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1.
J Nanobiotechnology ; 22(1): 265, 2024 May 17.
Artículo en Inglés | MEDLINE | ID: mdl-38760763

RESUMEN

BACKGROUND: Pulp regeneration is a novel approach for the treatment of immature permanent teeth with pulp necrosis. This technique includes the combination of stem cells, scaffolds, and growth factors. Recently, stem cell-derived extracellular vesicles (EVs) have emerged as a new methodology for pulp regeneration. Emerging evidence has proven that preconditioning is an effective scheme to modify EVs for better therapeutic potency. Meanwhile, proper scaffolding is of great significance to protect EVs from rapid clearance and destruction. This investigation aims to fabricate an injectable hydrogel loaded with EVs from pre-differentiated stem cells from human exfoliated deciduous teeth (SHEDs) and examine their effects on pulp regeneration. RESULTS: We successfully employed the odontogenic induction medium (OM) of SHEDs to generate functional EV (OM-EV). The OM-EV at a concentration of 20 µg/mL was demonstrated to promote the proliferation and migration of dental pulp stem cells (DPSCs). The results revealed that OM-EV has a better potential to promote odontogenic differentiation of DPSCs than common EVs (CM-EV) in vitro through Alizarin red phalloidin, alkaline phosphatase staining, and assessment of the expression of odontogenic-related markers. High-throughput sequencing suggests that the superior effects of OM-EV may be attributed to activation of the AMPK/mTOR pathway. Simultaneously, we prepared a photocrosslinkable gelatin methacryloyl (GelMA) to construct an OM-EV-encapsulated hydrogel. The hydrogel exhibited sustained release of OM-EV and good biocompatibility for DPSCs. The released OM-EV from the hydrogel could be internalized by DPSCs, thereby enhancing their survival and migration. In tooth root slices that were subcutaneously transplanted in nude mice, the OM-EV-encapsulated hydrogel was found to facilitate dentinogenesis. After 8 weeks, there was more formation of mineralized tissue, as well as higher levels of dentin sialophosphoprotein (DSPP) and dentin matrix protein-1 (DMP-1). CONCLUSIONS: The effects of EV can be substantially enhanced by preconditioning of SHEDs. The functional EVs from SHEDs combined with GelMA are capable of effectively promoting dentinogenesis through upregulating the odontogenic differentiation of DPSCs, which provides a promising therapeutic approach for pulp regeneration.


Asunto(s)
Diferenciación Celular , Pulpa Dental , Vesículas Extracelulares , Gelatina , Metacrilatos , Odontogénesis , Regeneración , Células Madre , Diente Primario , Pulpa Dental/citología , Humanos , Vesículas Extracelulares/química , Gelatina/química , Gelatina/farmacología , Diferenciación Celular/efectos de los fármacos , Odontogénesis/efectos de los fármacos , Animales , Células Madre/efectos de los fármacos , Células Madre/citología , Células Madre/metabolismo , Regeneración/efectos de los fármacos , Diente Primario/citología , Metacrilatos/química , Metacrilatos/farmacología , Ratones , Proliferación Celular/efectos de los fármacos , Ratones Desnudos , Células Cultivadas , Hidrogeles/química , Hidrogeles/farmacología , Movimiento Celular/efectos de los fármacos
2.
Int J Mol Sci ; 25(6)2024 Mar 21.
Artículo en Inglés | MEDLINE | ID: mdl-38542525

RESUMEN

Among the many lysosomal storage disorders (LSDs) that would benefit from the establishment of novel cell models, either patient-derived or genetically engineered, is mucopolysaccharidosis type II (MPS II). Here, we present our results on the establishment and characterization of two MPS II patient-derived stem cell line(s) from deciduous baby teeth. To the best of our knowledge, this is the first time a stem cell population has been isolated from LSD patient samples obtained from the dental pulp. Taking into account our results on the molecular and biochemical characterization of those cells and the fact that they exhibit visible and measurable disease phenotypes, we consider these cells may qualify as a valuable disease model, which may be useful for both pathophysiological assessments and in vitro screenings. Ultimately, we believe that patient-derived dental pulp stem cells (DPSCs), particularly those isolated from human exfoliated deciduous teeth (SHEDs), may represent a feasible alternative to induced pluripotent stem cells (iPSCs) in many labs with standard cell culture conditions and limited (human and economic) resources.


Asunto(s)
Enfermedades por Almacenamiento Lisosomal , Mucopolisacaridosis II , Humanos , Células Madre , Línea Celular , Diente Primario , Lisosomas , Pulpa Dental , Diferenciación Celular/fisiología , Proliferación Celular
3.
J Nanobiotechnology ; 21(1): 458, 2023 Nov 29.
Artículo en Inglés | MEDLINE | ID: mdl-38031158

RESUMEN

BACKGROUND: Microglial activation in the spinal trigeminal nucleus (STN) plays a crucial role in the development of trigeminal neuralgia (TN). The involvement of adenosine monophosphate-activated protein kinase (AMPK) and N-methyl-D-aspartate receptor 1 (NMDAR1, NR1) in TN has been established. Initial evidence suggests that stem cells from human exfoliated deciduous teeth (SHED) have a potential therapeutic effect in attenuating TN. In this study, we propose that SHED-derived exosomes (SHED-Exos) may alleviate TN by inhibiting microglial activation. This study sought to assess the curative effect of SHED-Exos administrated through the tail vein on a unilateral infraorbital nerve chronic constriction injury (CCI-ION) model in mice to reveal the role of SHED-Exos in TN and further clarify the potential mechanism. RESULTS: Animals subjected to CCI-ION were administered SHED-Exos extracted by differential ultracentrifugation. SHED-Exos significantly alleviated TN in CCI mice (increasing the mechanical threshold and reducing p-NR1) and suppressed microglial activation (indicated by the levels of TNF-α, IL-1ß and IBA-1, as well as p-AMPK) in vivo and in vitro. Notably, SHED-Exos worked in a concentration dependent manner. Mechanistically, miR-24-3p-upregulated SHED-Exos exerted a more significant effect, while miR-24-3p-inhibited SHED-Exos had a weakened effect. Bioinformatics analysis and luciferase reporter assays were utilized for target gene prediction and verification between miR-24-3p and IL1R1. Moreover, miR-24-3p targeted the IL1R1/p-p38 MAPK pathway in microglia was increased in CCI mice, and participated in microglial activation in the STN. CONCLUSIONS: miR-24-3p-encapsulated SHED-Exos attenuated TN by suppressing microglial activation in the STN of CCI mice. Mechanistically, miR-24-3p blocked p-p38 MAPK signaling by targeting IL1R1. Theoretically, targeted delivery of miR-24-3p may offer a potential strategy for TN.


Asunto(s)
Exosomas , MicroARNs , Neuralgia del Trigémino , Ratones , Humanos , Animales , Neuralgia del Trigémino/metabolismo , Exosomas/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Proteínas Quinasas Activadas por AMP/metabolismo , MicroARNs/genética , MicroARNs/metabolismo
4.
Oral Dis ; 29(2): 725-734, 2023 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-34510661

RESUMEN

OBJECTIVE: Stem cells from human exfoliated deciduous teeth (SHED) have bone regeneration ability and potential therapeutic applications. CD146, a cell adhesion protein expressed by vascular endothelial cells, is involved in osteoblastic differentiation of stem cells. The effect of CD146 on SHED-mediated bone regeneration in vivo remains unknown. We aimed to establish efficient conditions for SHED transplantation. MATERIALS AND METHODS: SHED were isolated from the pulp of an extracted deciduous tooth and cultured; CD146-positive (CD146+ ) and CD146-negative (CD146- ) populations were sorted. Heterogeneous populations of SHED and CD146+ and CD146- cells were transplanted into bone defects generated in the skulls of immunodeficient mice. Micro-computed tomography was performed immediately and 4 and 8 weeks later. Histological and immunohistochemical assessments were performed 8 weeks later. RESULTS: Bone regeneration was observed upon transplantation with CD146+ and heterogeneous populations of SHED, with significantly higher bone regeneration observed with CD146+ cells. Bone regeneration was higher in the CD146- group than in the control group, but significantly lower than that in the other transplant groups at 4 and 8 weeks. Histological and immunohistochemical assessments revealed that CD146+ cells promoted bone regeneration and angiogenesis. CONCLUSION: Transplantation of CD146+ SHED into bone defects may be useful for bone regeneration.


Asunto(s)
Regeneración Ósea , Células Endoteliales , Humanos , Ratones , Animales , Antígeno CD146 , Microtomografía por Rayos X , Cráneo/cirugía , Diferenciación Celular , Diente Primario , Pulpa Dental
5.
Int Endod J ; 56(10): 1284-1300, 2023 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-37485765

RESUMEN

AIM: Recently, miR-27b-5p was shown to be abundantly expressed in extracellular vehicles (EVs) from the inflammatory microenvironment. This study determined the role of miR-27b-5p in regulating osteogenic and odontogenic differentiation of stem cells from human exfoliated deciduous teeth (SHEDs) and further examined the regulatory mechanism of bone morphogenetic protein receptor type-1A (BMPR1A). METHODOLOGY: Characteristics of SHEDs and SHEDs-EVs derived from SHEDs were evaluated respectively. The expression of miR-27b-5p in SHEDs and EVs was detected during osteo-induction. Mechanically, SHEDs were treated with miR-27b-5p mimics or an inhibitor, and the osteogenic/odontogenic differentiation and proliferation were assessed. Bioinformatic analysis and luciferase reporter were utilized for target gene prediction and verification. Finally, BMPR1A-overexpressed plasmids were transfected into SHEDs to investigate the participation of the BMPR1A/SMAD4 pathway. Data were analysed using Student's t-test, one-way analysis of variance and Chi-square test. RESULTS: MiR-27b-5p was expressed in both SHEDs and EVs and was significantly increased at the initial stage of differentiation and then decreased in a time-dependent manner (p < .01). Upregulation of miR-27b-5p significantly suppressed osteogenic/odontogenic differentiation of SHEDs and inhibited proliferation (p < .05), whereas inhibition of miR-27b-5p enhanced the differentiation (p < .05). Dual-luciferase reporter assay and pull-down assay confirmed the binding site between miR-27b-5p and BMPR1A (p < .05). The overexpression of BMPR1A rescued the effect of miR-27b-5p, while contributed to the decrease of pluripotency (p < .05). Additionally, miR-27b-5p maintained pluripotency in BMPR1A-overexpressed SHEDs (p < .05). CONCLUSIONS: MiR-27b-5p in SHEDs/EVs was inversely associated with differentiation and suppressed the osteogenic and odontogenic differentiation of SHEDs and maintained the pluripotency of SHEDs partly by shuttering BMPR1A-targeting BMP signalling. Theoretically, inhibition of miR-27b-5p represents a potential strategy to promote osteanagenesis and dentinogenesis. However, miR-27b-5p capsuled EVs might maintain cell pluripotency and self-renewal for non-cell-targeted therapy.


Asunto(s)
MicroARNs , Humanos , Receptores de Proteínas Morfogenéticas Óseas de Tipo 1/genética , Receptores de Proteínas Morfogenéticas Óseas de Tipo 1/metabolismo , Diferenciación Celular/fisiología , Células Cultivadas , MicroARNs/metabolismo , Osteogénesis/genética , Células Madre , Diente Primario
6.
Clin Oral Investig ; 27(1): 125-137, 2023 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-36018448

RESUMEN

OBJECTIVES: To evaluate hydrogel-based scaffolds embedded with parathyroid hormone (PTH)-loaded mesoporous bioactive glass (MBG) on the enhancement of bone tissue regeneration in vitro. MATERIALS AND METHODS: MBG was produced via sol-gel technique followed by PTH solution imbibition. PTH-loaded MBG was blended into the hydrogels and submitted to a lyophilisation process associated with a chemical crosslinking reaction to the production of the scaffolds. Characterisation of the MBG and PTH-loaded MBG scaffolds, including the scanning electron microscope (SEM) connected with an X-ray detector (EDX), Fourier transform infrared (FTIR), compression strength, rheological measurements, swelling and degradation rates, and PTH release analysis, were performed. Also, bioactivity using simulated-body fluid (SBF), biocompatibility (MTT), and osteogenic differentiation analyses (von Kossa and Alizarin Red stainings, and µ-computed tomography, µCT) of the scaffolds were carried out. RESULTS: SEM images demonstrated MBG particles dispersed into the hydrogel-based scaffold structure, which was homogeneously porous and well interconnected. EDX and FTIR revealed large amounts of carbon, oxygen, sodium, and silica in the scaffold composition. Bioactivity experiments revealed changes on sample surfaces over the analysed period, indicating the formation of carbonated hydroxyapatite; however, the chemical composition remained stable. PTH-loaded hydrogel-based scaffolds were biocompatible for stem cells from human-exfoliated deciduous teeth (SHED). A high quantity of calcium deposits on the extracellular matrix of SHED was found for PTH-loaded hydrogel-based scaffolds. µCT images showed MBG particles dispersed into the scaffolds' structure, and a porous, lamellar, and interconnected hydrogel architecture. CONCLUSIONS: PTH-loaded hydrogel-based scaffolds demonstrated consistent morphology and physicochemical properties for bone tissue regeneration, as well as bioactivity, biocompatibility, and osteoinductivity in vitro. Thus, the scaffolds presented here are recommended for future studies on 3D printing. CLINICAL RELEVANCE: Bone tissue regeneration is still a challenge for several approaches to oral and maxillofacial surgeries, though tissue engineering applying SHED, scaffolds, and osteoinductive mediators might help to overcome this clinical issue.


Asunto(s)
Osteogénesis , Andamios del Tejido , Humanos , Andamios del Tejido/química , Hormona Paratiroidea/farmacología , Hidrogeles/farmacología , Regeneración Ósea , Vidrio/química , Porosidad , Materiales Biocompatibles/química
7.
Int J Mol Sci ; 24(4)2023 Feb 17.
Artículo en Inglés | MEDLINE | ID: mdl-36835460

RESUMEN

Regenerative therapy for tissues by mesenchymal stem cell (MSCs) transplantation has received much attention. The cluster of differentiation (CD)146 marker, a surface-antigen of stem cells, is crucial for angiogenic and osseous differentiation abilities. Bone regeneration is accelerated by the transplantation of CD146-positive deciduous dental pulp-derived mesenchymal stem cells contained in stem cells from human exfoliated deciduous teeth (SHED) into a living donor. However, the role of CD146 in SHED remains unclear. This study aimed to compare the effects of CD146 on cell proliferative and substrate metabolic abilities in a population of SHED. SHED was isolated from deciduous teeth, and flow cytometry was used to analyze the expression of MSCs markers. Cell sorting was performed to recover the CD146-positive cell population (CD146+) and CD146-negative cell population (CD146-). CD146 + SHED without cell sorting and CD146-SHED were examined and compared among three groups. To investigate the effect of CD146 on cell proliferation ability, an analysis of cell proliferation ability was performed using BrdU assay and MTS assay. The bone differentiation ability was evaluated using an alkaline phosphatase (ALP) stain after inducing bone differentiation, and the quality of ALP protein expressed was examined. We also performed Alizarin red staining and evaluated the calcified deposits. The gene expression of ALP, bone morphogenetic protein-2 (BMP-2), and osteocalcin (OCN) was analyzed using a real-time polymerase chain reaction. There was no significant difference in cell proliferation among the three groups. The expression of ALP stain, Alizarin red stain, ALP, BMP-2, and OCN was the highest in the CD146+ group. CD146 + SHED had higher osteogenic differentiation potential compared with SHED and CD146-SHED. CD146 contained in SHED may be a valuable population of cells for bone regeneration therapy.


Asunto(s)
Osteogénesis , Células Madre , Diente Primario , Humanos , Antígeno CD146/metabolismo , Diferenciación Celular , Proliferación Celular , Células Cultivadas , Pulpa Dental/metabolismo , Osteocalcina/metabolismo , Células Madre/citología , Diente Primario/citología
8.
J Oral Rehabil ; 49(2): 219-227, 2022 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-34386989

RESUMEN

BACKGROUND: Stem cells from human exfoliated deciduous teeth (SHED) have excellent immunomodulatory and neuroprotective abilities. It is possible that systemic SHED transplantation could ameliorate trigeminal neuralgia. The phosphorylation of c-Jun contributes to the development of hyperalgesia and allodynia. OBJECTIVE: The present study aimed to evaluate whether systemic SHED transplantation could lead to analgesic effects by regulating peripheral c-Jun in the trigeminal ganglia (TG) in a rat model of trigeminal neuralgia. METHODS: Chronic constriction injury of the infraorbital nerve (CCI-ION) was performed to establish a rat pain model. SHED were obtained from discarded exfoliated deciduous teeth from children and transplanted by a single infusion through the tail vein. SHED were labelled with the PKH26 red fluorescent cell linker mini kit for tract distribution. The mechanical threshold was determined using von Frey filaments. The mRNA levels of c-Jun in the ipsilateral TG were quantified. The phosphorylation of c-Jun in the ipsilateral TG was assessed by immunohistochemistry and Western blotting. RESULTS: PKH26-labelled SHED were distributed to both sides of TG, lung, liver and spleen. Systemic SHED transplantation significantly elevated the mechanical thresholds in CCI-ION rats and blocked the upregulation of c-Jun mRNA levels in the TG caused by nerve ligation. The activation of c-Jun in the TG was blocked by SHED transplantation. CONCLUSIONS: These findings demonstrate that systemic SHED administration reverts trigeminal neuralgia via downregulation of c-Jun in the TG.


Asunto(s)
Neuralgia del Trigémino , Animales , Regulación hacia Abajo , Humanos , Hiperalgesia , Dolor , Ratas , Ratas Sprague-Dawley , Células Madre , Diente Primario
9.
BMC Oral Health ; 22(1): 238, 2022 06 17.
Artículo en Inglés | MEDLINE | ID: mdl-35715777

RESUMEN

BACKGROUND: This in vivo experimental study investigated the effect of stem cells from human exfoliated deciduous teeth (SHEDs) on early osteogenesis around implants. METHODS: In four healthy adult male Beagle dogs, the left mandibular received implants and SHED as the experimental group, and the right mandibular received implants and phosphate-buffered saline as the control group. The Beagle dogs were randomly divided into groups A and B, which were sacrificed at 2 and 4 weeks after implantation. Micro-computed tomography and histological analysis were used to investigate the effect of SHED-loading on the early osseointegration around the implants. RESULTS: The total bone-to-implant contact (BIC%) and interthread bone improved significantly. The analysis of the bone volume fraction and trabecular thickness showed that the bone trabecula around the implants in the SHEDs group was thicker and denser than that in the control group, suggesting a better osseointegration. CONCLUSIONS: The application of implants pre-adhered with SHEDs improved and accelerated early osseointegration around the implant, resulting in thicker and denser trabecular bone.


Asunto(s)
Implantes Dentales , Oseointegración , Animales , Implantación Dental Endoósea/métodos , Perros , Humanos , Masculino , Mandíbula/patología , Mandíbula/cirugía , Células Madre , Diente Primario , Microtomografía por Rayos X
10.
Int J Mol Sci ; 22(5)2021 Feb 25.
Artículo en Inglés | MEDLINE | ID: mdl-33668763

RESUMEN

A subpopulation of mesenchymal stem cells, developmentally derived from multipotent neural crest cells that form multiple facial tissues, resides within the dental pulp of human teeth. These stem cells show high proliferative capacity in vitro and are multipotent, including adipogenic, myogenic, osteogenic, chondrogenic, and neurogenic potential. Teeth containing viable cells are harvested via minimally invasive procedures, based on various clinical diagnoses, but then usually discarded as medical waste, indicating the relatively low ethical considerations to reuse these cells for medical applications. Previous studies have demonstrated that stem cells derived from healthy subjects are an excellent source for cell-based medicine, tissue regeneration, and bioengineering. Furthermore, stem cells donated by patients affected by genetic disorders can serve as in vitro models of disease-specific genetic variants, indicating additional applications of these stem cells with high plasticity. This review discusses the benefits, limitations, and perspectives of patient-derived dental pulp stem cells as alternatives that may complement other excellent, yet incomplete stem cell models, such as induced pluripotent stem cells, together with our recent data.


Asunto(s)
Pulpa Dental/citología , Enfermedades Genéticas Congénitas/patología , Células Madre Mesenquimatosas/citología , Modelos Biológicos , Diferenciación Celular , Humanos
11.
J Clin Pediatr Dent ; 45(2): 104-111, 2021 Apr 01.
Artículo en Inglés | MEDLINE | ID: mdl-33951164

RESUMEN

BACKGROUND: Deciduous teeth undergo the physiologic process of resorption, during which the remnant pulp undergoes activation. However, the quality of stem cells obtained at various stages of root resorption has not been documented. OBJECTIVE: To isolate and characterize stem cells from deciduous teeth with varying levels of root resorption. STUDY DESIGN: Healthy primary anterior teeth were extracted according to the treatment needs of the patient. The teeth were categorized into SHED(1/3)- teeth with 0 to 1/3rd root resorption, SHED(2/3)- teeth with 1/3rd to 2/3rd root resorption, and SHED(COMP)- teeth with more than 2/3rd root resorption. SHED were characterized based on their morphology, viability, proliferation rate, population doubling time, expression of cell surface markers, and in vitro differentiation potential into osteocytes and adipocytes. RESULTS: SHED from all three groups demonstrated largely similar morphological and cellular characteristics. However, SHED(2/3) showed relatively better characteristics in terms of growth kinetics and phenotypic marker expression. Also, the differentiation ability for osteogenic and adipogenic cell lineages was slightly higher in SHED(1/3) and SHED(2/3) compared with SHED(COMP). CONCLUSION: Based on the cellular, phenotypic and biological characteristics, it is suggested that SHED (2/3) could be a useful source for tissue regeneration, and warrants further investigations.


Asunto(s)
Resorción Radicular , Proliferación Celular , Células Cultivadas , Pulpa Dental , Humanos , Células Madre , Diente Primario
12.
Biochem Cell Biol ; 98(3): 415-425, 2020 06.
Artículo en Inglés | MEDLINE | ID: mdl-31794246

RESUMEN

Mesenchymal stem cells (MSCs) have proven powerful potential for cell-based therapy both in regenerative medicine and disease treatment. Human umbilical cords and exfoliated deciduous teeth are the main sources of MSCs with no donor injury or ethical issues. The goal of this study was to investigate the differences in the biological characteristics of human umbilical cord mesenchymal stem cells (UCMSCs) and stem cells from human exfoliated deciduous teeth (SHEDs). UCMSCs and SHEDs were identified by flow cytometry. The proliferation, differentiation, migration, chemotaxis, paracrine, immunomodulatory, neurite growth-promoting capabilities, and acetaldehyde dehydrogenase (ALDH) activity were comparatively studied between these two MSCs in vitro. The results showed that both SHEDs and UCMSCs expressed cell surface markers characteristic of MSCs. Furthermore, SHEDs exhibited better capacity for proliferation, migration, promotion of neurite growth, and chondrogenic differentiation. Meanwhile, UCMSCs showed more outstanding adipogenic differentiation and chemotaxy. Additionally, there were no significant differences in osteogenic differentiation, immunomodulatory capacity, and the proportion of ALDHBright compartment. Our findings indicate that although both UCMSCs and SHEDs are mesenchymal stem cells and presented some similar biological characteristics, they also have differences in many aspects, which might be helpful for developing future clinical cellular therapies.


Asunto(s)
Células Madre Mesenquimatosas/citología , Diente Primario/citología , Cordón Umbilical/citología , Adipogénesis , Aldehído Oxidorreductasas/metabolismo , Animales , Diferenciación Celular , Movimiento Celular , Proliferación Celular , Quimiotaxis , Condrogénesis , Humanos , Ratones , Células 3T3 NIH , Neuritas/metabolismo , Osteogénesis
13.
IUBMB Life ; 72(4): 665-676, 2020 04.
Artículo en Inglés | MEDLINE | ID: mdl-31889420

RESUMEN

Stem cells from human exfoliated deciduous teeth (SHEDs) are highly proliferative, clonogenic, and multipotent stem cells with a neural crest cell origin. This property could be a desirable option for potential therapeutic applications. In this study, we focus on the effects of Rho kinase inhibitors Y-27632 and Noggin on the proliferation of SHEDs and their differentiation into neuron-like cells. SHEDs were extracted from 10 samples of deciduous teeth obtained from healthy children aged from 5 to 10. The passaged SHEDs were transfected with Noggin, Y-27632, or their combination. By means of MTT and colony formation assays, the effects of Y-27632 and Noggin on cell viability and colony formation were detected. Cellular morphology and neurosphere formation were observed under a microscope. Y-27632 transfection in SHEDs showed enhanced cell viability, colony formation, and neurosphere formation indicating that Y-27632 could promote cell proliferation of SHEDs. Furthermore, we observed that the SHEDs treated with Noggin in combination with Y-27632 displayed typical neuron-like cell morphology and reticular processes. Noggin or Y-27632 alone or in combination induced obviously increased NSE, Nestin, and GFAP levels, which were highest in SHEDs treated with the combination of Noggin and Y-27632. These findings suggest that Y-27632 promotes the proliferation of SHEDs, and Y-27632 and Noggin in combination have a synergistic effect on promoting differentiation of SHEDs into neuron-like cells.


Asunto(s)
Amidas/farmacología , Proteínas Portadoras/genética , Neuronas/citología , Piridinas/farmacología , Células Madre/efectos de los fármacos , Diente Primario/citología , Adipocitos/citología , Proteínas Portadoras/metabolismo , Diferenciación Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Separación Celular , Células Cultivadas , Niño , Preescolar , Expresión Génica/efectos de los fármacos , Humanos , Neuronas/fisiología , Osteoblastos/citología , Células Madre/citología , Quinasas Asociadas a rho/antagonistas & inhibidores
14.
Oral Dis ; 26(6): 1275-1283, 2020 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-32248596

RESUMEN

OBJECTIVES: We aimed to investigate whether the mesenchymal stem cell-endothelial cell crosstalk enhances angiogenic factor expression via nuclear factor-kappa B (NF-κB)-dependent mechanisms. MATERIALS AND METHODS: Human dermal microvascular endothelial cells (HDMECs) and stem cells from human exfoliated deciduous teeth (SHEDs) were cocultured for 96 hr, in the presence of NF-κB decoy oligodeoxynucleotides (ODNs) or scramble (control). Vascular endothelial cell growth factor (VEGF) and phospho-NF-κB p65 were measured with enzyme-linked immunosorbent assay. Angiogenesis-related gene expression was analyzed with microarray analysis followed by real-time polymerase chain reaction. Tube formation assay was conducted in the presence of NF-κB decoy. RESULTS: The VEGF and phospho-NF-κB p65 levels were significantly higher in the coculture with NF-κB decoy scramble than in single culture and coculture with NF-κB decoy ODN. Microarray analysis of SHEDs and HDMECs with NF-κB decoy scramble showed higher expression of proangiogenic genes, Bcl-2, NF-κB1, VEGFA, CXCL8, and CXCR1, and lower expression of proapoptotic genes, Bax and Caspase 9, compared to cells with NF-κB decoy ODN. Real-time PCR results for Bcl-2 and CXCL8 showed a similar trend. Tube formation assay showed more tube development in the presence of NF-κB decoy scramble. CONCLUSION: The SHED-HDMEC crosstalk enhanced proangiogenic factor expression via NF-κB-dependent pathways.

15.
Clin Oral Investig ; 24(1): 167-180, 2020 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-31069538

RESUMEN

OBJECTIVES: To investigate the effects of low-level laser irradiation (LLLI) on viability/proliferation, migration, osteo/odontogenic differentiation, and in vitro biomineralization of stem cells from human exfoliated deciduous teeth (SHED). MATERIALS AND METHODS: SHED cultures were established by enzymatic dissociation from pulps of deciduous teeth. SHED were irradiated with a diode laser (InGaAsP; 940 nm; 0.2 W, continuous mode) at energy fluences 4, 8, and 16 J/cm2 in the dark, while non-irradiated SHED served as control. Cell viability/proliferation was evaluated by MTT assay and cell mobilization by Transwell™ migration assay. Expression of osteo/odontogenesis-related genes (ALP, BMP-2, BGLAP, DSPP, MSX2, RUNX2) was assessed by real-time PCR, while in vitro biomineralization by Alizarin Red staining. Statistical analysis was performed by two-way ANOVA and Tukey's post hoc tests (*p < 0.05, **p < 0.01). RESULTS: Statistically significant stimulation of cell viability/proliferation was observed at all energy fluences, reaching the highest effect for the 4 and 16 J/cm2. Although the 8 J/cm2 fluence showed the lowest stimulatory effect on cell viability/proliferation, it was the most effective in inducing SHED migration, upregulation of odontogenesis-related genes (DSPP, ALP, BMP-2) at specific time-points, and the in vitro biomineralization potential of SHED compared to the other two energy fluences. CONCLUSIONS: LLLI proved beneficial in promoting SHED biological processes critical for pulp repair in deciduous teeth. Overall, the 8 J/cm2 energy fluence showed the most beneficiary effects. CLINICAL RELEVANCE: These results provide insights on a narrow "therapeutic window" of LLLI application in vital pulp therapies of deciduous teeth, paving the way for the establishment of effective clinical protocols.


Asunto(s)
Rayos Láser , Células Madre , Diente Primario , Diferenciación Celular , Proliferación Celular , Supervivencia Celular , Células Cultivadas , Pulpa Dental , Humanos
16.
Zhong Nan Da Xue Xue Bao Yi Xue Ban ; 45(6): 678-683, 2020 Jun 28.
Artículo en Inglés, Zh | MEDLINE | ID: mdl-32879125

RESUMEN

OBJECTIVES: To explore the difference in odontoblast differentiation capacity between stem cells from human exfoliated deciduous teeth (SHED) and dental pulp stem cells (DPSCs), and to examine the expression level of ephrinB1 in odontoblast differentiation of these stem cells. METHODS: The stems cells were divided into a SHED group and a DPSCs group. After odontoblast differentiation induction, the above 2 groups were also randomly divided into a 3 d group and a 7 d group, respectively.The calcium deposition was detected by alkaline phosphatase (ALP) staining and alizarin red staining.The mRNA and protein expressions of ephrinB1, dentin matrix protein-1 (DMP-1) and dentin sialophosphoprotein (DSPP) were detected by real-time PCR and Western blotting. RESULTS: ALP staining and alizarin red staining showed that there was stronger mineralization capacity in the SHED group than that in the DPSCs group. The relative mRNA and protein expressions of DMP-1, DSPP, and ephrinB1 in the SHED group were higher than those in the DPSCs group except for the protein expression of DMP-1 in the SHED 3 d group (all P<0.05). CONCLUSIONS: SHED has stronger odontoblast differentiation capacity than DPSCs. In addition, ephrinB1 may be involved in the processes of odontoblast differentiation in the SHED and DPSCs.


Asunto(s)
Odontoblastos , Osteogénesis , Diferenciación Celular , Proliferación Celular , Células Cultivadas , Pulpa Dental , Humanos , Células Madre , Diente Primario
17.
J Cell Biochem ; 120(2): 1464-1476, 2019 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-30317665

RESUMEN

Previous studies have revealed that long noncoding RNA (lncRNA) and microRNA play a crucial role in autism, which is a childhood neurodevelopmental disorder with complicated genetic origins. Hence, the study concerns whether lncRNA C21orf121/bone morphogenetic proteins 2 (BMP2)/miR-140-5p gene network affects directed differentiation of stem cells from human exfoliated deciduous teeth (SHED) to neuronal cells in rats with autism. Autism models were successfully established. The neuron cells that differentiated from SHED cell were identified. The expression of lncRNA C21orf121, miR-140-5p, BMP2, Nestin, ßIII-tubulin, and microtubule-associated protein 2 (MAP2) and the expression of neuron-specific enolase (NSE) were examined. Besides, the gap junction (GJ) function of SHED, the intracellular free Ca 2+ concentration, and the social behavior and repetitive stereotyped movements of rats in autism were detected. The target relationship between lncRNA C21orf121 and miR-140-5p and that between miR-140-5p and BMP2 were also verified. Firstly, we successfully isolated SHED and identified the differentiated neurons of SHED. Besides, the expression of BMP2, MAP2, Nestin, ßIII-tubulin, NSE positive rate, GJ function, and intracellular free Ca 2+ concentration were increased with the upregulation of C21orf121 and downregulation of miR-140-5p, and accumulated time of repetitive stereotyped movements decreased and the frequency of social behavior increased. The results indicate that lncRNA C21orf121 as a competing endogenous RNA competes with BMP2 binding to miR-140-5p, thereby promoting SHED to differentiate into neuronal cells via upregulating BMP2 expression.

18.
Biochem Biophys Res Commun ; 508(3): 850-856, 2019 01 15.
Artículo en Inglés | MEDLINE | ID: mdl-30528238

RESUMEN

Enzymatic antioxidant systems, mainly involving mitochondria, are critical for minimizing the harmful effects of reactive oxygen species, and these systems are enhanced by interactions with nonenzymatic antioxidant nutrients. Because fetal growth requires extensive mitochondrial respiration, pregnant women and fetuses are at high risk of exposure to excessive reactive oxygen species. The enhancement of the antioxidant system, e.g., by nutritional management, is therefore critical for both the mother and fetus. Folic acid supplementation prevents homocysteine accumulation and epigenetic dysregulation associated with one-carbon metabolism. However, few studies have examined the antioxidant effects of folic acid for healthy pregnancy outcomes. The purpose of this study was to elucidate the association between the antioxidant effect of folic acid and mitochondria in undifferentiated cells during fetal growth. Neural crest-derived dental pulp stem cells of human exfoliated deciduous teeth were used as a model of undifferentiated cells in the fetus. Pyocyanin induced excessive reactive oxygen species, resulting in a decrease in cell growth and migration accompanied by mitochondrial fragmentation and inactivation in dental pulp stem cells. This damage was significantly improved by folic acid, along with decreased mitochondrial reactive oxygen species, PGC-1α upregulation, DRP1 downregulation, mitochondrial elongation, and increased ATP production. Folic acid may protect undifferentiated cells from oxidative damage by targeting mitochondrial activation. These results provide evidence for a new benefit of folic acid in pregnant women and fetuses.


Asunto(s)
Antioxidantes/farmacología , Pulpa Dental/citología , Ácido Fólico/farmacología , Mitocondrias/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Células Madre/efectos de los fármacos , Diente Primario/citología , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Niño , Humanos , Piocianina/farmacología , Especies Reactivas de Oxígeno/metabolismo , Células Madre/citología , Células Madre/metabolismo
19.
Biochem Biophys Res Commun ; 516(1): 127-132, 2019 08 13.
Artículo en Inglés | MEDLINE | ID: mdl-31202461

RESUMEN

Orofacial clefts (OFCs) are among the most common congenital craniofacial malformations, including cleft lip with or without cleft palate as the core symptoms. Developmental or functional defects in neural crest cells (NCCs) that contribute to craniofacial morphogenesis are involved in OFC development. Previous studies have suggested that oxidative stress in NCCs is involved in the development of OFCs, suggesting that the anti-oxidative activity of folic acid (FA) could have protective effects. However, studies of human-derived NCCs are limited, as these cells are predominantly active during the embryonic stage. In this study, the effects of oxidative stress and FA were evaluated in human OFCs. In particular, NCC-derived stem cells from human exfoliated deciduous teeth (SHEDs) were obtained from 3 children with non-syndromic cleft lip with cleft palate (CLPs) and from 3 healthy children (CTRLs). Mitochondrial reactive oxygen species (ROS) levels were significantly higher in CLPs than in CTRLs and were associated with lower mRNA expression levels of superoxide dismutase 1 (SOD1) and decreased cell mobility. In addition, significantly greater vulnerability to pyocyanin-induced ROS, mimicking exogenous ROS, was observed in CLPs than in CTRLs. These vulnerabilities to endogenous and exogenous ROS in CLPs were significantly improved by FA. These results indicated that the transcriptional dysregulation of SOD1 in NCCs is an oxidative stress-related pathological factor in OFCs, providing novel evidence for the benefits of perinatal anti-oxidant supplementation, including FA, for the management of these common deformities.


Asunto(s)
Antioxidantes/uso terapéutico , Labio Leporino/tratamiento farmacológico , Fisura del Paladar/tratamiento farmacológico , Ácido Fólico/uso terapéutico , Estrés Oxidativo/efectos de los fármacos , Diente Primario/efectos de los fármacos , Células Cultivadas , Niño , Labio Leporino/metabolismo , Fisura del Paladar/metabolismo , Pulpa Dental/citología , Pulpa Dental/efectos de los fármacos , Pulpa Dental/metabolismo , Humanos , Células Madre/citología , Células Madre/efectos de los fármacos , Células Madre/metabolismo , Diente Primario/citología , Diente Primario/metabolismo , Complejo Vitamínico B/uso terapéutico
20.
Int J Mol Sci ; 20(8)2019 Apr 15.
Artículo en Inglés | MEDLINE | ID: mdl-30991705

RESUMEN

Oxidative stress causes severe tissue injury of the central nervous system in ischemic brain damage (IBD), traumatic brain injury (TBI) and neurodegenerative disorders. In this study, we used hydrogen peroxide (H2O2) to induce oxidative stress in organotypic brain slice cultures (OBSCs), and investigated the protective effects of oxidative stress-tolerant (OST) stem cells harvested from human exfoliated deciduous teeth (SHED) which were co-cultivated with OBSCs. Using presto blue assay and immunostaining, we demonstrated that both normal SHED and OST-SHED could prevent H2O2-induced cell death, and increase the numbers of mature neuron and neuronal progenitors in the hippocampus of OBSCs. During co-cultivation, OST-SHED, but not normal SHED, exhibited neuronal cell morphology and expressed neuronal markers. Results from ELISA showed that both normal SHED and OST-SHED significantly decreased oxidative DNA damage in H2O2-treated OBSCs. SHED could also produce neurotrophic factor BDNF (brain derived neurotrophic factor) and promoted the production of IL-6 in OBSCs. Although OST-SHED had lower cell viability, the neuronal protection of OST-SHED was significantly superior to that of normal SHED. Our findings suggest that SHED, especially OST-SHED, could prevent oxidative stress induced brain damage. OST-SHED can be explored as a new therapeutic tool for IBD, TBI and neurodegenerative disorders.


Asunto(s)
Encéfalo/citología , Técnicas de Cocultivo , Neuronas/citología , Neuroprotección , Estrés Oxidativo , Células Madre/citología , Diente Primario/citología , Animales , Muerte Celular , Supervivencia Celular , Células Cultivadas , Niño , Femenino , Humanos , Peróxido de Hidrógeno/metabolismo , Ratones Endogámicos ICR , Neurogénesis
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