Deposition Of Mineralized Tissue Research Articles

Effect of polydatin on the viability and odontogenic differentiation of human dental pulp stem cells: An in-vitro study

Background/purposeVarious materials have been used to promote human dental pulp stem cells (hDPSCs) differentiation to produce dentin bridge formation with less-than-optimal results. Polydatin (PD), a naturally present material with osteogenic properties can be a promising material in the pulp regeneration/repair process. The aim of this study was to evaluate the effect of (PD) on the viability and differentiation of human dental pulp stem cells. Materials and methodsPD effect on hDPSCs in terms of cellular viability, alkaline phosphatase (ALP) production, and messenger RNAs (mRNA) of odontogenic markers production using quantitative reverse transcription polymerase chain reaction (RT-qPCR) were evaluated. In addition, mineral deposits were detected with Alizarin red stain. ResultsThe viable hDPSCs in the presence of 0.01 μM and 0.1 μM PD were significantly higher than the control on days 3 and 7, respectively. In addition, ALP activity of hDPSCs was significantly increased with 0.01, 0.1, and 1 μM of PD. In addition, increased expression mRNAs of ALP, osteocalcin (OC), osteonectin (ON), osteopontin (OP), Runt-related transcription factor-2 (RUNX-2), dentin sialophosphoprotein (DSPP), and dentin matrix protein-1 (DMP-1) was observed after PD treatment, however, the difference was not statistically significant. Furthermore, increased size of mineral deposits was observed with PD. ConclusionPD promoted the expression of markers associated with odontogenic differentiation and mineralized tissue deposition in hDPSCs.

GelMA/TCP nanocomposite scaffold for vital pulp therapy

Pulp capping is a necessary procedure for preserving the vitality and health of the dental pulp, playing a crucial role in preventing the need for root canal treatment or tooth extraction. Here, we developed an electrospun gelatin methacryloyl (GelMA) fibrous scaffold incorporating beta-tricalcium phosphate (TCP) particles for pulp capping. A comprehensive morphological, physical-chemical, and mechanical characterization of the engineered fibrous scaffolds was performed. In vitro bioactivity, cell compatibility, and odontogenic differentiation potential of the scaffolds in dental pulp stem cells (DPSCs) were also evaluated. A pre-clinical in vivo model was used to determine the therapeutic role of the GelMA/TCP scaffolds in promoting hard tissue formation. Morphological, chemical, and thermal analyses confirmed effective TCP incorporation in the GelMA nanofibers. The GelMA+20%TCP nanofibrous scaffold exhibited bead-free morphology and suitable mechanical and degradation properties. In vitro, GelMA+20%TCP scaffolds supported apatite-like formation, improved cell spreading, and increased deposition of mineralization nodules. Gene expression analysis revealed upregulation of ALPL, RUNX2, COL1A1, and DMP1 in the presence of TCP-laden scaffolds. In vivo, analyses showed mild inflammatory reaction upon scaffolds’ contact while supporting mineralized tissue formation. Although the levels of Nestin and DMP1 proteins did not exceed those associated with the clinical reference treatment (i.e., mineral trioxide aggregate), the GelMA+20%TCP scaffold exhibited comparable levels, thus suggesting the emergence of differentiated odontoblast-like cells capable of dentin matrix secretion. Our innovative GelMA/TCP scaffold represents a simplified and efficient alternative to conventional pulp-capping biomaterials. Statement of significanceVital pulp therapy (VPT) aims to preserve dental pulp vitality and avoid root canal treatment. Biomaterials that bolster mineralized tissue regeneration with ease of use are still lacking. We successfully engineered gelatin methacryloyl (GelMA) electrospun scaffolds incorporated with beta-tricalcium phosphate (TCP) for VPT. Notably, electrospun GelMA-based scaffolds containing 20% (w/v) of TCP exhibited favorable mechanical properties and degradation, cytocompatibility, and mineralization potential indicated by apatite-like structures in vitro and mineralized tissue deposition in vivo, although not surpassing those associated with the standard of care. Collectively, our innovative GelMA/TCP scaffold represents a simplified alternative to conventional pulp capping materials such as MTA and Biodentine™ since it is a ready-to-use biomaterial, requires no setting time, and is therapeutically effective.

Histological evaluation of a novel phosphorylated pullulan-based pulp capping material: An in vivo study on rat molars.

To evaluate the dental pulp response to a novel mineral trioxide aggregate containing phosphorylated pullulan (MTAPPL) in rats after direct pulp capping. Ninety-six cavities were prepared in the maxillary first molars of 56male Wistar rats. The dental pulps were intentionally exposed and randomly divided into four groups according to the application of pulp capping materials: MTAPPL; phosphorylated pullulan (PPL); a conventional MTA (Nex-Cem MTA, NCMTA; positive control); and Super-Bond (SB; negative control). All cavities were restored with SB and observed for pulpal responses at 1-, 3-, 7- and 28-day intervals using a histological scoring system. Statistical analysis was performed using Kruskal-Wallis and Mann-Whitney U-test with Bonferroni's correction, and the level of significance was set at 0.05. DMP1 and CD34 antigen were used to evaluate odontoblast differentiation and pulpal vascularization, respectively. On day 1, mild inflammatory cells were present in MTAPPL and NCMTA groups; fewer inflammatory cells were present in the PPL, whereas SB was associated with a mild-to-moderate inflammatory response. A significant difference was observed between PPL and SB (p<.05). No mineralized tissue deposition was observed. On day 3, moderate-to-severe inflammatory cells were present in PPL and SB, whereas MTAPPL and NCMTA had a mild inflammatory response. Initial mineralized tissue deposition was observed in the NCMTA, MTAPPL and SB. A significant difference was observed between MTAPPL and PPL (p<.05). On day 7, a thin layer of mineralized tissue was observed in all tested groups with no or mild inflammatory response. On day 28, no inflammatory response was observed in MTAPPL, whereas NCMTA, PPL and SB had mild inflammatory responses. A significant difference was observed between MTAPPL and SB (p<.05). Complete mineralized tissue barrier formation was observed in MTAPPL, NCMTA and PPL with no significant difference (p>.05). SB exhibited incomplete mineralized tissue barriers, significantly different from NCMTA, MTAPPL and PPL (p<.05). The staining with CD34 was positive in all the groups on all observation days. The favourable pulpal responses and induction of mineralized tissue formation associated with MTAPPL indicate its potential application as a direct pulp capping material.

Regeneration of pulp-dentin complex using human stem cells of the apical papilla: in vivo interaction with two bioactive materials.

To compare the regenerative properties of human stem cells of the apical papilla (SCAPs) embedded in a platelet-rich plasma (PRP) scaffold, when implanted in vivo using an organotypic model composed of human root segments, with or without the presence of the bioactive cements - ProRoot MTA or Biodentine. SCAPs were isolated from third molars with incomplete rhizogenesis and expanded and characterized in vitro using stem cell and surface markers. The pluripotency of these cells was also assessed using adipogenic, chondrogenic, and osteogenic differentiation protocols. SCAPs together with a scaffold of PRP were added to the root segment lumen and the organotypic model implanted on the dorsal region of immunodeficient rats for a period of 4 months. Presence of SCAPs induced de novo formation of dentin-like and pulp-like tissue. A barrier of either ProRoot MTA or Biodentine did not significantly affect the fraction of sections from roots segments observed to contain deposition of hard material (P > 0.05). However, the area of newly deposited dentin was significantly greater in segments containing a barrier of Biodentine compared with ProRoot MTA (P < 0.001). SCAPs offer a viable alternative to other dental stem cells (DSCs) in their regenerative properties when enclosed in the microenvironment of human tooth roots. The present study also suggests that the presence of bioactive materials does not hinder or impede the formation of new hard tissues, but the presence of Biodentine may promote greater mineralized tissue deposition.

The Effect of Laser Photobiomodulation on Periodontal Ligament Stem Cells.

Photobiomodulation (PBM) is considered as a noninvasive procedure with the potential of inducing favorable changes in cellular behavior. In this study, we aimed to evaluate the effects of near-infrared low-intensity laser PBM on proliferation, viability and osteogenic differentiation of stem cells isolated from human periodontal ligament. A 940-nm diode laser with an energy density of 4Jcm-2 in a 100-mW continuous wave was used for irradiation in 3 sessions every 48h. Cell viability was measured 24, 48 and 72h after irradiation. The effects of laser on mineralized tissue deposition were evaluated by using Alizarin red staining after dividing cells into three groups of nonosteogenic medium (C-), an osteogenic medium without laser (C+), and an osteogenic medium with laser irradiation (L+). Gene expression levels were also evaluated by real-time PCR. Our results showed no significant difference between MTT levels of the study and control groups. After 14 and 21days, both L+ and C+ groups showed an increase in mineralized tissue formation compared to the C- group. There was an increase in VEGF and BMP expressions compared to C-. In conclusion, the irradiation setting used in this study may be able to improve mineralized tissue deposition.

Effect of Leptin on Odontoblastic Differentiation and Angiogenesis: An In Vivo Study

IntroductionLeptin is secreted as a peptide hormone from adipose tissues. The aim of this study was to evaluate the effects of leptin on reparative dentin formation and angiogenesis in the pulp tissue of teeth in vivo. MethodsTwenty-four 7-week-old male rats were anesthetized. Cavities were prepared in maxillary first molars. Pulp cappings were performed with collagen scaffold (Col) with a phosphate-buffered saline (PBS) vehicle (Col + PBS), leptin 1 μmol/L with Col (L1 + Col), or leptin 10 μmol/L with Col (L10 + Col). For the negative control group (no pulp capping), pulp capping was not performed. All cavities were sealed with resin-modified glass ionomer followed by a micro–computed tomographic scan, histologic examination, and immunohistochemical analysis. ResultsThe volume of newly formed mineralized tissue in the leptin group was significantly (P < .01) higher than that in the control group based on micro–computed tomographic analysis. In histologic examination, hard tissue formation was rarely shown in the no pulp capping and Col + PBS groups. However, significantly (P < .01) larger amounts of newly mineralized tissue deposition were observed in the leptin groups. In immunohistochemical analysis, reparative dentin and new vessels formed in the pulp cavity of the leptin groups. Vascular endothelial growth factor, dentin sialoprotein, and dentin sialophosphoprotein were expressed around the newly formed mineralized tissue area. ConclusionsLeptin showed the ability to induce angiogenesis, odontogenic differentiation, and mineralization in exposed rat pulps. Leptin also exhibited favorable inflammatory responses in the pulp tissue. Not only osteodentin but also tubular dentin and new vessels were observed in the pulp cavity.

Composite bilayered scaffolds with bio-functionalized ceramics for cranial bone defects: An in vivo evaluation

Despite having substantial regenerative capabilities, bone regeneration in critical injuries may be insufficient and require an additional intervention. With advancements in material science and production technology it is now possible to generate complex scaffolds with controlled architectures for repairing these injuries. Additionally, these materials can be functionalized with bioactive molecules to enhance osteoinductivity. In the present work, we developed a multifunctional composite bilayered scaffold (BS), integrating a ceramic nanocement (NC) and macroporous composite scaffold (CG) for cranial injuries, mimicking bone architecture. The scaffolds were functionalized with recombinant human bone morphogenetic protein-2 (rhBMP-2) (BMP) (2 μg/scaffold) and zoledronic acid (ZA) (10 μg/scaffold). We hypothesized that the composite scaffolds would support proliferation of osteoblast progenitor cells and provide controlled release of loaded bioactive molecules to induce bone regeneration. Higher amounts of mineralized tissue (MT) deposition was observed with functionalized scaffolds 12 weeks post in vivo implantation in 8.5 mm critical cranial defect in rats. Contrary to our expectations, BS + ZA functionalized scaffolds had highest MT deposition (13.9 mm3), followed by CG + ZA + BMP with 9.2 mm3 and BS + ZA + BMP with 7.6 mm3 of MT deposition, all significantly higher than non-functionalized CG (7.2 mm3) or BS (4.9 mm3) scaffolds and the empty [Teotia et al 2017 ACS Appl. Mater. Interfaces, 9, 6816–6828] groups. The results supported an osteopromotive multifunctional scaffold implantation in critical defects.

Effects of concentrated growth factor on proliferation, migration, and differentiation of human dental pulp stem cells in vitro

Concentrated growth factor, a novel autologous plasma extract, contained various growth factors which promoted tissue regeneration. In this study, we aimed to investigate the biological effects of concentrated growth factor on human dental pulp stem cells. The microstructure and biocompatibility of concentrated growth factor scaffolds were evaluated by scanning electron microscopy. Cell proliferation and migration, odontoblastic and endothelial cell differentiation potential were assessed after exposing dental pulp stem cells to different concentrations (5%, 10%, 20%, 50%, or 80%) of concentrated growth factor extracts. The results revealed that concentrated growth factor scaffolds possessed porous fibrin network with platelets and leukocytes, and showed great biocompatibility with dental pulp stem cells. Higher cell proliferation rates were detected in the concentrated growth factor–treated groups in a dose-dependent manner. Interestingly, in comparison to the controls, the low doses (<50%) of concentrated growth factor increased cell migration, alkaline phosphatase activity, and mineralized tissue deposition, while the cells treated in high doses (50% or 80%) showed no significant difference. After stimulating cell differentiation, the expression levels of dentin matrix protein-1, dentin sialophosphoprotein, vascular endothelial growth factor receptor-2 and cluster of differentiation 31 were significantly upregulated in concentrated growth factor–supplemented groups than those of the controls. Furthermore, the dental pulp stem cell–derived endothelial cells co-induced by 5% concentrated growth factor and vascular endothelial growth factor formed the most amount of mature tube-like structures on Matrigel among all groups, but the high-dosage concentrated growth factor exhibited no or inhibitory effect on cell differentiation. In general, our findings confirmed that concentrated growth factor promoted cell proliferation, migration, and the dental pulp stem cell–mediated dentinogenesis and angiogenesis process, by which it might act as a growth factor–loaded scaffold to facilitate dentin–pulp complex healing.

Pulp Revascularization after Root Canal Decontamination with Calcium Hydroxide and 2% Chlorhexidine Gel

IntroductionPulp revascularization may be considered a promising alternative for necrotic immature teeth. Many studies have accomplished passive decontamination associated with an antibiotic paste. To date, there is no report evaluating calcium hydroxide associated with 2% chlorhexidine gel for revascularization therapy. The aim of this case report was to describe a new proposal for pulp revascularization with mechanical decontamination and intracanal medication composed of calcium hydroxide and 2% chlorhexidine gel. MethodsThe patient, a 9-year-old girl, suffered an intrusion associated with pulp exposure caused by an enamel-dentin fracture in her maxillary left central incisor. After diagnosis, treatment consisted of revascularization therapy with gentle manual instrumentation of the cervical and medium thirds of the root in addition to intracanal medication with calcium hydroxide and 2% chlorhexidine gel for 21 days. In the second session, a blood clot was stimulated up to the cervical third of the root canal. Mineral trioxide aggregate (MTA; Angelus, Londrina, Paraná, Brazil) was used for cervical sealing of the canal. Coronal sealing was performed with temporary filling material and composite resin. ResultsDuring the follow-up period, the root canal space showed a progressive decrease in width, mineralized tissue deposition on root canal walls, and apical closure. A cone-beam computed tomography scan taken at the 2-year follow-up confirmed these findings and did not show complete root canal calcification. ConclusionsThis new proposal for revascularization therapy with 2% chlorhexidine gel may be used for the treatment of necrotic immature root canals.

Antimicrobial Photodynamic Therapy for the Treatment of Teeth with Apical Periodontitis: A Histopathological Evaluation

IntroductionThis study evaluated the in vivo response of apical and periapical tissues of dogs’ teeth with apical periodontitis after one-session endodontic treatment with and without antimicrobial photodynamic therapy (aPDT). MethodsSixty root canals with experimentally induced apical periodontitis were instrumented and assigned to 4 groups receiving aPDT and root canal filling (RCF) or not: group aPDT+/RCF+ (n = 20): aPDT (photosensitizer phenothiazine chloride at 10 mg/mL for 3 minutes and diode laser [λ = 660 nm, 60 mW/cm2] for 1 minute) and RCF in the same session; group aPDT+/RCF− (n = 10); group aPDT−/RCF+ (n = 20), and group aPDT−/RCF− (n = 10). Teeth were restored, and the animals were killed after 90 days. Sections from the maxillas and mandibles were stained with hematoxylin-eosin and Mallory trichrome and examined under light microscopy. Descriptive (ie, newly formed apical mineralized tissue, periapical inflammatory infiltrate, apical periodontal ligament thickness, and mineralized tissue resorption) and quantitative (ie, periapical lesion size and number of inflammatory cells) microscopic analysis was performed. Quantitative data were analyzed by the Kruskal-Wallis and Dunn tests (α = .05). ResultsIn the aPDT-treated groups, the periapical region was moderately/severely enlarged with no inflammatory cells, moderate neoangiogenesis and fibrogenesis, and the smallest periapical lesions. ConclusionsAlthough apical closure by mineralized tissue deposition was not achieved, the absence of inflammatory cells, moderate neoangiogenesis, and fibrogenesis in the periapical region in the groups treated with aPDT indicate that this can be a promising adjunct therapy to cleaning and shaping procedures in teeth with apical periodontitis undergoing one-session endodontic treatment.

Periapical repair after root canal filling with different root canal sealers

The aim of this study was to evaluate periapical repair after root canal filling with different endodontic sealers. Sixty-four root canals from dog s teeth were filled, divided into 4 groups (n=16). Root canals were instrumented with K-type files and irrigated with 1% sodium hypochlorite solution. Root canals were filled in the same session by active lateral condensation of the cones and sealers: Intrafill, AH Plus, Roeko Seal and Resilon/Epiphany System. After 90 days, the animals were euthanized and the tissues to be evaluated were processed and stained with hematoxylin and eosin. For histopathological analysis, the following parameters were evaluated: inflammatory process, mineralized tissue resorption, and apical mineralized tissue deposition. Histopathological analysis demonstrated that Intrafill had less favorable results in terms of apical and periapical repair, compared to the other sealers (p<0.05). AH Plus, Roeko Seal, and Epiphany sealers had similar and satisfactory results (p>0.05). In conclusion, AH Plus and the materials Roeko Seal and Epiphany are good options for clinical use in Endodontics.