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1.
PLoS One ; 17(9): e0267336, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36084086

RESUMO

BACKGROUND AND OBJECTIVE: There is evidence from previous studies that pregnancies and diseases are recorded in the tooth cementum. This study aims to assess the degree of concordance between signals based on irregular incremental lines (ILs) and reported pregnancies. MATERIAL AND METHODS: 23 recent and 24 archaeological human teeth with known birth history were included in this investigation. 129 histological sections of tooth roots were assessed for irregularities in appearance and width using a standardized protocol. Similarity of observed irregularities at the section level allowed us to define signals at the tooth level. The sensitivity of signals to detect pregnancies was determined and related to the signal prevalence. RESULTS: Pregnancy signals were frequently visually observed. However, applying a standardized process we could only reach signal sensitivities to identify pregnancies up to 20 percentage points above chance level. CONCLUSIONS: Based on a standardized and reproducible method it could be confirmed that some pregnancies leave visible signals in the tooth cementum. The results show the potential of the tooth cementum to support reconstruction of life courses in paleopathology. However, it seems that not all pregnancies affect the cementogenesis in such a way that irregular ILs are identifiable. Further research is needed to better understand which type of pregnancies and other conditions are recorded in the tooth cementum.


Assuntos
Cemento Dentário , Dente , Arqueologia , Cementogênese , Feminino , Humanos , Gravidez , Raiz Dentária/patologia
2.
Stem Cell Res Ther ; 13(1): 460, 2022 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-36068594

RESUMO

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.


Assuntos
Cementogênese , beta Catenina , Diferenciação Celular , Células Cultivadas , Cementogênese/fisiologia , Fator de Crescimento do Tecido Conjuntivo/genética , Conexina 43/genética , Cemento Dentário , Humanos , Ligamento Periodontal , RNA Interferente Pequeno , beta Catenina/genética
3.
Chin J Dent Res ; 25(2): 85-92, 2022 Jun 10.
Artigo em Inglês | MEDLINE | ID: mdl-35686588

RESUMO

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.


Assuntos
Cementogênese , Ligamento Periodontal , Cementogênese/fisiologia , Citocinas , Cemento Dentário/fisiologia , Periodonto
4.
J Clin Periodontol ; 49(9): 945-956, 2022 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-35634660

RESUMO

AIM: To date, controversies still exist regarding the exact cellular origin and regulatory mechanisms of periodontium development, which hinders efforts to achieve ideal periodontal tissue regeneration. Axin2-expressing cells in the periodontal ligament (PDL) have been shown to be a novel progenitor cell population that is essential for periodontal homeostasis. In the current study, we aimed to elucidate the regulatory role of bone morphogenetic protein receptor type 1A (BMPR1A)-mediated BMP signalling in Axin2-expressing cells during periodontium development. MATERIALS AND METHODS: Two strains of Axin2 gene reporter mice, Axin2lacZ/+ and Axin2CreERT2/+ ; R26RtdTomato/+ mice, were used. We next generated Axin2CreERT2/+ ; R26RDTA/+ ; R26RtdTomato/+ mice to genetically ablate of Axin2-lineage cells. Axin2CreERT2/+ ; Bmpr1afl/fl ; R26RtdTomato/+ mice were established to conditionally knock out Bmpr1a in Axin2-lineage cells. Multiple approaches, including micro-computed tomography, calcein green, and alizarin red double-labelling, scanning electron microscopy, and histological and immunostaining assays, were used to analyse periodontal phenotypes and molecular mechanisms. RESULTS: X-gal staining revealed that Axin2-expressing cells in the PDL were mainly distributed along the alveolar bone and cementum surface. Cell lineage tracing and cell ablation assays further demonstrated the indispensable role of Axin2-expressing cells in periodontium development. Next, we found that conditional knockout of Bmpr1a in Axin2-lineage cells led to periodontal defects, which were characterized by alveolar bone loss, impaired cementogenesis, and abnormal Sharpey's fibres. CONCLUSIONS: Our findings suggest that Axin2-expressing cells in the PDL are essential for periodontium development, which is regulated by BMP signalling.


Assuntos
Receptores de Proteínas Morfogenéticas Ósseas Tipo I/metabolismo , Ligamento Periodontal , Animais , Proteína Axina/genética , Proteínas Morfogenéticas Ósseas , Cementogênese , Cemento Dentário , Camundongos , Ligamento Periodontal/crescimento & desenvolvimento , Ligamento Periodontal/metabolismo , Periodonto , Transdução de Sinais , Microtomografia por Raio-X
5.
J Dent Res ; 101(9): 1092-1100, 2022 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-35311416

RESUMO

Stem and progenitor cells play important roles in the development and maintenance of teeth and bone. Surface markers expressed in bone marrow-derived mesenchymal stem cells are also expressed in dental tissue-derived stem cells. Mouse skeletal stem cells (mSSCs, CD45-Ter119-Tie2-CD51+Thy-6C3-CD105-CD200+) and human skeletal stem cells (hSSCs, CD45-CD235a-TIE2-CD31-CD146-PDPN+CD73+CD164+) have been identified in bone and shown to play important roles in skeletal development and regeneration. However, it is unclear whether dental tissues also harbor mSSC or hSSC populations. Here, we employed rainbow tracers and found that clonal expansion occurred in mouse dental tissues similar to that in bone. We sorted the mSSC population from mouse periodontal ligament (mPDL) tissue and mouse dental pulp (mDP) tissue in the lower incisors by fluorescence-activated cell sorting (FACS). In addition, we demonstrated that mPDL-derived skeletal stem cells (mPDL-SSCs) and mDP-derived skeletal stem cells (mDP-SSCs) have similar clonogenic capacity, as well as cementogenic and odontogenic potential, but not adipogenic potential, similar to the characteristics of mSSCs. Moreover, we found that the dental tissue-derived mSSC population plays an important role in repairing clipped incisors. Importantly, we sorted the hSSC population from human periodontal ligament (hPDL) and human dental pulp (hDP) tissue in molars and identified its stem cell characteristics. Finally, hPDL-like and hDP-like structures were generated after transplanting hPDL-SSCs and hDP-SSCs beneath the renal capsules. In conclusion, we demonstrated that mouse and human PDL and DP tissues harbor dental stem cells similar to mSSCs and hSSCs, respectively, providing a precise stem cell population for the exploration of dental diseases.


Assuntos
Células-Tronco Mesenquimais , Ligamento Periodontal , Adipogenia , Animais , Diferenciação Celular , Células Cultivadas , Cementogênese , Polpa Dentária , Humanos , Camundongos , Células-Tronco
6.
Int J Mol Sci ; 23(4)2022 Feb 20.
Artigo em Inglês | MEDLINE | ID: mdl-35216454

RESUMO

Spheroids reproduce the tissue structure that is found in vivo more accurately than classic two-dimensional (2D) monolayer cultures. We cultured human periodontal ligament stem cells (HPLSCs) as spheroids that were embedded in collagen gel to examine whether their cementogenic differentiation could be enhanced by treatment with recombinant human plasminogen activator inhibitor-1 (rhPAI-1). The upregulated expression of cementum protein 1 (CEMP1) and cementum attachment protein (CAP), established cementoblast markers, was observed in the 2D monolayer HPLSCs that were treated with rhPAI-1 for 3 weeks compared with that in the control and osteogenic-induction medium groups. In the embedded HPLSC spheroids, rhPAI-1 treatment induced interplay between the spheroids and collagenous extracellular matrix (ECM), indicating that disaggregated HPLSCs migrated and spread into the surrounding ECM 72 h after three-dimensional (3D) culture. Western blot and immunocytochemistry analyses showed that the CEMP1 expression levels were significantly upregulated in the rhPAI-1-treated embedded HPLSC spheroids compared with all the 2D monolayer HPLSCs groups and the 3D spheroid groups. Therefore, 3D collagen-embedded spheroid culture in combination with rhPAI-1 treatment may be useful for facilitating cementogenic differentiation of HPLSCs.


Assuntos
Ligamento Periodontal , Inibidor 1 de Ativador de Plasminogênio , Diferenciação Celular , Células Cultivadas , Cementogênese , Humanos , Inibidor 1 de Ativador de Plasminogênio/genética , Inibidor 1 de Ativador de Plasminogênio/metabolismo , Proteínas/metabolismo , Esferoides Celulares/metabolismo , Células-Tronco/metabolismo
7.
Biochem Biophys Res Commun ; 587: 9-15, 2022 01 08.
Artigo em Inglês | MEDLINE | ID: mdl-34861472

RESUMO

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.


Assuntos
Relógios Biológicos/genética , Calcificação Fisiológica/genética , Cemento Dentário/metabolismo , Membro 1 do Grupo D da Subfamília 1 de Receptores Nucleares/genética , Fosfatase Alcalina/genética , Fosfatase Alcalina/metabolismo , Animais , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Transformada , Proliferação de Células/efeitos dos fármacos , Cementogênese/efeitos dos fármacos , Cementogênese/genética , Cemento Dentário/citologia , Cemento Dentário/efeitos dos fármacos , Feminino , Regulação da Expressão Gênica , Humanos , Sialoproteína de Ligação à Integrina/genética , Sialoproteína de Ligação à Integrina/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Membro 1 do Grupo D da Subfamília 1 de Receptores Nucleares/metabolismo , Osteocalcina/genética , Osteocalcina/metabolismo , Pirrolidinas/farmacologia , Transdução de Sinais , Fator de Transcrição Sp7/genética , Fator de Transcrição Sp7/metabolismo , Tiofenos/farmacologia
8.
Int J Mol Sci ; 22(16)2021 Aug 11.
Artigo em Inglês | MEDLINE | ID: mdl-34445348

RESUMO

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.


Assuntos
Fibroblastos/fisiologia , Histonas/metabolismo , PPAR gama/fisiologia , Ligamento Periodontal/fisiologia , Acetilação , Diferenciação Celular/genética , Células Cultivadas , Cementogênese/genética , Cementogênese/fisiologia , Regulação da Expressão Gênica , Histona Acetiltransferases/metabolismo , Histonas/química , Humanos , Osteogênese/genética , Osteogênese/fisiologia , Ligamento Periodontal/citologia , Processamento de Proteína Pós-Traducional/genética
9.
Int J Mol Sci ; 22(10)2021 May 17.
Artigo em Inglês | MEDLINE | ID: mdl-34067898

RESUMO

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.


Assuntos
Cementogênese/efeitos dos fármacos , Cemento Dentário/metabolismo , Gases em Plasma/farmacologia , Animais , Calcificação Fisiológica/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/genética , Camundongos , Osteocalcina/metabolismo , Osteopontina/metabolismo , Gases em Plasma/metabolismo , Transcriptoma/genética
10.
J Dent Res ; 100(13): 1501-1509, 2021 12.
Artigo em Inglês | MEDLINE | ID: mdl-34009051

RESUMO

The WNT/ß-catenin signaling pathway plays a central role in the biology of the periodontium, yet the function of specific extracellular WNT ligands remains poorly understood. By using a Wnt1-inducible transgenic mouse model targeting Col1a1-expressing alveolar osteoblasts, odontoblasts, and cementoblasts, we demonstrate that the WNT ligand WNT1 is a strong promoter of cementum and alveolar bone formation in vivo. We induced Wnt1 expression for 1, 3, or 9 wk in Wnt1Tg mice and analyzed them at the age of 6 wk and 12 wk. Micro-computed tomography (CT) analyses of the mandibles revealed a 1.8-fold increased bone volume after 1 and 3 wk of Wnt1 expression and a 3-fold increased bone volume after 9 wk of Wnt1 expression compared to controls. In addition, the alveolar ridges were higher in Wnt1Tg mice as compared to controls. Nondecalcified histology demonstrated increased acellular cementum thickness and cellular cementum volume after 3 and 9 wk of Wnt1 expression. However, 9 wk of Wnt1 expression was also associated with periodontal breakdown and ectopic mineralization of the pulp. The composition of this ectopic matrix was comparable to those of cellular cementum as demonstrated by quantitative backscattered electron imaging and immunohistochemistry for noncollagenous proteins. Our analyses of 52-wk-old mice after 9 wk of Wnt1 expression revealed that Wnt1 expression affects mandibular bone and growing incisors but not molar teeth, indicating that Wnt1 influences only growing tissues. To further investigate the effect of Wnt1 on cementoblasts, we stably transfected the cementoblast cell line (OCCM-30) with a vector expressing Wnt1-HA and performed proliferation as well as differentiation experiments. These experiments demonstrated that Wnt1 promotes proliferation but not differentiation of cementoblasts. Taken together, our findings identify, for the first time, Wnt1 as a critical regulator of alveolar bone and cementum formation, as well as provide important insights for harnessing the WNT signal pathway in regenerative dentistry.


Assuntos
Cementogênese , Cemento Dentário , Animais , Camundongos , Osteogênese , Ligamento Periodontal , Microtomografia por Raio-X
11.
J Dent Res ; 100(11): 1289-1298, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-33853427

RESUMO

Cementum, a specialized bony layer covering an entire molar root surface, anchors teeth into alveolar bone. Gli1, a key transcriptional activator in Hedgehog signaling, has been identified as a mesenchymal progenitor cell marker in various tissues, including the periodontal ligament (PDL). To address the mechanisms by which Gli1+ progenitor cells contribute to cementogenesis, we used the Gli1lacZ/+ knock-in line to mark Gli1+ progenitors and the Gli1CreERT2/+; R26RtdTomato/+ line (named Gli1Lin) to trace Gli1 progeny cells during cementogenesis. Our data unexpectedly displayed a biphasic feature of Gli1+ PDL progenitor cells and cementum growth: a negative relationship between Gli1+ progenitor cell number and cementogenesis but a positive correlation between Gli1-derived acellular and cellular cementoblast cell number and cementum growth. DTA-ablation of Gli1Lin cells led to a cementum hypoplasia, including a significant reduction of both acellular and cellular cementoblast cells. Gain-of-function studies (by constitutive stabilization of ß-catenin in Gli1Lin cells) revealed a cementum hyperplasia. A loss of function (by conditional deletion of ß-catenin in Gli1+ cells) resulted in a reduction of postnatal cementum growth. Together, our studies support a vital role of Gli1+ progenitor cells in contribution to both types of cementum, in which canonical Wnt/ß-catenin signaling positively regulates the differentiation of Gli1+ progenitors to cementoblasts during cementogenesis.


Assuntos
Cementogênese , beta Catenina , Animais , Cemento Dentário/metabolismo , Proteínas Hedgehog , Camundongos Transgênicos , Via de Sinalização Wnt , Proteína GLI1 em Dedos de Zinco , beta Catenina/metabolismo
12.
J Cell Physiol ; 236(3): 2070-2086, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-32740946

RESUMO

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.


Assuntos
Cementogênese/efeitos dos fármacos , Efrina-B2/metabolismo , Hormônio Paratireóideo/farmacologia , Receptor EphB4/metabolismo , Transdução de Sinais , Animais , Linhagem Celular , Cemento Dentário/efeitos dos fármacos , Cemento Dentário/metabolismo , Masculino , Camundongos , Modelos Biológicos , Hormônio Paratireóideo/administração & dosagem , Ratos Wistar , Regeneração/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Tomografia Computadorizada por Raios X , Raiz Dentária/diagnóstico por imagem , Raiz Dentária/efeitos dos fármacos
13.
Genomics ; 113(1 Pt 1): 217-228, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33309767

RESUMO

The aim of this study was to explore the involvement of long noncoding RNAs (lncRNAs) during intermittent parathyroid hormone (PTH) induced cementogenesis. Expression profiles of lncRNAs and mRNAs were obtained using high-throughput microarray. Gene Ontology enrichment analysis, Kyoto Encyclopedia of Genes and Genomes pathway analysis, and coding-noncoding gene coexpression networks construction were performed. We identified 190 lncRNAs and 135 mRNAs that were differentially expressed during intermittent PTH-induced cementogenesis. In this process, the Wnt signaling pathway was negatively regulated, and eight lncRNAs were identified as possible core regulators of Wnt signaling. Based on the results of microarrray analysis, we further verified the repressed expression of Wnt signaling crucial components ß-catenin, APC and Axin2. Above all, we speculated that lncRNAs may play important roles in PTH-induced cementogenesis via the negative regulation of Wnt pathway.


Assuntos
Cementogênese , Hormônio Paratireóideo/metabolismo , RNA Longo não Codificante/metabolismo , Via de Sinalização Wnt , Proteína da Polipose Adenomatosa do Colo , Animais , Proteína Axina/genética , Proteína Axina/metabolismo , Linhagem Celular , Cemento Dentário/metabolismo , Camundongos , Osteoblastos/metabolismo , RNA Longo não Codificante/genética , Transcriptoma , beta Catenina/genética , beta Catenina/metabolismo
14.
J Cell Mol Med ; 24(14): 7939-7948, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32510818

RESUMO

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.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Calcificação Fisiológica/efeitos dos fármacos , Proteínas de Ciclo Celular/metabolismo , Cementogênese , NF-kappa B/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fator de Necrose Tumoral alfa/farmacologia , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Proteínas de Ciclo Celular/genética , Linhagem Celular , Cementogênese/efeitos dos fármacos , Citocinas/metabolismo , Imunofluorescência , Expressão Gênica , Técnicas de Silenciamento de Genes , Mediadores da Inflamação/metabolismo , Camundongos
15.
Bone ; 136: 115329, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32224162

RESUMO

Pyrophosphate (PPi) serves as a potent and physiologically important regulator of mineralization, with systemic and local concentrations determined by several key regulators, including: tissue-nonspecific alkaline phosphatase (ALPL gene; TNAP protein), the progressive ankylosis protein (ANKH; ANK), and ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1; ENPP1). Results to date have indicated important roles for PPi in cementum formation, and we addressed several gaps in knowledge by employing genetically edited mouse models where PPi metabolism was disrupted and pharmacologically modulating PPi in a PPi-deficient mouse model. We demonstrate that acellular cementum growth is inversely proportional to PPi levels, with reduced cementum in Alpl KO (increased PPi levels) mice and excess cementum in Ank KO mice (decreased PPi levels). Moreover, simultaneous ablation of Alpl and Ank results in reestablishment of functional cementum in dKO mice. Additional reduction of PPi by dual deletion of Ank and Enpp1 does not further increase cementogenesis, and PDL space is maintained in part through bone modeling/remodeling by osteoclasts. Our results provide insights into cementum formation and expand our knowledge of how PPi regulates cementum. We also demonstrate for the first time that pharmacologic manipulation of PPi through an ENPP1-Fc fusion protein can regulate cementum growth, supporting therapeutic interventions targeting PPi metabolism.


Assuntos
Cementogênese , Difosfatos , Animais , Cemento Dentário , Camundongos , Osteoclastos
16.
Arch Oral Biol ; 112: 104663, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31986333

RESUMO

OBJECTIVE: This study aims to uncover the role of interleukin-33 on cementoblast-mediated cementum repair. METHODS: 6-8-week-old C57BL/6 mice were used to establish the model of orthodontic tooth movement. Interleukin-33 and suppression of tumorigenicity2 (ST2) expressions were immunohistochemically detected in the periodontal tissue. In vitro, cementoblast-like (OCCM-30) cells were cultured in the presence of recombinant mouse interleukin-33 protein (rmIL-33) at a 1-14 d time frame. ST2 expressions were immunofluorescently labeled and quantitatively examined. The effects of interleukin-33 on cementoblast differentiation, mineralization and proliferation were examined by alkaline phosphatase, alizarin red staining and cell counting kit-8, respectively. To further clarify the effect of interleukin-33 on cementogenesis-related protein expressions, runt-related transcription factor 2 (RUNX2), osterix, osteopontin, bone sialoprotein(BSP), osteocalcin, osteoprotegerin (OPG) and receptor activator of NF-КB ligand (RANKL) expressions were examined by western blot. RESULTS: Orthodontic load of high magnitude induces external apical root resorption, and increases interleukin-33 expression in the periodontal tissue of mice. Cells in the cementum express ST2. Interleukin-33 initially down-regulates but later recovers ST2 mRNA and protein levels in OCCM-30 cells. Interleukin-33 abates cementoblast differentiation and mineralization, and suppresses RUNX2, osterix, BSP and osteopontin expressions in OCCM-30 cells at the later stage of the culture period. Interleukin-33 enhances RANKL expression, and reduces the ratio of OPG/RANKL in OCCM-30 cells. CONCLUSION: Orthodontic load of high magnitude induces interleukin-33 expression in the periodontal tissue. Interleukin-33 has a negative effect on cementogenesis via suppressing cementoblast differentiation, mineralization and cementogenesis-related protein expressions.


Assuntos
Cementogênese , Cemento Dentário/citologia , Interleucina-33/metabolismo , Técnicas de Movimentação Dentária , Animais , Diferenciação Celular , Células Cultivadas , Cemento Dentário/metabolismo , Proteína 1 Semelhante a Receptor de Interleucina-1/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Recombinantes
17.
J Cell Physiol ; 235(5): 4545-4558, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-31621902

RESUMO

Amelogenin isoforms, including full-length amelogenin (AMEL) and leucine-rich amelogenin peptide (LRAP), are major components of the enamel matrix, and are considered as signaling molecules in epithelial-mesenchymal interactions regulating tooth development and periodontal regeneration. Nevertheless, the molecular mechanisms involved are still poorly understood. The aim of the present study was to identify novel binding partners for amelogenin isoforms in the cementoblast (OCCM-30), using an affinity purification assay (GST pull-down) followed by mass spectrometry and immunoblotting. Protein-protein interaction analysis for AMEL and LRAP evidenced the plasminogen activation system (PAS) as a potential player regulating OCCM-30 response to amelogenin isoforms. For functional assays, PAS was either activated (plasmin) or inhibited (ε-aminocaproic acid [aminocaproic]) in OCCM-30 cells and the cell morphology, mineral nodule formation, and gene expression were assessed. PAS inhibition (EACA 100 mM) dramatically decreased mineral nodule formation and expression of OCCM-30 differentiation markers, including osteocalcin (Bglap), bone sialoprotein (Ibsp), osteopontin (Spp1), tissue-nonspecific alkaline phosphatase (Alpl) and collagen type I (Col1a1), and had no effect on runt-related transcription factor 2 (Runx2) and Osterix (Osx) mRNA levels. PAS activation (plasmin 5 µg/µl) significantly increased Col1a1 and decreased Bglap mRNA levels (p < .05). Together, our findings shed new light on the potential role of plasminogen signaling pathway in the control of the amelogenin isoform-mediated response in cementoblasts and provide new insights into the development of targeted therapies.


Assuntos
Amelogenina/metabolismo , Diferenciação Celular , Cementogênese , Cemento Dentário/metabolismo , Proteínas do Esmalte Dentário/metabolismo , Plasminogênio/metabolismo , Amelogenina/genética , Animais , Linhagem Celular , Ativação Enzimática , Regulação da Expressão Gênica , Redes Reguladoras de Genes , Camundongos , Ligação Proteica , Mapas de Interação de Proteínas , Transdução de Sinais
18.
J Periodontol ; 91(1): 110-119, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31347702

RESUMO

BACKGROUND: The periodontal ligament (PDL) comprises cells embedded in the extracellular matrix (ECM) and is required for periodontal healing upon tooth transplantation. However, the extent to which the ECM contributes to periodontal regeneration remains unknown. This study aimed to evaluate effects of separate PDL matrix without cells on periodontal healing. METHODS: Mandibular premolars extracted from six beagle dogs, were 1) left untreated, 2) decellularized, or 3) surface planed. Cytologic, histologic, scanning electron microscopy, and transmission electron microscopy analyses confirmed that detergents could be used to decellularize the PDL. An additional circumferential osteotomy was performed to replant dental roots into extraction sockets. Radiography and histology were used to evaluate periodontal regeneration 8 weeks later, with the data adjusted for multiple testing. RESULTS: In pristine extraction sites, total root resorption (P = 0.034), recovered PDL space (P = 0.012), and new cementum (P = 0.004) were greater in untreated teeth than in roots that underwent surface planing. There were no significant changes when comparing untreated teeth with teeth having a decellularized PDL (P = 0.081, P = 0.170, and P = 1.000, respectively), and decellularized teeth showed significant increase of new cementum compared to surface planed teeth (P = 0.048). In the defect area, only the recovered PDL space (P = 0.034) was greater in untreated teeth when comparing with denuded roots. CONCLUSION: These results suggest that in addition to untreated PDL, decellularized PDL also partially supports reattachment (particularly cementogenesis) in pristine extraction sites but not in defect areas.


Assuntos
Ligamento Periodontal , Reimplante Dentário , Animais , Cementogênese , Cemento Dentário , Cães , Raiz Dentária , Cicatrização
19.
J Dent ; 92: 103259, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31809792

RESUMO

OBJECTIVE: Human periodontal ligament stem cells (hPDLSCs) are promising for periodontal regeneration. However, to date, there has been no report of hPDLSC differentiation into the fibrogenic lineage. There has been no report demonstrating hPDLSC differentiation into all three (osteogenic, fibrogenic and cementogenic fibrogenic) lineages in the same report. The objectives of this study were to harvest hPDLSCs from the periodontal ligaments (PDL) of the extracted human teeth, and use the same vial of hPDLSCs to differentiate into all three (osteogenic, fibrogenic and cementogenic) lineages for the first time. METHODS: hPDLSCs were harvested from PDL tissues of the extracted premolars. The ability of hPDLSCs to form bone, cementum and collagen fibers was tested in culture mediums. Gene expressions were analyzed using quantitative real-time polymerase chain reaction (qRT-PCR). Immunofluorescence, alizarin red (ARS), Xylenol orange, picro sirius red staining (PSRS), alcian blue staining (ABS) and alkaline phosphatase (ALP) staining were evaluated. RESULTS: In osteogenic medium, hPDLSCs had high expressions of osteogenic genes (RUNX2, ALP, OPN and COL1) at 14 and 21 days (15-20 folds of that of control), and produced mineral nodules and ALP activity (5 and 10 folds those of the control). hPDLSCs in fibrogenic medium expressed high levels of PDL fibrogenic genes (COL1, COL3, FSP-1, PLAP-1 and Elastin) at 28 days (20-70 folds of control). They were stained strongly with F-actin and fibronection, and secreted PDL collagen fibers (5 folds of control). hPDLSCs in cementogenic medium showed high expressions of cementum genes (CAP, CEMP1 and BSP) at 21 days (10-15 folds of control) and synthesized mineralized cementum (50 folds via ABS, and 40 folds via ALP staining, compared to those of control). CONCLUSIONS: hPDLSCs differentiated into bone-, fiber- and cementum-forming cells, with potential for regeneration of periodontium to form the bone-PDL-cementum complex.


Assuntos
Cementogênese , Osteogênese , Ligamento Periodontal , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Humanos , Proteínas , Regeneração , Células-Tronco
20.
J Cell Biochem ; 121(3): 2606-2617, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31680324

RESUMO

Cementum regeneration is considered the gold standard for the treatment of periodontitis. As one of the most important primary proinflammatory cytokines, interleukin 1ß (IL1ß) plays an essential role during the early stage of periodontitis and its amounts simultaneously increase dramatically during this stage. Though promising, the differentiation of cementoblasts upon IL1ß-induced inflammation of the microenvironment and the relative interaction mechanism are still unknown. Here, we found that IL1ß inhibited cementoblast differentiation and microRNA-325-3p (miR-325-3p) was increased during IL1ß-stimulated cementoblasts. Bioinformatics analysis and luciferase reporter assay demonstrated miR-325-3p targeted runt-related transcription factor 2 directly. Transfection of miR-325-3p suppressed cementoblast differentiation in vitro and the formation of cementum-like tissues in vivo. The inhibitor of miR-325-3p could mitigate the above effects induced by IL1ß. Accordingly, our finding suggests a critical role of miR-325-3p in linking inflammation to impaired cementum regeneration and provides a potential possibility for applying miR-325-3p inhibitors in the treatment of periodontitis-related bone loss.


Assuntos
Diferenciação Celular , Cementogênese , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Cemento Dentário/citologia , Regulação da Expressão Gênica , Interleucina-1beta/farmacologia , MicroRNAs/genética , Animais , Proliferação de Células , Células Cultivadas , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Cemento Dentário/efeitos dos fármacos , Cemento Dentário/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL
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