Facilitating Reparative Dentin Formation Using Apigenin Local Delivery in the Exposed Pulp Cavity

Apigenin, a natural product belonging to the flavone class, affects various cell physiologies, such as cell signaling, inflammation, proliferation, migration, and protease production. In this study, apigenin was applied to mouse molar pulp after mechanically pulpal exposure to examine the detailed function of apigenin in regulating pulpal inflammation and tertiary dentin formation. In vitro cell cultivation using human dental pulp stem cells (hDPSCs) and in vivo mice model experiments were employed to examine the effect of apigenin in the pulp and dentin regeneration. In vitro cultivation of hDPSCs with apigenin treatment upregulated bone morphogenetic protein (BMP)- and osteogenesis-related signaling molecules such as BMP2, BMP4, BMP7, bone sialoprotein (BSP), runt-related transcription factor 2 (RUNX2), and osteocalcin (OCN) after 14 days. After apigenin local delivery in the mice pulpal cavity, histology and cellular physiology, such as the modulation of inflammation and differentiation, were examined using histology and immunostainings. Apigenin-treated specimens showed period-altered immunolocalization patterns of tumor necrosis factor (TNF)-α, myeloperoxidase (MPO), NESTIN, and transforming growth factor (TGF)-β1 at 3 and 5 days. Moreover, the apigenin-treated group showed a facilitated dentin-bridge formation with few irregular tubules after 42 days from pulpal cavity preparation. Micro-CT images confirmed obvious dentin-bridge structures in the apigenin-treated specimens compared with the control. Apigenin facilitated the reparative dentin formation through the modulation of inflammation and the activation of signaling regulations. Therefore, apigenin would be a potential therapeutic agent for regenerating dentin in exposed pulp caused by dental caries and traumatic injury.

Mouse Dspp frameshift model of human dentinogenesis imperfecta

Non-syndromic inherited defects of tooth dentin are caused by two classes of dominant negative/gain-of-function mutations in dentin sialophosphoprotein (DSPP): 5′ mutations affecting an N-terminal targeting sequence and 3′ mutations that shift translation into the − 1 reading frame. DSPP defects cause an overlapping spectrum of phenotypes classified as dentin dysplasia type II and dentinogenesis imperfecta types II and III. Using CRISPR/Cas9, we generated a Dspp−1fs mouse model by introducing a FLAG-tag followed by a single nucleotide deletion that translated 493 extraneous amino acids before termination. Developing incisors and/or molars from this mouse and a DsppP19L mouse were characterized by morphological assessment, bSEM, nanohardness testing, histological analysis, in situ hybridization and immunohistochemistry. DsppP19L dentin contained dentinal tubules but grew slowly and was softer and less mineralized than the wild-type. DsppP19L incisor enamel was softer than normal, while molar enamel showed reduced rod/interrod definition. Dspp−1fs dentin formation was analogous to reparative dentin: it lacked dentinal tubules, contained cellular debris, and was significantly softer and thinner than Dspp+/+ and DsppP19L dentin. The Dspp−1fs incisor enamel appeared normal and was comparable to the wild-type in hardness. We conclude that 5′ and 3′ Dspp mutations cause dental malformations through different pathological mechanisms and can be regarded as distinct disorders.

Secondary Dentin Formation Mechanism: The Effect of Attrition

Human dentin consists of a primary layer produced during tooth formation in early childhood and a second layer which first forms upon tooth eruption and continues throughout life, termed secondary dentin (SD). The effect of attrition on SD formation was considered to be confined to the area subjacent to attrition facets. However, due to a lack of three-dimensional methodologies to demonstrate the structure of the SD, this association could not be determined. Therefore, in the current study, we aimed to explore the thickening pattern of the SD in relation to the amount of occlusal and interproximal attrition. A total of 30 premolars (50–60 years of age) with varying attrition rates were evaluated using micro-computerized tomography. The results revealed thickening of the SD below the cementoenamel junction (CEJ), mostly in the mesial and distal aspects of the root (p < 0.05). The pattern of thickening under the tooth cervix, rather than in proximity to attrition facets, was consistent regardless of the attrition level. The amount of SD thickening mildly correlated with occlusal attrition (r = 0.577, p < 0.05) and not with interproximal attrition. The thickening of the SD below the CEJ coincided with previous finite element models, suggesting that this area is mostly subjected to stress due to occlusal loadings. Therefore, we suggest that the SD formation might serve as a compensatory mechanism aimed to strengthen tooth structure against deflection caused by mechanical loading. Our study suggests that occlusal forces may play a significant role in SD formation.

Secreted Frizzled-Related Protein 1 Promotes Odontoblastic Differentiation and Reparative Dentin Formation in Dental Pulp Cells

Direct pulp capping is an effective treatment for preserving dental pulp against carious or traumatic pulp exposure via the formation of protective reparative dentin by odontoblast-like cells. Reparative dentin formation can be stimulated by several signaling molecules; therefore, we investigated the effects of secreted frizzled-related protein (SFRP) 1 that was reported to be strongly expressed in odontoblasts of newborn molar tooth germs on odontoblastic differentiation and reparative dentin formation. In developing rat incisors, cells in the dental pulp, cervical loop, and inner enamel epithelium, as well as ameloblasts and preodontoblasts, weakly expressed Sfrp1; however, Sfrp1 was strongly expressed in mature odontoblasts. Human dental pulp cells (hDPCs) showed stronger expression of SFRP1 compared with periodontal ligament cells and gingival cells. SFRP1 knockdown in hDPCs abolished calcium chloride-induced mineralized nodule formation and odontoblast-related gene expression and decreased BMP-2 gene expression. Conversely, SFRP1 stimulation enhanced nodule formation and expression of BMP-2. Direct pulp capping treatment with SFRP1 induced the formation of a considerable amount of reparative dentin that has a structure similar to primary dentin. Our results indicate that SFRP1 is crucial for dentinogenesis and is important in promoting reparative dentin formation in response to injury.

Endodontic Treatment of a Traumatic Tusk Fracture With Exposed Pulp in an Asian Elephant (Elephas maximus)

Tusk fracture in elephants is a common incident often resulting in pulp exposure and pulpitis. Extensive lavage, endodontic therapy, direct pulp capping, or extraction are treatment options. In this report, the successful management of a broken tusk of a juvenile male Asian elephant (Elephas maximus) including morphological analysis of the tusk tip 2 years after surgery are presented. Treatment was carried out under barn conditions and included antimicrobial photodynamic therapy and partial pulpotomy with direct pulp capping. Immediate pain relief was reached. The fractured tusk was preserved and continued to grow. The therapeutic filling material remained intact for over 1 year but was absent 2 years after treatment. The former pulp cavity of the tusk tip was filled with reparative dentin, osteodentin, and bone, but the seal between these hard tissues and pulp chamber dentin was incomplete. Radiographs obtained 3 years after treatment showed no differences in pulp shape, pulp width, and secondary dentin formation between the treated right and the healthy left tusk. It can be concluded that in case of an emergency, the endodontic therapy of a broken elephant tusk can be attempted under improvised conditions with adequate success. Photodynamic therapy might contribute to prevent infection and inflammation of the pulp. The decision tree published by Steenkamp (2019) provides a valuable tool to make quick decisions regarding a suitable therapy of broken tusks.

Local Irradiation of Mouse Tooth Germ Gives Insight into the Direct Effects of Irradiation on Root Development

To elucidate the mechanism underlying the failure of root formation after irradiation, we established a method of local irradiation of the molar tooth germ and demonstrated that radiation directly affected dental root development. In the current study, to locally irradiate the lower first molars of 5-day-old C57BL/6J mice, we used lead glass containing a hole as a collimator. We confirmed that our local irradiation method targeted only the tooth germ. The irradiated root was immature in terms of apical growth, and dentin formation was irregular along the outside of the root apices. Moreover, calcified tissue apically surrounded Hertwig's epithelial root sheath, which disappeared abnormally early. This method using a local irradiation experimental model will facilitate research into radiation-induced disorders of dental root formation.

Dentin Growth after Direct Pulp Capping with the Different Fractions of Plasma Rich in Growth Factors (PRGF) vs. MTA: Experimental Study in Animal Model

Background: This study aimed to evaluate the area of dentin growth in rabbit incisors after pulp capping with plasma rich in growth factors (PRGF) compared with mineral trioxide aggregate (MTA) by fluorescence. Methods: twenty-seven upper and lower incisors of rabbits were divided into 4 groups: poor PRGF (F1) (n = 9 teeth), rich PRGF (F2) (n = 8 teeth), ProRoot MTA (positive control, n = 5 teeth), and untreated (NC) (negative control, n = 5). Fluorochrome markers were injected 24 h before surgery and the day before euthanasia, 28 days after the vital pulp therapy (VPT). Two transverse cuts were performed to every tooth: the first cut (A), 1 mm incisal to the gingival margin, and the second cut (B), 5 mm apical to the first cut. The sections were assessed with histomorphometric evaluation by fluorescence microscopy, comparing the dentin area between fluorescence marks and the total mineralized area. Results: The higher percentage of dentin growth was observed in the F2 group (B = 63.25%, A = 36.52%), followed by F1 (B = 57.63%, A = 30,12%) and MTA (B = 38.64%, A = 15,74%) The group with lowest percentage of dentin growth was the NC group (B = 29.22%, A = 7.82%). Significant difference (p < 0.05) was found between F2 group and MTA, also statistically significant difference has been observed comparing dentin growth areas of NC group with F1 and F2 groups. Conclusions: The application of PRGF rich and poor fraction as a pulp capping material stimulated dentin formation more intensively than MTA and NC.

Comparision of effectiveness of calcium hyroxide and MTA when used as an indirect pulp therapy (IPT) material- A clinical and radiological assessment

: The present study assessed and compared the success of an IPT procedure both clinically and radiographically when Dycal and MTA were used as an IPT material on primary molars.Children aged 4-9 years were screened and those who fulfilled the inclusion criteria were selected. Accordingly fiftychildren were divided into 2 groups with 25 patient in each group. Cavity preparation was done and the two test materials (Dycal and MTA) were placed at the base in their respective groups and restored with RMGIC. Post-operative radiograph was taken for baseline data. Patients were assessed at Subsequent at 1 and 6 months both clinically and radiographically. Both the test materials had formed a good biological seal, arrested further caries progression and did not cause any adverse pulpal reaction. However the amount of reparative dentin formed was highest in the Dycal group followed by MTA group. Both the experimental materials Dycal and MTA showed reparative dentin formation at the end of 1 and 6 months and also formed a good biological seal and maintained vitality of the pulp which indicates both are good IPT material.

Lithium Chloride Exerts Differential Effects on Dentinogenesis and Osteogenesis in Primary Pulp Cultures

Wnt/β-catenin signaling is known to play essential roles in odontoblast differentiation and reparative dentin formation. Various Wnt activators including LiCl have been increasingly studied for their effectiveness to induce repair of the dentin-pulp complex. LiCl is a simple salt thought to activate Wnt/β-catenin signaling by inhibiting GSK3β. Previous in vitro and in vivo studies showed that LiCl increased odontoblast differentiation and enhanced reparative dentin formation. However, the underlying molecular and cellular mechanisms by which LiCl regulates odontoblast and osteoblast differentiation during reparative dentinogenesis are not well-understood. Our in vitro studies show that exposure of early dental pulp progenitors to LiCl increased the survival and the pool of αSMA+ progenitors, leading to enhanced odontoblast and osteoblast differentiation. The positive effects of LiCl in the differentiation of osteoblasts and odontoblasts from αSMA+ progenitors are mediated by Wnt/β-catenin signaling. Our results also showed that continuous and late exposure of dental pulp cells to LiCl increased the expression of odontoblast markers through Wnt/β-catenin signaling, and the number of odontoblasts expressing DMP1-Cherry and DSPP-Cerulean transgenes. However, unlike the early treatment, both continuous and late treatments decreased the expression of Bsp and the expression of BSP-GFPtpz transgene. These observations suggest that prolonged treatment with LiCl in more mature cells of the dental pulp has an inhibitory effect on osteoblast differentiation. The inhibitory effects of LiCl on osteogenesis and Bsp were not mediated through Wnt/β-catenin signaling. These observations suggest that the effects of LiCl, and GSK3β antagonists on reparative dentinogenesis involve multiple pathways and are not specific to Wnt/β-catenin signaling.