Dental pulp-dentin complex defects remain a major unresolved problem in oral medicines. Clinical therapeutic methods including root canal therapy and vital pulp therapy are both considered as conservative strategies, which are incapable of repairing the pulp-dentin complex defects. Although biomaterial-based strategies show remarkable progress in antibacterial, anti-inflammatory, and pulp regeneration, the important modulatory effects of nerves within pulp cavity have been greatly overlooked, making it challenging to achieve functional pulp-dentin complex regeneration. In this study, we propose an injectable bioceramics-containing composite hydrogel in combination of Li-Ca-Si (LCS) bioceramics and gelatin methacrylate matrix with photo-crosslinking properties. Due to the sustained release of bioactive Li, Ca and Si ions from LCS, the composite hydrogels possess multiple functions of promoting the neurogenic differentiation of Schwann cells, odontogenic differentiation of dental pulp stem cells, and neurogenesis-odontogenesis couples in vitro. In addition, the in vivo results showed that LCS-containing composite hydrogel can significantly promote the pulp-dentin complex repair. More importantly, LCS bioceramics-containing composite hydrogel can induce the growth of nerve fibers, leading to the re-innervation of pulp tissues. Taken together, the study suggests that LCS bioceramics can induce the innervation of pulp-dentin complex repair, offering a referable strategy of designing multifunctional filling materials for functional periodontal tissue regeneration.
Dental Pulp/drug effects* ; Hydrogels/pharmacology* ; Animals ; Ceramics/pharmacology* ; Dentin/drug effects* ; Biocompatible Materials/pharmacology* ; Rats ; Gelatin ; Regeneration/drug effects* ; Cell Differentiation/drug effects* ; Injections ; Humans ; Odontogenesis/drug effects*

Tooth pulpitis is a prevalent oral disorder. Understanding the underlying mechanisms of pulpitis and developing effective treatment strategies hold great significance. Ferroptosis has recently emerged as a new form of cell death, but the role of ferroptosis in pulpitis remains largely unknown. In our study, single-cell RNA sequencing (scRNA-seq) was used to identify cellular heterogeneity between 3 pulpitis tissue and 3 healthy pulp tissue, and explored ferroptosis occurrence in pulpitis tissue and inflamed dental pulp cells (DPCs). In scRNA-seq, 40 231 cells (Pulpitis: 17 814; Healthy pulp: 22 417) were captured, and visualized into 12 distinct cell clusters. Differentially expressed ferroptosis-related genes (DE-FRGs) were almost presented in each cluster in pulpitis vs healthy pulp. ROS and Fe²⁺ levels significantly rose, and immunohistochemistry showed low expression of GPX4 and high expression of PTGS2 in pulpitis. In LPS-stimulated DPCs, thymosin α1 increased the expression of GPX4 and FTL, and decreased expression of TNF-α, IL-1β, IL-6, and Fe²⁺ levels. In rat pulpitis models, both prothymosin α (PTMA, precursor of thymosin α1) gelatin sponge placed at the hole of pulp (LPS-P(gs)) and PTMA injection in pulp (LPS-P(i)) significantly reduced infiltration of inflammatory cells and expression of PTGS2, and increased the expression of GPX4. In RNA sequencing, the expression of DE-FRGs were reversed when thymosin α1 were added in LPS-stimulated DPCs. Collectively, single-cell atlas reveals cellular heterogeneity between pulpitis and healthy pulp, and ferroptosis occurrence in pulpitis. Thymosin α1 may reduce ferroptosis in DPCs to alleviate pulpitis and thus potentially has the ability to treat pulpitis.
Ferroptosis/drug effects* ; Dental Pulp/drug effects* ; Animals ; Pulpitis/pathology* ; Rats ; Thymalfasin/pharmacology* ; Humans ; Male ; Thymosin/pharmacology* ; Disease Models, Animal ; Rats, Sprague-Dawley

We investigated the potential of human dental pulp stem cells (hDPSCs) to differentiate into dopaminergic neurons in vitro as an autologous stem cell source for Parkinson's disease treatment. The hDPSCs were expanded in knockout-embryonic stem cell (KO-ES) medium containing leukemia inhibitory factor (LIF) on gelatin-coated plates for 3-4 days. Then, the medium was replaced with KO-ES medium without LIF to allow the formation of the neurosphere for 4 days. The neurosphere was transferred into ITS medium, containing ITS (human insulin-transferrin-sodium) and fibronectin, to select for Nestin-positive cells for 6-8 days. The cells were then cultured in N-2 medium containing basic fibroblast growth factor (FGF), FGF-8b, sonic hedgehog-N, and ascorbic acid on poly-l-ornithine/fibronectin-coated plates to expand the Nestin-positive cells for up to 2 weeks. Finally, the cells were transferred into N-2/ascorbic acid medium to allow for their differentiation into dopaminergic neurons for 10-15 days. The differentiation stages were confirmed by morphological, immunocytochemical, flow cytometric, real-time PCR, and ELISA analyses. The expressions of mesenchymal stem cell markers were observed at the early stages. The expressions of early neuronal markers were maintained throughout the differentiation stages. The mature neural markers showed increased expression from stage 3 onwards. The percentage of cells positive for tyrosine hydroxylase was 14.49%, and the amount was 0.526 ± 0.033 ng/mL at the last stage. hDPSCs can differentiate into dopaminergic neural cells under experimental cell differentiation conditions, showing potential as an autologous cell source for the treatment of Parkinson's disease.
Animals ; Brain/pathology ; *Cell Differentiation/drug effects ; Cells, Cultured ; Culture Media/chemistry/pharmacology ; Dental Pulp/*cytology ; Dopaminergic Neurons/*cytology/*metabolism/pathology ; Enzyme-Linked Immunosorbent Assay ; Glial Fibrillary Acidic Protein/genetics/metabolism ; Humans ; Mice ; Mice, Inbred ICR ; Myelin Basic Protein/genetics/metabolism ; Real-Time Polymerase Chain Reaction ; Stage-Specific Embryonic Antigens/genetics/metabolism ; Stem Cells/*cytology/*metabolism/pathology ; Tubulin/genetics/metabolism ; Tyrosine 3-Monooxygenase/analysis/genetics/metabolism

We investigated the potential of human dental pulp stem cells (hDPSCs) to differentiate into dopaminergic neurons in vitro as an autologous stem cell source for Parkinson's disease treatment. The hDPSCs were expanded in knockout-embryonic stem cell (KO-ES) medium containing leukemia inhibitory factor (LIF) on gelatin-coated plates for 3-4 days. Then, the medium was replaced with KO-ES medium without LIF to allow the formation of the neurosphere for 4 days. The neurosphere was transferred into ITS medium, containing ITS (human insulin-transferrin-sodium) and fibronectin, to select for Nestin-positive cells for 6-8 days. The cells were then cultured in N-2 medium containing basic fibroblast growth factor (FGF), FGF-8b, sonic hedgehog-N, and ascorbic acid on poly-l-ornithine/fibronectin-coated plates to expand the Nestin-positive cells for up to 2 weeks. Finally, the cells were transferred into N-2/ascorbic acid medium to allow for their differentiation into dopaminergic neurons for 10-15 days. The differentiation stages were confirmed by morphological, immunocytochemical, flow cytometric, real-time PCR, and ELISA analyses. The expressions of mesenchymal stem cell markers were observed at the early stages. The expressions of early neuronal markers were maintained throughout the differentiation stages. The mature neural markers showed increased expression from stage 3 onwards. The percentage of cells positive for tyrosine hydroxylase was 14.49%, and the amount was 0.526 ± 0.033 ng/mL at the last stage. hDPSCs can differentiate into dopaminergic neural cells under experimental cell differentiation conditions, showing potential as an autologous cell source for the treatment of Parkinson's disease.
Animals ; Brain/pathology ; *Cell Differentiation/drug effects ; Cells, Cultured ; Culture Media/chemistry/pharmacology ; Dental Pulp/*cytology ; Dopaminergic Neurons/*cytology/*metabolism/pathology ; Enzyme-Linked Immunosorbent Assay ; Glial Fibrillary Acidic Protein/genetics/metabolism ; Humans ; Mice ; Mice, Inbred ICR ; Myelin Basic Protein/genetics/metabolism ; Real-Time Polymerase Chain Reaction ; Stage-Specific Embryonic Antigens/genetics/metabolism ; Stem Cells/*cytology/*metabolism/pathology ; Tubulin/genetics/metabolism ; Tyrosine 3-Monooxygenase/analysis/genetics/metabolism

This study aimed to present a new method based on numeric calculus to provide data on the theoretical volume ratio of voids when using the cold lateral compaction technique in canals with various diameters and tapers. Twenty-one simulated mathematical root canal models were created with different tapers and sizes of apical diameter, and were filled with defined sizes of standardized accessory gutta-percha cones. The areas of each master and accessory gutta-percha cone as well as the depth of their insertion into the canals were determined mathematically in Microsoft Excel. When the first accessory gutta-percha cone had been positioned, the residual area of void was measured. The areas of the residual voids were then measured repeatedly upon insertion of additional accessary cones until no more could be inserted in the canal. The volume ratio of voids was calculated through measurement of the volume of the root canal and mass of gutta-percha cones. The theoretical volume ratio of voids was influenced by the taper of canal, the size of apical preparation and the size of accessory gutta-percha cones. Greater apical preparation size and larger taper together with the use of smaller accessory cones reduced the volume ratio of voids in the apical third. The mathematical model provided a precise method to determine the theoretical volume ratio of voids in root-filled canals when using cold lateral compaction.
Dental Pulp Cavity ; drug effects ; Epoxy Resins ; therapeutic use ; Gutta-Percha ; therapeutic use ; Humans ; Models, Theoretical ; Root Canal Filling Materials ; therapeutic use ; Root Canal Preparation ; methods ; Surface Properties ; Titanium ; therapeutic use

OBJECTIVETo investigate the effect of epigallocatechin-3-gallate (EGCG) on biomodification of demineralized dentine substrate, in its permeability, hydrophobicity, and inhibition ability to collagen enzymatic degradation.

METHODSThe dentine substrates were treated with simulated pulpal pressure created by mixtures of 0.02%, 0.1% EGCG/bovine serum albumin (BSA) in acidic environment (pH4.4) for 48 h. A fluid-transport model was used to measure the fluid permeability through demineralized dentine substrate. Positive replicas of dentine substrate were fabricated before and after being subjected to acidic environment for scanning electron microscope (SEM) examination. The blank group contained no EGCG and the positive group were treated with Gluma desensitizer. Static contact angle measurements on demineralized dentin and 0.1% EGCG primed dentin were performed by contact angle analyzer. The priming time were 60 s, 120 s, 0.5 h, 1 h. Dentine specimens bonded with Adper single bond 2 were subjected to 100 mg/L collagenase and observed under SEM. Resin-bonded specimens (with 0.02%, 0.1%, 0.5% EGCG priming, or without EGCG priming) were created for micro-tensile bond strength evaluation (MTBS). Resin-bonded specimens after thermol cycling were created for MTBS evaluation.

RESULTSThe fluid permeability in the blank control group increased ([151.3±22.3]%), the fluid permeability in 0.1% EGCG/BSA group decreased ([23.7±6.3]%). Compared to the blank control group, the contact angle of 120 s, 0.5 h, 1 h groups increased by 31.0%, 53.5%, 57.8% in deep dentin and 37.4%, 59.3%, 62.4% in shallow dentin. The SEM examination showed that 0.1% and 0.5% EGCG priming for 120 s significantly increased dentin collagen's resistance to collagenase. The immediate MTBS of 0.1% and 0.5% EGCG groups were (29.4±4.8) and (19.8± 4.9) MPa. After thermol cycling, the MTBS of 0.1% and 0.5% EGCG groups were (19.9±5.1) and (15.3± 6.3) MPa.

CONCLUSIONSUnder acidic environment (pH4.4), the 0.1% EGCG can reduce dentine permeability under acidic environment. The 0.1% EGCG can increase hydrophobicity of dentin substrate, and strengthen dentin substrate's resistance to collagenase hydrolysis, thus increased the resin-dentin bonding durability.

Acid Etching, Dental ; Catechin ; analogs & derivatives ; pharmacology ; Collagen ; chemistry ; drug effects ; Collagenases ; pharmacology ; Composite Resins ; Dental Bonding ; Dental Cements ; Dental Pulp ; Dentin ; chemistry ; drug effects ; Dentin Permeability ; drug effects ; Dentin-Bonding Agents ; Glutaral ; pharmacology ; Hydrogen-Ion Concentration ; Hydrolysis ; Methacrylates ; pharmacology ; Microscopy, Electron, Scanning ; Pressure ; Resin Cements ; Serum Albumin, Bovine ; pharmacology ; Tensile Strength ; Time Factors

This study compared the biological changes of lipopolysaccharide (LPS)-treated dental pulp (DP) cells directly cultured on mineral trioxide aggregate (MTA) and calcium silicate (CS) cements. DP cells were treated with LPS for 24 h. Then, the LPS-treated DP cells were cultured on MTA or CS cements. Cell viability, cell death mechanism and interleukin (IL)-1β expressions were analysed. A one-way analysis of variance was used to evaluate the significance of the differences between the means. A significantly higher IL-1β expression (2.9-fold) was found for LPS-treated cells (P<0.05) compared with DP cells without LPS treatment at 24 h. Absorbance values of LPS-treated cells cultured on CS cement were higher than a tissue culture plate. A significant difference (P<0.05) in cell viability was observed between cells on CS and MTA cements 24 h after seeding. At 48 h, a high concentration of Si (5 mM) was released from MTA, which induced LPS-treated DP cell apoptosis. The present study demonstrates that CS cement is biocompatible with cultured LPS-treated DP cells. MTA stimulates inflammation in LPS-treated DP cells, which leads to greater IL-1β expression and apoptosis.
Calcium Compounds ; Dental Cements ; Dental Pulp ; drug effects ; metabolism ; Humans ; Inflammation ; chemically induced ; metabolism ; Interleukin-1beta ; metabolism ; Lipopolysaccharides ; pharmacology ; Silicates

Effective final irrigation regimen is an important step in order to achieve better disinfection and ensure residual antimicrobial effects after root canal preparation. The aim of this study was to compare the residual antimicrobial activity of 0.2% cetrimide, and 0.2% and 2% chlorhexidine in root canals infected with Enterococcus faecalis. Biofilms of E. faecalis were grown on uniradicular roots for 4 weeks. After root canal preparation, root canals were irrigated with 17% ethylenediaminetetraacetic acid (EDTA) to remove the smear layer. The roots were randomly divided into three experimental groups (n=26) according to the final irrigating solution: Group I, 5 mL 0.2% cetrimide; Group II, 5 mL 0.2% chlorhexidine; and Group III, 5 mL 2% chlorhexidine. Samples were collected for 50 days to denote the presence of bacterial growth. The proportion of ungrown specimens over 50 days was evaluated using the nonparametric Kaplan-Meier survival analysis. Differences among groups were tested using the log-rank test and the level of statistical significance was set at P<0.05. The highest survival value was found with 2% chlorhexidine, showing statistically significant differences from the other two groups. At 50 days, E. faecalis growth was detected in 69.23% specimens in Groups I and II, and in 34.61% specimens of Group III. There were no significant differences between 0.2% cetrimide and 0.2% chlorhexidine. Final irrigation with 2% chlorhexidine showed greater residual activity than 0.2% chlorhexidine and 0.2% cetrimide in root canals infected with E. faecalis.
Anti-Infective Agents, Local ; administration & dosage ; therapeutic use ; Bacterial Load ; drug effects ; Biofilms ; drug effects ; Cetrimonium Compounds ; therapeutic use ; Chlorhexidine ; administration & dosage ; therapeutic use ; Dental Pulp Cavity ; microbiology ; Edetic Acid ; therapeutic use ; Enterococcus faecalis ; drug effects ; Gram-Positive Bacterial Infections ; drug therapy ; Humans ; Microscopy, Electron, Scanning ; Root Canal Irrigants ; administration & dosage ; therapeutic use ; Root Canal Preparation ; methods ; Smear Layer ; Time Factors

All-trans retinoic acid (ATRA) inhibits matrix metalloproteinase (MMP)-2 and MMP-9 in synovial fibroblasts, skin fibroblasts, bronchoalveolar lavage cells and cancer cells, but activates MMP-9 in neuroblast and leukemia cells. Very little is known regarding whether ATRA can activate or inhibit MMPs in human dental pulp cells (HDPCs). The purpose of this study was to determine the effects of ATRA on the production and secretion of MMP-2 and -9 in HDPCs. The productions and messenger RNA (mRNA) expressions of MMP-2 and -9 were accessed by gelatin zymography and real-time polymerase chain reaction (PCR), respectively. ATRA was found to decrease MMP-2 level in a dose-dependent manner. Significant reduction in MMP-2 mRNA expression was also observed in HDPCs treated with 25 µmol⋅L(-1) ATRA. However, HDPCs treated with ATRA had no effect on the pattern of MMP-9 produced or secreted in either cell extracts or conditioned medium fractions. Taken together, ATRA had an inhibitory effect on MMP-2 expression in HDPCs, which suggests that ATRA could be a candidate as a medicament which could control the inflammation of pulp tissue in vital pulp therapy and regenerative endodontics.
Cell Culture Techniques ; Cell Survival ; drug effects ; Cells, Cultured ; Culture Media, Conditioned ; Dental Pulp ; cytology ; drug effects ; enzymology ; Dose-Response Relationship, Drug ; Humans ; Matrix Metalloproteinase 2 ; drug effects ; genetics ; Matrix Metalloproteinase 9 ; drug effects ; genetics ; RNA, Messenger ; drug effects ; Real-Time Polymerase Chain Reaction ; Transcription, Genetic ; drug effects ; Tretinoin ; administration & dosage ; pharmacology

OBJECTIVETo observe the effect of revascularization for treatment of immature teeth with endodontic infection mediated by calcium hydroxide.

METHODSSeventeen pediatric patients with endodontic infections of the permanent teeth were treated with routine root canal and pulp cavity irrigation and disinfection followed by application of calcium hydroxide paste to the root canal orifice to induce revascularization. Another 17 patients received conventional apexification procedures to serve as the control group. The patients were followed up to observe the therapeutic effect of the treatments.

RESULTSIn the revascularization treatment group, 4 cases showed healed periapical lesions 6 to 18 months after the surgery with thickened root canal walls and closure of the apical foramen; in 10 cases, the periapical lesions healed 12 to 18 months postoperatively with lengthened root, thickened root canal wall, and narrowed apical foramen. One patient reported pain and swelling at 2 months, and 2 patients showed the formation of gum fistula and ceased development of the roots at 7 and 8 months. In the control group, the periapical lesions healed in 1 cases at 12 months postoperatively with apical foramen closure; in 11 cases, hard tissues formed in the root apex without obviously lengthened roots 6 to 8 months after the surgery; in 5 cases, no apical barrier formed even 12 to 18 months after the surgery. The overall effective rates were similar between the two groups (P>0.05).

CONCLUSIONSRevascularization by calcium hydroxide sealing can promote root development of immature permanent teeth with pulpitis or periradicular periodontitis.

Adolescent ; Calcium Hydroxide ; therapeutic use ; Child ; Dental Pulp ; blood supply ; Dental Pulp Diseases ; therapy ; Dentition, Permanent ; Guided Tissue Regeneration, Periodontal ; Humans ; Neovascularization, Physiologic ; drug effects ; Treatment Outcome