RESUMEN
Finite element analysis (FEA) has been used to analyze the behavior of dental materials, mainly in implantology. However, FEA is a mechanical analysis and few studies have tried to simulate the biological characteristics of the healing process of loaded implants. This study used the rule of mixtures to simulate the biological healing process of immediate implants in an alveolus socket and bone-implant junction interface through FEA. Three-dimensional geometric models of the structures were obtained, and material properties were derived from the literature. The rule of mixtures was used to simulate the healing periods-immediate and early loading, in which the concentration of each cell type, based on in vivo studies, influenced the final elastic moduli. A 100 N occlusal load was simulated in axial and oblique directions. The models were evaluated for maximum and minimum principal strains, and the bone overload was assessed through Frost's mechanostat. There was a higher strain concentration in the healing regions and cortical bone tissue near the cervical portion. The bone overload was higher in the immediate load condition. The method used in this study may help to simulate the biological healing process and could be useful to relate FEA results to clinical practice.
Asunto(s)
Implantes Dentales , Módulo de Elasticidad , Análisis de Elementos Finitos , Carga Inmediata del Implante Dental , Alveolo Dental , Cicatrización de Heridas , Humanos , Alveolo Dental/fisiología , Cicatrización de Heridas/fisiología , Fenómenos Biomecánicos , Simulación por Computador , Interfase Hueso-Implante/fisiología , Estrés Mecánico , Proceso Alveolar/fisiología , Modelos Biológicos , Oseointegración/fisiología , Fuerza de la Mordida , Análisis del Estrés Dental/métodos , Osteoblastos/fisiología , Hueso Cortical/fisiología , Imagenología Tridimensional/métodosRESUMEN
It is known that cellular events underlying the processes of bone maintenance, remodeling, and repair have their basis in the embryonic production of bone. Shh signaling is widely described developing important morphogenetic control in bone by modifying the activity of osteoblast. Furthermore, identifying whether it is associated with the modulation of nuclear control is very important to be the basis for further applications. Experimentally, osteoblasts were exposed with cyclopamine (CICLOP) considering up to 1 day and 7 days, here considered an acute and chronic responses respectively. Firstly, we have validated the osteogenic model in vitro by exposing the osteoblasts to classical differentiating solution up to 7 days to allow the analysis of alkaline phosphatase and mineralization. Conversely, our data shows that differentiating osteoblasts present higher activity of inflammasome-related genes, while Shh signaling members were lower, suggesting a negative feedback between them. Thereafter, to better know about the role of Shh signaling on this manner, functional assays using CICLOP (5 µM) were performed and the data validates the previously hypothesis that Shh represses inflammasome related genes activities. Altogether, our data supports the anti-inflammatory effect of Shh signaling by suppressing Tnfα, Tgfß and inflammasome related genes during osteoblast differentiation, and this comprehension might support the understanding the molecular and cellular mechanisms related in bone regeneration by reporting molecular-related osteoblast differentiation.
Asunto(s)
Erizos , Inflamasomas , Animales , Inflamasomas/farmacología , Osteogénesis/genética , Osteoblastos/fisiologíaRESUMEN
This investigation is aimed to determine the effect of the modification of titanium surface with NaOH on the metabolism of osteoblasts treated with zoledronic acid (ZA). Machined and NaOH-treated titanium disks were used. Surfaces were characterized by scanning electron microscopy, confocal microscopy, and x-ray photoelectron spectroscopy (XPS) analysis. Human osteoblasts were seeded onto the disks. After 24 h, cells were treated with ZA at 5 µM for 7 days. At this point, cell viability, collagen synthesis, total protein production, alkaline phosphatase activity, and mineral nodule deposition were assessed. The results of surface roughness were descriptively and statistically analyzed (t-Student), while the XPS results were qualitatively described. Cell metabolism data were analyzed by the analysis of variance two-way and Tukey tests at a 5% significance level. The results demonstrated that NaOH-treatment increased surface roughness (p < .05) and confirmed the presence of sodium titanate and a pH switch on the NaOH-treated disks. This modification also resulted in higher cell viability, collagen synthesis, total protein production, and alkaline phosphatase by osteoblasts when compared to cells seeded onto machined disks (p < 0.05). In the presence of ZA, all cellular metabolism and differentiation parameters were significantly reduced for cells seeded on both surfaces (p < 0.05); however, the cells seeded onto modified surfaces showed higher values for these parameters, except for mineral nodule deposition (p < 0.05). NaOH modification improved cell adhesion and metabolism of osteogenic cells even in the presence of ZA. The surface modification of titanium with NaOH solution may be an interesting strategy to improve metabolism and differentiation of osteoblasts and accelerate osseointegration process, mainly for tissues exposed to ZA.
Asunto(s)
Fosfatasa Alcalina , Titanio , Fosfatasa Alcalina/metabolismo , Fosfatasa Alcalina/farmacología , Colágeno , Humanos , Osteoblastos/fisiología , Hidróxido de Sodio/farmacología , Propiedades de Superficie , Titanio/química , Titanio/farmacología , Ácido Zoledrónico/farmacologíaRESUMEN
The aim of this study was to evaluate biocompatibility of hydroxyapatite (HAP) from fish waste using in vitro and in vivo assays. Fish samples (whitemouth croaker - Micropogonias furnieri) from the biowaste was used as HAP source. Pre-osteoblastic MC3T3-E1 cells were used in vitro study. In addition, bone defects were artificially created in rat calvaria and filled with HAP in vivo. The results demonstrated that HAP reduced cytotoxicity in pre-osteoblast cells after 3 and 6 days following HAP exposure. DNA concentration was lower in the HAP group after 6 days. Quantitative RT-PCR did not show any significant differences (p > 0.05) between groups. In vivo study revealed that bone defects filled with HAP pointed out moderate chronic inflammatory cells with slight proliferation of blood vessels after 7 and 15 days. Chronic inflammatory infiltrate was absent after 30 days of HAP exposure. There was also a decrease in the amount of biomaterial, being followed by newly formed bone tissue. All experimental groups also demonstrated strong RUNX-2 immoexpression in the granulation tissue as well as in cells in close contact with biomaterial. The number of osteoblasts inside the defect area was lower in the HAP group when compared to control group after 7 days post-implantation. Similarly, the osteoblast surface as well as the percentage of bone surface was higher in control group when compared with HAP group after 7 days post-implantation. Taken together, HAP from fish waste is a promising possibility that should be explored more carefully by tissue-engineering or biotechnology.
Asunto(s)
Durapatita/aislamiento & purificación , Durapatita/farmacología , Productos Pesqueros , Animales , Regeneración Ósea/efectos de los fármacos , Sustitutos de Huesos/química , Sustitutos de Huesos/aislamiento & purificación , Sustitutos de Huesos/farmacología , Diferenciación Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Productos Pesqueros/análisis , Ensayo de Materiales , Ratones , Osteoblastos/citología , Osteoblastos/efectos de los fármacos , Osteoblastos/fisiología , Osteogénesis/efectos de los fármacos , Perciformes , Ratas , Cráneo/efectos de los fármacos , Cráneo/fisiología , Residuos Sólidos/análisisRESUMEN
MicroRNA-9 (miR-9) modulates gene expression and demonstrates high structural conservation and wide expression in the central nervous system. Bioinformatics analysis predicts almost 100 ion channels, membrane transporters and receptors, including genes linked to primary familial brain calcification (PFBC), as possible miR-9-5p targets. PFBC is a neurodegenerative disorder, characterized by bilateral and symmetrical calcifications in the brain, associated with motor and behavioral disturbances. In this work, we seek to study the influence of miR-9-5p in regulating genes involved in PFBC, in an osteogenic differentiation model with SaOs-2 cells. During the induced calcification process, solute carrier family 20 member 2 (SLC20A2) and platelet-derived growth factor receptor beta (PDGFRB) were downregulated, while platelet-derived growth factor beta (PDGFB) showed no significant changes. Significantly decreased levels of SLC20A2 and PDGFRB were caused by the presence of miR-9-5p, while PDGFB showed no regulation. We confirmed the findings using an miR-9-5p inhibitor and also probed the cells in electrophysiological analysis to assess whether such microRNA might affect a broader range of ion channels, membrane transporters and receptors. Our electrophysiological data show that an increase of the miR-9-5p in SaOs-2 cells decreased the density and amplitude of the output ionic currents, indicating that it may influence the activity, and perhaps the expression, of some ionic channels. Additional investigations should determine whether such an effect is specific to miR-9-5p, and whether it could be used, together with the miR-9-5p inhibitor, as a therapeutic or diagnostic tool.
Asunto(s)
Encefalopatías/metabolismo , Calcinosis/metabolismo , Diferenciación Celular , MicroARNs/metabolismo , Osteoblastos/metabolismo , Proteínas Cotransportadoras de Sodio-Fosfato de Tipo III/metabolismo , Potenciales de Acción , Encefalopatías/genética , Calcinosis/genética , Línea Celular Tumoral , Humanos , MicroARNs/genética , Osteoblastos/citología , Osteoblastos/fisiología , Proteínas Proto-Oncogénicas c-sis/genética , Proteínas Proto-Oncogénicas c-sis/metabolismo , Proteínas Cotransportadoras de Sodio-Fosfato de Tipo III/genéticaRESUMEN
Host inflammatory immune response comprises an essential element of the bone healing process, where M2 polarization allegedly contributes to a favorable healing outcome. In this context, immunoregulatory molecules that modulate host response, including macrophage polarization, are considered potential targets for improving bone healing. This study aims to evaluate the role of the immunoregulatory molecules VIP (Vasoactive intestinal peptide) and PACAP (Pituitary adenylate cyclase activating polypeptide), which was previously described to favor the development of the M2 phenotype, in the process of alveolar bone healing in C57Bl/6 (WT) mice. Experimental groups were submitted to tooth extraction and maintained under control conditions or treated with VIP or PACAP were evaluated by microtomographic (µCT), histomorphometric, immunohistochemical, and molecular analysis at 0, 3, 7, and 14 days to quantify tissue healing and host response indicators at the healing site. Gene expression analysis demonstrates the effectiveness of VIP or PACAP in modulating host response, evidenced by the early dominance of an M2-type response, which was paralleled by a significant increase in M2 (CD206+) in treated groups. However, despite the marked effect of M1/M2 balance in the healing sites, the histomorphometric analysis does not reveal an equivalent/corresponding modulation of the healing process. µCT reveals a slight increase in bone matrix volume and the trabecular thickness number in the PACAP group, while histomorphometric analyzes reveal a slight increase in the VIP group, both at a 14-d time-point; despite the increased expression of osteogenic factors, osteoblastic differentiation, activity, and maturation markers in both VIP and PACAP groups. Interestingly, a lower number of VIP and PACAP immunolabeled cells were observed in the treated groups, suggesting a reduction in endogenous production. In conclusion, while both VIP and PACAP treatments presented a significant immunomodulatory effect with potential for increased healing, no major changes were observed in bone healing outcome, suggesting that the signals required for bone healing under homeostatic conditions are already optimal, and additional signals do not improve an already optimal process. Further studies are required to elucidate the role of macrophage polarization in the bone healing process.
Asunto(s)
Proceso Alveolar/lesiones , Activación de Macrófagos/efectos de los fármacos , Polipéptido Hipofisario Activador de la Adenilato-Ciclasa/administración & dosificación , Péptido Intestinal Vasoactivo/administración & dosificación , Cicatrización de Heridas/inmunología , Proceso Alveolar/efectos de los fármacos , Proceso Alveolar/inmunología , Proceso Alveolar/cirugía , Animales , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/inmunología , Modelos Animales de Enfermedad , Femenino , Inmunomodulación/efectos de los fármacos , Masculino , Ratones , Osteoblastos/fisiología , Osteogénesis/efectos de los fármacos , Osteogénesis/inmunología , Extracción Dental/efectos adversos , Cicatrización de Heridas/efectos de los fármacos , Microtomografía por Rayos XRESUMEN
It has been shown that 17ß-estradiol (E2) helps to prevent bone loss. This study was undertaken to verify whether E2 action in human osteoblasts involves changes in the transcriptional profile of the TNF-α, IFN-γ, NF-κB, TRAIL, TGF-ß, MMP2, MMP9, RECK, TIMP1, TIMP2, CDK2, CDK4, SRC, RUNX2, and SHH genes. Infraphysiological doses of E2 elevated mRNAs in all genes except for INF-γ, TRAIL, and TGF-ß. Importantly, a significant increase in the CDKs -2 and -4 genes was found, which strongly suggests cell cycle progression, with a potential dependency of Src involvement, as well as a suppression of the osteoblast differentiation machinery, with ECM remodeling being involved. These data suggest that E2 plays an important role in bone formation and remodeling, and Src seems to play a pivotal role in driving cell proliferation and ECM remodeling. Taken together, these findings contribute to an understanding of the effects of infraphysiological E2 on modulating bone homeostasis, favoring bone resorption, and leading to osteoporosis.
Asunto(s)
Proliferación Celular/efectos de los fármacos , Estradiol/farmacología , Matriz Extracelular/metabolismo , Genes src/fisiología , Osteoblastos/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/genética , Proliferación Celular/genética , Células Cultivadas , Relación Dosis-Respuesta a Droga , Matriz Extracelular/efectos de los fármacos , Femenino , Humanos , Células Madre Mesenquimatosas/efectos de los fármacos , Células Madre Mesenquimatosas/fisiología , Osteoblastos/fisiología , Osteogénesis/efectos de los fármacos , Osteogénesis/genética , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genéticaRESUMEN
This study evaluated the influence of type 2 diabetes mellitus on bone loss, bone repair and cytokine production in hyperglycemic rats, treated or not with metformin. The animals were distributed as follow: Non-Hyperglycemic (NH), Non Hyperglycemic with Ligature (NH-L), Treated Non Hyperglycemic (TNH), Treated Non Hyperglycemic with Ligature Treated (TNH-L), Hyperglycemic (H), Treated Hyperglycemic (TH), Hyperglycemic with Ligature (H-L), Treated Hyperglycemic with Ligature (TH-L). At 40th day after induction of hyperglycemia, the groups NH-L, TNH-L, H-L, TH-L received a ligature to induce periodontitis. On the 69th, the TNH, TNH-L, TH, TH-L groups received metformin until the end of the study. Bone repair was evaluated at histometric and the expression levels of Sox9, RunX2 and Osterix. Analysis of the ex-vivo expression of TNF-α, IFN-γ, IL-12, IL-4, TGF-ß, IL-10, IL-6 and IL-17 were also evaluated. Metformin partially reverse induced bone loss in NH and H animals. Lower OPG/RANKL, increased OCN and TRAP expression were observed in hyperglycemic animals, and treatment with metformin partially reversed hyperglycemia on the OPG/RANKL, OPN and TRAP expression in the periodontitis. The expression of SOX9 and RunX2 were also decreased by hyperglycemia and metformin treatment. Increased ex vivo levels of TNF-α, IL-6, IL-4, IL-10 and IL-17 was observed. Hyperglycemia promoted increased IL-10 levels compared to non-hyperglycemic ones. Treatment of NH with metformin was able to mediate increased levels of TNF-α, IL-10 and IL-17, whereas for H an increase of TNF-α and IL-17 was detected in the 24- or 48-hour after stimulation with LPS. Ligature was able to induce increased levels of TNF-α and IL-17 in both NH and H. This study revealed the negative impact of hyperglycemia and/or treatment with metformin in the bone repair via inhibition of transcription factors associated with osteoblastic differentiation.
Asunto(s)
Pérdida de Hueso Alveolar/prevención & control , Diabetes Mellitus Experimental/complicaciones , Diabetes Mellitus Tipo 2/complicaciones , Hiperglucemia/complicaciones , Metformina/administración & dosificación , Periodontitis/prevención & control , Pérdida de Hueso Alveolar/etiología , Pérdida de Hueso Alveolar/metabolismo , Proceso Alveolar/citología , Proceso Alveolar/efectos de los fármacos , Proceso Alveolar/metabolismo , Proceso Alveolar/patología , Animales , Regeneración Ósea/efectos de los fármacos , Regeneración Ósea/genética , Diferenciación Celular/efectos de los fármacos , Citocinas/metabolismo , Diabetes Mellitus Experimental/inducido químicamente , Diabetes Mellitus Experimental/tratamiento farmacológico , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 2/inducido químicamente , Diabetes Mellitus Tipo 2/tratamiento farmacológico , Diabetes Mellitus Tipo 2/metabolismo , Humanos , Hiperglucemia/inducido químicamente , Hiperglucemia/tratamiento farmacológico , Hiperglucemia/metabolismo , Osteoblastos/fisiología , Periodontitis/etiología , Periodontitis/metabolismo , Ratas , Estreptozocina/toxicidad , Factores de Transcripción/metabolismoRESUMEN
La oxitocina (OXT) como la arginina-vasopresina (AVP) son dos hormonas primitivas secretadas por la hipófisis posterior. Sus receptores están mucho más ampliamente distribuidos en el organismo de lo que se pensaba originalmente, incluido el hueso. En los estudios preclínicos, la OXT ha mostrado ser anabólica para el hueso, promoviendo la osteogénesis sobre la adipogénesis y favoreciendo la actividad osteoblástica sobre la osteoclástica. Tanto los osteoblastos como los osteoclastos tienen receptores para la OXT, y los efectos de los estrógenos sobre la masa ósea en ratones está mediada por lo menos en parte por la OXT. El mecanismo preciso por el cual la activación de los receptores de oxitocina (OXTR) se traduce en un incremento de la formación ósea permanece poco claro. La AVP también podría afectar el esqueleto en forma directa. Dos de los receptores de la AVP, V1a y V2 están expresados en osteoblastos y osteoclastos. La inyección de AVP en ratones de tipo salvaje aumenta la formación osteoclastos que producen resorción y reduce los osteoblastos formadores de hueso. En forma opuesta, la exposición de precursores osteoblásticos a antagonistas de los receptores V1a o V2, incrementan la osteoblastogénesis, como también lo hace la deleción genética del receptor V1a. (AU)
Both oxytocin (OXT) and argininevasopressin (AVP) are primitive hormones secreted by the posterior pituitary gland. OXT receptors are much more widely distributed in the body than originally thought, including in bone. In preclinical studies, OXT has been shown to be anabolic for bone, promoting osteogenesis over adipogenesis and favoring osteoblastic over osteoclastic activity. Both osteoblasts and osteoclasts have receptors for OXT, and the effects of estrogen on bone mass in mice is mediated at least in part by OXT. The precise mechanism by which the activation of oxytocin receptors (OXTRs) results in an increase in bone formation remains unclear. AVP could also have direct actions on the skeleton. The two AVP receptors, V1a and V2, are expressed in osteoblasts and osteoclasts. Injection of AVP in wild-type mice increases the formation of osteoclasts increasing bone resorption, and reduces bone-forming osteoblasts. On the contrary, the exposure of osteoblastic precursors to V1a and V2 antagonists increase osteoblastogenesis, the same as the genetic deletion of the V1a receptor. (AU)
Asunto(s)
Humanos , Animales , Ratones , Hormonas Neurohipofisarias/biosíntesis , Arginina Vasopresina/efectos adversos , Oxitocina/uso terapéutico , Osteoblastos/fisiología , Osteoclastos/fisiología , Osteogénesis , Osteoporosis/terapia , Hormonas Neurohipofisarias/fisiología , Arginina Vasopresina/antagonistas & inhibidores , Arginina Vasopresina/biosíntesis , Arginina Vasopresina/fisiología , Arginina Vasopresina/uso terapéutico , Oxitocina/biosíntesis , Oxitocina/efectos adversos , Oxitocina/fisiología , Transducción de Señal , Densidad Ósea , Densidad Ósea/efectos de los fármacos , Receptores de Oxitocina/biosíntesis , Receptores de Oxitocina/fisiología , Estradiol/uso terapéutico , Estrógenos/fisiologíaRESUMEN
OBJECTIVE: The present study aimed to investigate the participation of focal adhesion kinases (FAK) in interactions between osteoblastic cells and titanium (Ti) surfaces with three different topographies, namely, untreated (US), microstructured (MS), and nanostructured (NS). METHODOLOGY: Osteoblasts harvested from the calvarial bones of 3-day-old rats were cultured on US, MS and NS discs in the presence of PF-573228 (FAK inhibitor) to evaluate osteoblastic differentiation. After 24 h, we evaluated osteoblast morphology and vinculin expression, and on day 10, the following parameters: gene expression of osteoblastic markers and integrin signaling components, FAK protein expression and alkaline phosphatase (ALP) activity. A smooth surface, porosities at the microscale level, and nanocavities were observed in US, MS, and NS, respectively. RESULTS: FAK inhibition decreased the number of filopodia in cells grown on US and MS compared with that in NS. FAK inhibition decreased the gene expression of Alp, bone sialoprotein, osteocalcin, and ALP activity in cells grown on all evaluated surfaces. FAK inhibition did not affect the gene expression of Fak, integrin alpha 1 ( Itga1 ) and integrin beta 1 ( Itgb1 ) in cells grown on MS, increased the gene expression of Fak in cells grown on NS, and increased the gene expression of Itga1 and Itgb1 in cells grown on US and NS. Moreover, FAK protein expression decreased in cells cultured on US but increased in cells cultured on MS and NS after FAK inhibition; no difference in the expression of vinculin was observed among cells grown on all surfaces. CONCLUSIONS: Our data demonstrate the relevance of FAK in the interactions between osteoblastic cells and Ti surfaces regardless of surface topography. Nanotopography positively regulated FAK expression and integrin signaling pathway components during osteoblast differentiation. In this context, the development of Ti surfaces with the ability to upregulate FAK activity could positively impact the process of implant osseointegration.
Asunto(s)
Proteína-Tirosina Quinasas de Adhesión Focal/antagonistas & inhibidores , Osteoblastos/efectos de los fármacos , Quinolonas/farmacología , Sulfonas/farmacología , Titanio/química , Animales , Diferenciación Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Proteína-Tirosina Quinasas de Adhesión Focal/análisis , Proteína-Tirosina Quinasas de Adhesión Focal/química , Expresión Génica , Integrinas/análisis , Microscopía Electrónica de Rastreo , Oseointegración/efectos de los fármacos , Osteoblastos/fisiología , Quinolonas/química , Ratas Wistar , Reacción en Cadena en Tiempo Real de la Polimerasa , Transducción de Señal , Sulfonas/química , Propiedades de SuperficieRESUMEN
Abstract Objective The present study aimed to investigate the participation of focal adhesion kinases (FAK) in interactions between osteoblastic cells and titanium (Ti) surfaces with three different topographies, namely, untreated (US), microstructured (MS), and nanostructured (NS). Methodology Osteoblasts harvested from the calvarial bones of 3-day-old rats were cultured on US, MS and NS discs in the presence of PF-573228 (FAK inhibitor) to evaluate osteoblastic differentiation. After 24 h, we evaluated osteoblast morphology and vinculin expression, and on day 10, the following parameters: gene expression of osteoblastic markers and integrin signaling components, FAK protein expression and alkaline phosphatase (ALP) activity. A smooth surface, porosities at the microscale level, and nanocavities were observed in US, MS, and NS, respectively. Results FAK inhibition decreased the number of filopodia in cells grown on US and MS compared with that in NS. FAK inhibition decreased the gene expression of Alp, bone sialoprotein, osteocalcin, and ALP activity in cells grown on all evaluated surfaces. FAK inhibition did not affect the gene expression of Fak, integrin alpha 1 ( Itga1 ) and integrin beta 1 ( Itgb1 ) in cells grown on MS, increased the gene expression of Fak in cells grown on NS, and increased the gene expression of Itga1 and Itgb1 in cells grown on US and NS. Moreover, FAK protein expression decreased in cells cultured on US but increased in cells cultured on MS and NS after FAK inhibition; no difference in the expression of vinculin was observed among cells grown on all surfaces. Conclusions Our data demonstrate the relevance of FAK in the interactions between osteoblastic cells and Ti surfaces regardless of surface topography. Nanotopography positively regulated FAK expression and integrin signaling pathway components during osteoblast differentiation. In this context, the development of Ti surfaces with the ability to upregulate FAK activity could positively impact the process of implant osseointegration.
Asunto(s)
Animales , Osteoblastos/efectos de los fármacos , Sulfonas/farmacología , Titanio/química , Quinolonas/farmacología , Proteína-Tirosina Quinasas de Adhesión Focal/antagonistas & inhibidores , Osteoblastos/fisiología , Sulfonas/química , Propiedades de Superficie , Microscopía Electrónica de Rastreo , Transducción de Señal , Expresión Génica , Integrinas/análisis , Diferenciación Celular/efectos de los fármacos , Células Cultivadas , Oseointegración/efectos de los fármacos , Ratas Wistar , Quinolonas/química , Proliferación Celular/efectos de los fármacos , Proteína-Tirosina Quinasas de Adhesión Focal/análisis , Proteína-Tirosina Quinasas de Adhesión Focal/química , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
Biological effects of titanium (Ti) alloys were analyzed on biofilms of Candida albicans, Enterococcus faecalis, Pseudomonas aeruginosa, Staphylococcus aureus, Streptococcus mutans, and Streptococcus sanguinis, as well as on osteoblast-like cells (MG63) and murine macrophages (RAW 264.7). Standard samples composed of aluminum and vanadium (Ti-6Al-4V), and sample containing niobium (Ti-35Nb) and zirconium (Ti-13Nb-13Zr) were analyzed. Monomicrobial biofilms were formed on the Ti alloys. MG63 cells were grown with the alloys and the biocompatibility (MTT), total protein (TP) level, alkaline phosphatase (ALP) activity, and mineralization nodules (MN) formation were verified. Levels of interleukins (IL-1ß and IL-17), tumor necrosis factor alpha (TNF-α), and oxide nitric (NO) were checked, from RAW 264.7 cells supernatants. Data were statically analyzed by one-way analysis of variance (ANOVA) and Tukey's test, or T-test (P ≤ 0.05). Concerning the biofilm formation, Ti-13Nb-13Zr alloy showed the best inhibitory effect on E. faecalis, P. aeruginosa, and S. aureus. And, it also acted similarly to the Ti-6Al-4V alloy on C. albicans and Streptococcus spp. Both alloys were biocompatible and similar to the Ti-6Al-4V alloy. Additionally, Ti-13Nb-13Zr alloy was more effective for cell differentiation, as observed in the assays of ALP and MN. Regarding the stimulation for release of IL-1ß and TNF-α, Ti-35Nb and Ti-13Nb-13Zr alloys inhibited similarly the synthesis of these molecules. However, both alloys stimulated the production of IL-17. Additionally, all Ti alloys showed the same effect for NO generation. Thus, Ti-13Nb-13Zr alloy was the most effective for inhibition of biofilm formation, cell differentiation, and stimulation for release of immune mediators.
Asunto(s)
Aleaciones/farmacología , Materiales Biocompatibles/farmacología , Biopelículas/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Osteogénesis/efectos de los fármacos , Titanio/farmacología , Aleaciones/química , Animales , Materiales Biocompatibles/química , Biopelículas/crecimiento & desarrollo , Candida albicans/efectos de los fármacos , Candida albicans/fisiología , Células Cultivadas , Ensayo de Materiales , Ratones , Pruebas de Sensibilidad Microbiana , Osteoblastos/citología , Osteoblastos/efectos de los fármacos , Osteoblastos/fisiología , Pseudomonas/efectos de los fármacos , Pseudomonas/fisiología , Células RAW 264.7 , Staphylococcus aureus/efectos de los fármacos , Staphylococcus aureus/fisiología , Streptococcus/efectos de los fármacos , Streptococcus/fisiología , Propiedades de Superficie , Titanio/químicaRESUMEN
Bone development and healing processes involve a complex cascade of biological events requiring well-orchestrated synergism with bone cells, growth factors, and other trophic signaling molecules and cellular structures. Beyond health processes, MMPs play several key roles in the installation of heart and blood vessel related diseases and cancer, ranging from accelerating metastatic cells to ectopic vascular mineralization by smooth muscle cells in complementary manner. The tissue inhibitors of MMPs (TIMPs) have an important role in controlling proteolysis. Paired with the post-transcriptional efficiency of specific miRNAs, they modulate MMP performance. If druggable, these molecules are suggested to be a platform for development of "smart" medications and further clinical trials. Thus, considering the pleiotropic effect of MMPs on mammals, the purpose of this review is to update the role of those multifaceted proteases in mineralized tissues in health, such as bone, and pathophysiological disorders, such as ectopic vascular calcification and cancer.
Asunto(s)
Remodelación Ósea/fisiología , Matriz Extracelular/fisiología , Metaloproteinasas de la Matriz/fisiología , Enfermedades Óseas/metabolismo , Enfermedades Óseas/fisiopatología , Progresión de la Enfermedad , Humanos , Inhibidores de la Metaloproteinasa de la Matriz/uso terapéutico , Neoplasias/metabolismo , Neoplasias/fisiopatología , Osteoblastos/fisiología , Inhibidores Tisulares de Metaloproteinasas/fisiología , Calcificación Vascular/metabolismo , Calcificación Vascular/fisiopatologíaRESUMEN
Some antioxidant compounds decrease the amount of intracellular reactive oxygen species (ROS) and consequently reduce the deleterious effects of ROS in osteoblasts. Thus, these compounds fight against osteoporosis. Brown seaweeds are a rich source of antioxidant fucose-containing sulfated polysaccharides (fucans and fucoidans). We obtained six fucoidans (FRFs)-F0.3, F0.5, F0.7, F1.0, F1.5, and F2.1-from Dictyota mertensii by proteolytic digestion followed by sequential acetone precipitation. Except for F0.3, all FRFs showed antioxidant activity in different in vitro tests. In pre- osteoblast-like cells (MC3T3-L1) exposed to H2O2-oxidative stress, caspase-3 and caspase-9 were activated, resulting in apoptosis of the cells. We also observed a decrease in superoxide dismutase (SOD) and alkaline phosphatase (ALP) activity. The antioxidant FRFs protected the cells from the oxidative damage caused by H2O2, decreasing intracellular ROS and caspase activation, and increasing SOD activity. The most effective protection against damage was provided by F0.7, F1.5, and F2.1. At 0.5 mg/mL, these FRFs also suppressed the H2O2-mediated inhibition of ALP activity. The data indicated that FRFs F0.7, F1.5, and F2.1 from D. mertensii were antioxidants that protected bone tissue from oxidative stress and could represent possible adjuvants for the treatment of bone fragility through counteracting oxidative phenomena.
Asunto(s)
Depuradores de Radicales Libres/farmacología , Phaeophyceae/química , Polisacáridos/farmacología , Algas Marinas/química , Células 3T3 , Animales , Apoptosis/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Depuradores de Radicales Libres/aislamiento & purificación , Depuradores de Radicales Libres/uso terapéutico , Humanos , Peróxido de Hidrógeno/toxicidad , Ratones , Osteoblastos/efectos de los fármacos , Osteoblastos/fisiología , Osteoporosis/tratamiento farmacológico , Oxidación-Reducción/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Polisacáridos/aislamiento & purificación , Polisacáridos/uso terapéuticoRESUMEN
Adipose-derived mesenchymal stem cells (ASCs) accelerate the osteointegration of bone grafts and improve the efficiency in the formation of uniform bone tissue, providing a practical and clinically attractive approach in bone tissue regeneration. In this work, the effect of nanofibrous biomimetic matrices composed of poly(ε-caprolactone) (PCL), nanometric hydroxyapatite (nHA) particles and 14-3-3 protein isoform epsilon on the initial stages of human ASCs (hASCs) osteogenic differentiation was investigated. The cells were characterized by flow cytometry and induction to differentiation to adipogenic and osteogenic lineages. The isolated hASCs were induced to differentiate to osteoblasts over all scaffolds, and adhesion and viability of the hASCs were found to be similar. However, the activity of alkaline phosphatase (ALP) as early osteogenic marker in the PCL-nHA/protein scaffold was four times higher than in PCL-nHA and more than five times than the measured in neat PCL.
Asunto(s)
Proteínas 14-3-3 , Durapatita , Células Madre Mesenquimatosas/efectos de los fármacos , Osteogénesis/efectos de los fármacos , Poliésteres , Andamios del Tejido/química , Proteínas 14-3-3/química , Proteínas 14-3-3/farmacología , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/fisiología , Células Cultivadas , Durapatita/química , Durapatita/farmacología , Galvanoplastia/métodos , Humanos , Ensayo de Materiales , Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/fisiología , Nanofibras/química , Osteoblastos/efectos de los fármacos , Osteoblastos/fisiología , Osteogénesis/fisiología , Poliésteres/química , Poliésteres/farmacología , Grasa Subcutánea Abdominal/citología , Propiedades de Superficie/efectos de los fármacos , Ingeniería de Tejidos/métodosRESUMEN
Bioactive glass (BG)-based scaffolds of 45S5 composition covered with hydroxyapatite nanoparticles loaded with Mg2+, Zn2+ and, both Mg2+ and Zn2+ ions, were developed and tested as materials for tissue engineering applications. The scaffolds were prepared by the foam replica technique and mono- and bi-metal loaded and unloaded hydroxyapatite nanoparticles (HA, Zn-HA, Mg-HA and Mg-Zn-HA) were obtained by an adaptation of the wet chemical deposition method. Coating of BG with these nanoparticles was performed by dip-coating to obtain HA-BG, Zn-HA-BG, Mg-HA-BG and Mg-Zn-HA-BG scaffolds. As predictor of the bone bonding ability of the produced scaffolds, in this study we investigated the formation of an apatite layer on the scaffold surfaces in the presence of simulated body fluid. The cytotoxicity and osteogenic properties of the materials in vitro was evaluated using human osteoblast-like MG-63 cell cultures. The mineralization assay following Kokubo's protocol indicated that bi-metal loaded Mg-Zn-HA-BG scaffolds exhibited higher/faster bioactivity than mono-metal loaded scaffolds while mineralization of HA-BG, Zn-HA-BG and Mg-HA-BG was similar to that of uncoated scaffolds. Moreover, an increase of proliferation of MG-63 cells after 48â¯h and 7 days was measured by BrdU assays for Mg-Zn-HA-BG scaffolds. In agreement with these results, SEM images confirmed increased interaction between these scaffolds and cells, in comparison to that observed for mono-metal-loaded HA-coated scaffolds. Altogether, the obtained results suggest that nanocrystalline Mg-Zn-HA coatings enhance the biological performance of standard scaffolds of 45S5 BG composition. Thus these novel ion doped HA coated scaffolds are attractive systems for bone tissue engineering.
Asunto(s)
Cerámica/química , Materiales Biocompatibles Revestidos/química , Durapatita/química , Vidrio/química , Magnesio/química , Osteoblastos/efectos de los fármacos , Andamios del Tejido , Zinc/química , Materiales Biomiméticos/química , Materiales Biomiméticos/farmacología , Líquidos Corporales/química , Adhesión Celular/efectos de los fármacos , Línea Celular , Proliferación Celular/efectos de los fármacos , Cerámica/farmacología , Materiales Biocompatibles Revestidos/farmacología , Durapatita/farmacología , Humanos , Nanopartículas/química , Osteoblastos/citología , Osteoblastos/fisiología , Osteogénesis/efectos de los fármacos , Ingeniería de Tejidos/métodosRESUMEN
Abstract Bone development and healing processes involve a complex cascade of biological events requiring well-orchestrated synergism with bone cells, growth factors, and other trophic signaling molecules and cellular structures. Beyond health processes, MMPs play several key roles in the installation of heart and blood vessel related diseases and cancer, ranging from accelerating metastatic cells to ectopic vascular mineralization by smooth muscle cells in complementary manner. The tissue inhibitors of MMPs (TIMPs) have an important role in controlling proteolysis. Paired with the post-transcriptional efficiency of specific miRNAs, they modulate MMP performance. If druggable, these molecules are suggested to be a platform for development of "smart" medications and further clinical trials. Thus, considering the pleiotropic effect of MMPs on mammals, the purpose of this review is to update the role of those multifaceted proteases in mineralized tissues in health, such as bone, and pathophysiological disorders, such as ectopic vascular calcification and cancer.
Asunto(s)
Humanos , Remodelación Ósea/fisiología , Metaloproteinasas de la Matriz/fisiología , Matriz Extracelular/fisiología , Osteoblastos/fisiología , Enfermedades Óseas/fisiopatología , Enfermedades Óseas/metabolismo , Progresión de la Enfermedad , Inhibidores Tisulares de Metaloproteinasas/fisiología , Calcificación Vascular/fisiopatología , Calcificación Vascular/metabolismo , Inhibidores de la Metaloproteinasa de la Matriz/uso terapéutico , Neoplasias/fisiopatología , Neoplasias/metabolismoRESUMEN
The main purpose of regenerative biology is to improve human health by exploiting cellular and molecular mechanisms favoring tissue repair. In recent years, non-mammalian vertebrates have emerged as powerful model organisms to tackle the problem of tissue regeneration. Here, we analyze the process of bone repair in metamorphosing Xenopus tropicalis tadpoles subjected to traumatic skull injury. Five days after skull perforation, a dense and highly vascularized mesenchymal is apparent over the injury site. Using an in vivo bone staining procedure based on independent pulses of Alizarin red and Calcein green, we show that the deposition of new bone matrix completely closes the wound in 15â¯days. The absence of cartilage implies that bone repair follows an intramembranous ossification route. Collagen second harmonic imaging reveals that while a well-organized lamellar type of bone is deposited during development, a woven type of bone is produced during the early-phase of the regeneration process. Osteoblasts lying against the regenerating bone robustly express fibrillar collagen 1a1, SPARC and Dlx5. These analyses establish Xenopus tropicalis as a new model system to improve traumatic skull injury recovery.