RESUMEN
OBJECTIVES: Root resorption in permanent teeth is a common pathological process that often follows dental trauma or orthodontic treatment. More rarely, root resorption is a feature of genetic disorders and can help with diagnosis. Thus, the present review aims to determine which genetic disorders could induce pathological root resorptions and thus which mutated genes could be associated with them. METHODS: We conducted a systematic review following the PRISMA guidelines. Articles describing root resorptions in patients with genetic disorders were included from PubMed, Embase, Web of Science, and Google Scholar. We synthesized the genetic disorder, the type, severity, and extent of the resorptions, as well as the other systemic and oral symptoms and histological features. RESULTS: The synthetic analysis included 25 studies among 937 identified records. We analyzed 21 case reports, three case series, and one cohort study. Overall, we highlighted 14 different pathologies with described root resorptions. Depending on the pathology, the sites of resorption, their extent, and their severity showed differences. CONCLUSION: With 14 genetic pathologies suspected to induce root resorptions, our findings are significant and enrich a previous classification. Among them, three metabolic disorders, three calcium-phosphorus metabolism disorders, and osteolysis disorders were identified.
Asunto(s)
Resorción Radicular , Humanos , Resorción Radicular/genética , Resorción Radicular/etiología , Enfermedades Genéticas Congénitas/genéticaRESUMEN
BACKGROUND: This study aimed to investigate the effects of various toll-like receptor (TLR) and C-type lectin receptor (CLR) ligands on osteogenic differentiation in human dental pulp stem cells (hDPSCs). METHODS: hDPSCs were cultured and treated with various concentrations (0.01, 0.1, 1.0, and 10 µg/mL) of TLR or CLR agonists (PG-LPS, E.coli LPS, poly(I:C), Pam3CSK4, Furfurman, and Zymosan). Cell viability was determined by MTT assay. The effects of TLR and CLR agonists on osteogenic differentiation of hDPSCs were measured by alkaline phosphatase (ALP) activity, Alizarin Red S staining, and Von Kossa staining. In addition, the mRNA expression of osteogenesis-related genes (ALP, COL1A1, RUNX2, OSX, OCN and DMP1) was examined by RT-qPCR. A non-parametric analysis was employed for the statistical analyses. The statistically significant difference was considered when p < 0.05. RESULTS: Treatment with TLR and CLR agonists was associated with an increase in hDPSCs' colony-forming unit ability. Compared with the control group, TLR and CLR agonists significantly inhibited the osteogenic differentiation of hDPSCs by decreasing the ALP activity, mineralised nodule formation, and mRNA expression levels of osteogenesis-related genes (ALP, COL1A1, RUNX2, OSX, OCN and DMP1). The inhibition of TRIF but not Akt signalling rescued the effects of TLR and CLR agonist attenuating hDPSCs' mineralisation. CONCLUSIONS: The activation of TLRs or CLRs exhibited an inhibitory effect on osteogenic differentiation of hDPSCs via the TRIF-dependent signalling pathway.
Asunto(s)
Pulpa Dental , Osteogénesis , Humanos , Subunidad alfa 1 del Factor de Unión al Sitio Principal/metabolismo , Lipopolisacáridos/farmacología , Lipopolisacáridos/metabolismo , Diferenciación Celular , Receptores Toll-Like/metabolismo , Células Madre , Proteínas Adaptadoras del Transporte Vesicular/metabolismo , Proteínas Adaptadoras del Transporte Vesicular/farmacología , ARN Mensajero/metabolismo , Células CultivadasRESUMEN
OBJECTIVE: To investigate the role of phosphatase and tensin homolog (PTEN) in dental pulp cells (hDPs) and adipose-derived mesenchymal stem cells (hADSCs). MATERIALS AND METHODS: Genetic variant was identified with exome sequencing. The hDPs isolated from a patient with Cowden syndrome were investigated for their proliferation, osteogenesis, adipogenesis, and gene expression compared with controls. The normal hDPs and hADSCs were treated with the PTEN inhibitor, VO-OHpic trihydrate (VOT), to investigate the effect of PTEN inhibition. RESULTS: A heterozygous nonsense PTEN variant, c.289C>T (p.Gln97*), was identified in the Cowden patient's blood and intraoral lipomas. The mutated hDPs showed significantly decreased proliferation, but significantly upregulated RUNX2 and OSX expression and mineralization, indicating enhanced osteogenic ability in mutated cells. The normal hDPs treated with VOT showed the decreases in proliferation, colony formation, osteogenic marker genes, alkaline phosphatase activity, and mineral deposition, suggesting that PTEN inhibition diminishes proliferation and osteogenic potential of hDPs. Regarding adipogenesis, the VOT-treated hADSCs showed a reduced number of cells containing lipid droplets, suggesting that PTEN inhibition might compromise adipogenic ability of hADSCs. CONCLUSIONS: PTEN regulates proliferation, enhances osteogenesis of hDPs, and induces adipogenesis of hADSCs. The gain-of-function PTEN variant, p.Gln97*, enhances osteogenic ability of PTEN in hDPs.
Asunto(s)
Adipogénesis , Células Madre Mesenquimatosas , Humanos , Adipogénesis/genética , Diferenciación Celular/genética , Tejido Adiposo , Osteogénesis/genética , Pulpa Dental , Células Madre Mesenquimatosas/metabolismo , Proliferación Celular/genética , Células Cultivadas , Fosfohidrolasa PTEN/genética , Fosfohidrolasa PTEN/metabolismo , Fosfohidrolasa PTEN/farmacologíaRESUMEN
AIM: To investigate the effect of Wnt3a on odonto/osteogenic differentiation of stem cells isolated from human exfoliated deciduous teeth (SHEDs) and reparative dentine formation in a rat model. METHODOLOGY: Stem cells isolated from human exfoliated deciduous teeth were cultured in media with Wnt3a (50-200 ng/ml). Wnt activation was confirmed by ß-catenin immunocytochemistry. Colony-forming unit assay (normalized percentage area), osteogenic gene expression analysis by real-time polymerase chain reaction and mineralization assays measured by the absorption at 540 nm were performed. Tertiary dentine formation in vivo was evaluated using 8-week-old, male Wistar rats. Cavities with pinpoint pulp exposure by a sharp instrument were prepared at the mesial surface of the first molars. Teeth were divided into (n = 6): (1) distilled water (negative control), (2) phosphate-buffered saline (PBS), (3) lithium chloride in DI (20 µM), and (4) Wnt3a in PBS (200 ng/ml). Collagen sponge was used as a scaffold. The cavity was sealed with glass ionomer restoration. Four weeks later, animals were euthanized by sodium pentobarbital (120 mg/kg body weight). Hard tissue formation was evaluated using micro-computerized tomography. Sixty consecutive slides from the initial plane were analysed and calculated as bone/dentine volume per total tissue volume. Paraffin sections (2 µm) were stained with haematoxylin and eosin and Masson's trichrome for morphological evaluation. Data are presented as the mean ± standard error. Mann-Whitney U test was used for two-group comparison. Kruskal Wallis followed by pairwise comparison was employed for three or more group comparisons. Statistical analysis was performed using GraphPad Prism 7. Differences were considered significant at p < .05. RESULTS: Wnt3a decreased SHEDs colony formation and increased OSX, BMP2, and DMP1 expression, corresponding to an increase in mineralization. Additionally, a significant increase in dentine/bone volume per total tissue volume was observed in Wnt3a treated defects. Dentine bridge formation at the exposure sites treated with Wnt3a demonstrated, while fibrous tissues were observed in the control. CONCLUSIONS: Wnt3a suppressed proliferation, increased osteogenic differentiation of SHEDs and promotes tertiary dentine formation. Wnt3a could be utilized as biological molecule for vital pulp therapy.
Asunto(s)
Dentina Secundaria , Osteogénesis , Animales , Humanos , Masculino , Ratas , Diferenciación Celular/fisiología , Diente Molar , Ratas Wistar , Proteína Wnt3ARESUMEN
OBJECTIVES: Minipigs present advantages for studying oral bone regeneration; however, standardized critical size defects (CSD) for alveolar bone have not been validated yet. The objectives of this study are to develop a CSD in the mandibular alveolar bone in Aachen minipigs and to further investigate the specific role of periosteum. MATERIALS AND METHODS: Three female Aachen minipigs aged 17, 24, and 84 months were used. For each minipig, a split-mouth design was performed: an osteotomy (2 cm height × 2.5 cm length) was performed; the periosteum was preserved on the left side and removed on the right side. Macroscopic, cone beam computed tomography (CBCT), microcomputed tomography (µCT), and histological analyses were performed to evaluate the bone defects and bone healing. RESULTS: In both groups, spontaneous healing was insufficient to restore initial bone volume. The macroscopic pictures and the CBCT results showed a larger bone defect without periosteum. µCT results revealed that BMD, BV/TV, and Tb.Th were significantly lower without periosteum. The histological analyses showed (i) an increased osteoid apposition in the crestal area when periosteum was removed and (ii) an ossification process in the mandibular canal area in response to the surgical that seemed to increase when periosteum was removed. CONCLUSIONS: A robust model of CSD model was developed in the alveolar bone of minipigs that mimics human mandibular bone defects. This model allows to further investigate the bone healing process and potential factors impacting healing such as periosteum. CLINICAL RELEVANCE: This model may be relevant for testing different bone reconstruction strategies for preclinical investigations.
Asunto(s)
Regeneración Ósea , Periostio , Animales , Femenino , Porcinos , Humanos , Periostio/cirugía , Porcinos Enanos , Proyectos Piloto , Microtomografía por Rayos X , Regeneración Ósea/fisiología , Mandíbula/diagnóstico por imagen , Mandíbula/cirugía , Mandíbula/patologíaRESUMEN
6-bromoindirubin-3'-oxime (BIO) is a candidate small molecule that effectively modulates Wnt signalling owing to its stable property. The present study investigated the influence of BIO on the odonto/osteogenic differentiation of human dental pulp stem cells (hDPSCs). hDPSCs were treated with 200, 400, or 800 nM BIO, and the effects on hDPSC responses and osteogenic differentiation were assessed. BIO-mediated Wnt activation was confirmed by ß-catenin nuclear translocation detected by immunofluorescence staining. BIO attenuated colony formation and cell migration determined by in vitro wound-healing assay. BIO increased early apoptotic cell population evaluated using flow cytometry. For osteogenic induction, BIO promoted alkaline phosphatase (ALP) activity and mineralisation in a dose-dependent manner. ALP, RUNX2, OCN, OSX, ANKH, DMP1, and DSPP mRNA expression were significantly upregulated. The OPG/RANKL expression ratio was also increased. Further, BIO attenuated adipogenic differentiation as demonstrated by decreased lipid accumulation and adipogenic-related gene expression. Bioinformatic analysis of RNA sequencing data from the BIO-treated hDPSCs revealed that BIO modulated pathways related to autophagy and actin cytoskeleton regulation. These findings demonstrated that BIO treatment promoted hDPSC osteogenic differentiation. Therefore, this small molecule is a strong candidate as a bioactive molecule to enhance dentin repair.
Asunto(s)
Osteogénesis , Células Madre , Apoptosis , Diferenciación Celular/genética , Proliferación Celular , Células Cultivadas , Pulpa Dental , Humanos , Indoles , Osteogénesis/genética , Oximas , Células Madre/metabolismoRESUMEN
Notch signaling is associated with many human malignancies, including oral squamous cell carcinoma (OSCC). However, the exact function of Notch signaling in OSCC remains unclear. Here, we investigated the effect of Notch signaling inhibition using a γ-secretase inhibitor (DAPT) on OSCC behaviours in vitro. Bioinformatic analysis of public-available gene expression profiles revealed the dysregulation of the Notch signaling pathway in OSCC compared with normal tissues, indicating the role of Notch signaling in OSCC regulation. RNA sequencing analysis of DAPT-treated human OSCC cells revealed the dysregulation of genes related to cell cycle-related pathways. Blocking Notch signaling significantly inhibited cell proliferation. DAPT-induced G0/G1 cell cycle arrest induced cell apoptosis. Furthermore, cell migration and invasion were also reduced in DAPT-treated cells. These findings indicate that Notch signaling activation participates in OSCC regulation by promoting cell growth, cell cycle progression, cell migration, and invasion. These mechanisms could facilitate OSCC progression. These results imply the potential use of Notch signaling inhibitors as a candidate adjuvant treatment in OSCC patients.
Asunto(s)
Carcinoma de Células Escamosas , Neoplasias de Cabeza y Cuello , Neoplasias de la Boca , Secretasas de la Proteína Precursora del Amiloide/metabolismo , Apoptosis , Carcinoma de Células Escamosas/tratamiento farmacológico , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/metabolismo , Ciclo Celular , Línea Celular Tumoral , Movimiento Celular , Proliferación Celular , Humanos , Neoplasias de la Boca/tratamiento farmacológico , Neoplasias de la Boca/genética , Neoplasias de la Boca/metabolismo , Inhibidores de Agregación Plaquetaria/farmacología , Carcinoma de Células Escamosas de Cabeza y CuelloRESUMEN
The indirect immobilisation of Jagged-1 (Jagged-1) promoted osteogenic differentiation of human dental pulp cells (hDPs). Furthermore, the analysis of the Reactome pathway of RNA sequencing data indicates the upregulated genes involved with the extracellular matrix (ECM). Hence, our objective was to investigate the effects of Jagged-1 on proteomic profiles of human dental pulp stem cells (hDPSC). hDPSCs were cultured on the surface coated with human IgG Fc fragment (hFc) and the surface coated with rhJagged1/Fc recombinant protein-coated surface. Cells were differentiated to the osteogenic lineage using an osteogenic differentiation medium (OM) for 14 days, and cells cultured in a growth medium were used as a control. The protein component of the cultured cells was extracted into the cytosol, membrane, nucleus, and cytoskeletal compartment. Subsequently, the proteomic analysis was performed using liquid chromatography-tandem mass spectrometry (LC-MS). Metascape gene list analysis reported that Jagged-1 stimulated the expression of the membrane trafficking protein (DOP1B), which can indirectly improve osteogenic differentiation. hDPSCs cultured on Jagged-1 surface under OM condition expressed COL27A1, MXRA5, COL7A1, and MMP16, which played an important role in osteogenic differentiation. Furthermore, common matrisome proteins of all cellular components were related to osteogenesis/osteogenic differentiation. Additionally, the gene ontology categorised by the biological process of cytosol, membrane, and cytoskeleton compartments was associated with the biomineralisation process. The gene ontology of different culture conditions in each cellular component showed several unique gene ontologies. Remarkably, the Jagged-1_OM culture condition showed the biological process related to odontogenesis in the membrane compartment. In conclusion, the Jagged-1 induces osteogenic differentiation could, mainly through the regulation of protein in the membrane compartment.
Asunto(s)
Osteogénesis , Proteómica , Humanos , Colágeno Tipo VII/metabolismo , Pulpa Dental/metabolismo , Matriz Extracelular/metabolismo , Colágenos Fibrilares/metabolismo , Proteína Jagged-1/genética , Proteína Jagged-1/metabolismo , Células Madre/metabolismoRESUMEN
OBJECTIVES: Dentin sialophosphoprotein (DSPP) plays an important role in the mineralization of both dentin and bones. The Dspp null mice developed periodontal diseases. Patients with DSPP mutations have dentinogenesis imperfecta (DGI), but very little is known about their bone characteristics. This study aims to characterize alveolar bone cells of a DGI patient with DSPP mutation. MATERIALS AND METHODS: Pathogenic variants were identified by whole exome and sanger sequencing. Cells isolated from the alveolar bones of a DSPP patient were investigated for their characteristics including cell morphology, attachment, spreading, proliferation, colony formation, mineralization, and osteogenic differentiation. RESULTS: We identified a Thai family with three members affected with autosomal dominant DGI harboring a heterozygous pathogenic missense mutation, c.50C > T, p.P17L, in exon 2 of the DSPP gene. The patients' phenotypes presented deteriorated opalescent teeth with periapical lesions, thickening of lamina dura, furcation involvement, alveolar bone loss, and bone exostoses. The alveolar bone cells isolated from DSPP patient exhibited compromised proliferation and colony formation. Scanning electron microscope revealed altered cellular morphology and spreading. The DSPP cells showed deviated mRNA levels of OCN, ALP, and COL1 but maintained in vitro mineralization ability compared to the control. CONCLUSIONS: We demonstrate that the DSPP p.P17L mutant alveolar bone cells had compromised cell spreading, proliferation, colony formation, and osteogenic induction, suggesting abnormal bone characteristics in the patient with DGI caused by DSPP mutation. CLINICAL RELEVANCE: DSPP mutation can induce the behavior alterations of alveolar bone cells.
Asunto(s)
Proceso Alveolar/citología , Dentinogénesis Imperfecta/genética , Proteínas de la Matriz Extracelular/genética , Mutación Missense , Fosfoproteínas/genética , Sialoglicoproteínas/genética , Adulto , Proliferación Celular , Células Cultivadas , Femenino , Humanos , Microscopía Electrónica de Rastreo , Linaje , Fenotipo , Reacción en Cadena en Tiempo Real de la Polimerasa , Células Madre , TailandiaRESUMEN
Dental pulp cells release adenosine triphosphate (ATP) in response to intrapulpal pressure and the amount released depends on the magnitude of the pressure. ATP regulates the differentiation of stem cells into adipocytes and osteoblasts. However, it is unknown whether extracellular ATP influences the stemness and osteogenic differentiation of stem cells from human exfoliated deciduous teeth (SHEDs). Therefore, this study investigated the effects of extracellular ATP at a low (0.1 µM) and high (10 µM) concentration on the stemness and osteogenic differentiation of SHEDs. Cells were cultured in either growth medium or osteogenic medium with or without 0.1-10 µM ATP. In growth medium, both concentrations of ATP increased the mRNA expression of pluripotent and osteogenic markers. In contrast, in osteogenic medium, 0.1 µM ATP enhanced in vitro mineralization, whereas 10 µM ATP inhibited this process. In addition, 10 µM ATP stimulated the mRNA expression and activity of ectonucleotide pyrophosphatase/phosphodiesterase (ENPP), an enzyme that regulates the phosphate/pyrophosphate ratio. Thus, depending on the growth condition and its concentration, ATP stimulated stemness and in vitro mineralization or inhibited mineralization. In growth medium, both ATP concentrations stimulated pluripotent and osteogenic marker gene expression. However, in osteogenic medium, a biphasic effect was found on in vitro mineralization; the low concentration stimulated, whereas the high concentration inhibited, mineralization. We propose that ATP released due to mechanical stress modulates the stemness and differentiation of SHEDs. J. Cell. Biochem. 119: 488-498, 2018. © 2017 Wiley Periodicals, Inc.
Asunto(s)
Adenosina Trifosfato/farmacología , Calcificación Fisiológica/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Pulpa Dental/metabolismo , Osteogénesis/efectos de los fármacos , Diente Primario/metabolismo , Células Cultivadas , Pulpa Dental/citología , Relación Dosis-Respuesta a Droga , Humanos , Diente Primario/citologíaRESUMEN
OBJECTIVE: To delineate orodental features, dental mineral density, genetic aetiology and cellular characteristics associated with amelogenesis imperfecta (AI). MATERIALS AND METHODS: Three affected patients in a family were recruited. Whole-exome sequencing was used to identify mutations confirmed by Sanger sequencing. The proband's teeth were subjected for mineral density analysis by microcomputerised tomography and characterisation of periodontal ligament cells (PDLCs). RESULTS: The patients presented yellow-brown, pitted and irregular enamel. A novel nonsense mutation, c.1261G>T, p.E421*, in exon 5 of the FAM83H was identified. The mineral density of the enamel was significantly decreased in the proband. The patient's PDLCs (FAM83H cells) exhibited reduced ability of cell proliferation and colony-forming unit compared with controls. The formation of stress fibres was remarkably present. Upon cultured in osteogenic induction medium, FAM83H cells, at day 7 compared to day 3, had a significant reduction of BSP, COL1 and OCN mRNA expression and no significant change in RUNX2. The upregulation of ALP mRNA levels and mineral deposition were comparable between FAM83H and control cells. CONCLUSIONS: We identified the novel mutation in FAM83H associated with autosomal dominant hypocalcified AI. The FAM83H cells showed reduced cell proliferation and expression of osteogenic markers, suggesting altered PDLCs in FAM83H-associated AI.
Asunto(s)
Amelogénesis Imperfecta/genética , Amelogénesis Imperfecta/patología , Ligamento Periodontal/patología , Proteínas/genética , Proliferación Celular , Células Cultivadas , Codón sin Sentido , Colágeno Tipo I/genética , Femenino , Humanos , Sialoproteína de Unión a Integrina/genética , Masculino , Osteocalcina/genética , Ligamento Periodontal/ultraestructura , Fibras de Estrés/ultraestructura , Secuenciación del Exoma , Adulto JovenRESUMEN
Basic fibroblast growth factor (bFGF) regulates pluripotent marker expression and cellular differentiation in various cell types. However, the mechanism by which bFGF regulates REX1 expression in stem cells, isolated from human exfoliated deciduous teeth (SHEDs) remains unclear. The aim of the present study was to investigate the regulation of REX1 expression by bFGF in SHEDs. SHEDs were isolated and characterized. Their mRNA and protein expression levels were determined using real-time polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. In some experiments, chemical inhibitors were added to the culture medium to impede specific signaling pathways. Cells isolated from human exfoliated deciduous tooth dental pulp tissue expressed mesenchymal stem cell surface markers (CD44, CD73, CD90, and CD105). These cells differentiated into osteogenic and adipogenic lineages, when appropriately induced. Treating SHEDs with bFGF induced REX1 mRNA expression and this effect was attenuated by pretreatment with FGFR or Akt inhibitors. Cycloheximide pretreatment also inhibited the bFGF-induced REX1 expression, implying the involvement of intermediate molecule(s). Further, the addition of an IL-6 neutralizing antibody attenuated the bFGF-induced REX1 expression by SHEDs. In conclusion, bFGF enhanced REX1 expression by SHEDs via the FGFR and Akt signaling pathways. Moreover, IL-6 participated in the bFGF-induced REX1 expression in SHEDs. J. Cell. Biochem. 118: 1480-1488, 2017. © 2016 Wiley Periodicals, Inc.
Asunto(s)
Factor 2 de Crecimiento de Fibroblastos/metabolismo , Interleucina-6/genética , Factores de Transcripción de Tipo Kruppel/genética , Células Madre/citología , Diente Primario/citología , Diferenciación Celular , Proliferación Celular , Células Cultivadas , Niño , Femenino , Factores de Crecimiento de Fibroblastos/genética , Regulación de la Expresión Génica , Humanos , Factores de Transcripción de Tipo Kruppel/metabolismo , Masculino , Proteínas Proto-Oncogénicas c-akt/genética , Transducción de Señal , Células Madre/metabolismo , Exfoliación Dental/genética , Exfoliación Dental/metabolismo , Diente Primario/metabolismoRESUMEN
Dental tissue-derived mesenchymal stem cells have been proposed as an alternative source for mesenchymal stem cells. Here, we investigated the differentiation ability toward insulin producing cells (IPCs) of human dental pulp stem cells (hDPSCs) and human periodontal ligament stem cells (hPDLSCs). These cells expressed mesenchymal stem cell surface markers and were able to differentiate toward osteogenic and adipogenic lineages. Upon 3 step-IPCs induction, hDPSCs exhibited more colony number than hPDLSCs. The mRNA upregulation of pancreatic endoderm/islet markers was noted. However, the significant increase was noted only for PDX-1, NGN-3, and INSULIN mRNA expression of hDPSCs. The hDPSCs-derived IPCs expressed PRO-INSULIN and released C-PEPTIDE upon glucose stimulation in dose-dependent manner. After IPCs induction, the Notch target, HES-1 and HEY-1, mRNA expression was markedly noted. Notch inhibition during the last induction step or throughout the protocol disturbed the ability of C-PEPTIDE release upon glucose stimulation. The results suggested that hDPSCs had better differentiation potential toward IPCs than hPDLSCs. In addition, the Notch signalling might involve in the differentiation regulation of hDPSCs into IPCs.
Asunto(s)
Pulpa Dental/citología , Células Secretoras de Insulina/citología , Células Madre Mesenquimatosas/citología , Ligamento Periodontal/citología , ARN Mensajero/genética , Adipocitos/citología , Adipocitos/efectos de los fármacos , Adipocitos/metabolismo , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Biomarcadores/metabolismo , Péptido C/genética , Péptido C/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Diferenciación Celular , Pulpa Dental/efectos de los fármacos , Pulpa Dental/metabolismo , Expresión Génica , Glucosa/metabolismo , Glucosa/farmacología , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Humanos , Células Secretoras de Insulina/efectos de los fármacos , Células Secretoras de Insulina/metabolismo , Péptidos y Proteínas de Señalización Intercelular/farmacología , Células Madre Mesenquimatosas/efectos de los fármacos , Células Madre Mesenquimatosas/metabolismo , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Osteocitos/citología , Osteocitos/efectos de los fármacos , Osteocitos/metabolismo , Ligamento Periodontal/efectos de los fármacos , Ligamento Periodontal/metabolismo , Cultivo Primario de Células , Proinsulina/genética , Proinsulina/metabolismo , ARN Mensajero/metabolismo , Receptores Notch/genética , Receptores Notch/metabolismo , Transducción de Señal , Extracción Dental , Transactivadores/genética , Transactivadores/metabolismo , Factor de Transcripción HES-1RESUMEN
Human dental pulp of exfoliated deciduous teeth contains the population of cells that exhibited mesenchymal stem cell (MSC) characters. Though, a cell amplification process is indeed required to secure an adequate cell number for such a potential employment. Several publications suggested the alteration of MSCs upon in vitro culture, for example, the decrease in proliferation and the loss of stem cell characters. Here, we investigated an influence of basic fibroblast growth factor (bFGF) on stem cells isolated from human exfoliated deciduous teeth (SHEDs) with respect to cell proliferation, colony forming unit efficiency and stem cell marker expression in both short- and long-term cultures. For short-term bFGF treatment, SHEDs were treated with bFGF for 48 h. While, in long-term bFGF supplementation, SHEDs were maintained in culture and continuous passage upon confluence in medium supplemented with bFGF. Cells at passage (P) 5 and 10 were employed for characterization. Our results showed that short-term bFGF treatment enhanced OCT4, REX1, and NANOG mRNA expression as well as colony forming unit ability. The FGFR inhibitor pretreatment was able to attenuate the influence of bFGF on pluripotent stem cell marker expression, confirming bFGF function. In addition, cells cultured in high passage number had decreased in cell proliferation, colony forming unit capacity, and pluripotent stem cell maker mRNA expression. However, bFGF supplementation in culture medium enhanced both pluripotent stem cell marker expression and colony forming unit capacity in later passage, though the effect was not robust. Together, these results indicate that high passage number may attenuate pluripotent properties of SHEDs and bFGF supplementation could be the beneficial approach to maintain SHEDs' stemness properties.
Asunto(s)
Factor 2 de Crecimiento de Fibroblastos/farmacología , Células Madre Mesenquimatosas/metabolismo , Células Madre Pluripotentes/metabolismo , Diente Primario/citología , Biomarcadores/metabolismo , Medios de Cultivo , Humanos , Células Madre Mesenquimatosas/citología , Células Madre Pluripotentes/citologíaRESUMEN
The decellularised extracellular matrix (dECM) of in vitro cell culture is a naturally derived biomaterial formed by the removal of cellular components. The compositions of molecules in the extracellular matrix (ECM) differ depending on various factors, including the culture conditions. Cell-derived ECM provides a 3-dimensional structure that has a complex influence on cell signalling, which in turn affects cell survival and differentiation. This review describes the effects of dECM derived from mesenchymal stem cells (MSCs) on cell responses, including cell migration, cell proliferation, and cell differentiation in vitro. Published articles were searched in the PubMed databases in 2005 to 2022, with assigned keywords (MSCs and decellularisation and cell culture). The 41 articles were reviewed, with the following criteria. (1) ECM was produced exclusively from MSCs; (2) decellularisation processes were performed; and (3) the dECM production was discussed in terms of culture systems and specific supplementations that are suitable for creating the dECM biomaterials. The dECM derived from MSCs supports cell adhesion, enhances cell proliferation, and promotes cell differentiation. Importantly, dECM derived from dental MSCs shows promise in regenerative dentistry applications. Therefore, the literature strongly supports cell-based dECMs as a promising option for innovative tissue engineering approaches for regenerative medicine.
Asunto(s)
Diferenciación Celular , Proliferación Celular , Células Madre Mesenquimatosas , Ingeniería de Tejidos , Humanos , Células Madre Mesenquimatosas/citología , Ingeniería de Tejidos/métodos , Medicina Regenerativa , Matriz Extracelular Descelularizada , Matriz Extracelular , Movimiento Celular , Adhesión Celular , Materiales Biocompatibles , Técnicas de Cultivo de CélulaRESUMEN
Nanostructured lipid carriers (NLC) represent the second generation of nanoparticles, offering numerous advantages over conventional delivery systems. These include improved stability, enhanced drug-loading capacity, and controlled release profiles, making them highly attractive candidates for a wide range of therapeutic applications. Their suitability for hydrophobic drugs like a traditional medicinal plant of Thailand as clove oil and alpha-mangostin. We investigated into nanostructured lipid carriers loaded with Alpha-Mangostin and clove oil (NLC-AMCO) into the physicochemical and biological characteristics to identify the formulation with the highest efficacy for treatment. The particle size, charge, polydispersity index, and other characterizations were recorded. The realtime ex vivo penetration was explored using canine gingival tissue. Drug sustained release was assessed by HPLC. Moreover, the antibacterial properties were tested by conventional methods. The NLC-AMCO can be stored at up to 40 °C for 60 days without any alterations in particle characteristics. Gingival tissue penetration and sustained drug release were superior compared to unencapsulated counterparts. It exhibited greater effectiveness in inhibiting bacterial growth than the antibiotics tested, particularly against bacteria from the oral cavities of dogs. Therefore, this alternative treatment approach offers cost-effectiveness and ease of administration for pet owners and reduces discomfort for the animals during restraint.
RESUMEN
Extracellular matrix (ECM) is an intricate structure providing the microenvironment niche that influences stem cell differentiation. This study aimed to investigate the efficacy of decellularized ECM derived from human dental pulp stem cells (dECM_DPSCs) and gingival-derived mesenchymal stem cells (dECM_GSCs) as an inductive scaffold for osteogenic differentiation of GSCs. The proteomic analysis demonstrated that common and signature matrisome proteins from dECM_DPSCs and dECM_GSCs were related to osteogenesis/osteogenic differentiation. RNA sequencing data from GSCs reseeded on dECM_DPSCs revealed that dECM_DPSCs upregulated genes related to the Hippo and Wnt signaling pathways in GSCs. In the inhibitor experiments, results revealed that dECM_DPSCs superiorly promoted GSCs osteogenic differentiation, mainly mediated through Hippo and Wnt signaling. The present study emphasizes the promising translational application of dECM_DPSCs as a bio-scaffold rich in favorable regenerative microenvironment for tissue engineering.
Asunto(s)
Osteogénesis , Vía de Señalización Wnt , Humanos , Osteogénesis/genética , Proteómica , Pulpa Dental , Matriz Extracelular/metabolismo , Diferenciación Celular , Células Madre/metabolismo , Proliferación Celular , Células CultivadasRESUMEN
Basic fibroblast growth factor (bFGF) and Notch signaling play critical roles in various cell behaviors. Here, we investigated the influence of bFGF and Notch signaling in alkaline phosphatase (ALP) expression and mineralization process in human periodontal ligament-derived mesenchymal stem cells (PDLSCs) and stem cells isolated from human exfoliated deciduous teeth (SHEDs). PDLSCs and SHEDs were cultured in osteogenic medium supplemented with bFGF or on the immobilized Notch ligands, JAGGED1. The ALP mRNA and protein expression were measured by quantitative reverse transcriptase polymerase chain reaction and enzymatic activity assay, respectively. Mineral deposition was determined using alizarin red S staining. The results showed that the addition of bFGF resulted in the decrease of ALP mRNA expression and enzymatic activity. In addition, the attenuation of mineralization was noted. These phenomenons were blocked by the addition of a fibroblast growth factor receptor inhibitor (SU5402) or a MEK inhibitor (PD98059). Interestingly, bFGF supplementation also decreased the Notch signaling component mRNA levels. Thus, to evaluate effect of Notch signaling in mineralization process, PDLSCs and SHEDs were exposed to JAGGED1 modified surface. The ALP mRNA and protein expression were significantly upregulated and the mineral deposition was markedly increased. These results could be reversed by the addition of a γ-secretase inhibitor. In addition, bFGF could attenuate the Notch-signaling-induced mineralization in both PDLSCs and SHEDs. These results suggest that mineralization was enhanced by Notch signaling but attenuated by bFGF signaling. This knowledge can be further utilized to control PDLSCs and SHEDs mineralization for tissue regeneration purpose.
Asunto(s)
Fosfatasa Alcalina/metabolismo , Proteínas de Unión al Calcio/farmacología , Factor 2 de Crecimiento de Fibroblastos/farmacología , Péptidos y Proteínas de Señalización Intercelular/farmacología , Proteínas de la Membrana/farmacología , Células Madre Mesenquimatosas/efectos de los fármacos , Células Madre Mesenquimatosas/metabolismo , Fosfatasa Alcalina/genética , Células Cultivadas , Humanos , Proteína Jagged-1 , Reacción en Cadena en Tiempo Real de la Polimerasa , Proteínas Serrate-JaggedRESUMEN
Asiaticoside, an active ingredient extracted from Centella asiatica, has been widely used to promote wound healing. In this study, the effects of asiaticoside on proliferation, protein synthesis, and osteogenic differentiation in human periodontal ligament cells (HPDLs) were investigated. HPDLs were treated with asiaticoside at concentrations of 25, 50, and 100 µg/mL. Cell number was determined by MTT assay. The mRNA expression was analyzed by reverse transcription-polymerase chain reaction. Western blot analysis and immunocytochemistry were used to confirm protein synthesis. Osteogenic differentiation was determined by alkaline phosphatase activity, osteoblast marker gene expression, and in vitro mineralization. The results showed that asiaticoside treatment, ranging from 25 to 100 mg/mL, had no effect on cytotoxicity or cell proliferation. When HPDLs were treated with asiaticoside in serum-free medium, dose-dependent increases in the levels of fibronectin and collagen type I mRNA and protein were observed at 72 h. Moreover, asiaticoside attenuated matrix metalloproteinase-1 but enhanced tissue inhibitor of metalloproteinase-1 mRNA expression. The addition of asiaticoside to osteogenic medium resulted in an increase in alkaline phosphatase enzymatic activity, up-regulation of osteoblast marker gene mRNA expression, and enhancement of mineralization by HPDLs. These results suggest the potential application of asiaticoside for enhancing periodontal tissue healing.
Asunto(s)
Diferenciación Celular/efectos de los fármacos , Colágeno Tipo I/biosíntesis , Osteogénesis/efectos de los fármacos , Ligamento Periodontal/citología , Triterpenos/farmacología , Proliferación Celular/efectos de los fármacos , Centella/química , Humanos , Metaloproteinasa 1 de la Matriz/metabolismo , Cultivo Primario de Células , Inhibidor Tisular de Metaloproteinasa-1/metabolismoRESUMEN
INTRODUCTION: Candida spp. has recently been introduced to interact with conventional carious bacteria, leading to dental caries progression and virulence ability. Evidence regarding the influence of Candida spp. on human dental pulp cell response remains unknown. This study aimed to investigate the effects of Candida albicans mannans on cytotoxicity, cell proliferation, osteogenic differentiation, and inflammatory-related gene expression in human dental pulp cells (hDPCs). METHODS: hDPCs were treated with cell wall mannans isolated from C. albicans, Candida krusei, Candida glabrata, Candida tropocalis, Candida parapsilosis, and Candida dubliniensis. Cell viability was performed using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide assay. Osteogenic differentiation- and inflammatory-related gene expression were determined using a real-time polymerase chain reaction. Mineralization was examined using alizarin red S staining. RESULTS: The treatment of mannans isolated from C. albicans, C. krusei, C. glabrata, C. tropocalis, C. parapsilosis, and C. dubliniensis at concentrations ranging from 10-100 µg/mL did not affect cytotoxicity or cell proliferation. Mannans isolated from C. albicans, C. glabrata, and C. tropocalis significantly attenuated mineralization. However, cell wall mannans isolated from C. krusei, C. parapsilosis, and C. dubliniensis did not significantly influence mineral deposition in hDPCs. C. albicans cell wall mannans significantly attenuated osteogenic differentiation-related gene expression (RUNX2, ALP, and ENPP1). Interestingly, IL12 messenger RNA expression was significantly upregulated when treated with C. albicans cell wall mannans. The addition of recombinant IL12 significantly decreased mineralization in hDPCs. CONCLUSIONS: C. albicans cell wall mannans attenuated osteogenic differentiation in hDPCs and up-regulated inflammatory-related gene IL12 expression.