Rat Dental Pulp Research Articles

MMP Inhibition, Marginal Integrity and Cytotoxicity of Zinc-Releasing GIC

ObjectivesThe degradation of the restorative-dentin interface due to endogenous dentin enzymes, such as matrix metalloproteinases (MMPs), is a significant issue that accelerates the deterioration of the dentin matrix and leads to the failure of restorative treatments. Caredyne Restore (CR), a novel glass ionomer cement (GIC) with zinc ions in its formulation, represents the latest effort to mitigate this issue. This investigation aimed to evaluate the MMPs inhibitory effect, marginal integrity, and cytotoxicity of CR compared to a conventional GIC, Fuji IX (FIX). MethodsThe inhibitory effect of CR on MMPs activity was evaluated using in-situ zymography to visualize the endogenous gelatinolytic activity in the GIC-dentin interface. Additionally, CR's sealing properties were investigated using dye-assisted confocal laser microscopy (CLSM) to assess marginal leakage across the GIC-dentin interface. Both in-situ zymography and CLSM observations were conducted 1 day and seven days after pH cycling. Finally, the cytotoxicity of the GIC eluates was examined on rat dental pulp cells (RPC-C2A). Assay measurements were performed after 24 and 48 h of incubation with test solutions prepared using various GIC eluate concentrations. ResultsThe MMPs activity of the CR specimens was significantly lower than that of FIX specimens after seven days of pH cycling. Signs of marginal leakage were also lower in the CR specimens. Comparison of CR and FIX eluates at the same concentration showed no significant difference in terms of biocompatibility. ConclusionsCaredyne restore is capable of inhibiting endogenous enzyme activity and improving sealing properties, while maintaining sufficient biocompatibility. Clinical SignificanceZinc ions released from Caredyne Restore offer a safe way to improve the quality of dental restorations and promote minimally invasive treatment, especially in lesions where the dentin matrix is susceptible to enzymatic degradation and recurrent caries.

The lipoteichoic acid of Lactobacillus plantarum effect on lymphocyte, VEGF-A and TGF-β expression in male rat dental pulp

ObjectiveLipoteichoic acid from Lactobacillus plantarum (L. plantarum) is a significant virulence factor that exacerbates pulp inflammation. Lipoteichoic acid plays a role in modulating the inflammatory to proliferative phase transition is crucial in determining the outcome of pulp healing or necrosis. This study explores the role of L. plantarum on lymphocytes and the expression of transforming growth factor β1 (TGF-β1) and vascular endothelial growth factor A (VEGF-A) in a male rat model of acute dental pulp injury. DesignThe acute dental pulp model was created in the upper molar of Rattus novergicus using a round bur. Then, the dental pulp was exposed to 10 µg/ml of the lipoteichoic acid of L. plantarum and filled with a temporary filling. In the next 24, 48, and 72 h, each animal was decapitated, and the expression of TGF-β1 and VEGF-A in dental pulp was analyzed using indirect immunohistochemistry, while the lymphocytes analyzed using hematoxyline-eosin staining. ResultLipoteichoic acid of L. plantarum induced acute dental pulp by increasing the lymphocyte number after 48 and 72 h of exposure (p < 0.05). While, inhibiting TGF-β1 expression after 48 and 72 h of exposure (p < 0.05), and VEGF-A was inhibiting after 72 h of exposure (p < 0.05). ConclusionExposure to lipoteichoic acid from L. plantarum significantly accelerates the inflammatory response in the dental pulp. However, this accelerated inflammation disrupts the proliferative phase, potentially leading to more extensive damage to the dental pulp.

Effects of Capsaicin on Migration and Alkaline Phosphatase Activity of Dental Pulp Cells.

Dental pulp, a specialized mesenchymal tissue within teeth, is pivotal in dental health and tissue repair. Capsaicin, the primary pungent component of chili peppers, is known for its diverse pharmacological properties. While capsaicin's effects on various cell types have been studied, its impact on dental pulp cells remains relatively unexplored. This study investigated the influence of pure capsaicin extract on dental pulp cell behavior, focusing on cell viability, proliferation, migration, and alkaline phosphatase (ALP) activity. Capsaicin solution was prepared and diluted to various concentrations (1 nM, 0.01 µM, 0.1 µM, 1 µM, 10 µM, and 100 µM), then was tested on rat dental pulp cells (RPC-C2A). Cell viability and proliferation were assessed using the MTT assay. Boyden chamber tests and wound healing were used for evaluating cell migration. The activity of ALP was determined to show cell function during dental pulp repair. The data were analyzed using a one-way analysis of variance or an independent-sample Kruskal-Wallis, followed by multiple comparison tests. Capsaicin of 100 µM exhibited cytotoxicity, whereas those with lower concentrations stimulated cell proliferation. Wound healing assays revealed increased cell migration, particularly when cultured with 1 nM capsaicin (p = 0.002). Boyden chamber assays demonstrated enhanced cell invasion without statistical significance. ALP activity of dental pulp cells increased significantly at 1 nM (p < 0.001) and 1 µM (p = 0.021) capsaicin concentrations, indicating potential dentinogenesis and pulp repair. Capsaicin of lower concentrations, less than 10 µM, is likely to promote proliferation, migration, and ALP activity of dental pulp cells. Our findings offer potential applications for capsaicin as a medication for dental pulp repair.

The road map to proper dental pulp experiments in animal models

Animal models are essential for the development and comparison of new dental materials. However, working on the dental pulp of animal models can be challenging. To provide guidance for conducting proper dental pulp experiments in animal models. The author conducted a literature search on the Google Scholar database to find papers related to pulp capping and animal modeling in dentistry. Based on her own experience in animal research and the findings from the literature review, the author provides practical advice for new researchers in this field. Dogs and monkeys have the size and morphology of teeth that facilitate accessibility while operating in the dental pulp. However, the response of rat dental pulp has been found to be identical to that of humans. Rat maxillary molars are often used for dental pulp studies. Anesthesia using a combination of ketamine and xylazine provides a suitable time for pulp procedures in rats, but it's important to note that anesthetized rats may remain alert despite receiving the correct dose. Attempting to increase the dosage can result in the rat being unable to recover from anesthesia. To ensure safe and effective anesthesia, it is recommended to administer only one-third of the ketamine dose. Xylazine re-dosing should be avoided. Dental researchers in the Middle East and Africa may face unique challenges, but working on rats is challenging worldwide. In the future, virtual reality and simulation may offer alternatives for experimental animals. Wister Albino is the more suitable animal model to be used in dental pulp experiments. The checklist and flow chart of PRAISE 2021 are mandatory for dental researchers experimenting with the dental pulp of animal models.

Autophagy induced by hypoxia in pulpitis is mediated by HIF-1α/BNIP3

ObjectiveHypoxia-inducible factor-1α (HIF-1α) and its downstream factor, 19 kDa BCL-2 interacting protein 3 (BNIP3), promote cellular autophagy under hypoxic conditions. However, their roles in pulpitis are unclear. Therefore, the changes in inflammatory response and autophagy levels caused by hypoxia during pulpitis were evaluated. Additionally, the regulatory mechanism of HIF-1α/BNIP3 in cellular autophagy in pulpitis was explored. DesignPulp from dental pulp tissues of healthy individuals and patients with pulpitis (n = 10) were exposed and combined with a low oxygen simulation chamber to construct pulpitis (n = 6), hypoxia (n = 6), and hypoxia+pulpitis (n = 6) rat models. Hematoxylin and eosin and immunohistochemical staining were used to detect the localization and expression levels of HIF-1α, BNIP3, and autophagy marker protein, LC3B. Transmission electron microscopy was used to confirm autophagosome formation. An in vitro hypoxic model of human dental pulp cells was established, and HIF-1α chemical inhibitor 3-(5′-hydroxymethyl-2′-furyl)− 1-benzylindazole (YC-1) was administered. Immunofluorescence and western blotting were used to detect the localization and protein levels of HIF-1α, BNIP3, and LC3B. ResultsAutophagy is significantly increased and HIF-1α and BNIP3 are elevated in inflamed dental pulp tissue. Both pulp exposure and hypoxia intervention cause inflammatory reactions in rat dental pulp tissue, accompanied by the autophagy activation. Hypoxia significantly enhances HIF-1α/BNIP3 and autophagy activation. BNIP3 downregulates and autophagy reduces after treatment with YC-1. ConclusionsIn pulpitis, activation of the HIF-1α/BNIP3 signaling pathway driven by hypoxia leads to increased autophagy. This provides a new molecular explanation for autophagy activation in apical periodontitis and new insights into the pathogenesis of the disease.

The Effects of Antimicrobial Photodynamic Therapy Used to Sterilize Carious Dentin on Rat Dental Pulp Tissue.

Antimicrobial photodynamic therapy (aPDT) used to sterilize carious dentin may irritate pulp tissues because of tissue-penetrating laser and singlet oxygen generation. This study aimed to assess the effects of aPDT on rat pulp tissues. A cavity formed in a rat maxillary first molar was treated with aPDT. The combined photosensitizer and laser irradiation conditions in the aPDT groups were as follows: methylene blue and 100 mW for 60 s, brilliant blue (BB) and 100 mW for 60 s, BB and 50 mW for 120 s, and BB and 200 mW for 30 s. Each cavity was treated with an all-in-one adhesive and filled with flowable resin. aPDT was not applied for the control. In each group, the rats were sacrificed on postoperative days 1 and 14, and thin sections of the treated teeth were prepared. Pulp tissue disorganization (PTD), inflammatory cell infiltration (ICI), and tertiary dentin formation (TDF) were evaluated. At 1-day evaluation, there were significant differences between the aPDT group and controls with respect to PTD and ICI (p < 0.01); 14 days later, almost all specimens showed tertiary dentin formation. The application of aPDT caused reversible damage to the rat pulp, while in the long term, healing occurred with the formation of tertiary dentin.

Age-dependent down-regulation of orexin receptors in trigeminal nucleus caudalis correlated with attenuation of orexinergic analgesia in rats

Aging is related to a variety of physiological organ changes, including central and peripheral nervous systems. It has been reported that the orexin signaling has a potential analgesic effect in different models of pain, especially inflammatory pulpal pain. However, the age-induced alteration in dental pain perception and orexin analgesia has not yet been fully elucidated.Here, we tested that how aging may change the effect of orexin-A on nociceptive behaviors in a rat dental pulp pain model. The expression levels of orexin receptors and the nociceptive neuropeptides substance P (SP) and calcitonin-related gene peptide (CGRP) were also assessed in the trigeminal nucleus caudalis (TNC) of young and aged rats. Dental pulp pain was induced by intradental application of capsaicin (100 μg). The immunofluorescence technique was used to evaluate the expression levels.The results show less efficiency of orexin-A to ameliorate pain perception in aged rats as compared to young rats. In addition, a significant decrease in the number of orexin 1 and 2 receptors was observed in the TNC of aged as compared to young rats. Dental pain-induced SP and CGRP overexpression was also significantly inhibited by orexin-A injection into the TNC of young animals. In contrast, orexin-A could not produce such effects in the aged animals.In conclusion, the older age-related reduction of the antinociceptive effect of orexin may be due to the downregulation of its receptors and inability of orexin signaling to inhibit the expression of nociceptive neuropeptides such as SP and CGRP in aged rats.

Effects of direct pulp capping with recombinant human erythropoietin and/or mineral trioxide aggregate on inflamed rat dental pulp.

This study aimed to evaluate the dental pulp responses to recombinant human erythropoietin (rhEPO) and/or mineral trioxide aggregate (MTA) in pulp capping of inflamed dental pulp in vivo. In accordance with ARRIVE guidelines, pulp inflammation was induced by exposing the maxillary first molars (n = 64) of Wistar rats (n = 32) to the oral environment for two days. The exposed pulps were randomly assigned four groups based on the pulp capping material: rhEPO, MTA, MTA + rhEPO, or an inert membrane. An additional eight rats formed the healthy control group. After four weeks, the animals were euthanized, and histological, qRT-PCR, and spectrophotometric techniques were employed to analyze the left maxillary segments, right first maxillary molars, and blood samples, respectively. Statistical significance was set at p < 0.05 and < 0.001. Pulp capping with rhEPO, MTA, or MTA + rhEPO resulted in lower inflammation and higher mineralization scores compared to untreated control. MTA + rhEPO group exhibited significantly decreased expression of tumor necrosis factor-alpha, and interleukin 1-beta, while MTA group showed substantially reduced expression of interferon-gamma. Both rhEPO and MTA + rhEPO groups presented elevated dentin matrix protein 1 levels compared to untreated control. Furthermore, pulp capping with rhEPO and/or MTA led to increased transforming growth factor-beta 1 expression and reductions of pro-inflammatory/immunoregulatory cytokine ratios and prooxidative markers. Pulp capping with rhEPO also resulted in increase of systemic antioxidative stress markers. Capping with rhEPO or MTA + rhEPO resulted in a favorable effect that was similar or even superior to that of MTA.

Inflammatory Response and Odontogenic Differentiation of Inflamed Dental Pulp Treated with Different Pulp Capping Materials: An In Vivo Study.

Previous studies have evaluated the pulpal responses to calcium silicate cements (CSCs) on normal dental pulp, but investigations on the effects of CSCs on inflamed pulp are limited. This study aimed to test the inflammatory response and odontogenic differentiation of inflamed rat dental pulp after direct pulp capping with CSCs. Wistar rat molars pulps were exposed for 48 hours to induce inflammation and then capped with ProRoot MTA (Dentsply, Tulsa, OK, USA), Biodentine (Septodont, Saint-Maur-des-Fossés, France), RetroMTA (Bio MTA, Seoul, Korea) and Dycal (Dentsply Caulk, Milford, DE, USA). The degree of pulpal inflammation and hard tissue formation was evaluated by histological analysis. Immunofluorescence staining for interleukin (IL)-6, osteocalcin (OCN), and runt-related transcription factor 2 (RUNX2) was also performed. After 4 weeks, complete recovery from inflammation was evident in 22%, 37.5%, 10%, and none of the ProRoot MTA, Biodentine, RetroMTA and Dycal samples, respectively. Heavy hard tissue deposition as a continuous hard tissue bridge was observed in 77.8%, 75%, 70%, and 60% of the ProRoot MTA, Biodentine, RetroMTA, and Dycal samples, respectively. IL-6, OCN, and RUNX2 were detected in all materials, mainly adjacent to areas of inflammation and reparative dentine formation. At 1, 2, and 4 weeks, significant differences were not observed between the inflammation and hard tissue formation scores of the four material groups (P > .05). In this study, pulpal inflammation was still present in most specimens at 4 weeks after pulp capping and a significant number of samples showed incomplete and discontinuous dentine bridge formation. The results of this study suggest that initial inflammatory conditions of the pulp may risk the prognosis of teeth treated with CSCs.

SrCuSi4 O10 /GelMA Composite Hydrogel-Mediated Vital Pulp Therapy: Integrating Antibacterial Property and Enhanced Pulp Regeneration Activity.

Vital pulp therapy (VPT) is considered a conservative treatment for preserving pulp viability in caries-induced dental pulp infections. However, bacterial contamination negatively affects dentine-pulp complex repair. The common capping materials show limited antimicrobial effects against some microorganisms. To improve the VPT efficacy, capping materials with increased antibacterial properties and enhanced odontogenic and angiogenic activities are needed. Herein, a SrCuSi4 O10 /gelatin methacrylate(SC/Gel) composite hydrogel has been proposed for infected dental pulp treatment. SrCuSi4 O10 (SC) is a microscale bioceramic composed of assembled multilayered nanosheets that possesses good near-infrared photothermal conversion ability and multiple bioactivities due to sustained Sr2+ , Cu2+ , and SiO3 2- ion release. It is shown that the SC/Gel composite hydrogel efficiently eliminates Streptococcus mutans and Lactobacillus casei and inhibits biofilm formation under photothermal heating, while the ion extract from SC promotes odontogenesis of rat dental pulp stem cellsand angiogenesis of human umbilical vein endothelial cells. The as-designed therapeutic effect of SC/Gel composite hydrogel-mediated VPT has been proven in a rat dental pulp infection model and yielded improved dentine-pulp complex repair compared with the commercially used iRoot® BP Plus. This study suggests that the SC/Gel composite hydrogel is a potential pulp-capping material with improved effects on dentine-pulp complex repair in infected pulp.

Biocompatibility enhancement via post-processing of microporous scaffolds made by optical 3D printer.

Providing a 3D environment that mimics the native extracellular matrix is becoming increasingly important for various applications such as cell function studies, regenerative medicine, and drug discovery. Among the most critical parameters to consider are the scaffold's complicated micro-scale geometry and material properties. Therefore, stereolithography based on photopolymerization is an emerging technique because of its ability to selectively form volumetric structures from liquid resin through localized polymerization reactions. However, one of the most important parameters of the scaffold is biocompatibility, which depends not only on the material but also on the exposure conditions and post-processing, which is currently underestimated. To investigate this systematically, microporous scaffolds with pore sizes of 0.05mm3 corresponding to a porosity of 16,4% were fabricated using the stereolithography printer Asiga PICO2 39 UV from the widely used resins FormLabs Clear and Flexible. The use of various polymers is usually limited for cells because, after wet chemical development, the non-negligible amount of remaining monomers intertwined in the photopolymerized structures is significantly toxic to cells. Therefore, the aim of this research was to find the best method to remove monomers from the 3D scaffold by additional UV exposure. For this purpose, a Soxhlet extractor was used for the first time, and the monomers were immersed in different alcohols. A Raman microspectroscopy was also used to investigate whether different post-processing methods affect DC (cross-linking) to find out if this specifically affects the biocompatibility of the scaffolds. Finally, mesenchymal stem cells from rat dental pulp were examined to confirm the increased biocompatibility of the scaffolds and their ability to support cell differentiation into bone tissue cells.

Impaired pulp healing associated with underlying disorders in the dental pulp of rats with type 2 diabetes

Background/purposeType 2 diabetes mellitus (T2DM), characterized by hyperglycemia, is a systematic disease affecting structure and healing ability in various tissues. This study aimed to investigate whether T2DM could impair the dental pulp healing and cause underlying pathological changes in the dental pulp before an injury occurred. Materials and methodsGoto-Kakizaki rats were used as T2DM model animals and performed with direct pulp capping procedures on the first maxillary molars. The molars at 1, 2, 4 weeks after operation and non-injured molars were examined using hematoxylin and eosin staining, immunohistochemical staining, immunofluorescence staining, and Masson's trichrome staining. The fresh dental pulp of maxillary incisors was collected for transmission electron microscopy and glucose content evaluation. ResultsThe T2DM rats showed deficient reparative dentin formation compared with the healthy rats. Before the occurrence of an injury, underlying pathological changes of major components in the pulp tissue were observed in T2DM rats, including vasculopathy; collagen abnormalities; decreased proliferation, decreased odontogenetic differentiation and damaged ultrastructure of dental pulp cells. High glucose content and advanced glycation end products accumulation were further found in the pulp tissue in T2DM rats. ConclusionT2DM can impede pulp healing process in rats, which is associated with underlying pathological changes in the non-injured pulp caused by the advanced glycation end products accumulation under high-glucose conditions.

VEGFA-modified DPSCs combined with LC-YE-PLGA NGCs promote facial nerve injury repair in rats

ObjectiveThe aim of this research was to investigate the effect of vascular endothelial growth factor A (VEGFA)-overexpressing rat dental pulp stem cells (rDPSCs) combined with laminin-coated and yarn-encapsulated poly(l-lactide-co-glycolide) (PLGA) nerve guidance conduit (LC-YE-PLGA NGC) in repairing 10 mm facial nerve injury in rats. Study DesignrDPSCs isolated from rat mandibular central incisor were cultured and identified in vitro and further transfected with the lentiviral vectors (Lv-VEGFA). To investigate the role and mechanisms of VEGFA in neurogenic differentiation in vitro, semaxanib (SU5416), Cell Counting Kit-8 (CCK-8), real-time quantitative polymerase chain reaction (qPCR) and Western blotting were performed. Ten-millimeter facial nerve defect models in rats were established and bridged by LC-YE-PLGA NGCs. The repair effects were detected by transmission electron microscopy (TEM), compound muscle action potential (CMAP), immunohistochemistry and immunofluorescence. ResultsExtracted cells exhibited spindle-shaped morphology, presented typical markers (CD44+CD90+CD34−CD45−), and presented multidirectional differentiation potential. The DPSCs with VEGFA overexpression were constructed successfully. VEGFA enhanced the proliferation and neural differentiation ability of rDPSCs, and the expression of neuron-specific enolase (NSE) and βIII-tubulin was increased. However, these trends were reversed with the addition of SU5416. This suggests that VEGFA mediates the above effects mainly through vascular endothelial growth factor receptor 2 (VEGFR2) binding. The LC-YE-NGC basically meet the requirements of facial nerve repair. For the in vivo experiment, the CMAP latency period was shorter in DPSCS-VEGFA-NGC group in comparison with other experimental groups, while the amplitude was increased. Such functional recovery correlated well with an increase in histological improvement. Further study suggested that VEGFA-modified DPSCs could increase the myelin number, thickness and axon diameter of facial nerve. NSE, βIII-tubulin and S100 fluorescence intensity and immunohistochemical staining intensity were significantly enhanced. ConclusionVEGFA-modified rDPSCs combined with LC-YE-PLGA NGCs have certain advantages in the growth and functional recovery of facial nerves in rats.

AR-A014418, a glycogen synthase kinase-3β inhibitor, mitigates lipopolysaccharide-induced inflammation in rat dental pulp stem cells via NLR family pyrin domain containing 3 inflammasome impairment

Background/purposeCell pyroptosis and gingival inflammation have been implicated in periodontitis progression. Our previous study revealed that AR-A014418, a pharmacological inhibitor of glycogen synthase kinase-3β (GSK-3β), can enhance the migratory and osteogenic differentiation abilities of rat dental pulp stem cells (rDPSCs). The present study aimed to explore the effect of AR on the inflammation of rDPSCs. Materials and methodsThe primary rDPSCs were isolated and identified by flow cytometry, as well as Oil red O and Alizarin Red S staining. The rDPSCs were cultured and exposed to lipopolysaccharide (LPS) before treating them with different concentrations of AR-A014418. The cell viability was detected using the CCK-8 assay. The generation and secretion of pro-inflammatory cytokines (IL-18, TNF-α, L-1β, and IL-6) were examined by qPCR and ELISA, respectively. To investigate the activation of the NLRP3 inflammasome, the expression levels of pro-caspase 1, cleaved caspase 1, as well as NLRP3 were analyzed by western blotting and immunofluorescence, respectively. ResultsIn the rDPSCs, LPS prohibited cell viability and enhanced the generation and secretion of pro-inflammatory cytokines. LPS upregulated NLRP3 and cleaved caspase-1 protein levels and promoted ASC speck formation in the rDPSCs. AR-A014418 administration effectively blocked the LPS-induced inflammation of the rDPSCs in a dose-dependent way. Mechanistically, AR-A014418 significantly restrained the up-regulation of NLRP3 and cleaved caspase-1 in LPS-treated rDPSCs. ConclusionCollectively, our findings suggest that AR-A014418 significantly mitigates LPS-induced inflammation of rDPSCs by blocking the activation of the NLRP3 inflammasome.

Cell Proliferation, Viability, Differentiation, and Apoptosis of Iron Oxide Labeled Stem Cells Transfected with Lipofectamine Assessed by MRI.

To assess in vitro and in vivo tracking of iron oxide labeled stem cells transfected by lipofectamine using magnetic resonance imaging (MRI), rat dental pulp stem cells (DPSCs) were characterized, labeled with iron oxide nanoparticles, and then transfected with lipofectamine to facilitate the internalization of these nanoparticles. Cell proliferation, viability, differentiation, and apoptosis were investigated. Prussian blue staining and MRI were used to trace transfected labeled cells. DPSCs were a morphologically spindle shape, adherent to culture plates, and positive for adipogenic and osteogenic inductions. They expressed CD73 and CD90 markers and lacked CD34 and CD45. Iron oxide labeling and transfection with lipofectamine in DPSCs had no toxic impact on viability, proliferation, and differentiation, and did not induce any apoptosis. In vitro and in vivo internalization of iron oxide nanoparticles within DPSCs were confirmed by Prussian blue staining and MRI tracking. Prussian blue staining and MRI tracking in the absence of any toxic effects on cell viability, proliferation, differentiation, and apoptosis were safe and accurate to track DPSCs labeled with iron oxide and transfected with lipofectamine. MRI can be a useful imaging modality when treatment outcome is targeted.

SVCT2-GLUT1-mediated ascorbic acid transport pathway in rat dental pulp and its effects during wound healing

Ascorbic acid (AA; vitamin C) plays a crucial role in the biosynthesis and secretion of collagen to produce the organic matrix of hard tissues. Nevertheless, the detailed mechanism by which AA induces reparative dentinogenesis is still unknown. This study aimed to investigate the pathway and function of AA during wound healing in a rat pulpotomy model. Sodium-dependent vitamin C transporter (SVCT) 2 and glucose transporter (GLUT) 1 were detected in odontoblasts, endothelial cells, and nerve fibers in normal pulp tissues. SVCT2 and GLUT1 were also expressed in odontoblast-like cells in pulpotomized tissues of Wistar rats, and immunopositive cells of SVCT2 were significantly increased at 5 days after pulpotomy (p < 0.05). By contrast, osteogenic disorder Shionogi (ODS) rats, which cannot generate AA, also expressed SVCT2 and GLUT1 in normal and wound healing conditions. However, in ODS rats, when compared with the AA-addition group, the formation of dentin bridges in the AA-loss group was not evident, a layer of osteopontin was significantly increased beneath the wound surface (p < 0.05), and alpha smooth muscle actin at the odontoblast-like cells observed along this layer was significantly increased (p < 0.05), but not Nestin. Moreover, the amounts of type 1 collagen generated in the reparative dentin and beneath the wound healing site were significantly diminished (p < 0.05). Macrophages expressing CD68 and CD206 increased beneath the wound site. Hence, AA may be involved in odontoblast-like cell differentiation and anti-inflammatory response during dental pulp wound healing. Our results provide new insights into the function of AA through SVCT2 and GLUT1 in reparative dentinogenesis and may help in developing new therapeutic targets for dental pulpal disease.

Influence of Direct Pulp Capping with Calcium Hydroxide and Mineral Trioxide Aggregate on Systemic Oxidative Stress in Rats

Abstract Direct pulp capping is a procedure where materials are placed on exposed vital pulp tissue in order to stimulate reparative dentinogenesis and preserve pulp vitality. Carious pulp exposure and direct pulp capping are usually accompanied by pulp inflammation which can affect pro- and anti-oxidative systems locally or systemically. Therefore, this study aimed to investigate the potential influence of direct capping of inflamed rat dental pulp with calcium hydroxide (CH) and mineral trioxide aggregate (MTA) on parameters of systemic oxidative status. Dental pulps of the first maxillary molars of Albino rats (n=32) were exposed and capped with either CH (n=8), MTA (n=8) or were left untreated (n=8). Animals with healthy pulp were used as a healthy control (n=8). After four weeks, animals were euthanized and blood samples were collected for biochemical analysis of parameters of systemic oxidative stress by spectrophotometric method. Untreated control had the significantly higher (p &lt;0.05) values of pro-oxidative parameters and lower (p &lt;0.05) values of anti-oxidative parameters (superoxide dismutase and reduced glutathione) compared to healthy control. CH and MTA groups showed reduced values of pro-oxidative parameters compared to untreated control and values of anti-oxidative parameters comparable to healthy control. Pulp exposure led to disbalance in systemic oxidative parameters while direct pulp capping with calcium hydroxide and mineral trioxide aggregate restored the levels of systemic oxidative parameters to that of animals with healthy dental pulp. These results indicate the importance of direct pulp capping and the potential influence of untreated inflamed pulp on systemic health.

COMPARISON OF THE NUMBER OF MACROPHAGES IN THE RAT REVERSIBLE PULPITIS AFTER APPLICATION OF BIODENTIN AND SIWAK EXTRACT

Background: Oral disease still need special attention, especially in dentistry. Inflammation of the pulp is caused by the presence of irritants. One of the mechanical irritants is iatrogenic factors caused by operator error during preparation so that roof of pulp chamber is exposed. This condition is called reversible pulpitis/inflammation of the pulp. Cells that play a role during inflammation are macrophages which are the second defense cell after neutrophil apoptosis. A direct pulp capping procedure is indicated for this condition. The material commonly used is biodentin, but because the price tends to be expensive, alternative materials are needed. Siwak contains flavonoids which are known to play an important anti-inflammatory role in the healing of exposed pulp. The purpose of this study was to compare the number of macrophages in the dental pulp of rats with reversible pulpitis after administration of biodentin and siwak extract. Method: This research is a true experimental with a post test only group design, consisting of 2 treatment groups, namely the 75% siwak extract group and the biodentin group. The material is applied after the tooth has been prepared. The research sample used male wistar rats and sacrificed on the 3rd day. Samples were stained with Hematoxylin Eosin to see macrophage cells. Result: The data obtained were analyzed using the independent T-test. The test results showed a significant difference, with a value of p = 0.008 (p &lt;0.05). Conclusion: The conclusion of this study that the siwak extract group showed a lower mean number of macrophage cells than the biodentin group.

Effects of vitamin B12 in culture medium for calcified nodule formation by rat dental pulp cells

Background/purposeTooth or bone regeneration requires large numbers of mesenchymal stem cells (MSCs). Although dental pulp is a suitable cell source, the number of MSCs present in this tissue is limited and they require a long period for regeneration. Therefore, the present study investigated vitamin B12 (Vb12) as an osteoinductive factor for MSCs obtained from dental pulp. Materials and methodsDental pulp tissue was removed from the root canals of the extracted mandibular incisors of three 6-week-old male Fischer 344/N Slc rats using an endodontic file and whole cells were harvested. After the primary culture, cells were sub-cultured for calcified nodule formation in MEM containing dexamethasone (Dex), β-glycerophosphate (β-GP), vitamin C (Vc), and Vb12. Calcified nodules were confirmed under an inverted phase-contrast microscope. The alkaline phosphatase (ALP) activity of cells and quantity of Ca2+ in calcified nodules were measured. Results were analyzed using the Tukey-Kramer test. ResultsDensely arranged calcified nodules were microscopically observed after the subculture of cells with Dex, β-GP, Vc, and Vb12. The ALP activity level was 0.077 ± 0.023 μmol/μg DNA in MEM supplemented with Vb12, which did not significantly differ from that without Vb12. A mass of calcium nodules formed in culture medium containing Dex, β-GP, Vc, and Vb12. The quantity of Ca2+ increased from 13.04 ± 0.44 to 20.91 ± 0.56 mg/dL (P < 0.01). ConclusionVb12 is effective for in vitro tooth or bone regeneration by the MSCs of rats and is useful as an osteoinductive factor for MSCs.

Upregulation of TACAN in the trigeminal ganglion affects pain transduction in acute pulpitis

ObjectiveAcute pulpitis is one of the common causes of tooth pain. TACAN (Tmem120a) is a newly identified ion channel that senses mechanical pain. In this experiment, we studied the expression of the TACAN ion channel in the trigeminal ganglia in a rat model of pulpitis to explore the correlation between the expression of this ion channel and inflammatory pain. DesignLipopolysaccharide was used to induce acute pulpitis in rats, and pulpitis was assessed histologically. The facial pain threshold of the rats was measured by the von Frey test. TACAN mRNA expression in rat dental pulp and the trigeminal nerve was measured by qPCR, and TACAN protein expression in the trigeminal ganglia was evaluated by western blot analysis and immunofluorescence. Antisense oligonucleotides were used to reduce TACAN protein expression in the trigeminal ganglia, and the change in the pain threshold in the rats with acute pulpitis was determined. ResultsThe results showed that the TACAN transcript level in rat pulp tissue increased under inflammatory conditions, and we proved that pulpitis increased TACAN protein expression in the rat ipsilateral trigeminal ganglia. The facial pain threshold was decreased in rats with pulpitis. A short-term decrease in TACAN protein expression could improve the pain threshold. ConclusionsWith the development of pulpitis after bacterial infection, the upregulation of TACAN expression in the trigeminal ganglia promoted pain sensitivity. A short-term reduction in TACAN expression relieved pain. Therefore, this study indicated that TACAN is a potential target channel for new analgesics.