Periodontal Tissue Development Research Articles

Unraveling Divergent Transcriptomic Profiles: A Comparative Single-Cell RNA Sequencing Study of Epithelium, Gingiva, and Periodontal Ligament Tissues.

The periodontium comprising periodontal ligament (PDL), gingiva, and epithelium play crucial roles in maintaining tooth integrity and function. Understanding tissue cellular composition and gene expression is crucial for illuminating periodontal pathophysiology. This study aimed to identify tissue-specific markers via scRNA-Seq. Primary human PDL, gingiva, and epithelium tissues (n = 7) were subjected to cell hashing and sorting. scRNA-Seq library preparation using 10× Genomics protocol and Illumina sequencing was conducted. The analysis was performed using Cellranger (v3.1.0), with downstream analysis via R packages Seurat (v5.0.1) and SCORPIUS (v1.0.9). Investigations identified eight distinct cellular clusters, revealing the ubiquitous presence of epithelial and gingival cells. PDL cells evolved in two clusters with numerical superiority. The other clusters showed varied predominance regarding gingival and epithelial cells or an equitable distribution of both. The cluster harboring most cells mainly consisted of PDL cells and was present in all donors. Some of the other clusters were also tissue-inherent, while the presence of others was environmentally influenced, revealing variability across donors. Two clusters exhibited genetic profiles associated with tissue development and cellular integrity, respectively, while all other clusters were distinguished by genes characteristic of immune responses. Developmental trajectory analysis uncovered that PDL cells may develop after epithelial and gingival cells, suggesting the inherent PDL cell-dominated cluster as a final developmental stage. This single-cell RNA sequencing study delineates the hierarchical organization of periodontal tissue development, identifies tissue-specific markers, and reveals the influence of environmental factors on cellular composition, advancing our understanding of periodontal biology and offering potential insights for therapeutic interventions.

Tissue engineering in periodontal regeneration: A future paradigm in inflammatory disorder

Periodontitis is an inflammatory disorder that results in tissue destruction when there are too many germs present or when the host’s normal inflammatory response is disrupted. The fight against periodontal regeneration is a titanic one. Tissue engineering was suggested by Langer and colleagues as a potential method for replacing the missing periodontal tissues. The development of functional and long-lasting periodontal tissues will take the role of sick tissues thanks to the science of tissue engineering, which combines engineering and the life sciences. Tissue engineering is an emerging specialisation in the realm of medical health and sciences. It is advancing, spreading its wings over numerous facets of molecular biology, physiology, surgery, regeneration, and molecular medicine. Cell biologists, molecular biologists, biomaterial engineers, experts in microscopic imaging, robotics engineers, computer-assisted designers, and developers of equipment such as bioreactors, where tissues are grown and nurtured, are just a few of the medical and technical specialties that make up the field. Finally, legal advisors and marketing research specialists (product development, and medical implementation) are also involved in the practical process of tissue engineering. New biological organs will be created in the future using engineering and biological techniques.

Genetic Factors of Teeth Impaction: Polymorphic and Haplotype Variants of PAX9, MSX1, AXIN2, and IRF6 Genes.

In recent research, there has been a growing awareness of the role of genetic factors in the positioning and eruption of teeth in the maxilla and mandible. This study aimed to evaluate the potential of specific polymorphic markers of single nucleotide polymorphisms (SNPs) located within the PAX9, MSX1, AXIN2, and IRF6 genes to determine the predisposition to tooth impaction. The study participants were divided into two groups: the first group consisted of individuals with at least one impacted secondary tooth. In contrast, the second group (control group) had no impacted teeth in their jaws. To analyze the genes, real-time PCR (polymerase chain reaction) and TaqMan probes were utilized to detect the selected polymorphisms. The findings suggest that disruptions in the structure and function of the mentioned genetic factors such as polymorphic and haplotype variants of PAX9, MSX1, AXIN2, and IRF6 genes, which play a direct role in tooth and periodontal tissue development, might be significant factors in tooth impaction in individuals with genetic variations. Therefore, it is reasonable to hypothesize that tooth impaction may be influenced, at least in part, by the presence of specific genetic markers, including different allelic variants of the PAX9, AXIN2, and IRF6 genes, and especially MSX1.

Apical Papilla Regulates Dental Follicle Fate via the OGN-Hh Pathway

Root apical complex, including Hertwig’s epithelial root sheath, apical papilla, and dental follicle (DF), is the germinal center of root development, wherein the DF constantly develops into periodontal tissue. However, whether DF development is regulated by the adjacent apical papilla remains largely unknown. In this study, we employed a transwell coculture system and found that stem cells from the apical papilla (SCAPs) inhibit the differentiation and maintain the stemness of dental follicle stem cells (DFSCs). Meanwhile, partial SCAP differentiation markers were upregulated after DFSC coculture. High-throughput RNA sequencing revealed that the Hedgehog (Hh) pathway was significantly downregulated in DFSCs cocultured with SCAPs. Upregulation or downregulation of the Hh pathway can respectively activate or inhibit the multidirectional differentiation of DFSCs. Osteoglycin (OGN) (previously known as mimecan) is highly expressed in the dental papilla, similarly to Hh pathway factors. By secreting OGN, SCAP regulated the stemness and multidirectional differentiation of DFSCs via the OGN-Hh pathway. Finally, Ogn–/– mice were established using the CRISPR/Cas9 system. We found that the root length growth rate was accelerated during root development from PN0 to PN30 in Ogn–/– mice. Moreover, the hard tissues (including dentin and cementum) of the root in Ogn–/– mice were thicker than those in wild-type mice. These phenotypes were likely due to Hh pathway activation and the increased cell proliferation and differentiation in both the apical papilla and DF. The current work elucidates the molecular regulation of early periodontal tissue development, providing a theoretical basis for future research on tooth root biology and periodontal tissue regeneration.

Semaphorin-RhoA signaling regulates HERS maintenance by acting against TGF-β-induced EMT.

Hertwig's epithelial root sheath (HERS) plays a role in root dentin formation. It produces the epithelial rests of Malassez (ERM) for the induction of periodontal tissue development during root formation. Although ERM is thought to be caused by epithelial-mesenchymal transition (EMT), the mechanism by which HERS is maintained as epithelium is unknown. Here, we aimed to elucidate the molecular mechanisms regulating the relationship between HERS maintenance and ERM development. To understand the relationship between HERS and ERM development during root formation, we observed the developing molar root using cytokeratin14 (CK14) Cre/tdTomato mice via stereomicroscopy. The relationship between semaphorin and transforming growth factor (TGF) signaling in the maintenance of HERS and ERM development was examined using CK14cre/R26-tdTomato mice and a HERS cell line. tdTomato-positive cells were observed on HERS and the migrating cells from HERS. The migrating cells showed reduced E-cadherin expression. In contrast, HERS cells expressed semaphorin receptors and active RhoA. Semaphorin signaling was associated with RhoA activation and cell-cell adhesion, while TGF-β induced decreased E-cadherin and active RhoA expression, and consequently enhanced cell migration. HERS induces root formation by controlling epithelial maintenance and EMT through the opposing effects of semaphorin and TGF-β signaling.

Development and regeneration of periodontal supporting tissues.

Periodontal tissues, including gingiva, cementum, periodontal ligament, and alveolar bone, play important roles in oral health. Under physiological conditions, periodontal tissues surround and support the teeth, maintaining the stability of the teeth and distributing the chewing forces. However, under pathological conditions, with the actions of various pathogenic factors, the periodontal tissues gradually undergo some irreversible changes, that is, gingival recession, periodontal ligament rupture, periodontal pocket formation, alveolar bone resorption, eventually leading to the loosening and even loss of the teeth. Currently, the regenerations of the periodontal tissues are still challenging. Therefore, it is necessary to study the development of the periodontal tissues, the principles and processes of which can be used to develop new strategies for the regeneration of periodontal tissues. This review summarizes the development of periodontal tissues and current strategies for periodontal healing and regeneration.

The Roles of SIBLING Proteins in Dental, Periodontal and Craniofacial Development

The majority of dental, periodontal, and craniofacial tissues are derived from the neural crest cells and ectoderm. Neural crest stem cells are pluripotent, capable of differentiating into a variety of cells. These cells can include osteoblasts, odontoblasts, cementoblasts, chondroblasts, and fibroblasts which are responsible for forming some of the tissues of the oral and craniofacial complex. The hard tissue forming cells deposit a matrix composed of collagen and non-collagenous proteins (NCPs) that later undergoes mineralization. The NCPs play a role in the mineralization of collagen. One such category of NCPs is the small integrin-binding ligand, N-linked glycoprotein (SIBLING) family of proteins. This family is composed of dentin sialophosphosprotein (DSPP), osteopontin (OPN), dentin matrix protein 1 (DMP1), bone sialoprotein (BSP), and matrix extracellular phosphoglycoprotein (MEPE). The SIBLING family is known to have regulatory effects in the mineralization process of collagen fibers and the maturation of hydroxyapatite crystals. It is well established that SIBLING proteins have critical roles in tooth development. Recent literature has described the expression and role of SIBLING proteins in other areas of the oral and craniofacial complex as well. The objective of the present literature review is to summarize and discuss the different roles the SIBLING proteins play in the development of dental, periodontal, and craniofacial tissues.

The role of Axin2+ cells in periodontal tissue development and regeneration

The role of Axin2+ cells in periodontal tissue development and regeneration

Proteoglycans in the periodontium: A review with emphasis on specific distributions, functions, and potential applications.

Proteoglycans (PGs) are largely glycosylated proteins, consisting of a linkage sugar, core proteins, and glycosaminoglycans (GAGs). To date, more than 40 kinds of PGs have been identified, and they can be classified as intracellular, cell surface, pericellular, and extracellular PGs according to cellular locations. To illustrate, extracellular PGs are known for regulating the homeostasis of the extracellular matrix; cell-surface PGs play a role in mediating cell adhesion and binding various growth factors. In the field of periodontology, PGs are implicated in cellular proliferation, migration, adhesion, contractility, and anoikis, thereby exerting a profound influence on periodontal tissue development, wound repair, the immune response, biomechanics, and pathological process. Additionally, the expression patterns of some PGs are dynamic and cell-specific. Therefore, determining the roles and spatial-temporal expression patterns of PGs in the periodontium could shed light on treatments for wound healing, tissue regeneration, periodontitis, and gingival overgrowth. In this review, close attention is paid to the distributions, functions, and potential applications of periodontal PGs. Related genetically modified animal experiments and involved signal transduction cascades are summarized for improved understanding of periodontal PGs. To date, however, there is a large amount of speculation on this topic that requires rigorous experiments for validation.

Evaluation of periostin levels in gingival crevicular fluid in association between coronary heart disease and chronic periodontitis

Periostin is a protein, which is essential for periodontal tissue integrity, development and maturity. The aim of this study was to evaluate the role of gingival crevicular fluid (GCF) periostin levels in the association between coronary heart disease (CHD) and chronic periodontitis (CP). This matched case-control study was conducted on 116 participants. The participants were matched for age, gender, and body mass index and divided into four groups as follows: (1) 29 patients with CHD and sever CP (CHD-CP), (2) 29 patients with CHD and without CP (CHD-H), (3) 29 patients without CHD and with sever CP (H-CP), and (4) 29 healthy participants (H-H). The GCF periostin was collected and evaluated using the enzyme-linked immunosorbent assay (ELASA). Finally, the data were analyzed by analysis of variance using the stata software. Significance was assigned at P < 0.05. The results showed that there was a significant difference in the GCF periostin levels in the four groups (P < 0.05). Moreover, according to the results of the Bonferroni's test, differences in the mean periostin levels were significant (P < 0.001) between CHD-CP and CHD-H, CHD-CP and H-CP, CHD-CP and H-H, CHD-H and H-H, and also between H-CP and H-H. The periostin levels reduced in the CHD patients, especially in the CHD-CP group. The findings reveal a probable role of periostin in the association between CHD and CP.

Role of Wnt5a in periodontal tissue development, maintenance, and periodontitis: Implications for periodontal regeneration (Review).

The periodontium is a highly dynamic microenvironment constantly adapting to changing external conditions. In the processes of periodontal tissue formation and remodeling, certain molecules may serve an essential role in maintaining periodontal homeostasis. Wnt family member 5a (Wnt5a), as a member of the Wnt family, has been identified to have extensive biological roles in development and disease, predominantly through the non-canonical Wnt signaling pathway or through interplay with the canonical Wnt signaling pathway. An increasing number of studies has also demonstrated that it serves crucial roles in periodontal tissues. Wnt5a participates in the development of periodontal tissues, maintains a non-mineralized state of periodontal ligament, and regulates bone homeostasis. In addition, Wnt5a is involved in the pathogenesis of periodontitis. Recently, it has been shown to serve a positive role in the regeneration of integrated periodontal complex. The present review article focuses on recent research studies of Wnt5a and its functions in development, maintenance, and pathological disorders of periodontal tissues, as well as its potential effect on periodontal regeneration.

Activation of β-catenin by TGF-β1 promotes ligament-fibroblastic differentiation and inhibits cementoblastic differentiation of human periodontal ligament cells.

TGF-β and Wnt/β-catenin signaling pathways are known to be essential for the development of periodontal tissue. In this study, we examined the crosstalk between TGF-β and Wnt/β-catenin signaling in ligament-fibroblastic differentiation of human periodontal ligament cells (hPDLCs). TGF-β1 treatment significantly increased the expression of ligament-fibroblastic markers, but such expression was preventing by treatment with SB431542, a TGF-β type I receptor inhibitor. As well as phosphorylation of Smad3, TGF-β1 increased β-catenin activation. The depletion of β-catenin reduced the expression of ligament-fibroblastic markers, suggesting that β-catenin is essential for ligament differentiation. The effect of TGF-β1 on β-catenin activation did not seem to be much correlated with Wnt stimuli, but endogenous DKK1 was suppressed by TGF-β1, indicating that β-catenin activation could be increased much more by TGF-β1. In addition to DKK1 suppression, Smad3 phosphorylation by TGF-β1 facilitated the nuclear translocation of cytoplasmic β-catenin. In contrast to ligament-fibroblastic differentiation, inhibition of TGF-β1 signaling was needed for cementoblastic differentiation of hPDLCs. BMP7 treatment accompanied by inhibition of TGF-β1 signaling had a synergistic effect on cementoblastic differentiation. In conclusion, β-catenin activation by TGF-β1 caused ligament-fibroblastic differentiation of hPDLCs, and the presence of TGF-β1 stimuli basically determined whether hPDLCs are differentiated into ligament progenitor or cementoblasts.

Temporal and spatial distribution of Gli1+ cells and their function during periodontal development

This study aimed to investigate the distribution of Gli1+ cells in the periodontal ligament (PDL) and to evaluate their contribution in the development of periodontal tissue by using transgenic mouse lines. Gli1lacZ/+ mice were harvested at different ages (3, 6, and 8 weeks), and the temporal and spatial distribution patterns of Gli1+ PDL cells were revealed by X-gal staining. Afterward, 3-week-old Gli1-CreERT2/+;R26RtdTomato/+ mice were administered with tamoxifen, and the fates of Gli1+ cells and their descendants were traced during periodontal development. A large number of Gli1+ cells were detected in the PDL of the 3-week-old mice; however, their number significantly decreased from 3 weeks to 8 weeks (P<0.05). Cell lineage tracing data showed that the descendants of Gli1+ cells dramatically increased from 3 weeks to 8 weeks (P<0.05) and gradually differentiated into fibroblasts, cementocytes, and osteocytes. The multi-differentiation potential of Gli1+ PDL cells was revealed, indicating that Gli1+ cells are an important cell source for periodontal development.

MiR-361-3p/Nfat5 Signaling Axis Controls Cementoblast Differentiation

The development of periodontal tissue is a complex process, including cementoblast proliferation and differentiation. Emerging reports suggest that microRNAs (miRNAs) play crucial roles in gene regulatory networks governing numerous biological processes. However, how miRNAs modulate cementoblast proliferation and differentiation remains largely unknown. In a previous study, we performed miRNA microarray profiling to fully reveal the expression patterns of miRNAs involved in cementoblast differentiation. We focused on miR-361-3p, which decreased during cementoblast differentiation. Overexpression of miR-361-3p resulted in decreased cementoblast differentiation, whereas the functional inhibition of miR-361-3p yielded the opposite effect. The bioinformatics approach identified nuclear factor of activated T-cell 5 (Nfat5) as a potential target of miR-361-3p, which was further verified by dual luciferase assay. Meanwhile, the expression pattern of Nfat5 was verified both in vitro and in vivo. Furthermore, knockdown of Nfat5 mimicked the inhibitory effect of overexpressing miR-361-3p in cementoblasts. Moreover, multiple signaling pathways, including the Erk1/2, JNK, p38, PI3K-Akt, and NF-κB pathways, were notably activated, and the Wnt/ß-catenin pathway was blocked by downregulation of Nfat5 or forced expression of miR-361-3p in cementoblast differentiation. Finally, the complementary approach demonstrated that miR-361-3p regulated cementoblast differentiation via or partially via Erk1/2 and PI3K-Akt. Overall, our study elucidated that the JNK, p38, NF-κB, and Wnt/ß-catenin pathways act as balancing players in the miR-361-3p/Nfat5 signaling axis during cementoblast differentiation.

HDAC6 regulates dental mesenchymal stem cells and osteoclast differentiation

BackgroundDental and periodontal tissue development is a complicated process involving a finely regulated network of communication among various cell types. Understanding the mechanisms involved in regulating dental mesenchymal stem cells (MSCs) and osteoclast cell differentiation is critical. However, it is still unclear whether histone deacetylase HDAC6 is involved in dental MSCs fate determination and osteoclast differentiation.MethodsWe used shRNA and siRNA knockdown to explore the role of HDAC6 in dental MSCs odontogenic differentiation and osteoclasts maturation.ResultsBased on HDAC6 knockdown dental MSCs, our data suggest that HDAC6 knockdown significantly increases alkaline phosphate activity and mineralized nodules formation. Additionally, mRNA expression of odontogenic marker genes (OSX, OCN, and OPN) was induced by HDAC6 knockdown. By using HDAC6 siRNA, we knocked down HDAC6 in osteoclast precursor RAW 264.7 cells. Our data suggests that HDAC6 knockdown significantly inhibited osteoclasts differentiation. Additionally, mRNA expression of osteoclast marker genes Trap, Mmp9, and Ctsk was decreased by HDAC6 knockdown.ConclusionsOur study demonstrated that HDAC6 plays an important role in regulating dental MSCs and osteoclasts differentiation.

Selective adipogenic differentiation of human periodontal ligament stem cells stimulated with high doses of glucose.

Periodontal tissue damage, accompanied by the degradation and destruction of periodontal tissue collagen, is one of the most clinically common complications and difficulty self-repair in patients with diabetes. Human periodontal ligament stem cells (PDLSC) are the undifferentiated mesenchymal cells that persist in the periodontal ligament after development of periodontal tissue and the ability of PDLSC osteogenic differentiation is responsible for repairing periodontal tissue defects. However, the reasons of high glucose environment in diabetic patients inhibiting PDLSC to repair periodontal tissues are unclear. To address these issues, we propose exposing PDLSC to high-sugar mimics the diabetic environment and investigating the activity of osteogenic differentiation and adipogenic differentiation of PDLSC. At the cellular level, high glucose can promote the adipogenic differentiation and inhibit osteogenic differentiation to decrease the self-repair ability of PDLSC in periodontal tissues. Mechanistically at the molecular level, these effects are elicited via regulating the mRNA and protein expression of C/EBPβ, PPAR-γ.

Comparative gene expression profiles of dental follicle at different stages of periodontal development: Combined use of laser capture microdissection and microarray

ObjectivesOdontogenesis is dependent on serial temporal and spatial epithelial-mesenchymal interaction. Multiple signaling networks have been identified in the inductive interactions of odontogenesis at the early stage. Technical obstacles and heterogeneity of the dental follicle (DF), which is the origin of the majority of periodontal tissues, have hindered the clarification of the molecular blueprint in early periodontal tissue development. This has in turn hindered studies of new/effective periodontal regeneration therapy. In this study, we comparatively analyzed the gene expression profiles of DF at E17 (when DF cells are histologically recognized) and PN2 (the initiation of periodontal development). MethodsGene expression profiles of DF at E17 and PN2 were assessed by the combined use of laser capture microdissection and microarray. ResultsComparative gene expression analysis of DF at E17 and PN2 during periodontal development revealed > 2-fold up-regulation and down-regulation of 2519 and 5060 genes, respectively. Bioinformatic analysis of the selected genes revealed that the temporally changed genes were mostly enriched in GO terms relative to the vasculature system, and were sometimes linked to multiple processes. RT-qPCR was used to verify the microarray data. ConclusionsThe delineation of the differential gene expressions between pre- and post-natal developmental stages of DF in vivo will increase the understanding of periodontal tissue development.

Modulation of microRNAs in Tooth Root and Periodontal Tissue Development.

Tooth root development begins after the completion of tooth crown development. Both the tooth root and crown undergo a series of interactions between the epithelium and adjacent mesenchymal cells. Although many studies have evaluated tooth crown formation, little is known about the regulatory mechanisms of tooth root development. MicroRNAs (miRNAs) are small noncoding RNAs that regulate protein expression through post-transcriptional mechanisms and participate in a broad range of biological processes, from development to tumorigenesis. The functional importance of miRNAs on the development of tooth root and periodontal tissues has been suggested in many studies. To summarize the functions of miRNAs on tooth root and periodontal tissue development. MicroRNAs are important to root odontogenesis, Hertwig's epithelial root sheath and periodontal tissue development, and have functions in stem cells from dental or periodontal tissues. The modulation of miRNAs in tooth root and periodontal tissue development is fine tuning.

Harmine promotes molar root development via SMAD1/5/8 phosphorylation

Tooth and bone are major tissues involved in physiological calcification in the body, and they use similar molecular pathways for development, homeostasis, and regeneration. Harmine (HMN) is a natural small compound that stimulates osteoblast differentiation in vitro and in vivo. Here we examined the biological effect of HMN on the postnatal development of molar tooth roots and periodontal tissues. HMN supported the formation of tooth roots and periodontal tissues in developing tooth germs. In tooth germ organ culture, HMN promoted the elongation of Hertwig's epithelial root sheath (HERS) and stimulated cell proliferation in HERS and dental follicle-derived tissues, including dental papillae and dental follicles. HMN stimulated cell proliferation and cell movement of HERS-derived cells without mesenchymal cells in vitro and directly induced the phosphorylation of SMAD1/5/8 protein in HERS-derived cells. Our results indicated that HMN was the first natural small compound to stimulate postnatal development of tooth germs.

Effects of cathepsin K on Emdogain-induced hard tissue formation by human periodontal ligament stem cells.

Recent studies have shown that patients with pycnodysostosis caused by cathepsin K (CTSK) genetic mutations exhibit significantly abnormal periodontal hard tissue structure. This finding suggests that CTSK may play a role in regulating the development of alveolar bone and cementum. However, the source of CTSK in the periodontal environment and the role of CTSK in periodontal regeneration, particularly hard tissue regeneration and development, remain unclear. After the isolation, cultivation, identification, and multi-lineage induction of human periodontal ligament stem cells (hPDLSCs), the present study used light and scanning electron microscopy, reverse-transcription quantitative polymerase chain reaction, western blotting, micro-computed tomography, immunohistochemical assays and ectopic hard tissue formation experiments to examine CTSK expression in hPDLSCs. The results indicated that CTSK expression was significantly upregulated in hPDLSCs during Emdogain induction but underwent minimal change during osteogenic or adipogenic induction. The present study also showed that the downregulation of CTSK expression inhibited osteogenic/cementogenic differentiation and ectopic hard tissue formation of hPDLSCs. It is therefore concluded that hPDLSCs expressed CTSK and that CTSK levels were significantly upregulated during Emdogain induction. Furthermore, CTSK promoted not only the osteogenic/cementogenic differentiation of hPDLSCs but also their ability to form ectopic hard tissue. These new findings may enhance the understanding of periodontal hard tissue development and functional regeneration. However, the specific underlying mechanisms require further investigation. Copyright © 2016 John Wiley & Sons, Ltd.