Cryopreservation of human teeth using vitrification method with cryoprotectant cocktails and N-acetylcysteine for banking and clinical applications.

Preserving freshly-extracted healthy human teeth offers an optional resource for potential tooth transplantation and cell therapy. This study aimed to assess the impact of vitrification, utilizing a blend of cryoprotectant agents and N-acetylcysteine (NAC), on the cryopreservation of periodontal ligament tissues, and investigate the underlying mechanisms of NAC on the tooth cryopreservation. Periodontal ligament cells were isolated from freshly-extracted healthy human permanent teeth, and cell sheets of PDLCs were fabricated. The samples including cell sheets, freshly-extracted human and rat teeth were cryopreserved with or without NAC for three months. The viability, ROS level, gene expressions and microstructure of PDLCs within cell sheets were assessed. The expression of SOD-2, Caspase3, LC3A/B and Catalase were evaluated through western blotting. Histological assessments of cryopreserved cell sheets and teeth were conducted. PDLCs were isolated from cryopreserved teeth, and their immunophenotype and differentiation ability were evaluated. The data was analyzed using one-way analysis of variance. The vitrification method showed good performance in preserving the viability and differentiation potential of PDLCs. Cryopreservation supplemented with NAC improved the survival rate of PDLCs, enhanced osteogenic differentiation ability, upregulated the expression of SOD-2 and Catalase, and inhibited cell apoptosis. Additionally, mRNA sequencing analysis revealed a significant activation of the PI3K-AKT pathway following cryopreservation via vitrification. Adding a PI3K-AKT activator improved the survival rates of PDLCs post-cryopreservation. The vitrification strategy combining various CPAs and NAC proved to be feasible for tooth cryopreservation. Targeting the PI3K-AKT pathway may improve the efficacy of tooth cryopreservation.

Cryopreservation of human dental roots using vitrification for autologous human tooth tissue banking.

This study aims to preliminarily evaluate the feasibility of autologous transplantation of tooth tissues cryopreserved with vitrification, by investigating the influence of cryopreservation with vitrification on human dental root, regarding the morphology, microhardness, cell apoptosis, proliferation and differentiation. Freshly extracted human permanent premolars were collected with crown removed. Dental roots were cryopreserved using a commercial vitrification medium (Kitazatousa). After six-month storage in liquid nitrogen, cryopreserved roots were thawed, and then evaluated using histological and immunohistochemical methods. Microhardness of dentine was measured with a Vickers indenter. Cells in periodontal ligament and dental pulp tissues were isolated and characterized. The proliferation, immunophenotype, apoptosis and differentiation ability of cells isolated from cryopreserved roots were evaluated. The data was analyzed using one-way analysis of variance (ANOVA) and Student's t-test. The gross and histological morphology of dental roots was not significantly changed after vitrification and thawing. A few tiny cracks were found in 3 of all 10 cryopreserved samples. No obvious changes were found in microstructure of dentine under SEM observation. Dental pulp cells and periodontal ligament cells were successfully isolated from tissues of cryopreserved human dental roots. There were also no significant differences of those periodontal ligament cells in the two groups regarding morphology, immunophenotype, viability, proliferation and apoptosis. The osteogenic and adipogenic differentiation capability of periodontal ligament cells was maintained by cryopreservation with vitrification. In the conditions of this study, cryopreservation with vitrification preserves cell survival, hardness and structural integrity of dental roots. Vitrification can be a potential way to preserve tooth tissue for future auto-transplantation and autologous cell therapy.

Characterization of cells in blood evoked from periapical tissues in immature teeth with pulp necrosis and their potential for autologous cell therapy in Regenerative Endodontics.

OBJECTIVE: The objectives of this study were to isolate, characterize progenitor cells from blood in the root canals of necrotic immature permanent teeth evoked from periapical tissues and evaluate the applicable potential of these isolated cells in Regenerative Endodontics. DESIGN: Ten necrotic immature permanent teeth from seven patients were included. Evoked bleeding from periapical tissues was induced after chemical instrumentation of the root canals. Cells were isolated from the canal blood and evaluated for cell surface marker expression, multilineage differentiation potential, proliferation ability, and target protein expression. Cell sheets formed from these cells were transferred into human root segments, and then transplanted into nude mice. Histological examination was performed after eight weeks. Data analysis was conducted using one-way ANOVA followed by Tukey's post-hoc comparison, considering p < 0.05 as statistically significant. RESULTS: The isolated cells exhibited characteristics typical of fibroblastic cells with colony-forming efficiency, and displayed Ki67 positivity and robust proliferation. Flow cytometry data demonstrated that at passage 3, these cells were positive for CD73, CD90, CD105, CD146, and negative for CD34 and CD45. Vimentin expression indicated a mesenchymal origin. Under differentiation media specific differentiation media, the cells demonstrated osteogenic, adipogenic, and chondrogenic differentiation potential. Subcutaneous root canals with cell sheets of isolated cells in nude mice showed the formation of pulp-like tissues. CONCLUSIONS: This study confirmed the presence of progenitor cells in root canals following evoked bleeding from periapical tissues of necrotic immature teeth. Isolated cells exhibited similar immunophenotype and regenerative potential with dental mesenchymal stromal cells in regenerative endodontic therapy.

Effect of conventional needle irrigation, passive ultrasonic irrigation, sonic irrigation and XP-endo finisher in removing intracanal Vitapex paste.

To compare the efficiency of conventional needle irrigation, passive ultrasonic irrigation, sonic irrigation and XP-endo Finisher in removing Vitapex paste from root canals. The root canals of human single-rooted teeth were prepared and obturated with Vitapex paste. After 2 weeks, the 48 teeth were randomly allocated into four groups: conventional needle irrigation (CNI) group, passive ultrasonic irrigation (PUI) group, sonic irrigation (SI) group and XP-endo Finisher (XP) group. The specimens of four groups were scanned using a micro-computed tomography after the Vitapex pastes was removed. The results showed that more residual paste was left in the CNI group than in the other three groups (p < 0.05). There was no significant difference among the PUI, SI and XP group (p > 0.05). And, the majority of the residual Vitapex was found in the apical third, with a small amount in the middle third after using any of the three agitated irrigation techniques.

Human Digested Dentin Matrix for Dentin Regeneration and the Applicative Potential in Vital Pulp Therapy.

INTRODUCTION: Human dentin is a natural acellular matrix with excellent reported biocompatibility. The aim was to fabricate a novel dentin matrix material from human dentin and investigate its applicative potential for vital pulp therapy. METHODS: Digested dentin matrix extract (DDME) was fabricated using controlled enzymatic digestion under acidic conditions. The surfaces and biocompatibility of DDME were then investigated, with its effects on the odontogenic differentiation of human dental pulp cells (hDPCs) also studied. The ability of DDME to induce mineralization was assessed in a nude mouse model. The performance of DDME as a pulp capping agent was evaluated in an in situ rat model. The molecular mechanism was verified by mRNA sequencing. RESULTS: A novel type of dentin matrix material with a uniform size of 8 µm was fabricated. DDME had a similar band compared with grinded dentin matrix, with a smaller size, and more uneven surface, as detected by Fourier-transform infrared spectrometer and X-ray photoelectron spectroscopy. DDME at low concentrations did not affect hDPC viability or proliferation, but enhanced runt-related transcription factor 2, dentin matrix acidic phosphoprotein 1, and COL1A1 (collagen type I alpha 1 chain) expression in hDPCs in vitro. DDME was superior to HA-TCP (hydroxyapatite-tricalcium phosphate) in dentin-like mineralized tissue formation after subcutaneous transplantation. In the rat model of pulpotomy, DDME showed visible curative effects. The underlying mechanism may be the inhibition of Hippo signaling following DDME treatment. DDME promoted Yes-associated protein (YAP) 1 nuclear influx, thereby enhancing the expression of DMP-1 (dentin matrix acidic phosphoprotein 1), which was reversed by YAP inhibitor treatment. CONCLUSIONS: Human DDME can be used as a biomaterial for dentin regeneration. The combined application of DDME and current pulp capping agents is a potential choice for vital pulp therapy.

Construction of a CQDs/Ag3PO4/BiPO4 Heterostructure Photocatalyst with Enhanced Photocatalytic Degradation of Rhodamine B under Simulated Solar Irradiation.

A carbon quantum dot (CQDs)/Ag3PO4/BiPO4 heterostructure photocatalyst was constructed by a simple hydrothermal synthesis method. The as-prepared CQDs/Ag3PO4/BiPO4 photocatalyst has been characterized in detail by X-ray diffraction, field-emission scanning electron microscopy, transmission electron microscopy, X-ray photoelectron spectroscopy, ultraviolet-visible spectroscopy, and photoelectrochemical measurements. It is demonstrated that the CQDs/Ag3PO4/BiPO4 composite is constructed by assembling Ag3PO4 fine particles and CQDs on the surface of rice-like BiPO4 granules. The CQDs/Ag3PO4/BiPO4 heterostructure photocatalyst exhibits a higher photocatalytic activity for the degradation of the rhodamine B dye than that of Ag3PO4, BiPO4, and Ag3PO4/BiPO4. The synergistic effects of light absorption capacity, band edge position, separation, and utilization efficiency of photogenerated carriers play the key role for the enhanced photodegradation of the rhodamine B dye.

A novel Bi4Ti3O12/Ag3PO4 heterojunction photocatalyst with enhanced photocatalytic performance.

In this work, we integrated Ag3PO4 with Bi4Ti3O12 to form Bi4Ti3O12/Ag3PO4 heterojunction nanocomposites by an ion-exchange method. The as-prepared Bi4Ti3O12/Ag3PO4 composites were systematically characterized by means of XRD, SEM, TEM, BET, XPS, UV-vis DRS, EIS, PL spectroscopy, and photocurrent response. SEM, TEM, and XPS results demonstrate the creation of Bi4Ti3O12/Ag3PO4 heterojunction with obvious interfacial interaction between Bi4Ti3O12 and Ag3PO4. PL spectra, EIS spectra, and photocurrent responses reveal that the composites display an enhanced separation efficiency of photogenerated electron-hole pairs, which is due to the charge transfer between Bi4Ti3O12 and Ag3PO4. Rhodamine B (RhB) was chosen as the target organic pollutant to evaluate its degradation behavior over Bi4Ti3O12/Ag3PO4 composites under simulated sunlight irradiation. Compared to bare Bi4Ti3O12 and Ag3PO4 nanoparticles, the composites exhibit a significantly enhanced photocatalytic activity. The highest photocatalytic activity is observed for the 10% Bi4Ti3O12/Ag3PO4 composite with 10% Bi4Ti3O12 content, which is about 2.6 times higher than that of bare Ag3PO4. The photocatalytic mechanism involved was investigated and discussed in detail.