RESUMEN
This study aimed to illustrate the biological behavior and changes in cell function during the progression of apical periodontitis in deciduous teeth and to explore the underlying molecular mechanism. Deciduous teeth periodontal ligament stem cells (DePDLSCs) were derived and their identity was confirmed. The viability, inflammation, and osteogenic ability of cells were tested by exposing them to various concentrations of lipopolysaccharide (LPS) (0-100 µg/mL) using the cell counting kit-8 (CCK-8) assay, reverse transcription polymerase chain reaction (real-time PCR), alkaline phosphatase (ALP) staining, and ALP activity assay. In addition, osteogenic-induced cells with and without 10 µg/mL LPS were harvested for high-throughput sequencing. Based on sequencing data, proinflammatory factors and ALP expression were measured after interference with the PI3K-AKT signaling pathway activator, 740Y-P. LPS biphasically affected the proliferation and osteogenesis of DePDLSCs. Low concentrations of LPS showed stimulatory effects, whereas inhibitory effects were observed at high concentrations. Sequencing analysis showed that the PI3K-AKT signaling pathway was significantly downregulated when DePDLSCs were treated with 10 µg/mL LPS. The LPS-induced inflammation and osteogenesis inhibition of DePDLSCs were partially rescued by 740Y-P treatment. In conclusion, LPS affected DePDLSCs proliferation and osteogenesis in a biphasic manner. Moderate activation of PI3K-AKT signaling pathway was beneficial for osteogenic differentiation and anti-inflammatory effect in DePDLSCs. This research may provide etiological probes for apical periodontitis and its treatment.
Asunto(s)
Osteogénesis , Ligamento Periodontal , Células Madre , Diente Primario , Ligamento Periodontal/citología , Ligamento Periodontal/efectos de los fármacos , Humanos , Osteogénesis/efectos de los fármacos , Osteogénesis/fisiología , Diente Primario/citología , Células Madre/efectos de los fármacos , Lipopolisacáridos/farmacología , Diferenciación Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Inflamación , Transducción de Señal/efectos de los fármacos , Células Cultivadas , Reacción en Cadena en Tiempo Real de la Polimerasa , Fosfatasa Alcalina/metabolismo , Fosfatasa Alcalina/análisisRESUMEN
Periodontal regeneration is a challenge, and tissue engineering based on periodontal ligament stem cells (PDLSCs) has been shown to be a promising alternative to this process. However, the need for scaffolds has limited the therapeutic use of PDLSCs. In this context, scaffold-free tissue engineering using the cell sheet (CS) technique has been developed as an alternative approach to improve tissue regeneration. Previously, we showed that Protease-activated receptor-1 (PAR1) can regulate PDLSCs. Herein, we evaluate whether PAR1 influences osteogenesis in CSs produced from PDLSCs, without the use of scaffolds. PDLSCs were isolated and immunophenotyped. Then, CSs were obtained by supplementing the culture medium with ascorbic acid (50 µg/mL), and PAR1 was activated through its agonist peptide (100 nM). Scaffold-free 3D CSs were successfully produced from PDLSCs, and they showed higher proliferation potential than isolated PDLSCs. Also, PAR1 activation decreased senescence and improved osteogenic differentiation of CSs by increasing mineralized nodule deposition and alkaline phosphatase concentration; PAR1 also modulated osteogenic markers at the gene and protein levels. We further demonstrated that this effect was regulated by Wnt, TGF-ßI, MEK, p38 MAPK, and FGF/VEGF signaling pathways in PDLSCs (p < 0.05%). Overall, PAR1 activation increased osteogenic activity in CSs, emerging as a promising scaffold-free therapeutic approach for periodontal regeneration.
Asunto(s)
Diferenciación Celular , Proliferación Celular , Osteogénesis , Ligamento Periodontal , Receptor PAR-1 , Células Madre , Ingeniería de Tejidos , Ligamento Periodontal/citología , Osteogénesis/efectos de los fármacos , Osteogénesis/fisiología , Humanos , Diferenciación Celular/efectos de los fármacos , Células Madre/fisiología , Células Madre/efectos de los fármacos , Células Cultivadas , Proliferación Celular/efectos de los fármacos , Ingeniería de Tejidos/métodos , Fosfatasa Alcalina/análisis , Fosfatasa Alcalina/metabolismo , Reproducibilidad de los Resultados , Adolescente , Factores de Tiempo , Reacción en Cadena en Tiempo Real de la Polimerasa , Inmunofenotipificación , Análisis de VarianzaRESUMEN
OBJECTIVE: Several materials have been developed to preserve pulp vitality. They should have ideal cytocompatibility characteristics to promote the activity of stem cells of human exfoliated deciduous teeth (SHED) and thus heal pulp tissue. OBJECTIVE: To evaluate the cytotoxicity of different dilutions of bioceramic material extracts in SHED. METHODOLOGY: SHED were immersed in αMEM + the material extract according to the following experimental groups: Group 1 (G1) -BBio membrane, Group 2 (G2) - Bio-C Repair, Group 3 (G3) - MTA Repair HP, Group 4 (G4) - TheraCal LC, and Group 5 (G5) - Biodentine. Positive and negative control groups were maintained respectively in αMEM + 10% FBS and Milli-Q Water. The methods to analyze cell viability and proliferation involved MTT and Alamar Blue assays at 24, 48, and 72H after the contact of the SHED with bioceramic extracts at 1:1 and 1:2 dilutions. Data were analyzed by the three-way ANOVA, followed by Tukey's test (p<0.05). RESULTS: At 1:1 dilution, SHED in contact with the MTA HP Repair extract showed statistically higher cell viability than the other experimental groups and the negative control (p<0.05), except for TheraCal LC (p> 0.05). At 1:2 dilution, BBio Membrane and Bio-C showed statistically higher values in intra- and intergroup comparisons (p<0.05). BBio Membrane, Bio-C Repair, and Biodentine extracts at 1:1 dilution showed greater cytotoxicity than 1:2 dilution in all periods (p<0.05). CONCLUSION: MTA HP Repair showed the lowest cytotoxicity even at a 1:1 dilution. At a 1:2 dilution, the SHED in contact with the BBio membrane extract showed high cell viability. Thus, the BBio membrane would be a new non-cytotoxic biomaterial for SHED. Results offer possibilities of biomaterials that can be indicated for use in clinical regenerative procedures of the dentin-pulp complex.
Asunto(s)
Compuestos de Aluminio , Materiales Biocompatibles , Compuestos de Calcio , Proliferación Celular , Supervivencia Celular , Cerámica , Pulpa Dental , Combinación de Medicamentos , Ensayo de Materiales , Óxidos , Silicatos , Células Madre , Diente Primario , Humanos , Diente Primario/efectos de los fármacos , Silicatos/química , Silicatos/toxicidad , Silicatos/farmacología , Supervivencia Celular/efectos de los fármacos , Compuestos de Calcio/química , Compuestos de Calcio/farmacología , Compuestos de Calcio/toxicidad , Células Madre/efectos de los fármacos , Factores de Tiempo , Óxidos/química , Óxidos/toxicidad , Proliferación Celular/efectos de los fármacos , Pulpa Dental/efectos de los fármacos , Pulpa Dental/citología , Cerámica/química , Cerámica/toxicidad , Compuestos de Aluminio/química , Compuestos de Aluminio/toxicidad , Materiales Biocompatibles/química , Materiales Biocompatibles/farmacología , Análisis de Varianza , Reproducibilidad de los Resultados , Bismuto/química , Bismuto/toxicidad , Bismuto/farmacología , Células Cultivadas , Valores de Referencia , Sales de Tetrazolio , Xantenos/química , OxazinasRESUMEN
Osteoporosis is the most common metabolic bone disorder and is associated with a high incidence of fractures. Angiogenesis and adequate blood flow are important during bone repair and maintenance. Estrogens play a key role in bone formation, in the prevention of bone resorption and vasculature maintenance. Hormone replacement therapy (HRT) has been used with great benefits for bone fracture prevention but has been linked to the development of serious important side effects, including cancer and stroke. Phytoestrogens are an attractive alternative to HRT because their chemical structure is similar to estradiol but, they could behave as selective modulators: acting as antagonists of estrogen receptors in the breast and endometrium and as agonists in the vascular endothelium and bone. Hops contain a wide variety of phytoestrogens that have individually been shown to possess estrogenic activity by either blocking or mimicking. In this study we have to evaluate the in vitro effects and mechanisms of action of hops extracts on the osteogenic and adipogenic capacity of bone marrow progenitor cells (BMPCs), and the angiogenic potential of EA.hy926 endothelial cells. We show that hops extracts increase the proliferative capacity of BMPCs and promote their osteogenic differentiation while decreasing their pro-osteoclastogenic capacity; and that these effects are mediated by the MAPK pathway. Additionally, hops extracts prevent the adipogenic differentiation of BMPCs and promote endothelial cell activity, by mechanisms also partially mediated by MAPK.
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Células de la Médula Ósea , Diferenciación Celular , Proliferación Celular , Células Endoteliales , Humulus , Osteogénesis , Extractos Vegetales , Humulus/química , Osteogénesis/efectos de los fármacos , Humanos , Células Endoteliales/efectos de los fármacos , Células Endoteliales/metabolismo , Extractos Vegetales/farmacología , Proliferación Celular/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Animales , Células de la Médula Ósea/efectos de los fármacos , Células de la Médula Ósea/metabolismo , Células de la Médula Ósea/citología , Células Madre/efectos de los fármacos , Células Madre/metabolismo , Células Madre/citología , Neovascularización Fisiológica/efectos de los fármacos , Fitoestrógenos/farmacología , Adipogénesis/efectos de los fármacos , Ratones , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Células Cultivadas , Línea CelularRESUMEN
OBJECTIVES: To evaluate the effects of dentin biomodification agents (Proanthocyanidin (PAC), Cardol (CD) and Cardol-methacrylate (CDMA) on dentin hydrophilicity by contact angle measurement, viability of dental pulp stem cells (DPSCs) and nanomechanical properties of the hybrid layer (HL). METHODS: CDMA monomer was synthesized from cardol through methacrylic acid esterification. Human extracted third molars were used for all experiments. For nanomechanical tests, specimens were divided in four groups according to the primer solutions (CD, CDMA, PAC and control) were applied before adhesive and composite coating. Nanomechanical properties of the HL were analyzed by nanoindentation test using a Berkovich probe in a nanoindenter. Wettability test was performed on dentin surfaces after 1 min biomodification and measured by contact angle analysis. Cytotoxicity was assessed by a MTT assay with DPSCs after 48 and 72 h. Data were analyzed with Student's t test or Two-way ANOVA and Tukey HSD test (p < 0.05). RESULTS: CD and CDMA solutions achieved greater hydrophobicity and increased the water-surface contact angles when compared to PAC and control groups (p < 0.05). PAC group showed a greater reduction of elastic modulus in nanoindentation experiments when compared to CD and CDMA groups (p < 0.05) after 4 months of aging. CD inhibited cell proliferation compared to all further materials (p < 0.05), whilst CDMA and PAC indicated no cell cytotoxicity to human DPSCs. SIGNIFICANCE: Cardol-methacrylate provided significantly higher hydrophobicity to dentin and demonstrated remarkable potential as collagen crosslinking, attaining the lowest decrease of HL's mechanical properties. Furthermore, such monomer did not affect pulp cytotoxicity, thereby highlighting promising feasibility for clinical applications.
Asunto(s)
Supervivencia Celular , Dentina , Metacrilatos , Humectabilidad , Humanos , Supervivencia Celular/efectos de los fármacos , Metacrilatos/química , Metacrilatos/farmacología , Dentina/química , Proantocianidinas/farmacología , Proantocianidinas/química , Ensayo de Materiales , Pulpa Dental/citología , Tercer Molar , Células Madre/efectos de los fármacos , Propiedades de Superficie , Interacciones Hidrofóbicas e Hidrofílicas , Técnicas In VitroRESUMEN
The dental implant surface plays a crucial role in osseointegration. The topography and physicochemical properties will affect the cellular functions. In this research, four distinct titanium surfaces have been studied: machined acting (MACH), acid etched (AE), grit blasting (GBLAST), and a combination of grit blasting and subsequent acid etching (GBLAST + AE). Human amniotic mesenchymal (hAMSCs) and epithelial stem cells (hAECs) isolated from the amniotic membrane have attractive stem-cell properties. They were cultured on titanium surfaces to analyze their impact on biological behavior. The surface roughness, microhardness, wettability, and surface energy were analyzed using interferometric microscopy, Vickers indentation, and drop-sessile techniques. The GBLAST and GBLAST + AE surfaces showed higher roughness, reduced hydrophilicity, and lower surface energy with significant differences. Increased microhardness values for GBLAST and GBLAST + AE implants were attributed to surface compression. Cell viability was higher for hAMSCs, particularly on GBLAST and GBLAST + AE surfaces. Alkaline phosphatase activity enhanced in hAMSCs cultured on GBLAST and GBLAST + AE surfaces, while hAECs showed no mineralization signals. Osteogenic gene expression was upregulated in hAMSCs on GBLAST surfaces. Moreover, α2 and ß1 integrin expression enhanced in hAMSCs, suggesting a surface-integrin interaction. Consequently, hAMSCs would tend toward osteoblastic differentiation on grit-blasted surfaces conducive to osseointegration, a phenomenon not observed in hAECs.
Asunto(s)
Amnios , Implantes Dentales , Propiedades de Superficie , Titanio , Humanos , Titanio/química , Amnios/citología , Amnios/metabolismo , Osteogénesis , Diferenciación Celular , Células Cultivadas , Oseointegración , Células Madre/citología , Células Madre/metabolismo , Células Madre Mesenquimatosas/metabolismo , Células Madre Mesenquimatosas/citología , Supervivencia Celular , Fosfatasa Alcalina/metabolismoRESUMEN
El tejido adiposo es un tipo de tejido conectivo especializado que cumple diversas funciones en el organismo. Además de ser una reserva de energía, el tejido adiposo también actúa como aislante térmico y ayuda a proteger los órganos internos. Sobre la diversidad funcional del tejido adiposo, se vienen realizado importantes descubrimientos como: Reserva de energía, Aislamiento térmico, Mantenimiento de funciones metabólicas, Protección de órganos, Respuesta inmunológica con regulación del sistema inmune, Regulación hormonal a través de secreción con señalización de factores paracrinos, autocrinos y endocrinos conocido como adipocinas o adipoquinas (del griego adipo = grasa; y kinos = movimiento) que son una serie de moléculas de señalización celular secretadas por el adipocito que al aumentar su tamaño estimula la secreción de citocinas con características proinflamatorias que influyen en el control de su celularidad, la angiogénesis, la migración de células inmunológicas, cambios en la homeostasis, que modifican los procesos de coagulación y fibrinolisis Actualmente la terapia celular y la medicina regenerativa son áreas de gran desarrollo en la investigación biomédica. El uso de células madre/estromales mesenquimales (MSCs) tienen un gran potencial terapéutico como herramienta para la regeneración de tejidos y el tratamiento de enfermedades inflamatorias de distintos tipos. OBJETIVO: revisar y resumir las funciones del tejido adiposo y su papel último en la terapia celular y la medicina regenerativa de aislamiento de células madre derivadas del tejido adiposo y su aplicación en diversas terapias de regeneración celular. MÉTODOS: la metodología utilizada para la revisión bibliográfica fue de tipo descriptivo mediante la revisión sistemática de artículos, consultando diversas bases de datos que finalmente se seleccionaron 51 de los últimos años, en español e inglés RESULTADOS: esta revisión bibliográfica muestra nuevos conocimientos sobre el tejido adiposo, incluyendo la anatomía-fisiología en el humano. Las múltiples propiedades de las células madre/estromales mesenquimales MSCs incrementan su potencial terapéutico en la regeneración de tejidos y el tratamiento de enfermedades inflamatorias. No obstante, es necesario generar más información acerca de los mecanismos involucrados en la regeneración tisular, la fuente óptima para aislarlas, los efectos que ejercen, como potencial terapéutico en los usos clínicos en todos los campos de la cirugía de estas células madre/estromales mesenquimales MSCs CONCLUSIÓN: las células madre/estromales mesenquimales MSCs tienen un gran potencial terapéutico en medicina, ya que pueden diferenciarse en diferentes tipos celulares y tienen propiedades inmunológicas moduladoras. Se están desarrollando investigaciones en diversas áreas, como la medicina regenerativa, enfermedades autoinmunes, enfermedades cardiovasculares y cáncer, con el objetivo de aprovechar al máximo estas células en el tratamiento de las mismas y sobre todo en artritis y ortopedia Estas investigaciones abren nuevos caminos para la anatomo/histo/fisio/patología y el desarrollo de terapias dirigidas al tejido adiposo y su papel en las enfermedades.
Adipose tissue, a specialized type of connective tissue, serves various functions in the human body. Beyond being an energy reservoir, adipose tissue acts as thermal insulation and helps protect internal organs. Recent discoveries have shed light on the multifunctional aspects of adipose tissue, including its role in energy storage, thermal regulation, maintenance of metabolic functions, organ protection, and immune response. Adipose tissue also secretes signaling molecules known as adipokines, which play a crucial role in immune regulation, hormonal balance, and cellular homeostasis. These factors influence processes such as angiogenesis, immune cell migration, and coagulation. Currently, cellular therapy and regenerative medicine are rapidly advancing fields in biomedical research. Mesenchymal stem/stromal cells (MSCs) derived from adipose tissue hold significant therapeutic potential for tissue regeneration and the treatment of various inflammatory conditions. OBJECTIVE: this review aims to summarize the functions of adipose tissue and explore its ultimate role in cellular therapy and regenerative medicine. Specifically, we focus on isolating MSCs from adipose tissue and their application in various cellular regeneration therapies. METHODS: we conducted a descriptive literature review by systematically analyzing articles from diverse databases. A total of 51 relevant articles published in both Spanish and English over recent years were selected. RESULTS: this literature review provides novel insights into adipose tissue, including its anatomy and physiology in humans. The diverse properties of MSCs enhance their therapeutic potential for tissue regeneration and inflammatory disease treatment. However, further research is needed to understand the mechanisms involved in tissue regeneration, optimal MSC isolation methods, and their clinical applications across surgical fields. CONCLUSION: mesenchymal stem/stromal cells (MSCs) offer significant therapeutic promise in Medicine due to their ability to differentiate into various cell types and modulate immune responses. Ongoing investigations span areas such as regenerative medicine, autoimmune diseases, cardiovascular conditions, and cancer. These studies pave the way for anatomopathological, and therapeutic advancements related to adipose tissue and its role in diseases.
Asunto(s)
Células Madre , Tejido Adiposo , Células Madre MesenquimatosasRESUMEN
The aim of this study was to evaluate the effect of adipose-derived stem cells (ADSCs) in the treatment of acute rupture of the Achilles tendon. It was a cross-sectional study involving 15 patients. Patients were randomly divided: group 1-rupture; group 2-suture; group 3-rupture + ADSCs. In the AOFAS score, the score was higher in group 3 with a significant difference. In the ATRS score, the score was higher in groups 2 and 3, also with a significant difference. As for the ultrasound score, there was a significant difference between the experimental groups in relation to this score, however, in the multiple comparisons test, comparing two groups at a time, it was possible to observe a significant difference of the experimental groups. It can be concluded that cell therapy in this condition may be a treatment option due to tissue regeneration and significant recovery of function.
Asunto(s)
Tendón Calcáneo , Tejido Adiposo , Humanos , Tendón Calcáneo/lesiones , Masculino , Femenino , Rotura/terapia , Adulto , Tejido Adiposo/citología , Estudios Transversales , Trasplante de Células Madre , Persona de Mediana Edad , Células Madre/citología , Traumatismos de los Tendones/terapia , Tratamiento Basado en Trasplante de Células y Tejidos/métodos , Resultado del TratamientoRESUMEN
Bone tissue regeneration strategies have incorporated the use of natural polymers, such as hydroxyapatite (nHA), chitosan (CH), gelatin (GEL), or alginate (ALG). Additionally, platelet concentrates, such as platelet-rich fibrin (PRF) have been suggested to improve scaffold biocompatibility. This study aimed to develop scaffolds composed of nHA, GEL, and CH, with or without ALG and lyophilized PRF, to evaluate the scaffold's properties, growth factor release, and dental pulp stem cells (DPSC), and osteoblast (OB) derived from DPSC viability. Four scaffold variations were synthesized and lyophilized. Then, degradation, swelling profiles, and morphological analysis were performed. Furthermore, PDGF-BB and FGF-B growth factors release were quantified by ELISA, and cytotoxicity and cell viability were evaluated. The swelling and degradation profiles were similar in all scaffolds, with pore sizes ranging between 100 and 250 µm. FGF-B and PDGF-BB release was evidenced after 24 h of scaffold immersion in cell culture medium. DPSC and OB-DPSC viability was notably increased in PRF-supplemented scaffolds. The nHA-CH-GEL-PRF scaffold demonstrated optimal physical-biological characteristics for stimulating DPSC and OB-DPSC cell viability. These results suggest lyophilized PRF improves scaffold biocompatibility for bone tissue regeneration purposes.
Asunto(s)
Alginatos , Supervivencia Celular , Quitosano , Pulpa Dental , Durapatita , Gelatina , Osteoblastos , Fibrina Rica en Plaquetas , Células Madre , Andamios del Tejido , Humanos , Pulpa Dental/citología , Quitosano/química , Quitosano/farmacología , Gelatina/química , Fibrina Rica en Plaquetas/química , Fibrina Rica en Plaquetas/metabolismo , Andamios del Tejido/química , Células Madre/efectos de los fármacos , Células Madre/citología , Células Madre/metabolismo , Supervivencia Celular/efectos de los fármacos , Durapatita/química , Durapatita/farmacología , Alginatos/química , Alginatos/farmacología , Osteoblastos/efectos de los fármacos , Osteoblastos/citología , Adhesión Celular/efectos de los fármacos , Ingeniería de Tejidos/métodos , Células CultivadasRESUMEN
OBJECTIVE: To explore the feasibility of injectable platelet-rich fibrin (i-PRF) in regenerative endodontics by comparing the effect of i-PRF and platelet-rich fibrin (PRF) on the biological behavior and angiogenesis of human stem cells from the apical papilla (SCAPs). METHODOLOGY: i-PRF and PRF were obtained from venous blood by two different centrifugation methods, followed by hematoxylin-eosin (HE) staining and scanning electron microscopy (SEM). Enzyme-linked immunosorbent assay (ELISA) was conducted to quantify the growth factors. SCAPs were cultured with different concentrations of i-PRF extract (i-PRFe) and PRF extract (PRFe), and the optimal concentrations were selected using the Cell Counting Kit-8 (CCK-8) assay. The cell proliferation and migration potentials of SCAPs were then observed using the CCK-8 and Transwell assays. Mineralization ability was detected by alizarin red staining (ARS), and angiogenesis ability was detected by tube formation assay. Real-time quantitative polymerase chain reaction (RT-qPCR) was performed to evaluate the expression of genes related to mineralization and angiogenesis. The data were subjected to statistical analysis. RESULTS: i-PRF and PRF showed a similar three-dimensional fibrin structure, while i-PRF released a higher concentration of growth factors than PRF ( P <.05). 1/4× i-PRFe and 1/4× PRFe were selected as the optimal concentrations. The cell proliferation rate of the i-PRFe group was higher than that of the PRFe group ( P <.05), while no statistical difference was observed between them in terms of cell mitigation ( P >.05). More importantly, our results showed that i-PRFe had a stronger effect on SCAPs than PRFe in facilitating mineralization and angiogenesis, with the consistent result of RT-qPCR ( P <.05). CONCLUSION: This study revealed that i-PRF released a higher concentration of growth factors and was superior to PRF in promoting proliferation, mineralization and angiogenesis of SCAPs, which indicates that i-PRF could be a promising biological scaffold for application in pulp regeneration.
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Proliferación Celular , Ensayo de Inmunoadsorción Enzimática , Péptidos y Proteínas de Señalización Intercelular , Microscopía Electrónica de Rastreo , Neovascularización Fisiológica , Fibrina Rica en Plaquetas , Reacción en Cadena en Tiempo Real de la Polimerasa , Endodoncia Regenerativa , Humanos , Proliferación Celular/efectos de los fármacos , Neovascularización Fisiológica/efectos de los fármacos , Endodoncia Regenerativa/métodos , Células Cultivadas , Reproducibilidad de los Resultados , Movimiento Celular/efectos de los fármacos , Células Madre/efectos de los fármacos , Factores de Tiempo , Estudios de Factibilidad , Análisis de Varianza , Papila Dental/efectos de los fármacos , Papila Dental/citología , Valores de ReferenciaRESUMEN
Lichen planopilaris (LPP) and frontal fibrosing alopecia (FFA) are primary cicatricial alopecia that cause a major impact on quality of life due to irreversible hair loss and symptoms as itching, burning and pain. They are characterized by permanent loss of hair follicle stem cells (HFSCs) by pathomechanisms still poorly understood, resulting in poor efficacy of currently available treatments. Caveolae are flask-shaped lipid rafts invaginated within the plasma membrane of multiple cell types. Although their role in the HF physiology and pathophysiology is relatively unknown, we have previously demonstrated that the primary structural component of caveolae (caveolin-1 or Cav1) is upregulated in FFA. Thus, we propose to investigate the expression and localization of caveolae-associated structural proteins (Cav1, Cav2, and Cavin-1) and HFSCs (identified by K15) in both LPP and FFA. We analyzed 4 patients with LPP biopsied in affected and non-affected (NA) scalp, 4 patients with FFA biopsied in affected scalp and 4 healthy controls. Affected scalp of LPP and FFA demonstrated increased levels of Cav1 and Cavin-1 compared with HC and LPP-NA. Moreover, Cav1, Cav2 and Cavin1 all exhibit high colocalization with K15 and their expression appears to be negatively correlated, supporting the hypothesis that these proteins are important players in LPP/FFA and may serve as therapeutic targets in future treatments.
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Alopecia , Caveolas , Caveolina 1 , Folículo Piloso , Liquen Plano , Regulación hacia Arriba , Humanos , Alopecia/patología , Alopecia/metabolismo , Folículo Piloso/patología , Folículo Piloso/metabolismo , Liquen Plano/metabolismo , Liquen Plano/patología , Persona de Mediana Edad , Femenino , Caveolina 1/metabolismo , Masculino , Caveolas/metabolismo , Cuero Cabelludo/patología , Adulto , Queratina-15/metabolismo , Anciano , Biopsia , Fibrosis , Células Madre/metabolismo , Células Madre/patología , Proteínas de Unión al ARN/metabolismoRESUMEN
Studies regarding cytotoxic effects attributed to the use of adhesive bonding agents on pulp tissue are not conclusive. To point out whether these materials are safe for clinical use, in vivo exposure of dental pulp to adhesive bonding agents was simulated using an experimental setup in which Human Dental Pulp Stem Cells (hDPSC) are exposed to the action of two kinds of adhesives: self-etching adhesives and two-step bonding agents through a dentine barrier. Cytotoxic effects on these cells were evaluated by MTT assay protocol and fluorescence microscopy, and their results were contrasted to those obtained through Raman spectra taken on single hDPSCs. Overall, no significant cytotoxic effects were observed by combining all the techniques, and cell viability close to 90% was achieved for a dentine barrier of at least 1 mm thick. Moreover, Raman spectroscopy was able to detect structural DNA damage in some dental pulp cells when exposed to two-step bonding agents, suggesting that this technique could be considered a complementary tool with the potential to evaluate cell toxicity beyond cell viability.
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Supervivencia Celular , Pulpa Dental , Recubrimientos Dentinarios , Espectrometría Raman , Células Madre , Humanos , Pulpa Dental/citología , Pulpa Dental/efectos de los fármacos , Células Madre/efectos de los fármacos , Recubrimientos Dentinarios/toxicidad , Supervivencia Celular/efectos de los fármacos , Microscopía Fluorescente , Células CultivadasRESUMEN
Insufficient osseointegration of titanium-based implants is a factor conditioning their long-term success. Therefore, different surface modifications, such as multifunctional oxide coatings, calcium phosphates, and the addition of molecules such as peptides, have been developed to improve the bioactivity of titanium-based biomaterials. In this work, we investigate the behavior of human oral mucosal stem cells (hOMSCs) cultured on amorphous titanium oxide (aTiO2), surfaces designed to simulate titanium (Ti) surfaces, biofunctionalized with a novel sequence derived from cementum attachment protein (CAP-p15), exploring its impact on guiding hOMSCs towards an osteogenic phenotype. We carried out cell attachment and viability assays. Next, hOMSCs differentiation was assessed by red alizarin stain, ALP activity, and western blot analysis by evaluating the expression of RUNX2, BSP, BMP2, and OCN at the protein level. Our results showed that functionalized surfaces with CAP-p15 (1 µg ml-1) displayed a synergistic effect increasing cell proliferation and cell attachment, ALP activity, and expression of osteogenic-related markers. These data demonstrate that CAP-p15 and its interaction with aTiO2surfaces promote osteoblastic differentiation and enhanced mineralization of hOMSCs when compared to pristine samples. Therefore, CAP-p15 shows the potential to be used as a therapeutical molecule capable of inducing mineralized tissue regeneration onto titanium-based implants.
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Adhesión Celular , Diferenciación Celular , Proliferación Celular , Mucosa Bucal , Osteogénesis , Células Madre , Titanio , Titanio/química , Humanos , Osteogénesis/efectos de los fármacos , Mucosa Bucal/citología , Mucosa Bucal/metabolismo , Células Madre/citología , Células Madre/metabolismo , Propiedades de Superficie , Células Cultivadas , Osteoblastos/citología , Osteoblastos/metabolismo , Materiales Biocompatibles Revestidos/química , Materiales Biocompatibles Revestidos/farmacología , Supervivencia Celular , Oseointegración/efectos de los fármacos , Materiales Biocompatibles/químicaRESUMEN
MicroRNAs (miRNAs) are non-coding RNAs involved in the regulation of gene expression associated with cell differentiation, proliferation, adhesion, and important biological functions such as inflammation. miRNAs play roles associated with the pathogenesis of chronic degenerative disorders including cardiovascular diseases. Understanding the influence of miRNAs and their target genes can effectively streamline the identification of key biologically active pathways that are important in the development of vascular grafts through the tissue engineering of blood vessels. To determine miRNA expression levels and identify miRNA target genes and pathways with biological roles in scaffolds that have been repopulated with adipose-derived stem cells (ASCs) generated through tissue engineering for the construction of blood vessels. miRNA quantification assays were performed in triplicate to determine miRNA expression in a total of 20 samples: five controls (natural inferior vena cava), five scaffolds recellularized with ASCs and differentiated into the endothelium (luminal layer), five samples of complete scaffolds seeded with ASCs differentiated into the endothelium (luminal layer) and smooth muscle (extraluminal layer), and five samples of ASC without cell differentiation. Several differentially expressed miRNAs were identified and predicted to modulate target genes with roles in key pathways associated with angiogenesis, vascular system control, and endothelial and smooth muscle regulation, including migration, proliferation, and growth. These findings underscore the involvement of these pathways in the regulatory mechanisms that are essential for vascular scaffold production through tissue engineering. Our research contributes to the knowledge of miRNA-regulated mechanisms, which may impact the design of vascular substitutes, and provide valuable insights for enhancing clinical practice. The molecular pathways regulated by miRNAs in tissue engineering of blood vessels (TEBV) allowed us to elucidate the main phenomena involved in cellular differentiation to constitute a blood vessel, with the main pathways being essential for angiogenesis, cellular differentiation, and differentiation into vascular smooth muscle.
Asunto(s)
Diferenciación Celular , MicroARNs , Ingeniería de Tejidos , Andamios del Tejido , MicroARNs/genética , MicroARNs/metabolismo , Ingeniería de Tejidos/métodos , Humanos , Andamios del Tejido/química , Diferenciación Celular/genética , Tejido Adiposo/metabolismo , Tejido Adiposo/citología , Vasos Sanguíneos/metabolismo , Vasos Sanguíneos/crecimiento & desarrollo , Regulación de la Expresión Génica , Neovascularización Fisiológica/genética , Células Madre/metabolismo , Células Madre/citología , Proliferación Celular/genética , Transducción de SeñalRESUMEN
OBJECTIVES: This study aimed to assess antimicrobial efficacy, cytotoxicity, and cytokine release (IL-1b, IL-6, IL-10, TNF-α) from human dental pulp stem cells (hDPSCs) of chitosan (CH) and hydroxyapatite (HAp)-modified glass ionomer cements (GIC). METHODS: GICs with varied CH and HAp concentrations (0 %, 0.16 %, 2 %, 5 %, 10 %) were tested against S. mutans for 24 h or 7 days. Antimicrobial activity was measured using an MTT test. Cytotoxicity evaluation followed for optimal concentrations, analyzing mitochondrial activity and apoptosis in hDPSCs. Cytokine release was assessed with MAGPIX. Antimicrobial analysis used Shapiro-Wilk, Kruskal-Wallis, and Dunnett tests. Two-way ANOVA, Tukey, and Dunnett tests were applied for hDP metabolism and cytokine release. RESULTS: CH 2 % and HAp 5 % significantly enhanced GIC antimicrobial activity, especially after seven days. In immediate analysis, all materials showed reduced mitochondrial activity compared to the control. After 24 h, CH demonstrated mitochondrial metabolism similar to the control. All groups exhibited mild cytotoxicity (â¼30 % cell death). Only IL-6 was influenced, with reduced release in experimental groups. SIGNIFICANCE: CH 2 % and HAp 5 % were most effective for antibacterial effects. GIC-CH 2 % emerged as the most promising formula, displaying significant antibacterial effects with reduced hDPSC toxicity.
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Quitosano , Citocinas , Pulpa Dental , Durapatita , Cementos de Ionómero Vítreo , Quitosano/química , Quitosano/farmacología , Cementos de Ionómero Vítreo/toxicidad , Cementos de Ionómero Vítreo/farmacología , Cementos de Ionómero Vítreo/química , Humanos , Durapatita/química , Durapatita/farmacología , Pulpa Dental/citología , Pulpa Dental/efectos de los fármacos , Citocinas/metabolismo , Streptococcus mutans/efectos de los fármacos , Antiinfecciosos/farmacología , Antiinfecciosos/química , Ensayo de Materiales , Células Cultivadas , Células Madre/efectos de los fármacos , Apoptosis/efectos de los fármacosRESUMEN
Dental pulp stem cells (DPSCs) and periodontal ligament stem cells (PDLSCs) can differentiate into osteoblasts, indicating that both are potential candidates for bone tissue engineering. Osteogenesis is influenced by many environmental factors, one of which is lipopolysaccharide (LPS). LPS-induced NF-κB activity affects the osteogenic potencies of different types of MSCs differently. This study evaluated the effect of LPS-induced NF-κB activity and its inhibition in DPSCs and PDLSCs. DPSCs and PDLSCs were cultured in an osteogenic medium, pretreated with/without NF-κB inhibitor Bay 11-7082, and treated with/without LPS. Alizarin red staining was performed to assess bone nodule formation, which was observed under an inverted light microscope. NF-κB and alkaline phosphatase (ALP) activities were measured to examine the effect of Bay 11-7082 pretreatment and LPS supplementation on osteogenic differentiation of DPSCs and PDLSCs. LPS significantly induced NF-κB activity (p = 0.000) and reduced ALP activity (p = 0.000), which inhibited bone nodule formation in DPSCs and PDLSCs. Bay 11-7082 inhibited LPS-induced NF-κB activity, and partially maintained ALP activity and osteogenic potency of LPS-supplemented DPSCs and PDLSCs. Thus, inhibition of LPS-induced NF-κB activity can maintain the osteogenic potency of DPSCs and PDLSCs.
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Pulpa Dental , Lipopolisacáridos , FN-kappa B , Nitrilos , Osteogénesis , Ligamento Periodontal , Sulfonas , Humanos , Antraquinonas/química , Células Cultivadas , Pulpa Dental/citología , Pulpa Dental/metabolismo , Lipopolisacáridos/metabolismo , Lipopolisacáridos/farmacología , FN-kappa B/antagonistas & inhibidores , FN-kappa B/metabolismo , Nitrilos/farmacología , Osteogénesis/efectos de los fármacos , Ligamento Periodontal/citología , Células Madre/efectos de los fármacos , Células Madre/metabolismo , Sulfonas/farmacologíaAsunto(s)
Liposomas , Melanosis , Extractos Vegetales , Humanos , Melanosis/tratamiento farmacológico , Extractos Vegetales/farmacología , Extractos Vegetales/administración & dosificación , Femenino , Resultado del Tratamiento , Adulto , Coffea/química , Persona de Mediana Edad , Extractos Celulares/farmacología , Células Madre/efectos de los fármacosRESUMEN
Adipose tissue metabolism is actively involved in the regulation of energy balance. Adipose-derived stem cells (ASCs) play a critical role in maintaining adipose tissue function through their differentiation into mature adipocytes (Ad). This study aimed to investigate the impact of an obesogenic environment on the epigenetic landscape of ASCs and its impact on adipocyte differentiation and its metabolic consequences. Our results showed that ASCs from rats on a high-fat sucrose (HFS) diet displayed reduced adipogenic capacity, increased fat accumulation, and formed larger adipocytes than the control (C) group. Mitochondrial analysis revealed heightened activity in undifferentiated ASC-HFS but decreased respiratory and glycolytic capacity in mature adipocytes. The HFS diet significantly altered the H3K4me3 profile in ASCs on genes related to adipogenesis, mitochondrial function, inflammation, and immunomodulation. After differentiation, adipocytes retained H3K4me3 alterations, confirming the upregulation of genes associated with inflammatory and immunomodulatory pathways. RNA-seq confirmed the upregulation of genes associated with inflammatory and immunomodulatory pathways in adipocytes. Overall, the HFS diet induced significant epigenetic and transcriptomic changes in ASCs, impairing differentiation and causing dysfunctional adipocyte formation.NEW & NOTEWORTHY Obesity is associated with the development of chronic diseases like metabolic syndrome and type 2 diabetes, and adipose tissue plays a crucial role. In a rat model, our study reveals how an obesogenic environment primes adipocyte precursor cells, leading to epigenetic changes that affect inflammation, adipogenesis, and mitochondrial activity after differentiation. We highlight the importance of histone modifications, especially the trimethylation of histone H3 to lysine 4 (H3K4me3), showing its influence on adipocyte expression profiles.
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Adipocitos , Adipogénesis , Tejido Adiposo , Dieta Alta en Grasa , Epigénesis Genética , Histonas , Transcriptoma , Animales , Ratas , Adipocitos/metabolismo , Dieta Alta en Grasa/efectos adversos , Histonas/metabolismo , Masculino , Adipogénesis/genética , Adipogénesis/fisiología , Tejido Adiposo/metabolismo , Diferenciación Celular/genética , Células Madre/metabolismo , Obesidad/metabolismo , Obesidad/genética , Reprogramación Celular/fisiología , Células Cultivadas , Ratas Wistar , Ratas Sprague-DawleyRESUMEN
BACKGROUND: Genetically modified pigs are considered ideal models for studying human diseases and potential sources for xenotransplantation research. However, the somatic cell nuclear transfer (SCNT) technique utilized to generate these cloned pig models has low efficiency, and fetal development is limited due to placental abnormalities. RESULTS: In this study, we unprecedentedly established putative porcine trophoblast stem cells (TSCs) using SCNT and in vitro-fertilized (IVF) blastocysts through the activation of Wing-less/Integrated (Wnt) and epidermal growth factor (EGF) pathways, inhibition of transforming growth factor-ß (TGFß) and Rho-associated protein kinase (ROCK) pathways, and supplementation with ascorbic acid. We also compared the transcripts of putative TSCs originating from SCNT and IVF embryos and their differentiated lineages. A total of 19 porcine TSCs exhibiting typical characteristics were established from SCNT and IVF blastocysts (TSCsNT and TSCsIVF). Compared with the TSCsIVF, TSCsNT showed distinct expression patterns suggesting unique TSCsNT characteristics, including decreased mRNA expression of genes related to apposition, steroid hormone biosynthesis, angiopoiesis, and RNA stability. CONCLUSION: This study provides valuable information and a powerful model for studying the abnormal development and dysfunction of trophoblasts and placentas in cloned pigs.
Asunto(s)
Blastocisto , Técnicas de Transferencia Nuclear , Trofoblastos , Animales , Trofoblastos/metabolismo , Porcinos , Diferenciación Celular , Femenino , Células Madre , Fertilización In Vitro/métodosRESUMEN
Periodontal ligament stem cells (PDLSCs) show plasticity towards the adipogenic lineage; however, little has been done on the participation of epigenetic mechanisms. Histone acetylation is a dynamic process, though balanced by histone acetyltransferases (HATs) and histone deacetylases (HDACs) activities. This process can be halted by HDACs inhibitors, such as trichostatin A (TSA) and valproic acid (VPA). This study aimed to determine the role of HDACs class I in adipogenic differentiation of PDL cells. PDLSCs were treated with TSA at concentrations of 100, 200, and 250 nM, or VPA at 1, 4 and 8 mM. Cell viability was assessed using MTT assays. Gene expression of pluripotency markers (NANOG, OCT4, SOX2), HAT genes (p300, GCN5), and HDACs genes (HDAC1-3) was analyzed by RT-qPCR. Adipogenic differentiation was evaluated via oil red O staining, and acetylation of histone H3 lysine 9 (H3K9ac) was examined by Western blot. VPA treatment resulted in a 60% reduction in cell proliferation, compared to a 50% when using TSA. Cell viability was not affected by either inhibitor. Furthermore, both TSA and VPA induced adipogenic differentiation, through an increase in the deposition of lipid droplets and in GCN5 and p300 expression were observed. Western blot analysis showed that TSA increased H3K9ac levels on adipogenic differentiation of PDLSCs. These findings highlight the potential of HDAC inhibitors as a tool for modulating H3K9 acetylation status and thus influencing adipogenic differentiation of PDLCs.