CXXC5 mitigates P. gingivalis-inhibited cementogenesis by influencing mitochondrial biogenesis.

BACKGROUND: Cementoblasts on the tooth-root surface are responsible for cementum formation (cementogenesis) and sensitive to Porphyromonas gingivalis stimulation. We have previously proved transcription factor CXXC-type zinc finger protein 5 (CXXC5) participates in cementogenesis. Here, we aimed to elucidate the mechanism in which CXXC5 regulates P. gingivalis-inhibited cementogenesis from the perspective of mitochondrial biogenesis. METHODS: In vivo, periapical lesions were induced in mouse mandibular first molars by pulp exposure, and P. gingivalis was applied into the root canals. In vitro, a cementoblast cell line (OCCM-30) was induced cementogenesis and submitted for RNA sequencing. These cells were co-cultured with P. gingivalis and examined for osteogenic ability and mitochondrial biogenesis. Cells with stable CXXC5 overexpression were constructed by lentivirus transduction, and PGC-1α (central inducer of mitochondrial biogenesis) was down-regulated by siRNA transfection. RESULTS: Periapical lesions were enlarged, and PGC-1α expression was reduced by P. gingivalis treatment. Upon apical inflammation, Cxxc5 expression decreased with Il-6 upregulation. RNA sequencing showed enhanced expression of osteogenic markers, Cxxc5, and mitochondrial biogenesis markers during cementogenesis. P. gingivalis suppressed osteogenic capacities, mitochondrial biogenesis markers, mitochondrial (mt)DNA copy number, and cellular ATP content of cementoblasts, whereas CXXC5 overexpression rescued these effects. PGC-1α knockdown dramatically impaired cementoblast differentiation, confirming the role of mitochondrial biogenesis on cementogenesis. CONCLUSIONS: CXXC5 is a P. gingivalis-sensitive transcription factor that positively regulates cementogenesis by influencing PGC-1α-dependent mitochondrial biogenesis. Video Abstract.

Loss of ß-catenin causes cementum hypoplasia by hampering cementogenic differentiation of Axin2-expressing cells.

BACKGROUND AND OBJECTIVE: Although cementum plays an essential role in tooth attachment and adaptation to occlusal force, the regulatory mechanisms of cementogenesis remain largely unknown. We have previously reported that Axin2-expressing (Axin2+ ) mesenchymal cells in periodontal ligament (PDL) are the main cell source for cementum growth, and constitutive activation of Wnt/ß-catenin signaling in Axin2+ cells results in hypercementosis. Therefore, the aim of the present study was to further evaluate the effects of ß-catenin deletion in Axin2+ cells on cementogenesis. MATERIALS AND METHODS: We generated triple transgenic mice to conditionally delete ß-catenin in Axin2-lineage cells by crossing Axin2CreERT2/+ ; R26RtdTomato/+ mice with ß-cateninflox/flox mice. Multiple approaches, including X-ray analysis, micro-CT, histological stainings, and immunostaining assays, were used to analyze cementum phenotypes and molecular mechanisms. RESULTS: Our data revealed that loss of ß-catenin in Axin2+ cells led to a cementum hypoplasia phenotype characterized by a sharp reduction in the formation of both acellular and cellular cementum. Mechanistically, we found that conditional removal of ß-catenin in Axin2+ cells severely impaired the secretion of cementum matrix proteins, for example, bone sialoprotein (BSP), dentin matrix protein 1 (DMP1) and osteopontin (OPN), and markedly inhibited the differentiation of Axin2+ mesenchymal cells into osterix+ cementoblasts. CONCLUSIONS: Our findings confirm the vital role of Axin2+ mesenchymal PDL cells in cementum growth and demonstrate that Wnt/ß-catenin signaling shows a positive correlation with cementogenic differentiation of Axin2+ cells.

Connective tissue growth factor promotes cementogenesis and cementum repair via Cx43/ß-catenin axis.

BACKGROUND: Orthodontic tooth movement inevitably induces cementum resorption, which is an urgent problem for orthodontists to confront. Human periodontal ligament stem cells (hPDLSCs) exert an important role in the orthodontic tooth movement and exhibit multidirectional differentiation ability in cementum regeneration. Connective tissue growth factor (CTGF) is an important extracellular matrix protein for bone homeostasis and cell differentiation. The purpose of our study was to explore the role of CTGF in cementum repair and cementogenesis and to elucidate its underlying mechanism. METHODS: A cementum defect model was established by tooth movement with heavy forces, and the cementum repair effect of CTGF was observed via micro-CT, HE staining and immunohistochemical staining. RT‒qPCR, western blotting (WB), alizarin red staining and ALP activity experiments verified the mineralization ability of hPDLSCs stimulated with CTGF. The expression of Cx43 in periodontal ligament cells was detected by WB and immunofluorescence (IF) experiments after CTGF stimulation in vivo and in vitro. Subsequently, the mineralization ability of hPDLSCs was observed after application of CTGF and the small interfering RNA Si-Cx43. Additionally, co-intervention via application of the small interfering RNA Si-CTGF and the Cx43 agonist ATRA in hPDLSCs was performed to deepen the mechanistic study. Next, WB, IF experiments and co-immunoprecipitation were conducted to confirm whether CTGF triggers the Cx43/ß-catenin axis to regulate cementoblast differentiation of hPDLSCs. RESULTS: Local oral administration of CTGF to the cementum defects in vivo facilitated cementum repair. CTGF facilitated the cementogenesis of hPDLSCs in a concentration-dependent manner. Cx43 acted as a downstream effector of CTGF to regulate cementoblast differentiation. Si-Cx43 reduced CTGF-induced cementoblast differentiation. The Cx43 agonist ATRA restored the low differentiation capacity induced by Si-CTGF. Further mechanistic studies showed that CTGF triggered the activation of ß-catenin in a dose-dependent manner. In addition, co-localization IF analysis and co-immunoprecipitation demonstrated that Cx43 interacted with ß-catenin at cell‒cell connections. Si-Cx43 attenuated the substantial expression of ß-catenin induced by CTGF. The Cx43 agonist reversed the inhibition of ß-catenin induced by Si-CTGF. IF demonstrated that the nuclear importation of ß-catenin was related to the immense expression of Cx43 at cell‒cell junctions. CONCLUSIONS: Taken together, these data demonstrate that CTGF promotes cementum repair and cementogenesis through activation of the Cx43/ß-catenin signalling axis.

Growth Factors in Periodontal Complex Regeneration.

The ultimate goal of periodontal treatments is the regeneration of all lost periodontal tissues including bone, cementum and the periodontal ligament (PDL). Until now, the clinical methods for periodontal regeneration have been associated with significant failure or incomplete success. Various studies have reported the promising effects of growth factors/cytokines on periodontal regeneration. Growth factors/cytokines include proteins or steroid hormones that bind to cellular receptors, known as signalling molecules, and that trigger cellular responses that eventually stimulate cell proliferation and differentiation. The present review aims to provide an overview of the main growth factors that play an important role in and have been used in the regeneration of periodontal components.

PPARγ-Induced Global H3K27 Acetylation Maintains Osteo/Cementogenic Abilities of Periodontal Ligament Fibroblasts.

The periodontal ligament is a soft connective tissue embedded between the alveolar bone and cementum, the surface hard tissue of teeth. Periodontal ligament fibroblasts (PDLF) actively express osteo/cementogenic genes, which contribute to periodontal tissue homeostasis. However, the key factors maintaining the osteo/cementogenic abilities of PDLF remain unclear. We herein demonstrated that PPARγ was expressed by in vivo periodontal ligament tissue and its distribution pattern correlated with alkaline phosphate enzyme activity. The knockdown of PPARγ markedly reduced the osteo/cementogenic abilities of PDLF in vitro, whereas PPARγ agonists exerted the opposite effects. PPARγ was required to maintain the acetylation status of H3K9 and H3K27, active chromatin markers, and the supplementation of acetyl-CoA, a donor of histone acetylation, restored PPARγ knockdown-induced decreases in the osteo/cementogenic abilities of PDLF. An RNA-seq/ChIP-seq combined analysis identified four osteogenic transcripts, RUNX2, SULF2, RCAN2, and RGMA, in the PPARγ-dependent active chromatin region marked by H3K27ac. Furthermore, RUNX2-binding sites were selectively enriched in the PPARγ-dependent active chromatin region. Collectively, these results identified PPARγ as the key transcriptional factor maintaining the osteo/cementogenic abilities of PDLF and revealed that global H3K27ac modifications play a role in the comprehensive osteo/cementogenic transcriptional alterations mediated by PPARγ.

Recombinant amelogenin regulates the bioactivity of mouse cementoblasts in vitro.

Amelogenin (AMG) is a cell adhesion molecule that has an important role in the mineralization of enamel and regulates events during dental development and root formation. The purpose of the present study was to investigate the effects of recombinant human AMG (rhAMG) on mineralized tissue-associated genes in cementoblasts. Immortalized mouse cementoblasts (OCCM-30) were treated with different concentrations (0.1, 1, 10, 100, 1000, 10,000, 100,000 ng · mL-1) of recombinant human AMG (rhAMG) and analyzed for proliferation, mineralization and mRNA expression of bone sialoprotein (BSP), osteocalcin (OCN), collagen type I (COL I), osteopontin (OPN), runt-related transcription factor 2 (Runx2), cementum attachment protein (CAP), and alkaline phosphatase (ALP) genes using quantitative RT-PCR. The dose response of rhAMG was evaluated using a real-time cell analyzer. Total RNA was isolated on day 3, and cell mineralization was assessed using von Kossa staining on day 8. COL I, OPN and lysosomal-associated membrane protein-1 (LAMP-1), which is a cell surface binding site for amelogenin, were evaluated using immunocytochemistry. F-actin bundles were imaged using confocal microscopy. rhAMG at a concentration of 100,000 ng · mL-1 increased cell proliferation after 72 h compared to the other concentrations and the untreated control group. rhAMG (100,000 ng · mL-1) upregulated BSP and OCN mRNA expression levels eightfold and fivefold, respectively. rhAMG at a concentration of 100,000 ng · mL-1 remarkably enhanced LAMP-1 staining in cementoblasts. Increased numbers of mineralized nodules were observed at concentrations of 10,000 and 100,000 ng · mL-1 rhAMG. The present data suggest that rhAMG is a potent regulator of gene expression in cementoblasts and support the potential application of rhAMG in therapies aimed at fast regeneration of damaged periodontal tissue.

Osteopontin regulates dentin and alveolar bone development and mineralization.

The periodontal complex is essential for tooth attachment and function and includes the mineralized tissues, cementum and alveolar bone, separated by the unmineralized periodontal ligament (PDL). To gain insights into factors regulating cementum-PDL and bone-PDL borders and protecting against ectopic calcification within the PDL, we employed a proteomic approach to analyze PDL tissue from progressive ankylosis knock-out (Ank-/-) mice, featuring reduced PPi, rapid cementogenesis, and excessive acellular cementum. Using this approach, we identified the matrix protein osteopontin (Spp1/OPN) as an elevated factor of interest in Ank-/- mouse molar PDL. We studied the role of OPN in dental and periodontal development and function. During tooth development in wild-type (WT) mice, Spp1 mRNA was transiently expressed by cementoblasts and strongly by alveolar bone osteoblasts. Developmental analysis from 14 to 240days postnatal (dpn) indicated normal histological structures in Spp1-/- comparable to WT control mice. Microcomputed tomography (micro-CT) analysis at 30 and 90dpn revealed significantly increased volumes and tissue mineral densities of Spp1-/- mouse dentin and alveolar bone, while pulp and PDL volumes were decreased and tissue densities were increased. However, acellular cementum growth was unaltered in Spp1-/- mice. Quantitative PCR of periodontal-derived mRNA failed to identify potential local compensators influencing cementum in Spp1-/- vs. WT mice at 26dpn. We genetically deleted Spp1 on the Ank-/- mouse background to determine whether increased Spp1/OPN was regulating periodontal tissues when the PDL space is challenged by hypercementosis in Ank-/- mice. Ank-/-; Spp1-/- double deficient mice did not exhibit greater hypercementosis than that in Ank-/- mice. Based on these data, we conclude that OPN has a non-redundant role regulating formation and mineralization of dentin and bone, influences tissue properties of PDL and pulp, but does not control acellular cementum apposition. These findings may inform therapies targeted at controlling soft tissue calcification.

Yes-associated protein 1 promotes the differentiation and mineralization of cementoblast.

Yes-associated protein 1 (YAP1) transcriptional coactivator is a mediator of mechanosensitive signaling. Cementum, which covers the tooth root surface, continuously senses external mechanical stimulation. Cementoblasts are responsible for the mineralization and maturation of the cementum. However, the effect of YAP1 on cementoblast differentiation remains largely unknown. In this study, we initially demonstrated that YAP1 overexpression enhanced the mineralization ability of cementoblasts. YAP1 upregulated the mRNA and protein expression of several cementogenesis markers, such as alkaline phosphatase (ALP), runt-related transcription factor 2 (Runx2), osteocalcin (OCN), and dentin matrix acidic phosphoprotein 1 (DMP1). The YAP1 overexpression group showed higher intensities of ALP and Alizarin red stain than the YAP1-knockdown group. Unexpectedly, a sharp increase in the expression of dentin sialophosphoprotein (DSPP) was induced by the overexpression of YAP1. Knockdown of YAP1 suppressed DSPP transcriptional activity. YAP1 overexpression activated Smad-dependent BMP signaling and slightly inhibited Erk1/2 signaling pathway activity. Treatment with specific BMP antagonist (LDN193189) prevented the upregulation of the mRNA levels of ALP, RUNX2, and OCN, as well as intensity of ALP-stained and mineralized nodules in cementoblasts. The Erk1/2 signaling pathway inhibitor (PD 98,059) upregulated these cementogenesis markers. Thus, our study suggested that YAP1 enhanced cementoblast mineralization in vitro. YAP1 exerted its effect on the cementoblast partly by regulating the Smad-dependent BMP and Erk1/2 signaling pathways.

MicroRNA-155-3p Mediates TNF-α-Inhibited Cementoblast Differentiation.

Periodontitis is a prevalent and chronic inflammatory disease that is interrelated with systemic health. Periodontitis can be promoted by tumor necrosis factor α (TNF-α). Cementum, a vital part of the periodontium, is a bone-like mineralized tissue that is produced by cementoblasts. Our laboratory previously revealed that TNF-α inhibits cementoblast differentiation and mineralization. However, how TNF-α modulates cementoblast differentiation and mineralization remains largely unknown. MicroRNA-155 (miR-155) is induced and regulates TNF-α-inhibited osteogenic differentiation. In this study, we found that miR-155-3p was increased during TNF-α-stimulated OCCM-30 cells and involved in cementoblast differentiation and mineralization. Overexpression of miR-155-3p suppressed cementoblast mineralization. Bioinformatics analysis revealed that potassium channel tetramerization domain containing 1 ( Kctd1) is a candidate target gene of miR-155-3p. Moreover, miR-155-3p overexpression suppressed KCTD1 levels. Meanwhile, its knockdown increased KCTD1 expression. Transfection with miR-155-3p also inhibited the luciferase activity of 3'-untranslated regions in the Kctd1 wild type but not the mutant. These data indicated that Kctd1 is a direct and novel target of miR-155-3p. The Wnt signaling pathway inhibits cementoblast differentiation, and we further demonstrated that miR-155-3p partially modulates cementoblast differentiation through the canonical Wnt signaling pathway. In addition to the gain/loss function assay of miR-155-3p, the luciferase activity assay of canonical Wnt signaling was performed. The assays revealed that miR-155-3p increased ß-catenin-mediated transcriptional activation. Overall, our data clarified that miR-155-3p mediated TNF-α-inhibited cementoblast differentiation by targeting Kctd1, at least partially through canonical Wnt signaling pathway. These findings reveal the expanded function of miRNAs in cementoblast differentiation and mineralization.

Efficacy of stem cells on periodontal regeneration: Systematic review of pre-clinical studies.

This systematic review aims to evaluate mesenchymal stem cells (MSC) periodontal regenerative potential in animal models. MEDLINE, EMBASE and LILACS databases were searched for quantitative pre-clinical controlled animal model studies that evaluated the effect of local administration of MSC on periodontal regeneration. The systematic review was conducted according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses statement guidelines. Twenty-two studies met the inclusion criteria. Periodontal defects were surgically created in all studies. In seven studies, periodontal inflammation was experimentally induced following surgical defect creation. Differences in defect morphology were identified among the studies. Autogenous, alogenous and xenogenous MSC were used to promote periodontal regeneration. These included bone marrow-derived MSC, periodontal ligament (PDL)-derived MSC, dental pulp-derived MSC, gingival margin-derived MSC, foreskin-derived induced pluripotent stem cells, adipose tissue-derived MSC, cementum-derived MSC, periapical follicular MSC and alveolar periosteal cells. Meta-analysis was not possible due to heterogeneities in study designs. In most of the studies, local MSC implantation was not associated with adverse effects. The use of bone marrow-derived MSC for periodontal regeneration yielded conflicting results. In contrast, PDL-MSC consistently promoted increased PDL and cementum regeneration. Finally, the adjunct use of MSC improved the regenerative outcomes of periodontal defects treated with membranes or bone substitutes. Despite the quality level of the existing evidence, the current data indicate that the use of MSC may provide beneficial effects on periodontal regeneration. The various degrees of success of MSC in periodontal regeneration are likely to be related to the use of heterogeneous cells. Thus, future studies need to identify phenotypic profiles of highly regenerative MSC populations.

Cementogenesis is inhibited under a mechanical static compressive force via Piezo1.

OBJECTIVE: To investigate whether Piezo1, a mechanotransduction gene mediates the cementogenic activity of cementoblasts under a static mechanical compressive force. MATERIALS AND METHODS: Murine cementoblasts (OCCM-30) were exposed to a 2.0 g/cm2 static compressive force for 3, 6, 12, and 24 hours. Then the expression profile of Piezo1 and the cementogenic activity markers osteoprotegerin (Opg), osteopontin (Opn), osteocalcin (Oc), and protein tyrosine phosphataselike member A (Ptpla) were analyzed. Opg, Opn, Oc, and Ptpla expression was further measured after using siRNA to knock down Piezo1. Real-time PCR, Western blot, and cell proliferation assays were performed according to standard procedures. RESULTS: After mechanical stimulation, cell morphology and proliferation did not change significantly. The expression of Piezo1, Opg, Opn, Oc, and Ptpla was significantly decreased, with a high positive correlation between Opg and Piezo1 expression. After Piezo1 knockdown, the expression of Opg, Opn, Oc, and Ptpla was further decreased under mechanical stimulation. CONCLUSIONS: Cementogenic activity was inhibited in OCCM-30 cells under static mechanical force, a process that was partially mediated by the decrease of Piezo1. This study provides a new viewpoint of the pathogenesis mechanism of orthodontically induced root resorption and repair.

Wnt activity is associated with cementum-type transition.

BACKGROUND AND OBJECTIVE: Cellular and acellular cementum and the cells that form them are postulated to have different characteristics, and the relationship between these two tissues is not well understood. Based on the hypothesis that Wnt signaling is involved in the determination of cementum type, we examined Wnt activity along the tooth root and analyzed cementum formation in genetic mutant models. MATERIAL AND METHODS: We generated mutant models with Wnt signaling upregulation (OC Catnblox(ex3)/+ ), downregulation (OC Wlsfl/fl ), and a compound mutant (Enpp1asj/asj ;OC Catnblox(ex3)/+ ) to compare cementum apposition patterns of ectonucleotide diphosphatase/phosphodiesterase (Enpp1) mutant (Enpp1asj/asj ). The analysis of structural morphology and histology was performed with hematoxylin and eosin and immunohistochemical staining and scanning electron microscopy. RESULTS: The cementum type of upper apical region of tooth roots in the molar is altered from the cellular form at the initial stage to the acellular form at the late stage of cementum formation. However, the basal part of this apical region is not altered and retains cellular cementum characters with strong Wnt activity. In the genetic mutant models for Wnt upregulation, cellular cementum is formed at the cervical region instead of acellular cementum. However, Enpp1 mutant mice have clearly different characteristics with cellular-type cementum even with dramatically increased cervical cementum matrix. In addition, we found that acellular-type formation could be altered into cellular-type formation by analyzing Wnt upregulation and compound mutant models. CONCLUSIONS: Cementum type is not determined by its specific location and could be transformed with Wnt activity during cementum formation.

Bifunctional bioceramics stimulating osteogenic differentiation of a gingival fibroblast and inhibiting plaque biofilm formation.

Gingival recession is a common clinical problem that results in esthetic deficiencies and poor plaque control and predominantly occurs in aged patients. In order to restore the cervical region, ideal biomaterials should possess the ability to stimulate proliferation and osteogenesis/cementogenesis of human gingival fibroblasts (HGF) and have a strong antibiofilm effect. The aim of the present study was to investigate the interactions of HGF and oral multispecies biofilms with Ca, Mg and Si-containing bredigite (BRT, Ca7MgSi4O16) bioceramics. BRT extract induced osteogenic/cementogenic differentiation of HGF and its inhibition of plaque biofilm formation were systematically studied. BRT extract in concentrations lower than <200 mg mL(-1) presented high biocompatibility to HGF cells in 3 days. Ion extracts from BRT also stimulated a series of bone-related gene and protein expressions in HGF cells. Furthermore, BRT extract significantly inhibited oral multispecies plaque biofilm growth on its surface and contributed to over 30% bacterial cell death without additional antibacterial agents in two weeks. A planktonic killing test showed that BRT suppressed 98% plaque bacterial growth compared to blank control in 3 days. The results also revealed that BRT extract has an osteostimulation effect on HGF. The suppression effect on plaque biofilms suggested that BRT might be used as a bioactive material for cervical restoration and that the synergistic effect of bioactive ions, such as Ca, Mg and Si ions, played an important role in the design and construction of bifunctional biomaterials in combination with tissue regeneration and antibiofilm activity.

Simulation of Low-Intensity Ultrasound Propagating in a Beagle Dog Dentoalveolar Structure to Investigate the Relations between Ultrasonic Parameters and Cementum Regeneration.

The therapeutic effect of low-intensity pulsed ultrasound on orthodontically induced inflammatory root resorption is believed to be brought about through mechanical signals induced by the low-intensity pulsed ultrasound. However, the stimulatory mechanism triggering dental cell response has not been clearly identified yet. The aim of this study was to evaluate possible relations between the amounts of new cementum regeneration and ultrasonic parameters such as pressure amplitude and time-averaged energy density. We used the finite-element method to simulate the previously published experiment on ultrasonic wave propagation in the dentoalveolar structure of beagle dogs. Qualitative relations between the thickness of the regenerated cementum in the experiment and the ultrasonic parameters were observed. Our results indicated that the areas of the root surface with greater ultrasonic pressure were associated with larger amounts of cementum regeneration. However, the establishment of reliable quantitative correlations between ultrasound parameters and cementum regeneration requires more experimental data and simulations.

Periodontal regeneration employing gingival margin-derived stem/progenitor cells in conjunction with IL-1ra-hydrogel synthetic extracellular matrix.

AIM: This study investigated the periodontal regenerative potential of gingival margin-derived stem/progenitor cells (G-MSCs) in conjunction with IL-1ra-releasing hyaluronic acid synthetic extracellular matrix (HA-sECM). MATERIALS AND METHODS: Periodontal defects were induced at four sites in eight miniature pigs in the premolar/molar area (-4 weeks). Autologus G-MSCs were isolated from the free gingival margin and magnetically sorted, using anti-STRO-1 antibodies. Colony formation and multilineage differentiation potential were tested. The G-MSCs were expanded and incorporated into IL-1ra-loaded/unloaded HA-sECM. Within every miniature pig, four periodontal defects were randomly treated with IL-1ra/G-MSCs/HA-sECM (test group), G-MSCs/HA-sECM (positive-control), scaling and root planing (SRP; negative control-1) or left untreated (no-treatment group; negative control 2). Differences in clinical attachment level (ΔCAL), probing depth (ΔPD), gingival recession (ΔGR), radiographic defect volume (ΔRDV), and changes in bleeding on probing (BOP) between baseline and 16 weeks post-transplantation, as well as periodontal attachment level (PAL), junctional epithelium length (JE), connective tissue adhesion (CTA), cementum regeneration (CR) and bone regeneration (BR) at 16 weeks post-transplantation were evaluated. RESULTS: Isolated G-MSCs showed stem/progenitor cell characteristics. IL-1ra loaded and unloaded G-MSCs/HA-sECM showed higher ΔCAL, ΔPD, ΔGR, PAL, CR and BR as well as a lower JE compared to their negative controls and improved BOP. CONCLUSION: G-MSCs in conjunction with IL-1ra-loaded/unloaded HA-sECM show a significant periodontal regenerative potential.

Cell-to-cell communication--periodontal regeneration.

BACKGROUND: Although regenerative treatment options are available, periodontal regeneration is still regarded as insufficient and unpredictable. AIM: This review article provides scientific background information on the animated 3D film Cell-to-Cell Communication - Periodontal Regeneration. RESULTS: Periodontal regeneration is understood as a recapitulation of embryonic mechanisms. Therefore, a thorough understanding of cellular and molecular mechanisms regulating normal tooth root development is imperative to improve existing and develop new periodontal regenerative therapies. However, compared to tooth crown and earlier stages of tooth development, much less is known about the development of the tooth root. The formation of root cementum is considered the critical element in periodontal regeneration. Therefore, much research in recent years has focused on the origin and differentiation of cementoblasts. Evidence is accumulating that the Hertwig's epithelial root sheath (HERS) has a pivotal role in root formation and cementogenesis. Traditionally, ectomesenchymal cells in the dental follicle were thought to differentiate into cementoblasts. According to an alternative theory, however, cementoblasts originate from the HERS. What happens when the periodontal attachment system is traumatically compromised? Minor mechanical insults to the periodontium may spontaneously heal, and the tissues can structurally and functionally be restored. But what happens to the periodontium in case of periodontitis, an infectious disease, after periodontal treatment? A non-regenerative treatment of periodontitis normally results in periodontal repair (i.e., the formation of a long junctional epithelium) rather than regeneration. Thus, a regenerative treatment is indicated to restore the original architecture and function of the periodontium. Guided tissue regeneration or enamel matrix proteins are such regenerative therapies, but further improvement is required. As remnants of HERS persist as epithelial cell rests of Malassez in the periodontal ligament, these epithelial cells are regarded as a stem cell niche that can give rise to new cementoblasts. Enamel matrix proteins and members of the transforming growth factor beta (TGF-ß) superfamily have been implicated in cementoblast differentiation. CONCLUSION: A better knowledge of cell-to-cell communication leading to cementoblast differentiation may be used to develop improved regenerative therapies to reconstitute periodontal tissues that were lost due to periodontitis.

Novel MesoPorous BioGlass/silk scaffold containing adPDGF-B and adBMP7 for the repair of periodontal defects in beagle dogs.

AIM: The local delivery of growth factors via gene therapy has gained tremendous awareness in recent years due to their sustained growth factor delivery to target tissues. The aim of this study was to fabricate and investigate a scaffold able to release growth factors via gene therapy for the repair of periodontal tissues. MATERIALS AND METHODS: Novel mesoporous bioglass (MBG)/silk fibrin scaffold combined with BMP7 and/or PDGF-B adenovirus was fabricated and tested in vitro for cell migration, proliferation and differentiation. Furthermore, acute-type buccal dehiscence periodontal defects (mesiodistal width × depth: 5 × 5 mm) were created on the buccal portion of the maxillary premolars in five normal male beagle dogs (12 months old, 15.0 ± 2.0 kg) and histologically examined for periodontal regeneration following implantation of the following five groups: (1) no scaffold, (2) MBG/silk scaffold alone, (3) scaffold + adPDGF-B, (4) scaffold + adBMP7, (5) scaffold + adPDGF-b + adBMP7. RESULTS: In vitro findings demonstrated that adPDGF-B was able to rapidly recruit periodontal ligament (PDL) cells over sixfold more effectively than adBMP7, whereas adBMP7 was more able to induce osteoblast differentiation of PDL cells. In vivo findings demonstrate that scaffolds loaded with adPDGF-B were able to partially regenerate the periodontal ligament while adBMP7 scaffolds primarily improved new bone formation. The combination of both adPDGF-B and adBMP7 synergistically promoted periodontal regeneration by allowing up to two times greater regeneration of the periodontal ligament, alveolar bone and cementum when compared to each adenovirus used alone. CONCLUSIONS: Although both PDGF-B and BMP7 are individually capable of promoting periodontal regeneration to some degree, their combination synergistically promotes wound healing in acute-type buccal dehiscence periodontal defects when delivered simultaneously. This study demonstrates the promise for successful delivery of low-cost, effective growth factor delivery via gene therapy for the treatment of periodontal defects.

Periodontal ligament-derived cells for periodontal regeneration in animal models: a systematic review.

BACKGROUND AND OBJECTIVE: Implantation of periodontal ligament stem cells is emerging as a potential periodontal regenerative procedure. This systematic review considers the evidence from animal models investigating the use of periodontal ligament stem cells for successful periodontal regeneration. MATERIAL AND METHODS: PubMed, Embase, MEDLINE and Google Scholar were searched to December 2013 for quantitative studies examining the outcome of implanting periodontal ligament stem cells into experimental periodontal defects in animals. Inclusion criteria were: implantation of periodontal ligament stem cells into surgically created periodontal defects for periodontal regeneration; animal models only; source of cells either human or animal; and published in English. We followed the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. RESULTS: From the literature search, 43 studies met the inclusion criteria. A wide variety of surgical defects were created in four species of animal (dog, rat, pig and sheep). Owing to wide variability in defect type, cell source and cell scaffold, no meta-analysis was possible. Outcome measures included new bone, new cementum and new connective tissue formation. In 70.5% of the results, statistically significant improvements of these measures was recorded. CONCLUSION: These results are notable in that they indicate that irrespective of the defect type and animal model used, periodontal ligament stem cell implantation can be expected to result in a beneficial outcome for periodontal regeneration. It is recommended that there is sufficient evidence from preclinical animal studies to warrant moving to human studies to examine the efficacy, safety, feasibility (autologous vs. allogeneic transplantation) and delivery of periodontal ligament stem cells for periodontal regeneration.

Effect of a tunnel-structured ß-tricalcium phosphate graft material on periodontal regeneration: a pilot study in a canine one-wall intrabony defect model.

BACKGROUND AND OBJECTIVE: Tissue regeneration is affected by the porosity, chemical properties and geometric structure of graft materials. Regeneration of severe periodontal defects, such as one-wall intrabony defects, is difficult because of reduced tissue support, and bone grafts are commonly used in such cases. In the present study, a tunnel-structured ß-tricalcium phosphate (tunnel ß-TCP) graft material designed to stimulate bone formation was fabricated. The objective of this pilot study was to evaluate the effect of this graft material on periodontal regeneration in one-wall intrabony defects in dogs. MATERIAL AND METHODS: Six male beagle dogs were used in this study. First, the mandibular second and third incisors were extracted. Experimental surgery was performed 12 wk after tooth extraction. Bilateral 4 × 8 mm (width × depth) one-wall intrabony defects were created in the mesial side of the mandibular canines. At the experimental sites, the defects were filled with tunnel ß-TCP, whereas the control defects were left empty. Twelve weeks after surgery, qualitative and quantitative histological analyses were performed. RESULTS: There were no signs of clinical inflammation 12 wk after surgery. Coronal extension indicative of new bone formation was higher at the experimental sites than at the control sites, although the differences between both the sites in the newly formed cementum and connective tissue attachment were not significant. Newly formed periodontal ligament and cementum-like tissue were evident along the root surface at the experimental sites. The inner surface of the tunnels was partially resorbed and replaced with new bone. New blood vessels were observed inside the lumens of tunnel ß-TCP. CONCLUSION: Tunnel ß-TCP serves as a scaffold for new bone formation in one-wall intrabony defects.

Cementum proteins: role in cementogenesis, biomineralization, periodontium formation and regeneration.

Destruction of the periodontium is normally associated with periodontal disease, although many other factors, such as trauma, aging, infections, orthodontic tooth movement and systemic and genetic diseases, can contribute to this process. Strategies (such as guided tissue regeneration) have been developed to guide and control regeneration using bioresorbable membranes and bone grafts. Although effective to a certain point, these strategies have the problem that they are not predictable and do not completely restore the architecture of the original periodontium. To achieve complete repair and regeneration it is necessary to recapitulate the developmental process with complete formation of cementum, bone and periodontal ligament fibers. Detailed knowledge of the biology of cementum is key for understanding how the periodontium functions, identifying pathological issues and for developing successful therapies for repair and regeneration of damaged periodontal tissue. It is the purpose of this review to focus on the role of cementum and its specific components in the formation, repair and regeneration of the periodontium. As cementum is a matrix rich in growth factors that could influence the activities of various periodontal cell types, this review will examine the characteristics of cementum, its composition and the role of cementum components, especially the cementum protein-1, during the process of cementogenesis, and their potential usefulness for regeneration of the periodontal structures in a predictable therapeutic manner.