoxicity on odontoblast-like cells, and long-term exposure can stimulate their dentinogenic-related mineralization activity. The transdentinal elution of ions from S-PRG fillers is not expected to be harmful to the dental pulp and may exert bioactive effects by inducing dentin matrix deposition through the metabolism of underlying odontoblasts. [ABSTRACT FROM AUTHOR]

This study aimed to assess antimicrobial efficacy, cytotoxicity, and cytokine release (IL-1b, IL-6, IL-10, TNF-α) from human dental pulp stem cells (hDPSCs) of chitosan (CH) and hydroxyapatite (HAp)-modified glass ionomer cements (GIC). GICs with varied CH and HAp concentrations (0 %, 0.16 %, 2 %, 5 %, 10 %) were tested against S. mutans for 24 h or 7 days. Antimicrobial activity was measured using an MTT test. Cytotoxicity evaluation followed for optimal concentrations, analyzing mitochondrial activity and apoptosis in hDPSCs. Cytokine release was assessed with MAGPIX. Antimicrobial analysis used Shapiro-Wilk, Kruskal-Wallis, and Dunnett tests. Two-way ANOVA, Tukey, and Dunnett tests were applied for hDP metabolism and cytokine release. CH 2 % and HAp 5 % significantly enhanced GIC antimicrobial activity, especially after seven days. In immediate analysis, all materials showed reduced mitochondrial activity compared to the control. After 24 h, CH demonstrated mitochondrial metabolism similar to the control. All groups exhibited mild cytotoxicity (∼30 % cell death). Only IL-6 was influenced, with reduced release in experimental groups. CH 2 % and HAp 5 % were most effective for antibacterial effects. GIC-CH 2 % emerged as the most promising formula, displaying significant antibacterial effects with reduced hDPSC toxicity. • GICs with 2 % Chitosan and 5 % hydroxyapatite have higher antibacterial properties. • IL-1b, IL-10 and TNF-a released from hDPSCs are not modulated by modified GICs. • GIC containing 2 % Chitosan has low cytotoxicity with higher antibacterial properties. [ABSTRACT FROM AUTHOR]

Historical reports of unpredictable outcomes associated with vital pulpal therapies, particularly direct pulp capping (DPC), have contributed to clinicians' skepticism of the procedure. Contemporary reports highlight more predictable outcomes of vital pulpal therapies, inclusive of DPC. There is a dearth of reported patient-centered outcomes of these procedures. Insurance claims were used in an observational, retrospective cohort study to evaluate outcomes of DPC performed on permanent teeth. Statistical analyses included Kaplan-Meier survival estimates and Cox proportional hazards regression. Log-rank tests were used to evaluate unadjusted differences in survival. Cox proportional hazard regression was used to evaluate the adjusted hazard of adverse event occurrence. The analytic cohort included 4,136 teeth from 3,716 patients. DPC procedures were identified in public-payer (85.5%) and private-payer (13.4%) insurance claims databases. After DPC, procedure survival rate was 83% and tooth survival rate was 93% during a mean follow-up time of 52 months. Molar tooth type, same-day permanent restoration placement, and amalgam restoration type were significant positive predictors of procedure (DPC) survival. Age was not a statistically significant predictor of procedure survival after controlling for tooth type, gender, time to restoration, and restoration type. Nonmolar tooth type and younger age were significant positive predictors of tooth survival after DPC. Failures were most likely to occur within the first year. DPC has favorable patient-centered outcomes and contributes to long-term tooth survival. The favorable patient-centered outcomes of DPC bolster calls to consider cost-effectiveness and access to care for endodontic procedures. [ABSTRACT FROM AUTHOR]

Periapical lesions are common pathologies affecting the alveolar bone, often initiated by intraradicular lesions resulting from microbial exposure to dental pulp. These microorganisms trigger inflammatory and immune responses. When endodontic treatment fails to eliminate the infection, periapical lesions persist, leading to bone loss. The RANK/RANKL/OPG pathway plays a crucial role in both the formation and the destruction of the bone. In this study, the objective was to inhibit the RANK/RANKL pathway in vitro within exposed Thp-1 macrophages to endodontic microorganisms, specifically Enterococcus faecalis, which was isolated from root canals of 20 patients with endodontic secondary/persistent infection, symptomatic and asymptomatic, and utilizing an α-IRAK-4 inhibitor, we introduced endodontic microorganisms and/or lipoteichoic acid from Streptococcus spp. to cellular cultures in a culture plate, containing thp-1 cells and/or PBMC from patients with apical periodontitis. Subsequently, we assessed the percentages of RANK+, RANKL+, and OPG+ cells through flow cytometry and measured the levels of several inflammatory cytokines (IL-1β, TNF-α, IL-6, IL-8, IL-10, and IL-12p70) in the cellular culture supernatant through a CBA kit and performed analysis by flow cytometry. A significant difference was observed in the percentages of RANK+RANKL+, OPG+ RANKL+ cells in thp-1 cells and PBMCs from patients with apical periodontitis. The findings revealed significant differences in the percentages of the evaluated cells, highlighting the novel role of the IRAK-4 inhibitor in addressing this oral pathology, apical periodontitis, where bone destruction is observed. [ABSTRACT FROM AUTHOR]

Although the postmortem interval estimation still represents one of the main goals of forensic medicine, there are still several limitations that weigh on the methods most used for its determination: for this reason, even today, precisely estimating the postmortem interval remains one of the most important challenges in the forensic pathology field. To try to overcome these limitations, in recent years, numerous studies have been conducted on the potential use of the mRNA degradation time for reaching a more precise post mortem interval (PMI) estimation. An evidence-based systematic review of the literature has been conducted to evaluate the state of the art of the knowledge focusing on the potential correlation between mRNA degradation and PMI estimation. The research has been performed using the electronic databases PubMed and Scopus. The analysis conducted made it possible to confirm the potential applicability of mRNA for reaching a more precise PMI estimation. The analysis of the results highlighted the usefulness of some mRNAs, such as β-actin and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) mRNA, especially in short time frames, within a few hours or days of death. The matrices on which these analyses were conducted were also analyzed, resulting in less exposure to the external environment, including the heart, brain, and dental pulp. The major limitations were also reported, including the short time intervals analyzed in most of the articles, the lack of mathematical models, and the failure to report the error rate between the mRNA degradation time and PMI. Given the still small number of published articles, the lack of globally recognized standardized methods, and the numerous techniques used to evaluate the mRNA degradation times, numerous and larger studies are still necessary to reach more solid and shared evidence. [ABSTRACT FROM AUTHOR]

Mesenchymal stem-cell-derived extracellular vesicles (MSC-EVs) have been increasingly investigated for cancer therapy and drug delivery, and they offer an advanced cell-free therapeutic option. However, their overall effects and efficacy depend on various factors, including the MSC source and cargo content. In this study, we isolated EVs from the conditioned medium of human immature dental pulp stem cells (hIDPSC-EVs) and investigated their effects on two papillary thyroid cancer (PTC) cell lines (BCPAP and TPC1). We observed efficient uptake of hIDPSC-EVs by both PTC cell lines, with a notable impact on gene regulation, particularly in the Wnt signaling pathway in BCPAP cells. However, no significant effects on cell proliferation were observed. Conversely, hIDPSC-EVs significantly reduced the invasive capacity of both PTC cell lines after 120 h of treatment. These in vitro findings suggest the therapeutic potential of hIDPSC-EVs in cancer management and emphasize the need for further research to develop novel and effective treatment strategies. Furthermore, the successful internalization of hIDPSC-EVs by PTC cell lines underscores their potential use as nanocarriers for anti-cancer agents. [ABSTRACT FROM AUTHOR]

This review article describes the methods and clinical recommendations for reinforcing traumatized anterior immature teeth with pulp necrosis treated with mineral trioxide aggregate (MTA) apexification. Traumatic injury can cause pulp necrosis and incomplete root formation in immature teeth. MTA apexification is the treatment of choice for necrotic immature teeth, particularly during the middle or late stages of root development. MTA apexification has a high success rate; however, failures due to cervical or root fractures occasionally occur. The risk of fracture is higher in immature teeth with thin root dentin, particularly those with external root resorption. Furthermore, the loading force from any parafunctional habit also increases fracture risk. Therefore, intra‐radicular reinforcement may be necessary after MTA apexification. In vitro, intraradicular restoration with a resin composite/core build‐up material or a prefabricated fiber post demonstrated better root reinforcement than root canal obturation materials (i.e., gutta‐percha and sealer). However, the root‐reinforcement effect of MTA orthograde filling in the entire root canal remains unclear. In vivo, the survival of fractured teeth with intraradicular restorations (resin composite/core build‐up material or prefabricated fiber posts) is extremely high. Moreover, the survival of teeth with gutta‐percha/sealer obturation or MTA orthograde filling and restoration with resin composite extending into the cervical third of the root canal approximately 1–2 mm below the cemento‐enamel junction is acceptably high. Based on this evidence, the remaining tooth/root structure and loading force should be carefully examined when considering intra‐radicular reinforcement of immature anterior teeth treated with MTA apexification. [ABSTRACT FROM AUTHOR]

Background: Symptomatic irreversible pulpitis often results in heightened reactions to thermal stimuli such as pain evoked by a cold stimulus, and spontaneous odontogenic pain (unprovoked pain). Objective: This study primarily compared the clinical manifestations of odontogenic spontaneous pain and pain provoked by cold stimulus specifically focusing on their sensory discriminative characteristics (intensity, duration and quality) between symptomatic irreversible pulpitis patients with and without referred pain. Methods: Twenty‐three patients with symptomatic irreversible pulpitis with referred pain and 12 patients without referred pain were included in this cross‐sectional study. The following outcomes were assessed: odontogenic spontaneous pain and its descriptors; pain evoked by cold stimulus and qualitative sensory testing before and after local anaesthesia; referred pain location; use of analgesic medication; complementary anaesthesia efficacy. T‐test, chi‐squared and McNemar tests were applied to the data (p <.50). Results: Patients with referred pain presented a greater odontogenic pain intensity (p <.05) when considered the average of the last 24 h. These patients also showed higher pain rating and pain descriptors (p <.05). Intensity and duration of the pain evoked by cold stimulus in the non‐affected contralateral tooth at baseline were higher for patients with referred pain (p <.05). Conclusion: Symptomatic irreversible pulpitis patients with referred pain present greater odontogenic spontaneous pain and a heightened pain sensitivity. Therefore, patients with referred pain seem more complex from a pain severity perspective, supporting the clinical utility of discriminating symptomatic irreversible pulpitis with and without referred pain. [ABSTRACT FROM AUTHOR]

Heparan sulfate (HS) is a major component of dental pulp tissue. We previously reported that inhibiting HS biosynthesis impedes endothelial differentiation of dental pulp stem cells (DPSCs). However, the underlying mechanisms by which exogenous HS induces DPSC differentiation and pulp tissue regeneration remain unknown. This study explores the impact of exogenous HS on vasculogenesis and dentinogenesis of DPSCs both in vitro and in vivo. Human-derived DPSCs were cultured in endothelial and odontogenic differentiation media and treated with HS. Endothelial differentiation of DPSCs was investigated by real-time polymerase chain reaction and capillary sprouting assay. Odontogenic differentiation was assessed through real-time polymerase chain reaction and detection of mineralized dentin-like deposition. Additionally, the influence of HS on pulp tissue was assessed with a direct pulp capping model, in which HS was delivered to exposed pulp tissue in rats. Gelatin sponges were loaded with either phosphate-buffered saline or 101–102 μg/mL HS and placed onto the pulp tissue. Following a 28-day period, tissues were investigated by histological analysis and micro–computed tomography imaging. HS treatment markedly increased expression levels of key endothelial and odontogenic genes, enhanced the formation of capillary-like structures, and promoted the deposition of mineralized matrices. Treatment of exposed pulp tissue with HS in the in vivo pulp capping study induced formation of capillaries and reparative dentin. Exogenous HS effectively promoted vasculogenesis and dentinogenesis of DPSCs in vitro and induced reparative dentin formation in vivo , highlighting its therapeutic potential for pulp capping treatment. [ABSTRACT FROM AUTHOR]

Estimating the post-mortem interval (PMI) of human remains based on the histomorphology of dental pulp parameters is promising, but available evidence is scarce and sometimes contradictory without a scientific model. The aim of the study is to characterise the histomorphological changes of dental pulp associated with the decomposition of human remains by a qualitative and quantitative approach. The main aim is to establish a correlation based on post-mortem (PM) dental pulp histomorphology and the PMI, and whether pulp degradation could be an available medico-legal tool for PMI estimation beyond the first week after death (late PMI). The eligible sample consisted of 27 sound teeth from 16 healthy patients aged 16 to 72 years due to orthodontic or oral surgery treatment, to create PMI’s simulating the death of the subject as the time elapsed from tooth avulsion. Data collected from patients (sex, date of birth, tooth position, date and hour of the avulsion, date and hour of pulp extraction) were anonymised in accordance with the requirements of Faculty of Dental Medicine of the University of Lisbon. The sample was divided into 9 groups of 3 teeth according to different PMI sets from T0 (baseline) up to 2 weeks (T0, 7, 12, 24, 36, 48, and 72 hours, 1 and 2 weeks). All the dental samples were stored at room temperature up to the time of pulp extraction and then prepared with haematoxylin and eosin stain. High-resolution microscopy was performed to obtain histological images. An operator performed the qualitative evaluation of blood vessels, collagen fibres, and the extra-cellular matrix (ECM) in PM pulps and measured the variation in cells/ nuclei density by counting 6 different ROIs (Regions of Interest) for each pulp manually and automatically (quantitative analysis). Qualitative results showed that the degeneration of dental pulp appears 7 hours after death but histological changes in vessels, fibres, and ECM in PM dental pulp are characterised by high variability, consequently it is not possible to generalise the results for early PMIs. Quantitative measurements proved that cell count cannot be standardised due to the presence of superimposed layers of cells and nuclei fragmentation. Odontoblasts did not demonstrate evidence of cellular or nuclear lysis up to 14 PM suggesting their applicability in late PMIs. Future research will focus on late PMIs and different techniques of tooth preparation. [ABSTRACT FROM AUTHOR]

Introduction: There is a paucity of studies evaluating the preservation and recovery vitality of teeth associated with cystic lesions in the jaws. The purpose of the present study was to evaluate whether the use of the marsupialization treatment method in the large cystic lesions of the jaws associated within involved teeth would save the pulp vitality. Materials and methods: It was a cohort study of patients treated at the Department of Oral and Maxillofacial Surgery from April 2019 to April 2023. Patients with primary cystic lesions associated teeth, with a cyst diameter of >3.4 cm, along with dental root resorption of less than half the root length, and treated by marsupialization were included. Patient age, gender, location, associated teeth, size, management, dental pulp vitality before and after marsupialization, and the duration of treatment were evaluated. Pulp vitality of involved teeth was assessed with electrical pulp tester. Predictor Variable: It was the treatment method. Main Outcome Variable(s): The primary outcome variable was the recovery or preservation of pulp vitality. Covariates: The study variables were demographic, clinical features of the cysts, clinical features of associated teeth, and treatment method. Analyses: Categorical data were presented in mean with standard deviation, and set the level of statistical significance at p < 0.05. Results: The sample included 73 patients with large cystic lesions; 47 (64.4%) were male and 26 (35.6%) were female. The age range was 9 to 68 years (mean ± standard deviation, 22.4 ± 4). The 73 patients had 305 teeth associated with large cystic lesions. Of these dentitions, 85.9% of teeth had normal pulp vitality and 14.1% were negative before marsupialization. Following marsupialization, preservation of pulp vitality was noted in 61.3% and 18.0% of teeth lost their vitality. Of those without normal dental pulp vitality, 70.9% of teeth recovered their vitality of which 46.5% had recovered their vitality 9 to 12 months following marsupialization. Conclusion: Marsupialization followed by second surgery maybe a treatment option for preserving and recovering the pulp vitality of teeth associated with large cystic jaw lesions. [ABSTRACT FROM AUTHOR]

(1) Background: The aim of this study was to compare the cytotoxicity of selected resin-modified materials used in direct contact with the dental pulp (TheraCal LC, TheraCal PT, and ApaCal ART) with calcium silicate cement (Biodentine). (2) Methods: The mouse fibroblast Balb/3T3 cell line and the extracts of tested materials in four concentrations were used for the testing. An MTT assay was performed in three independent experiments with six replicates for each concentration of tested material. The cell viability (%) and cytotoxicity were expressed (cytotoxic effect is considered in cases where the cell viability is lower than 70%). The mean of the cell viability and the standard deviation were expressed for each material at all concentrations. ANOVA and Dunnet's post hoc tests were used for the statistical analysis. All of these tests were performed at the 0.05 significance level. (3) Results: At all concentrations, the cell viability was statistically significantly lower (p ≤ 0.002) for all tested materials compared to Biodentine. ApaCal ART showed a high level of cytotoxicity at all concentrations (cell viability lower than 47.71%, p < 0.0001). The same result was found for TheraCal LC at concentrations of 100%, 50% and 25% and TheraCal PT at concentrations of 100% and 50%. TheraCal LC at a 10% concentration (cell viability 68.18%) and TheraCal PT at a 25% concentration (cell viability 60.63%) indicated potential cytotoxicity. TheraCal PT at a 10% concentration was not found to be cytotoxic (cell viability 79.18%, p = 0.095). (4) Conclusion: The resin-modified calcium silicate and calcium phosphate materials showed higher cytotoxic potential, so they should be used with caution when in direct contact with the dental pulp. [ABSTRACT FROM AUTHOR]

Dens invaginatus (DI) is a developmental anomaly of the teeth characterised by the in‐folding of the enamel into the dentin. Oehlers' Type III DI is the most serious form, in which the inherently invaginated channels communicate with periodontal and dental pulp tissue, increasing the risk of bacterial contamination. However, varying and complex anatomical features make diagnosis and treatment challenging. Conventional endodontic therapies promote healing by avoiding unnecessary interventions (e.g., surgical or other invasive treatments). Radiographic examination can reveal the structural details of such malformations. We obtained multiple procedural details for treating Type III DI based on radiographic analyses from our clinical experience. In addition, we introduce a new classification strategy for the management of Type III DI that is more applicable to treatment needs. This study aimed to discuss the anatomical features and current treatment considerations of Type III DI. [ABSTRACT FROM AUTHOR]

This study aimed to assess the association between genetic polymorphisms in BMP2 (rs1005464 and rs235768), BMP4 (rs17563), SMAD6 (rs2119261 and rs3934908) and RUNX2 (rs59983488 and rs1200425) and pulp stones (PS). A total of 117 participants, consisting of 63 individuals with PS and 54 without PS, were included. Digital radiographs and a demographic/clinical questionnaire were used. Genomic DNA from salivary cells was genotyped via real‐time polymerase chain reaction. Statistical analyses, including Chi‐Square, Fisher's exact tests, Poisson regression and dimensionality reduction, were conducted. The rs2119261 polymorphism in the SMAD6 gene showed an association with genotype distribution in the recessive model (p = 0.049). The T–T haplotype in the SMAD6 gene (rs2119261 and rs3934908) was more prevalent in the control group and significantly linked with PS (p = 0.029). No associations were found between PS risk and genetic polymorphisms in BMP2, BMP4 and RUNX2. Polymorphisms in the SMAD6 gene were associated with PS. [ABSTRACT FROM AUTHOR]

Background: Insulin has been known to regulate bone metabolism, yet its specific molecular mechanisms during the proliferation and osteogenic differentiation of dental pulp stem cells (DPSCs) remain poorly understood. This study aimed to explore the effects of insulin on the bone formation capability of human DPSCs and to elucidate the underlying mechanisms. Methods: Cell proliferation was assessed using a CCK-8 assay. Cell phenotype was analyzed by flow cytometry. Colony-forming unit-fibroblast ability and multilineage differentiation potential were evaluated using Toluidine blue, Oil red O, Alizarin red, and Alcian blue staining. Gene and protein expressions were quantified by real-time quantitative polymerase chain reaction and Western blotting, respectively. Bone metabolism and biochemical markers were analyzed using electrochemical luminescence and chemical colorimetry. Cell adhesion and growth on nano-hydroxyapatite/collagen (nHAC) were observed with a scanning electron microscope. Bone regeneration was assessed using micro-CT, fluorescent labeling, immunohistochemical and hematoxylin and eosin staining. Results: Insulin enhanced the proliferation of human DPSCs as well as promoted mineralized matrix formation in a concentration-dependent manner. 10− 6 M insulin significantly up-regulated osteogenic differentiation-related genes and proteins markedly increased the secretion of bone metabolism and biochemical markers, and obviously stimulated mineralized matrix formation. However, it also significantly inhibited the expression of genes and proteins of receptors and receptor substrates associated with insulin/insulin-like growth factor-1 signaling (IIS) pathway, obviously reduced the expression of the phosphorylated PI3K and the ratios of the phosphorylated PI3K/total PI3K, and notably increased the expression of the total PI3K, phosphorylated AKT, total AKT and mTOR. The inhibitor LY294002 attenuated the responsiveness of 10− 6 M insulin to IIS/PI3K/AKT/mTOR pathway axis, suppressing the promoting effect of insulin on cell proliferation, osteogenic differentiation and bone formation. Implantation of 10− 6 M insulin treated DPSCs into the backs of severe combined immunodeficient mice and the rabbit jawbone defects resulted in enhanced bone formation. Conclusions: Insulin induces insulin resistance in human DPSCs and effectively promotes their proliferation, osteogenic differentiation and bone formation capability through gradually inducing the down-regulation of IIS/PI3K/AKT/mTOR pathway axis under insulin resistant states. [ABSTRACT FROM AUTHOR]