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1.
Stem Cells ; 42(2): 116-127, 2024 Feb 08.
Artigo em Inglês | MEDLINE | ID: mdl-37952104

RESUMO

Human dental pulp stem cells (hDPSCs) play a vital role in the regeneration of the pulp-dentin complex after pulp disease. While the regeneration efficiency relies on the odontoblastic differentiation capacity of hDPSCs, this is difficult to regulate within the pulp cavity. Although nicotinamide riboside (NR) has been found to promote tissue regeneration, its specific role in pulp-dentin complex regeneration is not fully understood. Here, we aimed to explore the role of NR in the odontoblastic differentiation of hDPSCs and its underlying molecular mechanism. It was found that NR enhanced the viability and retarded senescence in hDPSCs with higher NAD+/NADH levels. In contrast to the sustained action of NR, the multi-directional differentiation of hDPSCs was enhanced after NR pre-treatment. Moreover, in an ectopic pulp regeneration assay in nude mice, transplantation of hDPSCs pretreated with NR promoted the formation of a dentin-like structure surrounded by cells positively expressing DMP-1 and DSPP. RNA-Seq demonstrated inhibition of the HIF-1 signaling pathway in hDPSCs pretreated with NR. The number of HIF-1α-positive cells was significantly decreased in hDPSCs pretreated by NR in vivo. Similarly, NR significantly downregulated the expression of HIF-1α in vitro. The findings suggested that NR could potentially regulate hDPSC odontoblastic differentiation and promote the development of innovative strategies for dental pulp repair.


Assuntos
Polpa Dentária , Niacinamida , Odontoblastos , Compostos de Piridínio , Animais , Humanos , Camundongos , Diferenciação Celular , Células Cultivadas , Camundongos Nus , Niacinamida/análogos & derivados , Regeneração , Transdução de Sinais , Células-Tronco/metabolismo
2.
Cell Signal ; 73: 109695, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32565162

RESUMO

Illuminating the mechanisms of odontoblast differentiation of human dental pulp stem cells (hDPSCs) is the key to find therapeutic clues to promote odontogenesis. LncRNAs play a regulatory role in odontoblast differentiation. Here, we identified a novel lncRNA, named lncRNA CALB2. It was up-regulated in odontoblast-differentiated hDPSCs and potentially interacted with miR-30b-3p and RUNX2. Via gain- and loss-of-function approaches, we found lncRNA CALB2 significantly promoted the odontoblast differentiation of hDPSCs. Then, dual luciferase reporter assay and RNA immunoprecipitation assay revealed that both lncRNA CALB2 and RUNX2 mRNA could directly bind to miR-30b-3p via the same binding sites. Interestingly, miR-30b-3p in hDPSCs was down-regulated and RUNX2 was up-regulated during odontoblast differentiation. Moreover, lncRNA CALB2 knockdown significantly reduced the protein level of RUNX2, DSPP and DMP-1, while miR-30b-3p inhibitor rescued the reduction. Furthermore, miR-30b-3p exerted an inhibitory effect on odontoblast differentiation, which could be reversed by lncRNA CALB2. Collectively, these findings indicate that the newly identified lncRNA CALB2 acts as a miR-30b-3p sponge to regulate RUNX2 expression, thus promoting the odontoblast differentiation of hDPSCs. LncRNA CALB2/miR-30b-3p/RUNX2 axis could be a novel therapeutic target for accelerating odontogenesis.


Assuntos
Calbindina 2/fisiologia , Polpa Dentária , MicroRNAs/fisiologia , Odontoblastos , Odontogênese , Células-Tronco , Diferenciação Celular , Células Cultivadas , Polpa Dentária/citologia , Polpa Dentária/metabolismo , Humanos , Odontoblastos/citologia , Odontoblastos/metabolismo , Células-Tronco/citologia , Células-Tronco/metabolismo
3.
Arch Oral Biol ; 117: 104794, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32553945

RESUMO

OBJECTIVE: This study aims to investigate the expression pattern of nucleotide-binding oligomerization domain-like receptor family pyrin domain containing 6 (NLRP6) in human dental pulp tissues and cells, and roughly explore the role of NLRP6 in dental pulp immunity. METHODS: Immunohistochemistry and immunofluorescence double staining were performed to determine the expression and localization of NLRP6 in healthy and inflamed pulp tissues. The expression of NLRP6 in human dental pulp cells (HDPCs) was investigated by immunocytofluorescence. Furthermore, reverse transcription polymerase chain reaction (RT-PCR) and western blot were used to evaluate the impact of lipopolysaccharide (LPS) stimulation on NLRP6 expression in HDPCs. Last, NLRP6 gene was silenced by lentiviral short hairpin RNA to explore the impact of NLRP6 on LPS-induced interleukin (IL)-1ß. RESULTS: NLRP6 was predominantly expressed in odontoblasts layer and blood vessels of healthy dental pulp, as well as infiltrated immune cells and fibroblasts of inflamed pulp. Further immunofluorescence double staining showed that pericytes and endothelial cells in the dental pulp blood vessels, macrophages and T cells as well as fibroblasts in the inflamed pulp expressed NLRP6. NLRP6 was also basically expressed in cultured HDPCs and upregulated by LPS stimulation. Knockdown of NLRP6 in HDPCs significantly inhibited the LPS-induced IL-1ß expression. CONCLUSIONS: Our study revealed the expression and distribution of NLRP6 in human dental pulp tissues. Furthermore, NLRP6 was also basically expressed in cultured HDPCs, which could be upregulated by LPS stimulation, indicating the involvement of NLRP6 in dental pulp immune response.


Assuntos
Polpa Dentária/metabolismo , Células Endoteliais/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Odontoblastos/metabolismo , Células Cultivadas , Polpa Dentária/imunologia , Humanos , Lipopolissacarídeos/farmacologia
4.
ACS Biomater Sci Eng ; 5(7): 3561-3571, 2019 Jul 08.
Artigo em Inglês | MEDLINE | ID: mdl-33405738

RESUMO

Exosomes are important mediators of intercellular communication and have a vital part in the diagnosis and treatment of various diseases in humans. Here, we investigated the benefits and underlying mechanism of exosomes secreted via stem cells from human exfoliated deciduous teeth (SHED-derived exosomes) in promoting alveolar bone regeneration, thus providing new insights into exosome-based therapy for periodontitis. SHED-derived exosomes were isolated by ultracentrifugation. The impacts of SHED-derived exosomes on the angiogenic ability of human umbilical vein endothelial cells (HUVECs) and the osteogenic capability of rat bone marrow mesenchymal stem cells (BMSCs) were evaluated in vitro. Compound C, a pharmacological blocker of adenosine monophosphate-activated protein kinase (AMPK), was used to examine the role of the AMPK signaling cascade in these processes. Periodontal defect rat models were established and treated with PBS, ß-tricalcium phosphate (ß-TCP), or a grouping of exosomes/ß-TCP. Microcomputed tomography (micro-CT) scanning, hematoxylin and eosin (HE) staining, Masson staining, and immunofluorescence staining were done to inspect the impacts of the exosomes/ß-TCP combination on periodontal bone regeneration. Our outcomes indicated that the expression of angiogenesis-related genes (KDR, SDF-1, and FGF2), osteogenesis-related genes (COL1, RUNX2, and OPN), and phosphorylated (p)-AMPK were upregulated after treatment with exosomes, while the positive impacts of SHED-derived exosomes on HUVECs and BMSCs were partially reversed by compound C. Micro-CT analysis demonstrated that the exosomes/ß-TCP group exhibited better bone regeneration than either the ß-TCP group or the control group. Additionally, the results of HE and Masson staining as well as immunofluorescence staining showed neovascularization and new bone formation in the exosomes/ß-TCP group but only limited new bone formation in the other two groups. Thus, SHED-derived exosomes contribute to periodontal bone regeneration by promoting neovascularization and new bone formation, possibly through the AMPK signaling pathway.

5.
Inflammation ; 41(2): 409-417, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29178062

RESUMO

The AIM2 inflammasome pathway has been determined to play an important role in cellular immune defense against bacterial and viral infections; however, its function and regulatory mechanism in human dental pulp cells (HDPCs) during pulpitis remains poorly understood. In this study, we explored whether the AIM2 inflammasome pathway was activated in HDPCs in response to dsDNA and defined its role in regulating IL-1ß secretion. We demonstrated that stimulation with IFN-γ and cytoplasmic DNA significantly activated the AIM2 inflammasome and increased IL-1ß secretion in HDPCs. Moreover, AIM2 overexpression significantly up-regulated both cleaved Caspase-1 expression and IL-1ß release in HDPCs, while suppression of ASC and Caspase-1 resulted in down-regulation of cleaved Caspase-1 and IL-1ß secretion. These results suggest that Caspase-1-dependent IL-1ß processing and secretion require the AIM2 inflammasome pathway in HDPCs and that the AIM2 inflammasome pathway is critical for regulation of the dental pulp immune response.


Assuntos
Proteínas de Ligação a DNA/metabolismo , DNA/fisiologia , Polpa Dentária/metabolismo , Inflamassomos/metabolismo , Interleucina-1beta/metabolismo , Caspase 1 , Células Cultivadas , Proteínas de Ligação a DNA/fisiologia , Polpa Dentária/citologia , Polpa Dentária/imunologia , Humanos , Inflamassomos/fisiologia , Pulpite
6.
Exp Ther Med ; 14(1): 73-78, 2017 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-28672895

RESUMO

Pulp capping is a restorative technique employed in an attempt to maintain pulpal vitality and generate reparative dentin. Ca2+ released from capping materials is suggested to promote reparative dentin formation. Transient receptor potential channel 6 (TRPC6) is a receptor-operated Ca2+ channel that serves an important role in Ca2+ influx in the majority of non-excitable cells, and influences the calcium signaling and cell respond. Therefore, the purpose of the present study was to gain an insight into the role of TRPC6 in the odontoblastic differentiation of human dental pulp cells (HDPCs). Human dental pulp tissues and HDPCs were obtained from healthy third molars. By immunohistochemical staining, TRPC6 was observed to be highly expressed in the dental pulp tissue, particularly in the odontoblast layer. In addition, the protein level of TRPC6 was increased in a time-dependent manner during odontogenic differentiation of HDPCs. Downregulation of TRPC6 by a lentivirus vector containing TRPC6 shRNA inhibited the process of odontogenic differentiation in HDPCs. In conclusion, the current data demonstrated that TRPC6 served a significant role in the odontogenic differentiation of HDPCs, suggesting it may be a promising therapeutic target in regenerative endodontics.

7.
J Endod ; 43(2): 315-320, 2017 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-28041683

RESUMO

INTRODUCTION: Calcium ions (Ca2+) actively participate in reparative dentin formation by promoting cellular proliferation and differentiation of human dental pulp cells (hDPCs). Transient receptor potential cation channel, subfamily C, member 1 (TRPC1) activates Ca2+ entry upon store depletion in a variety of cell types. However, the function of TRPC1 in hDPCs has not been reported. Therefore, we aimed to analyze the role of TRPC1 in hDPCs undergoing odontoblast-like differentiation. METHODS: Immunohistochemical staining was used to determine the distribution of TRPC1 in pulp tissues. Western blot analysis was used to detect the protein level of TRPC1 in the odontoblast-like differentiation of hDPCs. Knockdown of TRPC1 was performed with an adenoviral vector to evaluate the role of TRPC1 in hDPCs during odontoblast-like differentiation. RESULTS: The results showed that TRPC1 was highly expressed in the cytoplasm of dental pulp cells, especially in the odontoblast layer of the healthy pulp. Moreover, the protein level of TRPC1 increased in a time-dependent manner during the odontoblast-like differentiation of hDPCs. Importantly, knockdown of TRPC1 attenuated the process of odontoblast-like differentiation as indicated by the reduction in mineralized nodules and the down-regulation of dentin sialophosphoprotein and dentin matrix protein 1. Moreover, knockdown of TRPC1 decreased Ca2+ entry to the cytoplasm of hDPCs. CONCLUSIONS: Our data indicated a pivotal role of TRPC1 in the odontoblastlike differentiation of hDPCs, which may be a therapeutic target to enhance reparative dentin formation.


Assuntos
Diferenciação Celular/fisiologia , Polpa Dentária/citologia , Odontoblastos/fisiologia , Canais de Cátion TRPC/fisiologia , Adolescente , Adulto , Western Blotting , Cálcio/metabolismo , Células Cultivadas , Polpa Dentária/fisiologia , Humanos , Reação em Cadeia da Polimerase em Tempo Real , Adulto Jovem
8.
Cell Tissue Res ; 364(2): 309-18, 2016 05.
Artigo em Inglês | MEDLINE | ID: mdl-26646542

RESUMO

Long noncoding RNAs (lncRNAs) have recently emerged as an important class of regulatory molecules in diverse biological processes, although lncRNA involvement in the odontoblast-like differentiation of human dental pulp cells (hDPCs) is poorly understood. We investigate the expression of lncRNAs in this differentiation and explore their underlying role and the involved mechanism. Integrated comparative lncRNA microarray profiling was used to examine lncRNA expression during this differentiation. The differential expression of lncRNAs was validated by quantitative real-time reverse transcription plus the polymerase chain reaction. Differential lncRNA overexpression was performed with an adenoviral vector and the role and mechanism was then investigated in odontoblast-like differentiation. We identified 139 differentially expressed lncRNAs during this differentiation. Among them, five lncRNAs differentially expressed in microarray analysis were validated. Notably, lncRNA DANCR expression was significantly downregulated during hDPC differentiation to odontoblast-like cells in a time-dependent manner. Moreover, lncRNA DANCR overexpression blocked mineralized nodule formation and the expression of DSPP and DMP-1 in hDPCs after 14 days of odontogenic induction. Importantly, the upregulation of DANCR significantly decreased the expression levels of p-GSK-3ß and ß-catenin expression indicating that lncRNA DANCR can inhibit the activation of the Wnt/ß-catenin signal pathway during the odontoblast-like differentiation of hDPCs. Thus, the modulation of Wnt/ß-catenin signaling by lncRNA DANCR represents a potential therapeutic option for reparative dentin formation and regenerative endodontics.


Assuntos
Diferenciação Celular/genética , Polpa Dentária/citologia , Odontoblastos/citologia , Odontogênese/genética , RNA Longo não Codificante/genética , Proteínas Wnt/antagonistas & inibidores , Via de Sinalização Wnt/genética , beta Catenina/antagonistas & inibidores , Células Cultivadas , Dentina/metabolismo , Glicogênio Sintase Quinase 3 beta/biossíntese , Humanos , beta Catenina/biossíntese
9.
Inflammation ; 38(2): 566-75, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24986444

RESUMO

The inflammasome has been determined to play an important role in inflammatory diseases in recent years. Absent in melanoma 2 (AIM2), an inflammasome that recognizes cytoplasmic DNA, has recently been identified as a critical regulator of immune responses. In this study, we explored whether AIM2 was expressed in human dental pulp and defined the role of AIM2 in regulating interleukin (IL)-1ß secretion. We demonstrated that AIM2 was only detected in the odontoblast layer of healthy dental pulp, whereas strong expression was observed in inflamed dental pulp. Stimulation with interferon gamma (IFN-γ) and cytoplasmic DNA significantly activated the AIM2 inflammasome and increased IL-1ß secretion in human dental pulp cells (HDPCs) in a time- and dose-dependent manner. Moreover, the knockdown of AIM2 downregulated both cleaved-caspase-1 expression and IL-1ß release in HDPCs. These results suggest that AIM2 expressed in human dental pulp plays an important role in the immune defense by activating the inflammasome signaling pathway.


Assuntos
Proteínas de Ligação a DNA/genética , DNA/metabolismo , Polpa Dentária/metabolismo , Inflamassomos/imunologia , Interleucina-1beta/metabolismo , Adolescente , Adulto , Caspase 1/biossíntese , Células Cultivadas , DNA/genética , Proteínas de Ligação a DNA/biossíntese , Polpa Dentária/citologia , Humanos , Inflamação/imunologia , Interferon gama/farmacologia , Dente Serotino/metabolismo , Odontoblastos/citologia , Odontoblastos/metabolismo , Interferência de RNA , RNA Interferente Pequeno , Transdução de Sinais/imunologia , Adulto Jovem
10.
Connect Tissue Res ; 55(3): 217-24, 2014 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-24409810

RESUMO

Bone morphogenetic protein-2 (BMP-2) is a multi-functional growth factor belonging to the transforming growth factor ß superfamily that has a broad range of activities that affect many different cell types. BMP-2 induces odontoblastic differentiation of human dental pulp cells (DPCs), but the underlying mechanism remains unclear. In this study, we investigated the potential role of the JNK mitogen-activated protein kinases (MAPK) pathway in BMP-2-induced odontoblastic differentiation of DPCs. The levels of phosphorylated and unphosphorylated JNK MAPK were quantified by Western blot analysis following treatment with BMP-2 and the JNK inhibitor SP600125. The role of JNK MAPK in the BMP-2-induced odontoblastic differentiation of DPCs was determined by measuring alkaline phosphatase (ALP) activity and by examining the expression of odontoblastic markers using quantitative real-time polymerase chain reaction analysis. The effect of JNK MAPK silencing on odontoblastic differentiation was also investigated. BMP-2 upregulated the phosphorylation of JNK in DPCs in a dose- and time-dependent manner. Early markers of odontoblastic differentiation, including ALP activity, osteopontin and dentin matrix protein-1, were not inhibited by the JNK inhibitor. However, the JNK inhibitor, SP600125, significantly inhibited late-stage differentiation of odontoblasts, including the gene expression of osteocalcin, dentin sialophosphoprotein and bone sialoprotein, and also reduced the formation of mineralized nodules in BMP-2-treated DPCs. Consistent with this observation, silencing of JNK MAPK also decreased late-stage odontoblastic differentiation. Taken together, these findings suggest that JNK activity is required for late-stage odontoblastic differentiation induced by BMP-2.


Assuntos
Proteína Morfogenética Óssea 2/metabolismo , Diferenciação Celular/fisiologia , Polpa Dentária/citologia , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Odontoblastos/citologia , Polpa Dentária/metabolismo , Humanos , Odontoblastos/metabolismo , Transdução de Sinais/fisiologia , Fator de Crescimento Transformador beta/metabolismo
11.
PLoS One ; 8(10): e77058, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-24204734

RESUMO

BACKGROUND: The origin of hadrosaurid dinosaurs is far from clear, mainly due to the paucity of their early Late Cretaceous close relatives. Compared to numerous Early Cretaceous basal hadrosauroids, which are mainly from Eastern Asia, only six early Late Cretaceous (pre-Campanian) basal hadrosauroids have been found: three from Asia and three from North America. METHODOLOGY/PRINCIPAL FINDINGS: Here we describe a new hadrosauroid dinosaur, Yunganglong datongensis gen. et sp. nov., from the early Late Cretaceous Zhumapu Formation of Shanxi Province in northern China. The new taxon is represented by an associated but disarticulated partial adult skeleton including the caudodorsal part of the skull. Cladistic analysis and comparative studies show that Yunganglong represents one of the most basal Late Cretaceous hadrosauroids and is diagnosed by a unique combination of features in its skull and femur. CONCLUSIONS/SIGNIFICANCE: The discovery of Yunganglong adds another record of basal Hadrosauroidea in the early Late Cretaceous, and helps to elucidate the origin and evolution of Hadrosauridae.


Assuntos
Osso e Ossos/anatomia & histologia , Dinossauros/anatomia & histologia , Fósseis , Esqueleto , Crânio/anatomia & histologia , Animais , Evolução Biológica , China , Dinossauros/classificação
12.
J Endod ; 39(5): 632-7, 2013 May.
Artigo em Inglês | MEDLINE | ID: mdl-23611381

RESUMO

INTRODUCTION: Enterococcus faecalis is commonly detected in persistent apical periodontitis characterized by unimproved periradicular bone resorption. The aim of the present study was to examine the effect of lipoteichoic acid (LTA), a major virulence factor of E. faecalis, on apoptosis of osteoblasts. METHODS: Human osteoblast-like MG63 cells were treated with LTA from E. faecalis at a series of concentrations for 48 hours. The proliferation of the MG63 cells was assessed by 3-(4,5-dimethyl-thiazol-2-yl)-2,5- diphenyl-tetrazolium bromide assay. To examine the apoptosis, the LTA-treated cells were analyzed by flow cytometry by using annexin V-fluorescein isothiocyanate and propidium iodide double staining. Hoechst 33258 staining was performed to observe the morphologic changes of the cells. To investigate the apoptosis at the molecular level, the protein levels of Bcl-2 and Bax were determined by Western blot. Meanwhile, the caspase-3 activity was detected with caspase colorimetric protease assay. RESULTS: The proliferation of MG63 cells was inhibited by E. faecalis LTA in a dose-dependent manner. Flow cytometry assay indicated that LTA had a stimulating effect on MG63 cell apoptosis. Typical morphologies of apoptotic cells were observed under fluorescence microscope. Furthermore, the cell apoptosis was confirmed by (1) the down-regulation of the antiapoptotic protein (Bcl-2), (2) the up-regulation of the proapoptotic protein (Bax), and (3) the elevated caspase-3 activity. CONCLUSIONS: LTA of E. faecalis could inhibit the proliferation and induce apoptosis of human osteoblast-like MG63 cells.


Assuntos
Apoptose/efeitos dos fármacos , Enterococcus faecalis/fisiologia , Lipopolissacarídeos/farmacologia , Osteoblastos/efeitos dos fármacos , Ácidos Teicoicos/farmacologia , Caspase 3/efeitos dos fármacos , Técnicas de Cultura de Células , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Forma Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Corantes Fluorescentes , Humanos , Proteínas Proto-Oncogênicas c-bcl-2/efeitos dos fármacos , Fatores de Virulência/farmacologia , Proteína X Associada a bcl-2/efeitos dos fármacos
13.
J Endod ; 38(12): 1592-7, 2012 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-23146643

RESUMO

INTRODUCTION: One of the best-characterized Nod-like receptor (NLR) family members is pyrin domain containing 3 (NLRP3). Intracellular NLRP3 is the most versatile innate immune receptor. On activation, NLRP3 assembles into a multiprotein complex, termed an inflammasome, which regulates the secretion and bioactivity of interleukin-1 family cytokines. NLRP3 has broad specificity for mediating an immune response to a wide range of microbial stimuli or danger signals. Therefore, we hypothesize that NLRP3 plays an essential role in the detection of bacterial pathogens and the initiation of inflammation within the dental pulp. Thus, the aim of this study was to evaluate the expression of NLRP3 in normal human dental pulp cells (HDPCs) and pulp tissues. METHODS: Pulp tissues were collected from freshly extracted human third molars, and HDPCs were prepared from the explants of normal dental pulp tissues. Reverse transcription-polymerase chain reaction and Western blotting were performed to detect the levels of NLRP3 mRNA and protein, respectively. In addition, immunohistochemical staining was used to determine the distribution of NLRP3 in pulp tissues. RESULTS: Normal human dental pulp tissues displayed high levels of NLRP3 mRNA and protein. NLRP3 proteins were principally expressed in odontoblasts and some pulp vascular endothelial cells. Moreover, HDPCs also expressed NLRP3 but at a relatively low level in comparison with that of dental pulp tissues. CONCLUSIONS: The expression of NLRP3 in HDPCs and pulp tissues suggests that NLRP3-mediated signaling pathways may play an important role in dental immune defense.


Assuntos
Proteínas de Transporte/análise , Polpa Dentária/imunologia , Adolescente , Adulto , Western Blotting , Proteínas de Transporte/genética , Técnicas de Cultura de Células , Corantes , Citoplasma/ultraestrutura , Polpa Dentária/citologia , Células Endoteliais/citologia , Endotélio Vascular/citologia , Feminino , Fibroblastos/patologia , Humanos , Imunidade Inata/imunologia , Imuno-Histoquímica , Masculino , Proteína 3 que Contém Domínio de Pirina da Família NLR , Odontoblastos/citologia , Pulpite/imunologia , Pulpite/patologia , RNA Mensageiro/análise , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Adulto Jovem
14.
J Endod ; 38(10): 1348-54, 2012 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-22980176

RESUMO

INTRODUCTION: MicroRNAs (miRNAs) play momentous roles in various biological processes including cell differentiation. However, little is known about the role of miRNAs in human dental pulp cells (hDPCs) during odontogenic differentiation. The aims of this study were to investigate the expression of miRNAs in the primary culture of hDPCs when incubated in odontogenic medium. METHODS: The potential characteristics of hDPCs were investigated by miRNA microarray and real-time reverse transcriptase polymerase chain reaction. Bioinformatics (ie, target prediction, Gene Ontology analysis, and Kyoto Encyclopedia of Genes and Genomes mapping tools) were applied for predicting the complementary target genes of miRNAs and their biological functions. RESULTS: A total of 22 miRNAs were differentially expressed in which 12 miRNAs up-regulated and 10 miRNAs down-regulated in differentiated hDPCs compared with the control. The target genes of differential miRNAs were predicted to associate with several biological functions and signaling pathways including the mitogen-activated protein kinase (MAPK) and the Wnt signaling pathway. CONCLUSIONS: The differential expression miRNAs may be involved in governing hDPC odontogenic differentiation, thus contributing to the future investigations of regulatory mechanisms in reparative dentin formation and dental pulp regeneration.


Assuntos
Polpa Dentária/citologia , MicroRNAs/biossíntese , MicroRNAs/fisiologia , Odontogênese/fisiologia , Transdução de Sinais , Adolescente , Adulto , Fosfatase Alcalina/genética , Análise de Variância , Diferenciação Celular , Proteínas da Matriz Extracelular/genética , Perfilação da Expressão Gênica , Humanos , Sialoproteína de Ligação à Integrina/genética , Sistema de Sinalização das MAP Quinases , Odontoblastos/citologia , Odontogênese/genética , Análise de Sequência com Séries de Oligonucleotídeos , Osteocalcina/genética , Fosfoproteínas/genética , Cultura Primária de Células , Análise de Componente Principal , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sialoglicoproteínas/genética , Via de Sinalização Wnt , Adulto Jovem
15.
J Endod ; 38(1): 66-71, 2012 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-22152623

RESUMO

INTRODUCTION: Bone morphogenetic protein-2 (BMP-2) is a member of the transforming growth factor-ß (TGF-ß) superfamily, which has a broad range of activities that affect many different cell types. Previous research has suggested that BMP-2 induces the differentiation of human dental pulp cells (DPCs) into odontoblast-like cells. However, the mechanism by which BMP-2 induces odontoblastic differentiation has not yet been established. In the present study, we examined the involvement of the BMP/Smad pathway in mediating odontoblastic differentiation in DPCs. METHODS: Levels of phosphorylated and unphosphorylated Smad1/5 were quantified by Western blot analysis in response to BMP-2 and the BMP signaling inhibitor noggin. Some nuclear translocation of Smad1/5 was also observed by immunofluorescence staining in isolated DPCs treated with BMP-2. The effects of noggin on the BMP-2-induced odontoblastic differentiation of DPCs were determined by alkaline phosphatase activity assay, and the expression of odontoblastic markers was evaluated by reverse transcription polymerase chain reaction analysis and Western blotting. RESULTS: We found that BMP-2 induced the phosphorylation and nuclear translocation of Smad 1/5. In addition, noggin significantly inhibited alkaline phosphatase activity and odontoblastic differentiation and reduced the formation of mineralized nodules in BMP-2-treated DPCs. CONCLUSIONS: These findings suggest that Smad 1/5 is involved in BMP-2-induced odontoblastic differentiation in DPCs.


Assuntos
Proteína Morfogenética Óssea 2/farmacologia , Polpa Dentária/citologia , Odontoblastos/efeitos dos fármacos , Proteína Smad1/farmacologia , Proteína Smad5/farmacologia , Fosfatase Alcalina/análise , Fosfatase Alcalina/efeitos dos fármacos , Biomarcadores/análise , Western Blotting , Proteína Morfogenética Óssea 2/antagonistas & inibidores , Proteínas Morfogenéticas Ósseas/antagonistas & inibidores , Calcificação Fisiológica/efeitos dos fármacos , Proteínas de Transporte/farmacologia , Técnicas de Cultura de Células , Diferenciação Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Polpa Dentária/efeitos dos fármacos , Técnica Indireta de Fluorescência para Anticorpo , Humanos , Fosforilação , Transporte Proteico , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/efeitos dos fármacos , Proteína Smad1/metabolismo , Proteína Smad5/metabolismo
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