Periodontal tissue regeneration with cementogenesis after application of brain-derived neurotrophic factor in 3-wall inflamed intra-bony defect.

OBJECTIVE: The purpose of this study is to investigate regenerative process by immunohistochemical analysis and evaluate periodontal tissue regeneration following a topical application of BDNF to inflamed 3-wall intra-bony defects. BACKGROUND: Brain-derived neurotrophic factor (BDNF) plays a role in the survival and differentiation of central and peripheral neurons. BDNF can regulate the functions of non-neural cells, osteoblasts, periodontal ligament cells, endothelial cells, as well as neural cells. Our previous study showed that a topical application of BDNF enhances periodontal tissue regeneration in experimental periodontal defects of dog and that BDNF stimulates the expression of bone (cementum)-related proteins and proliferation of human periodontal ligament cells. METHODS: Six weeks after extraction of mandibular first and third premolars, 3-wall intra-bony defects were created in mandibular second and fourth premolars of beagle dogs. Impression material was placed in all of the artificial defects to induce inflammation. Two weeks after the first operation, BDNF (25 and 50 µg/mL) immersed into atelocollagen sponge was applied to the defects. As a control, only atelocollagen sponge immersed in saline was applied. Two and four weeks after the BDNF application, morphometric analysis was performed. Localizations of osteopontin (OPN) and proliferating cell nuclear antigen (PCNA)-positive cells were evaluated by immunohistochemical analysis. RESULTS: Two weeks after application of BDNF, periodontal tissue was partially regenerated. Immunohistochemical analyses revealed that cells on the denuded root surface were positive with OPN and PCNA. PCNA-positive cells were also detected in the soft connective tissue of regenerating periodontal tissue. Four weeks after application of BDNF, the periodontal defects were regenerated with cementum, periodontal ligament, and alveolar bone. Along the root surface, abundant OPN-positive cells were observed. Morphometric analyses revealed that percentage of new cementum length and percentage of new bone area of experimental groups were higher than control group and dose-dependently increased. CONCLUSION: These findings suggest that BDNF could induce cementum regeneration in early regenerative phase by stimulating proliferation of periodontal ligament cells and differentiation into periodontal tissue cells, resulting in enhancement of periodontal tissue regeneration in inflamed 3-wall intra-bony defects.

REV-ERBs negatively regulate mineralization of the cementoblasts.

OBJECTIVE: The role of circadian clock in cementogenesis is unclear. This study examines the role of REV-ERBs, one of circadian clock proteins, in proliferation, migration and mineralization of cementoblasts to fill the gap in knowledge. METHODS: Expression pattern of REV-ERBα in cementoblasts was investigated in vivo and in vitro. CCK-8 assay, scratch wound healing assay, alkaline phosphatase (ALP) and alizarin red S (ARS) staining were performed to evaluate the effects of REV-ERBs activation by SR9009 on proliferation, migration and mineralization of OCCM-30, an immortalized cementoblast cell line. Furthermore, mineralization related markers including osterix (OSX), ALP, bone sialoprotein (BSP) and osteocalcin (OCN) were evaluated. RESULTS: Strong expression of REV-ERBα was found in cellular cementum around tooth apex. Rev-erbα mRNA oscillated periodically in OCCM-30 and declined after mineralization induction. REV-ERBs activation by SR9009 inhibited proliferation but promoted migration of OCCM-30 in vitro. Results of ALP and ARS staining suggested that REV-ERBs activation negatively regulated mineralization of OCCM-30. Mechanically, REV-ERBs activation attenuated the expression of OSX and its downstream targets including ALP, BSP and OCN. CONCLUSIONS: REV-ERBs are involved in cementogenesis and negatively regulate mineralization of cementoblasts via inhibiting OSX expression. Our study provides a potential target regarding periodontal and cementum regeneration.

Intermittent parathyroid hormone promotes cementogenesis via ephrinB2-EPHB4 forward signaling.

Intermittent parathyroid hormone (PTH) promotes periodontal repair, but the underlying mechanisms remained unclear. Recent studies found that ephrinB2-EPHB4 forward signaling mediated the anabolic effect of PTH in bone homeostasis. Considering the similarities between cementum and bone, we aimed to examine the therapeutic effect of PTH on resorbed roots and explore the role of forward signaling in this process. In vivo experiments showed that intermittent PTH significantly accelerated the regeneration of root resorption and promoted expression of EPHB4 and ephrinB2. When the signaling was blocked, the resorption repair was also delayed. In vitro studies showed that intermittent PTH promoted the expression of EPHB4 and ephrinB2 in OCCM-30 cells. The effects of PTH on the mineralization capacity of OCCM-30 cells was mediated through the ephrinB2-EPHB4 forward signaling. These results support the premise that the anabolic effects of intermittent PTH on the regeneration of root resorption is via the ephrinB2-EPHB4 forward signaling pathway.

Cold Atmospheric Plasma Promotes Regeneration-Associated Cell Functions of Murine Cementoblasts In Vitro.

The aim of the study was to examine the efficacy of cold atmospheric plasma (CAP) on the mineralization and cell proliferation of murine dental cementoblasts. Cells were treated with CAP and enamel matrix derivates (EMD). Gene expression of alkaline phosphatase (ALP), bone gamma-carboxyglutamate protein (BGLAP), periostin (POSTN), osteopontin (OPN), osterix (OSX), collagen type I alpha 1 chain (COL1A1), dentin matrix acidic phosphoprotein (DMP)1, RUNX family transcription factor (RUNX)2, and marker of proliferation Ki-67 (KI67) was quantified by real-time PCR. Protein expression was analyzed by immunocytochemistry and ELISA. ALP activity was determined by ALP assay. Von Kossa and alizarin red staining were used to display mineralization. Cell viability was analyzed by XTT assay, and morphological characterization was performed by DAPI/phalloidin staining. Cell migration was quantified with an established scratch assay. CAP and EMD upregulated both mRNA and protein synthesis of ALP, POSTN, and OPN. Additionally, DMP1 and COL1A1 were upregulated at both gene and protein levels. In addition to upregulated RUNX2 mRNA levels, treated cells mineralized more intensively. Moreover, CAP treatment resulted in an upregulation of KI67, higher cell viability, and improved cell migration. Our study shows that CAP appears to have stimulatory effects on regeneration-associated cell functions in cementoblasts.

Yes-associated protein promotes tumour necrosis factor α-treated cementoblast mineralization partly by inactivating NF-κB pathway.

Cementum regeneration, as one of the most difficult challenges of periodontal regeneration, is influenced by inflammatory factors. Inflammation may hamper or promote periodontal tissue repair under different circumstances, as it is found to do in dentin-pulp complex and bone tissue. Our team demonstrated that YAP promotes mineralization of OCCM, a cementoblast cell line. However, the effect of YAP on its mineralization under inflammatory microenvironment is unclear. In this study, cementogenesis in vitro was up-regulated after transient TNF-α treatment for 30 minutes. YAP expression also was increased by TNF-α treatment. YAP overexpression promoted OCCM mineralization after the cells were transiently treated with TNF-α because YAP overexpression inhibited NF-κB pathway activity, while YAP knockdown elevated it. The inhibited mineralization potential and activated NF-κB pathway activity by YAP knockdown also were partly rescued by the application of the NF-κB inhibitor Bay 11-7082. These results demonstrated that YAP plays a positive role in the mineralization of TNF-α transiently treated cementoblast, partly by inhibiting the NF-κB pathway activity.

Lithium chloride attenuates suppressed differentiation induced by mechanical strain in cementoblasts.

Aim: The purpose of this study was to investigate the influence of mechanical strain on OCCM-30 cementoblast differentiation and Wnt/ß-catenin pathway activity. Materials and Methods: Mechanical tension in the form of 2500-µ strain was applied to the cells using the Forcel four-point bending system, with or without the Wnt signaling activator, lithium chloride. Changes in cell differentiation and the expression of Wnt/ß-catenin pathway components in response to strain and lithium chloride were assessed by real-time PCR, immunofluorescence, and western blotting. Results: The mRNA expression levels of the cementoblastogenesis-related genes alkaline phosphatase, runt-related transcription factor 2, and collagen 1, were decreased with mechanical strain. Similarly, the Wnt signaling pathway component genes LRP5, AXIN2, and LEF1 were decreased. The immunofluorescence assay demonstrated that scant ß-catenin underwent nuclear translocation after the cells were subjected to mechanical strain. Moreover, western blotting showed that the protein levels of both ß-catenin and phosphorylated ß-catenin were increased after mechanical strain. In the presence of lithium chloride, the differentiation that was suppressed by mechanical strain was attenuated. Conclusions: 2500-µ strain mechanical strain inhibited cementoblast differentiation activity in vitro, which could be alleviated by actviating Wnt/ß-catenin signaling using lithium chloride.

The effect of CPNE7 on periodontal regeneration.

Introduction: Preameloblast-conditioned medium (PA-CM), as a mixture of dental epithelium-derived factors, has been reported to regenerate dentin and periodontal tissues in vitro and in vivo. The aim of this study was to investigate the biological effect of Cpne7 on the proliferation, migration, and cementoblast differentiation of periodontal cells in vitro, and on the regeneration of periodontal tissue using periodontal defect model with canine in vivo. Materials and methods: The effect of Cpne7 on cell proliferation, migration, and cementoblast differentiation of periodontal cells were evaluated in vitro. A periodontal defect canine model was designed and the defects were divided into five groups: Group 1: No treatment (negative control), Group 2: Collagen carrier only, Group 3: PA-CM with collagen carrier (positive control), Group 4: PA-CM + CPNE7 Antibody (Ab) with collagen carrier, and Group 5: recombinant CPNE7 (rCPNE7) protein with collagen carrier. Results: Cpne7 was expressed in HERS cells and periodontal ligament (PDL) fibers. By real-time PCR, Cpne7 increased expression of Cap compared to the control. In the periodontal defect canine model, rCPNE7 or PA-CM regenerated periodontal complex, and the arrangement of the newly formed PDL-like fibers were perpendicular to the newly formed cementum and alveolar bone like Sharpey's fibers in natural teeth, while PA-CM + CPNE7 Ab showed irregular arrangement of the newly formed PDL-like fibers compared to the rCPNE7 or PA-CM group. Conclusion: These findings suggest that Cpne7 may have a functional role in periodontal regeneration by supporting periodontal cell attachment to cementum and facilitating physiological arrangement of PDL fibers.

Intermittent parathyroid hormone (PTH) promotes cementogenesis and alleviates the catabolic effects of mechanical strain in cementoblasts.

BACKGROUND: External root resorption, commonly starting from cementum, is a severe side effect of orthodontic treatment. In this pathological process and repairing course followed, cementoblasts play a significant role. Previous studies implicated that parathyroid hormone (PTH) could act on committed osteoblast precursors to promote differentiation, and inhibit apoptosis. But little was known about the role of PTH in cementoblasts. The purpose of this study was to investigate the effects of intermittent PTH on cementoblasts and its influence after mechanical strain treatment. RESULTS: Higher levels of cementogenesis- and differentiation-related biomarkers (bone sialoprotein (BSP), osteocalcin (OCN), Collagen type I (COL1) and Osterix (Osx)) were shown in 1-3 cycles of intermittent PTH treated groups than the control group. Additionally, intermittent PTH increased alkaline phosphatase (ALP) activity and mineralized nodules formation, as measured by ALP staining, quantitative ALP assay, Alizarin red S staining and quantitative calcium assay. The morphology of OCCM-30 cells changed after mechanical strain exertion. Expression of BSP, ALP, OCN, osteopontin (OPN) and Osx was restrained after 18 h mechanical strain. Furthermore, intermittent PTH significantly increased the expression of cementogenesis- and differentiation-related biomarkers in mechanical strain treated OCCM-30 cells. CONCLUSIONS: Taken together, these data suggested that intermittent PTH promoted cementum formation through activating cementogenesis- and differentiation-related biomarkers, and attenuated the catabolic effects of mechanical strain in immortalized cementoblasts OCCM-30.

Redefining the induction of periodontal tissue regeneration in primates by the osteogenic proteins of the transforming growth factor-ß supergene family.

The molecular bases of periodontal tissue induction and regeneration are the osteogenic proteins of the transforming growth factor-ß (TGF-ß) supergene family. These morphogens act as soluble mediators for the induction of tissues morphogenesis sculpting the multicellular mineralized structures of the periodontal tissues with functionally oriented ligament fibers into newly formed cementum. Human TGF-ß3 (hTGF-ß3 ) in growth factor-reduced Matrigel® matrix induces cementogenesis when implanted in class II mandibular furcation defects surgically prepared in the non-human primate Chacma baboon, Papio ursinus. The newly formed periodontal ligament space is characterized by running fibers tightly attached to the cementoid surface penetrating as mineralized constructs within the newly formed cementum assembling and initiating within the mineralized dentine. Angiogenesis heralds the newly formed periodontal ligament space, and newly sprouting capillaries are lined by cellular elements with condensed chromatin interpreted as angioblasts responsible for the rapid and sustained induction of angiogenesis. The inductive activity of hTGF-ß3 in Matrigel® matrix is enhanced by the addition of autogenous morcellated fragments of the rectus abdominis muscle potentially providing myoblastic, pericytic/perivascular stem cells for continuous tissue induction and morphogenesis. The striated rectus abdominis muscle is endowed with stem cell niches in para/perivascular location, which can be dominant, thus imposing stem cell features or stemness to the surrounding cells. This capacity to impose stemness is morphologically shown by greater alveolar bone induction and cementogenesis when hTGF-ß3 in Matrigel® matrix is combined with morcellated fragments of autogenous rectus abdominis muscle. The induction of periodontal tissue morphogenesis develops as a mosaic structure in which the osteogenic proteins of the TGF-ß supergene family singly, synergistically and synchronously initiate and maintain tissue induction and morphogenesis. In primates, the presence of several homologous yet molecularly different isoforms with osteogenic activity highlights the biological significance of this apparent redundancy and indicates multiple interactions during embryonic development and bone regeneration in postnatal life. Molecular redundancy with associated different biological functionalities in primate tissues may simply represent the fine-tuning of speciation-related molecular evolution in anthropoid apes at the early Pliocene boundary, which resulted in finer tuning of the bone induction cascade.

Periodontal healing with a preameloblast-conditioned medium in dogs.

BACKGROUND AND OBJECTIVE: The predictability of conventional periodontal treatments for damaged periodontal tissue is limited, particularly on the regeneration of new cementum. As signaling molecules, a range of growth factors has been used to promote periodontal regeneration on periodontal ligament (PDL) and cementum defects. A preameloblast-conditioned medium (PA-CM) was prepared from cultured murine apical bud cells, which can differentiate into ameloblasts. We examined the effect of PA-CM on PDL cells and cementoblasts in vitro and evaluated histologically the effects of PA-CM on the regeneration of experimentally induced periodontal defects in vivo. MATERIAL AND METHODS: In vitro, the effects of PA-CM on the migration of human PDL cells were examined using a scratch wound healing assay and a transwell assay. The differentiation and mineralization potential of PA-CM-treated human PDL cells and murine cementoblastic OCCM-30 cells was examined by real-time polymerase chain reaction and Alizarin red-S staining. In vivo, six mongrel dogs (12-16 kg; 6-8 mo old) were used. Twenty-four roots were replanted with either, (i) only periodontal defects (n = 12; control group), or (ii) periodontal defects and PA-CM treatment (n = 12; experimental group). In the experimental group, the PDL and cementum between notches was removed using a Gracey curette and soaked in 0.08 mL water containing 80 µg of a PA-CM for 2 min. The dogs were killed at 4 and 8 wk post-surgery. RESULTS: The in vitro results showed that PA-CM stimulated the migration of PDL cells and promoted the differentiation and mineralization of PDL cells and cementoblasts. Real-time polymerase chain reaction analysis revealed stronger expression of Runx2, Osx, OC, Bsp and Cap mRNAs in the PA-CM-treated PDL cells and cementoblasts than those in the control cells. In vivo, newly formed PDL-like tissue and cementum-like tissue were observed partially between the root surfaces and newly formed bone in the experimental group. The regenerated PDL-like tissue in the experimental group was significantly higher than that in the control group at 8 wk (p < 0.05). The replacement resorption on the experimental group was significantly lower than that in the control group at 8 wk (p < 0.05). In addition, the amount of newly formed cementum-like tissue in the experimental group was significantly higher than that in the control group at 4 and 8 wk (p < 0.05). CONCLUSION: These results suggest that PA-CM has the potential to regenerate periodontal tissues in PDL and cementum defects.

Fibroblast growth factor-2 promotes healing of surgically created periodontal defects in rats with early, streptozotocin-induced diabetes via increasing cell proliferation and regulating angiogenesis.

AIM: To evaluate the effects of fibroblast growth factor (FGF)-2 on the healing of surgical periodontal defects in rats with early, streptozotocin-induced diabetes. MATERIALS AND METHODS: Fifty Wistar rats were assigned to streptozotocin-induced diabetes or non-diabetes group. Periodontal defects were surgically created at maxillary first molars. Defects were treated with hydroxypropyl cellulose (HPC) or FGF-2 with HPC. Defect fill was evaluated by microcomputed tomography. Histological and immunohistochemical analyses were performed. RESULTS: Compared to vehicle alone, FGF-2 treatment yielded significantly greater bone volume and trabecular thickness in diabetes group. Diabetes group displayed reduced new bone formation and significantly longer epithelial down-growth compared to non-diabetes group. In diabetes group, FGF-2 treatment increased PCNA-positive cells and new bone formation after 2 weeks and suppressed epithelial down-growth, but new cementum formation was minimal even after 4 weeks. In diabetes group, overexpression of vascular endothelial growth factor was evident in cells within connective tissue, and no significant enhancement was observed by FGF-2 treatment. FGF-2 increased the expression of α-smooth muscle actin in diabetes group. CONCLUSIONS: Treatment of surgical periodontal defects in diabetic rats with the single application of FGF-2 provided beneficial effects primarily on new bone formation via increasing cell proliferation and regulating angiogenesis.

Periodontal healing after application of enamel matrix derivative in surgical supra/infrabony periodontal defects in rats with streptozotocin-induced diabetes.

BACKGROUND AND OBJECTIVE: Epidemiologic and clinical studies have indicated that diabetes is a risk factor for periodontal disease progression and healing. The aim of the present study was to evaluate short-term healing after enamel matrix derivative (EMD) application in combined supra/infrabony periodontal defects in diabetic rats. MATERIAL AND METHODS: Thirty male Wistar rats were initially divided into two groups, one with streptozotocin-induced diabetes and another one with healthy (non-diabetic) animals. Bony defects were surgically created on the mesial root of the first maxillary molars. After root surface planing and EDTA conditioning, EMD was applied to the roots at one side of the maxillae, while those on the contralateral sides were left untreated. Animals were killed 3 wk after surgery, and block sections were prepared for histologic and histomorphometric analysis. RESULTS: There was statistically significant more gingival recession in diabetic animals than in non-diabetic animals. The length of the junctional epithelium was significantly shorter in the EMD-treated sites in both diabetic and normoglycemic rats. Sulcus depth and length of supracrestal soft connective tissue showed no statistically significant differences between groups. In all animals, new bone formation was observed. Although new bone occurred more frequently in healthy animals, the extent of new bone was not significantly different between groups. In none of the teeth, a layer of new cementum was detectable. EMD had no influence on bone or cementum regeneration. Adverse reactions such as excessive inflammation due to bacterial root colonization, ankylosis and bone fractures were exclusively observed in diabetic animals, irrespective of EMD treatment. CONCLUSION: Within the limits of the present study, it can be concluded that periodontal healing was impaired in streptozotocin-induced diabetic rats. EMD had no beneficial effects on new bone and cementum formation during short-term healing in this defect model and could not ameliorate the adverse effects in the systemically compromised animals.

Acemannan sponges stimulate alveolar bone, cementum and periodontal ligament regeneration in a canine class II furcation defect model.

BACKGROUND AND OBJECTIVE: Periodontal disease is a common infectious disease, found worldwide, causing the destruction of the periodontium. The periodontium is a complex structure composed of both soft and hard tissues, thus an agent applied to regenerate the periodontium must be able to stimulate periodontal ligament, cementum and alveolar bone regeneration. Recent studies demonstrated that acemannan, a polysaccharide extracted from Aloe vera gel, stimulated both soft and hard tissue healing. This study investigated effect of acemannan as a bioactive molecule and scaffold for periodontal tissue regeneration. MATERIAL AND METHODS: Primary human periodontal ligament cells were treated with acemannan in vitro. New DNA synthesis, expression of growth/differentiation factor 5 and runt-related transcription factor 2, expression of vascular endothelial growth factor, bone morphogenetic protein-2 and type I collagen, alkaline phosphatase activity, and mineralized nodule formation were determined using [(3)H]-thymidine incorporation, reverse transcription-polymerase chain reaction, enzyme-linked immunoabsorbent assay, biochemical assay and alizarin red staining, respectively. In our in vivo study, premolar class II furcation defects were made in four mongrel dogs. Acemannan sponges were applied into the defects. Untreated defects were used as a negative control group. The amount of new bone, cementum and periodontal ligament formation were evaluated 30 and 60 d after the operation. RESULTS: Acemannan significantly increased periodontal ligament cell proliferation, upregulation of growth/differentiation factor 5, runt-related transcription factor 2, vascular endothelial growth factor, bone morphogenetic protein 2, type I collagen and alkaline phosphatase activity, and mineral deposition as compared with the untreated control group in vitro. Moreover, acemannan significantly accelerated new alveolar bone, cementum and periodontal ligament formation in class II furcation defects. CONCLUSION: Our data suggest that acemannan could be a candidate biomolecule for periodontal tissue regeneration.

Premixed calcium silicate-based ceramic sealers promote osteogenic/cementogenic differentiation of human periodontal ligament stem cells: A microscopy study.

To evaluate the effects of premixed calcium silicate based ceramic sealers on the viability and osteogenic/cementogenic differentiation of human periodontal ligament stem cells (hPDLSCs). The materials evaluated were TotalFill BC Sealer (TFbc), AH Plus Bioceramic Sealer (AHPbc), and Neosealer Flo (Neo). Standardized discs and 1:1, 1:2, and 1:4 eluates of the tested materials were prepared. The following in vitro experiments were carried out: ion release, cell metabolic activity 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, cell migration, immunofluorescence experiment, cell attachment, gene expression, and mineralization assay. Statistical analyses were performed using one-way ANOVA followed by Tukey's post hoc test (p < .05). Increased Ca2+ release was detected in TFbc compared to AHPbc and Neo (*p < .05). Biological assays showed a discrete cell metabolic activity and cell migration in Neo-treated cell, whereas scanning electronic microscopy assay exhibited that TFbc group had a better cell adhesion process of substrate attachment, spreading, and cytoskeleton development on the niche-like structures of the cement than AHPbc and Neo. The sealers tested were able to induce overexpression of the CEMP-1, ALP, and COL1A1 genes in the first days of exposure, particularly in the case of TFbc (***p < .001). All materials tested significantly increased the mineralization of hPDLSCs when compared to the negative control, although more pronounced calcium deposition was observed in the TFbc-treated cells (***p < .001). Our results suggested that TFbc promotes cell differentiation, both by increasing the expression of key osteo/odontogenic genes and by promoting mineralization of the extracellular matrix, whereas this phenomenon was less evident in Neo and AHPbc. RESEARCH HIGHLIGHTS: TFbc group had a better cell adhesion process of substrate attachment, spreading, and cytoskeleton development on the niche-like structures of the cement than AHPbc and Neo. The sealers tested were able to induce overexpression of the CEMP-1, ALP, and COL1A1 genes in the first days of exposure, particularly in the case of TFbc. All materials tested significantly increased the mineralization of hPDLSCs when compared to the negative control, although more pronounced calcium deposition was observed in the TFbc-treated cells.

Effects of EMD in combination with bone swaging and calcium phosphate bone cement on periodontal regeneration in one-wall intrabony defects in dogs.

BACKGROUND AND OBJECTIVE: Although the application of EMD is a widely accepted periodontal-regenerative therapy, its effects on noncontained intrabony defects are unpredictable because of the lack of a space-making property. The combined use of EMD and autogenous bone grafts reportedly stimulates significant periodontal regeneration in intrabony defects. The aim of the present study was to evaluate the effects of EMD in combination with bone swaging (BS) and injectable calcium phosphate bone cement (CPC), which was placed into the spaces between the grafted swaged bone and the proximal host bone, on periodontal healing in one-wall intrabony defects in dogs. MATERIAL AND METHODS: One-wall intrabony defects (3 mm wide and 5 mm deep) were surgically created on the mesial and distal sides of the bilateral mandibular premolars in four dogs. The 16 defects were assigned to one of the following treatments: EMD only, BS only, EMD with BS (EMD + BS), or EMD with BS and CPC (EMD + BS + CPC). The animals were killed 8 wk after surgery for histologic evaluation. RESULTS: The height of newly formed bone was significantly greater in the EMD + BS + CPC group (3.73 ± 0.30 mm) than in the BS-only (2.74 ± 0.33 mm; p < 0.05) and EMD + BS (2.88 ± 0.98 mm; p < 0.05) groups. The area of newly formed bone was significantly larger in the EMD + BS + CPC group (5.68 ± 1.66 mm(2)) than in the EMD-only (3.68 ± 0.33 mm(2); p < 0.05), BS-only (3.48 ± 1.26 mm(2); p < 0.05) and EMD + BS (3.38 ± 1.37 mm(2); p < 0.05) groups. The EMD-only (4.63 ± 0.42 mm), EMD + BS (4.67 ± 0.30 mm) and EMD + BS + CPC (4.78 ± 0.54 mm) groups showed significantly greater cementum formation than did the BS-only group (3.93 ± 0.56 mm; p < 0.05). CONCLUSION: These results indicate that treatment with EMD + BS + CPC promotes favorable periodontal healing in one-wall intrabony defects in dogs.

Histological responses of the periodontium to MTA: a systematic review.

AIM: The purpose of this systematic review was to investigate whether a mineral trioxide aggregate (MTA) restoration of an endodontic-periodontal communication leads to regeneration of the adjacent periodontal tissues. METHODOLOGY: The databases MEDLINE-PubMed, Cochrane-CENTRAL, and EMBASE were searched, up to July 2012. In vivo studies that reported on the histological response of the periodontium to MTA were selected. RESULTS: The screening of 98 title-abstracts, full-text reading, and hand searches in literature lists yielded 24 papers. All of them involved animals. There were no studies reporting on human histology. Study protocols presented heterogeneity regarding treated lesions, intervention, and reported outcomes. The histological results of the animal studies showed minimal inflammatory reactions, bone healing, periodontal ligament presence, and consistent cementum formation. Time lapse after mixing, bacterial contamination, root canal disinfection, and inflammation influenced MTA's cementoconductive properties. CONCLUSIONS: Within the limitations of the selected papers concerning inhomogeneous study protocols and low methodological quality scores, their findings were consistent with regard to MTA's biocompatibility and cementogenic ability. Experimental animal studies show that MTA can promote healing towards regeneration. There is now a distinct need to examine the clinical performance of MTA in well-controlled prospective human cohort studies.

Effects of bone morphogenetic protein-6 on periodontal wound healing/regeneration in supraalveolar periodontal defects in dogs.

OBJECTIVE: Application of a synthetic BMP-6 polypeptide in a rat periodontal fenestration defect model enhanced periodontal wound healing/regeneration including new bone and cementum formation. The purpose of this study was to translate the relevance of these initial observations into a discriminating large animal model. METHODS: Critical-size (4-5 mm) supraalveolar periodontal defects were created at the 2(nd) and 3(rd) mandibular premolar teeth in 11 Beagle dogs. Experimental sites received BMP-6 at 0.25, 1.0 and 2.0 mg/ml soak-loaded onto an absorbable collagen sponge (ACS) carrier or ACS alone (control) each condition repeated in four jaw quadrants. The animals were euthanized at 8 weeks when block biopsies were collected and processed for histologic/histometric analysis. RESULTS: BMP-6 at 0.25, 1.0 and 2.0 mg/ml soak-loaded onto the ACS yielded significantly enhanced new bone (0.99 ± 0.07 versus 0.23 ± 0.13 mm/BMP-6 at 0.25 mg/ml) and cementum (2.45 ± 0.54 versus 0.73 ± 0.15 mm/BMP-6 at 0.25 mg/ml) formation including a functionally oriented periodontal ligament compared with control (p < 0.05). A significant inverse linear association between BMP-6 dose and new bone (ß = -0.21 ± 0.09 mm, p = 0.016) and cementum height (ß = -0.34 ± 0.15 mm, p = 0.023) was observed. Minimal root resorption was observed without significant differences between groups. Ankylosis was not observed for any of the experimental groups. CONCLUSIONS: Surgical application of BMP-6/ACS onto critical-size supraalveolar defects enhanced periodontal wound healing/regeneration, in particular cementogenesis including a functionally oriented periodontal ligament; the low BMP-6 0.25 mg/ml concentration apparently providing the most effective dose.

Wound healing of dehiscence defects following different root conditioning modalities: an experimental study in dogs.

OBJECTIVE: The purpose of this study was to investigate the periodontal healing pattern of dehiscence-type defects following different chemical root conditioning modalities. MATERIALS AND METHODS: Buccal osseous dehiscence defects were created on six teeth of seven dogs. After dental plaque accumulation, defects were treated with sterile saline solution (control group) or one chemical conditioning modality: citric acid (CA group), ethylenediaminetetraacetic acid (EDTA group), tetracycline (TTC group), citric acid + tetracycline (CA + TTC group), or tetracycline + citric acid (TTC + CA group). After 3 months of healing, clinical parameters were evaluated, and the animals were killed. Histological sections were processed, and a computer-assisted histometric analysis was used to evaluate the formation of new cementum, new bone, and epithelial apical migration. RESULTS: All treatments yielded significant improvements in terms of probing depth decrease and clinical attachment level gain compared to baseline values; however, without significant differences among the groups (p > 0.05; one-way ANOVA). The highest amount of new cementum was noted in the EDTA group (3.72 ± 0.83 mm, 77.6 %), while the lowest amount of new bone was observed in the TTC group (0.7 ± 0.94 mm, 14.3 %). However, no statistically significant differences could be observed among the groups regarding epithelial apical migration, new cementum, and alveolar bone formation (p > 0.05). CONCLUSION: Chemical root surface conditioning did not promote any significant improvement in periodontal healing pattern of dehiscence-type defects in dogs. CLINICAL RELEVANCE: Chemical root surface conditioning after surgical debridement did not promote positive or negative effects on periodontal healing pattern of dehiscence-type defects.

Synergistic induction of periodontal tissue regeneration by binary application of human osteogenic protein-1 and human transforming growth factor-ß3 in Class II furcation defects of Papio ursinus.

BACKGROUND AND OBJECTIVE: Binary applications of recombinant human osteogenic protein-1 (hOP-1) and transforming growth factor-ß3 (hTGF-ß3) synergize to induce pronounced bone formation. To induce periodontal tissue regeneration, binary applications of hOP-1 and hTGF-ß(3) were implanted in Class II furcation defects of the Chacma baboon, Papio ursinus. MATERIAL AND METHODS: Defects were created bilaterally in the furcation of the first and second mandibular molars of three adult baboons. Single applications of 25 µg hOP-1 and 75 µg hTGF-ß(3) in Matrigel(®) matrix were compared with 20:1 binary applications, i.e. 25 µg hOP-1 and 1.25 µg hTGF-ß(3). Morcellated fragments of autogenous rectus abdominis striated muscle were added to binary applications. Sixty days after implantation, the animals were killed and the operated tissues harvested en bloc. Undecalcified sections were studied by light microscopy, and regenerated tissue was assessed by measuring volume and height of newly formed alveolar bone and cementum. RESULTS: The hOP-1 and hTGF-ß(3) induced periodontal tissue regeneration and cementogenesis. Qualitative morphological analysis of binary applications showed clear evidence for considerable periodontal tissue regeneration. Quantitatively, the differences in the histomorphometric values did not reach statistical significance for the group size chosen for this primate study. The addition of morcellated muscle fragments did not enhance tissue regeneration. Binary applications showed rapid expansion of the newly formed bone against the root surfaces following fibrovascular tissue induction in the centre of the treated defects. CONCLUSION: Binary applications of hOP-1 and hTGF-ß(3) in Matrigel(®) matrix in Class II furcation defects of P. ursinus induced substantial periodontal tissue regeneration, which was tempered, however, by the anatomy of the furcation defect model, which does not allow for the rapid growth and expansion of the synergistic induction of bone formation, particularly when additionally treated with responding myoblastic stem cells.

Enhanced adipogenic differentiation and reduced collagen synthesis induced by human periodontal ligament stem cells might underlie the negative effect of recombinant human bone morphogenetic protein-2 on periodontal regeneration.

BACKGROUND AND OBJECTIVE: Recombinant human bone morphogenetic protein-2 (rhBMP-2) is a potent inducer for the regeneration of mineralized tissue, but has a limited effect on the regeneration of cementum and periodontal ligament (PDL). The aim of the present study was to determine the effects of rhBMP-2 on the in vitro and in vivo biologic activity of well-characterized human PDL stem cells (hPDLSCs) and to elucidate the underlying mechanism of minimal periodontal regeneration by rhBMP-2. MATERIAL AND METHODS: hPDLSCs were isolated and cultured, and then transplanted into an ectopic subcutaneous mouse model using a carrier treated either with or without rhBMP-2. Comprehensive histologic, histometric and immunohistochemical analyses were performed after an 8-wk healing period. The effects of rhBMP-2 on the adipogenic and osteogenic/cementogenic differentiation of hPDLSCs were also evaluated. The effect of rhBMP-2 on both soluble and insoluble collagen synthesis was analyzed, and the expression of mRNA and protein for collagen types I, II, III and V was assessed. RESULTS: In the present study, rhBMP-2 promoted both adipogenic and osteogenic/cementogenic differentiation of hPDLSCs in vitro, and the in vivo potential of hPDLSCs to form mineralized cementum and organized PDL tissue was down-regulated following treatment with rhBMP-2. Collagen synthesis, which plays a crucial role in the regeneration of cementum and the periodontal attachment, was significantly reduced, with associated modification of the relevant mRNA and protein expression profiles. CONCLUSION: In summary, the findings of the present study suggest that enhanced adipogenic differentiation and inhibition of collagen synthesis by hPDLSCs appear to be partly responsible for the minimal effect of rhBMP-2 on cementum and PDL tissue regeneration by hPDLSCs.