RESUMO
A bioactive borate glass, 13-93B3 (B3), has been used successfully in the clinic to treat chronic, nonhealing wounds without scarring. However, the mechanism by which B3 stimulates wound healing is poorly understood. Because adipose stem cells (ASCs) have been shown to have multiple roles in wound repair, we hypothesized that B3 triggers ASCs. In this study, we evaluate the effects of B3 on ASC survival, migration, differentiation, and protein secretion in vitro. In concentrations ≤10 mg/ml, B3 did not affect ASC viability under static conditions. B3 promoted the migration of ASCs but did not increase differentiation into bone or fat. B3 also decreased ASCs secretion of collagen I, PAI-1, MCP-1, DR6, DKK-1, angiogenin, IL-1, IGFBP-6, VEGF, and TIMP-2; increased expression of IL-1R and E-selectin; had a transient decrease in IL-6 secretion; and had a transient increase in bFGF secretion. Together, these results show that B3 alters the protein secretion of ASCs.
Assuntos
Tecido Adiposo/citologia , Boratos/química , Diferenciação Celular , Vidro/química , Células-Tronco/efeitos dos fármacos , Materiais Biocompatíveis , Movimento Celular , Sobrevivência Celular , Regulação da Expressão Gênica , Humanos , Teste de MateriaisRESUMO
Bioactive glasses have transformed healthcare due to their versatility. Bioactive borate glass, in particular, has shown remarkable healing properties for both hard and soft tissues. Incorporating dopants into the composition of bioactive glass helps to control mechanical properties, and it increases their usefulness for clinical applications. Using a bioactive borate glass, 13-93B3 (B3), we investigated eleven dopants on the viability and migration potential of adipose stem cells (ASCs), a therapeutic source of cells used in tissue engineering and cell therapy. Our results show that under standard cell culture conditions, only Cu-doped B3 decreased cell viability, while only Y-doped B3 attracted ASCs as it dissolved in cell culture media. Using a transwell invasion assay, priming ASCs with Co, Fe, Ga, I, Sr, or Zn-doped B3 increased their homing capacity. Because there is widespread interest in optimizing and enhancing the homing efficiency of ASCs and other therapeutic cells, we then tested if priming bone marrow mesenchymal stem cells (BMSCs) with dopants also increased their homing capacity. In the case of BMSCs, there was a significant increase in invasion when cells were primed with any of the doped-B3 glasses. This work shows that incorporating dopants into borate glasses can provide a platform for a safe and efficient method that stimulates endogenous cells and healing mechanisms.
Assuntos
Células-Tronco Adultas/fisiologia , Boratos/química , Vidro/química , Teste de Materiais , Células-Tronco Mesenquimais/fisiologia , Técnicas de Cultura de Células/instrumentação , Movimento Celular , Sobrevivência Celular , Humanos , Propriedades de Superfície , Engenharia Tecidual/métodosRESUMO
Glucosyltransferases (Gtfs) from S. mutans play critical roles in the development of virulent oral biofilms associated with dental caries disease. Gtfs adsorbed to the tooth surface produce glucans that promote local microbial colonization and provide an insoluble exopolysaccharides (EPS) matrix that facilitates biofilm initiation. Moreover, agents that inhibit the enzymatic activity of Gtfs in solution often have reduced or no effects on surface-adsorbed Gtfs. This study elucidated the mechanisms responsible for the differences in functionality that GtfB exhibits in solution vs surface-adsorbed. Upon adsorption to planar fused-quartz substrates, GtfB displayed a 37% loss of helices and 36% increase of ß-sheets, as determined by circular dichroism (CD) spectroscopy, and surface-induced conformational changes were more severe on substrates modified with CH3- and NH2-terminated self-assembled monolayers. GtfB also underwent substantial conformation changes when adsorbing to hydroxyapatite (HA) microspheres, likely due to electrostatic interactions between negatively charged GtfB and positively charged HA crystal faces. Conformational changes were lessened when HA surfaces were coated with saliva (sHA) prior to GtfB adsorption. Furthermore, GtfB remained highly active on sHA, as determined by in situ attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy, producing glucans that were structurally different than GtfB in solution and known to increase the accumulation and virulence of biofilms. Our data provide the first insight into the structural underpinnings governing Gtf conformation and enzymatic function that occur on tooth surfaces in vivo, which may lead to designing potent new inhibitors and improved strategies to combat the formation of pathogenic oral biofilms.
Assuntos
Durapatita/química , Glucanos/biossíntese , Glucosiltransferases/química , Glucosiltransferases/metabolismo , Saliva/química , Adsorção , Durapatita/metabolismo , Glucanos/química , Conformação Molecular , Tamanho da Partícula , Saliva/metabolismo , Streptococcus mutans/enzimologia , Streptococcus mutans/metabolismo , Propriedades de SuperfícieRESUMO
The aim of this study was to propose and validate a new unified method for testing dissolution rates of bioactive glasses and their variants, and the formation of calcium phosphate layer formation on their surface, which is an indicator of bioactivity. At present, comparison in the literature is difficult as many groups use different testing protocols. An ISO standard covers the use of simulated body fluid on standard shape materials but it does not take into account that bioactive glasses can have very different specific surface areas, as for glass powders. Validation of the proposed modified test was through round robin testing and comparison to the ISO standard where appropriate. The proposed test uses fixed mass per solution volume ratio and agitated solution. The round robin study showed differences in hydroxyapatite nucleation on glasses of different composition and between glasses of the same composition but different particle size. The results were reproducible between research facilities. Researchers should use this method when testing new glasses, or their variants, to enable comparison between the literature in the future.
Assuntos
Apatitas/química , Materiais Biomiméticos/química , Materiais Biomiméticos/normas , Líquidos Corporais/química , Cerâmica/química , Vidro/química , Teste de Materiais/normas , Apatitas/normas , Cerâmica/análise , Cerâmica/normas , Vidro/análise , Vidro/normas , Internacionalidade , Teste de Materiais/métodos , Tamanho da Partícula , Padrões de Referência , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
PURPOSE: To test whether iron oxide (IO)-containing yttrium aluminosilicate (YAS) microparticles (MPs) can generate localized therapeutic hyperthermia (≥ 43°C) when injected intratumorally in an animal model of liver cancer and whether MP distributions could be visualized with magnetic resonance (MR) imaging. MATERIALS AND METHODS: Twenty-one Sprague-Dawley rats implanted with N1-S1 liver tumors were assigned to alternating magnetic field (AMF) exposure following intratumoral injection with IO-YAS MPs (n = 7), sham surgery (n = 7), or baseline iron quantification (n = 7). Three fiberoptic probes allowed spatial and temporal monitoring of temperatures during 24 minutes of AMF exposure. T2-weighted turbo spin-echo MR imaging was performed within 1 hour after the procedure to detect signal voids caused by IO-YAS deposition. Hematoxylin and eosin-stained pathologic slides were also obtained, and the presence of IO-YAS was evaluated with inductively coupled plasma optical emission spectroscopy. RESULTS: Following AMF exposure, intratumoral temperatures after IO-YAS MP injection achieved therapeutic hyperthermia whereas those after sham surgery did not (46.6°C ± 1.3 vs 36.8°C ± 0.4; P < .0001). Within the treated group, the normal hepatic parenchyma (NHP) and rectal temperatures were 37.4°C ± 0.9 and 36.5°C ± 1.0 (P = .0809) at the conclusion of AMF exposure, respectively. A T2-weighted signal void at the tumor site was observed in all seven treated animals, and intratumoral IO-YAS was visualized on subsequent histopathologic examination in each case. The mean ratio of tumor:NHP Fe concentrations attributable to IO-YAS MPs was 108:1. CONCLUSIONS: AMF exposure of intratumoral IO-YAS MPs generates localized therapeutic hyperthermia in an animal model of liver cancer. MR detectability and potential for combination brachytherapy warrants further investigation for thermoradiotherapy in liver cancer.
Assuntos
Compostos Férricos/uso terapêutico , Hipertermia Induzida/métodos , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas/terapia , Imagem por Ressonância Magnética Intervencionista/métodos , Ítrio/uso terapêutico , Animais , Braquiterapia/métodos , Linhagem Celular Tumoral , Terapia Combinada/métodos , Estudos de Viabilidade , Masculino , Microesferas , Radioterapia Guiada por Imagem/métodos , Ratos , Ratos Sprague-Dawley , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Resultado do TratamentoRESUMO
Microfibrous bioactive glasses are showing a considerable capacity to heal soft tissue wounds, but little information is available on the mechanism of healing. In the present study, the conversion of microfibrous borate bioactive glass (diameter = 0.2-5 µm) with the composition designated 13-93B3 (5.5 Na2O, 11.1 K2O, 4.6 MgO, 18.5 CaO, 3.7 P2O5, 56.6 B2O3 wt%) was evaluated in vitro as a function of immersion time in a simulated body fluid (SBF) at 37 °C using structural and chemical techniques. Silicate 45S5glass microfibers (45 SiO2, 24.5 Na2O, 24.5 CaO, 6 P2O5 wt%) were also studied for comparison. Microfibrous 13-93B3 glass degraded almost completely and converted to a calcium phosphate material within 7-14 days in SBF, whereas >85 % of the silica remained in the 45S5 microfibers, forming a silica gel phase. An amorphous calcium phosphate (ACP) product that formed on the 13-93B3 microfibers crystallized at a slower rate to hydroxyapatite (HA) when compared to the ACP that formed on the 45S5 fibers. For immersion times >3 days, the 13-93B3 fibers released a higher concentration of Ca into the SBF than the 45S5 fibers. The fast and more complete degradation, slow crystallization of the ACP product, and higher concentration of dissolved Ca in SBF could contribute to the capacity of the microfibrous borate 13-93B3 glass to heal soft tissue wounds.
Assuntos
Líquidos Corporais , Boratos/química , Vidro , Silicatos/química , Microscopia Eletrônica de Varredura , Microscopia Eletrônica de Transmissão , Espectroscopia de Infravermelho com Transformada de Fourier , Difração de Raios XRESUMO
The primary objective of this study was to evaluate in vitro responses of MLO-A5 osteogenic cells to two modifications of the bioactive glass 13-93. The modified glasses, which were designed for use as cell support scaffolds and contained added boron to form the glasses 13-93 B1 and 13-93 B3, were made to accelerate formation of a bioactive hydroxyapatite surface layer and possibly enhance tissue growth. Quantitative MTT cytotoxicity tests revealed no inhibition of growth of MLO-A5 cells incubated with 13-93 glass extracts up to 10 mg/ml, moderate inhibition of growth with 13-93 B1 glass extracts, and noticeable inhibition of growth with 13-93 B3 glass extracts. A morphology-based biocompatibility test was also performed and yielded qualitative assessments of the relative biocompatibilities of glass extracts that agree with those obtained by the quantitative MTT test. However, as a proof of concept experiment, when MLO-A5 cells were seeded onto 13-93 B3 scaffolds in a dynamic in vitro environment, cell proliferation occurred as evidenced by qualitative and quantitative MTT labeling of scaffolds. Together these results demonstrate the in vitro toxicity of released borate ion in static experiments; however borate ion release can be mitigated in a dynamic environment similar to the human body where microvasculature is present. Here we argue that despite toxicity in static environments, boron-containing 13-93 compositions may warrant further study for use in tissue engineering applications.
Assuntos
Boro/química , Vidro/química , Osteoblastos/efeitos dos fármacos , Animais , Boro/farmacologia , Compostos de Boro/síntese química , Compostos de Boro/química , Compostos de Boro/farmacologia , Proliferação de Células/efeitos dos fármacos , Forma Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Formazans/farmacologia , Camundongos , Osteoblastos/citologia , Osteoblastos/fisiologia , Sais de Tetrazólio/farmacologia , Engenharia Tecidual/instrumentação , Alicerces Teciduais/efeitos adversosRESUMO
Recent advances in additive manufacturing have led to the development of innovative solutions for tissue regeneration. Hydrogel materials have gained significant attention for burn wound treatment in clinical practice among various advanced dressings due to their soothing and moisturizing activity. However, prolonged healing, pain, and traumatic removal due to the lack of long-term wound hydration are some of the challenges in the treatment of second-degree burn wounds. In this study, 3D-printed dressings were fabricated using gelatin, alginate, and bioactive borate glass (BBG) using an extrusion-based bioprinter. After ionic crosslinking, the 3D-printed dressings were characterized for mechanical properties, degradation rate, hydration activity, and in vitro cell viability using human fibroblasts. The results demonstrated that in 3D-printed dressings with 20 wt% BBG, Young's modulus increased by 105%, and 10-day degradation rate decreased by 62%. Addition of BBG prevented the burst release of water from hydrogel dressings and enabled the continuous water release for up to 10 days, which is crucial in treating second-degree burn wounds. 3D-printed hydrogel dressings with BBG showed long-term cell viability that can be a result of the accumulative release of therapeutic ions from BBG particulate. The in vivo wound healing functionality of the dressings was investigated using a rat model with a second-degree burn wound. Our animal study showed that the 3D-printed dressings with BBG exhibited faster wound closure, non-adhesive contact, non-invasive debridement, and non-traumatic dressing removal. Histological analysis suggested that 3D-printed dressings contributed to more uniform re-epithelialization and tissue remodeling compared to the non-printed hydrogels of the same compositions. Critically, 3D-printed dressings with BBG led to significant regeneration of hair follicles compared to the 3D-printed hydrogel, non-printed hydrogel, and the control groups. The superior outcome of the 3D-printed hydrogel-BBG20 dressings can be attributed to the bioactive formulation, which promotes moist wound healing for longer time periods, and the non-adhesive porous texture of the 3D-printed dressings with increased wound-dressing interactions. Our findings provided proof of concept for the synergistic effect of bioactive formulation and the porous texture of the 3D-printed hydrogel dressings incorporated with BBG on continuous water release and, consequently, on second-degree burn wound healing.
RESUMO
The conversion of 45S5 glass and glass-ceramics to a hydroxyapatite (HA)-like material in vitro has been studied extensively, but only for short reaction times (typically <3 months). In this paper, we report for the first time on the long-term conversion of 45S5 glass-ceramic microspheres (designated 45S5c) in an aqueous phosphate solution. Microspheres of 45S5c (75-150 µm) were immersed for 10 years at room temperature (~25 °C) in K(2)HPO(4) solution with a concentration of 0.01 M or 1.0 M, and with a starting pH of 7.0 or 9.5. The reacted 45S5c microspheres and solutions were analyzed using structural and analytical techniques. Only 25-45 vol% of the 45S5c microspheres were converted to an HA-like material after the 10 year reaction. In solutions with a starting pH of 9.5, an increase in the K(2)HPO(4) concentration from 0.01 to 1.0 M resulted in a doubling of the volume of the microspheres converted to an HA-like material but had little effect on the composition of the HA-like product. In comparison, reaction of the 45S5c microspheres in the solution with a starting pH of 7.0 resulted in an HA-like product in the 0.01 M K(2)HPO(4) solution but a calcium pyrophosphate product, Ca(10)K(4)(P(2)O(7))(6).9H(2)O, in the 1.0 M solution. The consequences of these results for the long-term use of 45S5 glass-ceramics in biomedical applications are discussed.
Assuntos
Cerâmica/química , Vidro/química , Microesferas , Transição de Fase , Fosfatos/farmacologia , Água/farmacologia , Difusão , Estabilidade de Medicamentos , Microquímica , Microscopia Eletrônica de Varredura , Concentração Osmolar , Fosfatos/química , Compostos de Potássio/química , Compostos de Potássio/farmacologia , Soluções/química , Soluções/farmacologia , Espectroscopia de Infravermelho com Transformada de Fourier , Fatores de Tempo , Água/química , Difração de Raios XRESUMO
Hollow hydroxyapatite (HA) microspheres were prepared by reacting solid microspheres of Li(2)O-CaO-B(2)O(3) glass (106-150 µm) in K(2)HPO(4) solution, and evaluated as a controlled delivery device for a model protein, bovine serum albumin (BSA). Reaction of the glass microspheres for 2 days in 0.02 M K(2)HPO(4) solution (pH = 9) at 37°C resulted in the formation of biocompatible HA microspheres with a hollow core diameter equal to 0.6 the external diameter, high surface area (~100 m(2)/g), and a mesoporous shell wall (pore size ≈ 13 nm). After loading with a solution of BSA in phosphate-buffered saline (PBS) (5 mg BSA/ml), the release kinetics of BSA from the HA microspheres into a PBS medium were measured using a micro bicinchoninic acid (BCA) protein assay. Release of BSA initially increased linearly with time, but almost ceased after 24-48 h. Modification of the BSA release kinetics was achieved by modifying the microstructure of the as-prepared HA microspheres using a controlled heat treatment (1-24 h at 600-900°C). Sustained release of BSA was achieved over 7-14 days from HA microspheres heated for 5 h at 600°C. The amount of BSA released at a given time was dependent on the concentration of BSA initially loaded into the HA microspheres. These hollow HA microspheres could provide a novel inorganic device for controlled local delivery of proteins and drugs.
Assuntos
Materiais Biocompatíveis/química , Durapatita/química , Microesferas , Proteínas/química , Células 3T3 , Animais , Bovinos , Sistemas de Liberação de Medicamentos , Temperatura Alta , Concentração de Íons de Hidrogênio , Cinética , Camundongos , Microscopia Eletrônica de Varredura/métodos , Soroalbumina Bovina/química , Espectroscopia de Infravermelho com Transformada de Fourier/métodos , Temperatura , Fatores de Tempo , Difração de Raios XRESUMO
A solid freeform fabrication technique, freeze extrusion fabrication (FEF), was investigated for the creation of three-dimensional bioactive glass (13-93) scaffolds with pre-designed porosity and pore architecture. An aqueous mixture of bioactive glass particles and polymeric additives with a paste-like consistency was extruded through a narrow nozzle, and deposited layer-by-layer in a cold environment according to a computer-aided design (CAD) file. Following sublimation of the ice in a freeze dryer, the construct was heated according to a controlled schedule to burn out the polymeric additives (below ~500°C), and to densify the glass phase at higher temperature (1 h at 700°C). The sintered scaffolds had a grid-like microstructure of interconnected pores, with a porosity of ~50%, pore width of ~300 µm, and dense glass filaments (struts) with a diameter or width of ~300 µm. The scaffolds showed an elastic response during mechanical testing in compression, with an average compressive strength of 140 MPa and an elastic modulus of 5-6 GPa, comparable to the values for human cortical bone. These bioactive glass scaffolds created by the FEF method could have potential application in the repair of load-bearing bones.
Assuntos
Materiais Biocompatíveis/química , Osso e Ossos/patologia , Substitutos Ósseos/química , Osso e Ossos/metabolismo , Força Compressiva , Elasticidade , Consolidação da Fratura , Vidro/química , Humanos , Teste de Materiais , Polímeros/química , Porosidade , Pressão , Estresse Mecânico , Temperatura , Termogravimetria , Alicerces Teciduais/químicaRESUMO
The conversion of glass to a hydroxyapatite (HA) material in an aqueous phosphate solution is used as an indication of the bioactive potential of the glass, as well as a low temperature route for preparing biologically useful materials. In this work, the effect of varying concentrations of pyrophosphate ions in the phosphate solution on the conversion of a calcium-lithium-borate glass to HA was investigated. Particles of the glass (150-355 µm) were immersed for up to 28 days in 0.25 M K(2)HPO(4) solution containing 0-0.1 M K(4)P(2)O(7). The kinetics of degradation of the glass particles and their conversion to HA were monitored by measuring the weight loss of the particles and the ionic concentration of the solution. The structure and composition of the conversion products were analyzed using X-ray diffraction, scanning electron microscopy, and Fourier transform infrared spectroscopy. For K(4)P(2)O(7) concentrations of up to 0.01 M, the glass particles converted to HA, but the time for complete conversion increased from 2 days (no K(4)P(2)O(7)) to 10 days (0.01 M K(4)P(2)O(7)). When the K(4)P(2)O(7) concentration was increased to 0.1 M, the product consisted of an amorphous calcium phosphate material, which eventually crystallized to a pyrophosphate product (predominantly K(2)CaP(2)O(7) and Ca(2)P(2)O(7)). The consequences of the results for the formation of HA materials and devices by the glass conversion route are discussed.
Assuntos
Materiais Biocompatíveis/síntese química , Durapatita/síntese química , Materiais Biocompatíveis/química , Boratos , Cálcio , Difosfatos , Durapatita/química , Vidro , Lítio , Teste de Materiais , Microscopia Eletrônica de Varredura , Fosfatos , Soluções , Espectroscopia de Infravermelho com Transformada de Fourier , Água , Difração de Raios XRESUMO
Solid microspheres (diameter = 106-150 µm) of a Li(2)O-CaO-B(2)O(3) glass were reacted in a K(2)HPO(4) solution to form hollow hydroxyapatite (HA) microspheres. The effect of the temperature (25°-60°C), K(2)HPO(4) concentration (0.01-0.25M), and pH (9-12) of the solution on the diameter (d) of the hollow core normalized to the diameter (D) of the HA microspheres, the surface area, and the pore size of the microsphere wall was studied. The statistically significant process variables that influenced these microstructural characteristics were evaluated using a factorial design approach. While the pH had little effect, the concentration of the solution had a marked effect on d/D, surface area, and pore size, whereas temperature markedly influenced d/D and pore size, but not the surface area. The largest hollow core size (d/D value ≈ 0.6) was obtained at the lowest temperature (25°C) or the lowest K(2)HPO(4) concentration (0.02M), while microspheres with the highest surface area (140 m(2)/g), with pores of size 10-12 nm were obtained at the highest concentration (0.25M). The consequences of these results for potential application of these hollow HA microspheres as devices for local delivery of proteins, such as drugs or growth factors, are discussed.
RESUMO
Osteons are the repeating unit throughout cortical bone, consisting of canals filled with blood and nerve vessels surrounded by concentric lamella of hydroxyapatite-containing collagen fibers, providing mechanical strength. Creating a biodegradable scaffold that mimics the osteon structure is crucial for optimizing cellular infiltration and ultimately the replacement of the scaffold with native cortical bone. In this study, a modified air-gap electrospinning setup was exploited to continuously wrap highly aligned polycaprolactone polymer nanofibers around individual 1393 bioactive glass microfibers, resulting in a synthetic structure similar to osteons. By varying the parameters of the device, scaffolds with polymer fibers wrapped at angles between 5-20° to the glass fiber were chosen. The scaffold indicated increased cell migration by demonstrating unidirectional cell orientation along the fibers, similar to recent work regarding aligned nerve and muscle regeneration. The wrapping decreased the porosity from 90% to 80%, which was sufficient for glass conversion through ion exchange validated by inductively coupled plasma. Scaffold degradation was not cytotoxic. Encapsulating the glass with polymer nanofibers caused viscoelastic deformation during three-point bending, preventing typical brittle glass fracture, while maintaining cell migration. This scaffold design structurally mimics the osteon, with the intent to replace its material compositions for better regeneration.
RESUMO
Three-dimensional (3D) bioprinting technologies have shown great potential in the fabrication of 3D models for different human tissues. Stem cells are an attractive cell source in tissue engineering as they can be directed by material and environmental cues to differentiate into multiple cell types for tissue repair and regeneration. In this study, we investigate the viability of human adipose-derived mesenchymal stem cells (ASCs) in alginate-gelatin (Alg-Gel) hydrogel bioprinted with or without bioactive glass. Highly angiogenic borate bioactive glass (13-93B3) in 50 wt% is added to polycaprolactone (PCL) to fabricate scaffolds using a solvent-based extrusion 3D bioprinting technique. The fabricated scaffolds with 12 × 12 × 1 mm3 in overall dimensions are physically characterized, and the glass dissolution from PCL/glass composite over a period of 28 days is studied. Alg-Gel composite hydrogel is used as a bioink to suspend ASCs, and scaffolds are then bioprinted in different configurations: Bioink only, PCL+bioink, and PCL/glass+bioink, to investigate ASC viability. The results indicate the feasibility of the solvent-based bioprinting process to fabricate 3D cellularized scaffolds with more than 80% viability on day 0. The decrease in viability after 7 days due to glass concentration and static culture conditions is discussed. The feasibility of modifying Alg-Gel with 13-93B3 glass for bioprinting is also investigated, and the results are discussed.
RESUMO
Bioactive glasses have recently gained attention in tissue engineering and three-dimensional (3D) bioprinting because of their ability to enhance angiogenesis. Some challenges for developing biological tissues with bioactive glasses include incorporation of glass particles and achieving a 3D architecture mimicking natural tissues. In this study, we investigate the fabrication of scaffolds with a polymer/bioactive glass composite using near-field electrospinning (NFES). An overall controlled 3D scaffold with pores, containing random fibers, is created and aimed to provide superior cell proliferation. Highly angiogenic borate bioactive glass (13-93B3) in 20 wt.% is added to polycaprolactone (PCL) to fabricate scaffolds using the NFES technique. Scaffolds measuring 5 mm × 5 mm × 0.2 mm3 in overall dimensions were seeded with human adipose-derived mesenchymal stem cells to investigate the cell viability. The cell viability on PCL and PCL+glass scaffolds fabricated using NFES technique and 3D printing is compared and discussed. The results indicated higher cell proliferation on 3D biomimetic scaffolds fabricated by NFES technique.
RESUMO
This in vitro study was conducted to evaluate the ability of two types of constructs of bioactive, silica-based 13-93 glass fibers to support the growth and differentiation of MC3T3-E1 osteoblastic cells. The two types of constructs tested included single-layer 13-93 glass fiber rafts and three-dimensional porous scaffolds formed from sintered 13-93 fibers. Scanning electron micrographs showed a closely adhering, well-spread morphology of MC3T3-E1 cells seeded on both types of constructs. The scanning electron microscopy images also showed a continuous increase in cell densities during a 6 day incubation on 13-93 glass fiber rafts and scaffolds. Quantitative fluorescence measurements of DNA also revealed a linear increase in cell density during a 6 day incubation on both types of 13-93 constructs. Examination of scaffolds incubated in MTT containing medium showed the presence of metabolically active viable cells within the interior of the scaffold. The addition of ascorbic acid to MC3T3-E1 cells cultured on the 13-93 glass fibers triggered a threefold increase in alkaline phosphatase, a key indicator of osteoblast differentiation. The sintered scaffolds were found to have open, interconnected pores favorable for tissue ingrowth with a compressive strength similar to cancellous bone. Collectively, the results indicate that 13-93 glass fiber scaffolds are a favorable substrate for the growth and differentiation of osteoblasts and a promising material for bone tissue engineering and repair of bone defects.
Assuntos
Substitutos Ósseos , Cerâmica , Osteoblastos/citologia , Células 3T3 , Fosfatase Alcalina/metabolismo , Animais , Diferenciação Celular , Proliferação de Células , Sobrevivência Celular , DNA/metabolismo , Teste de Materiais , Camundongos , Microscopia Eletrônica de Varredura , Osteoblastos/metabolismo , Engenharia Tecidual , Difração de Raios XRESUMO
The silicate-based 45S5 bioactive glass, typically in particulate form, has been widely investigated for bone repair. However, its application as a scaffold for bone tissue engineering is limited due to the difficulty of forming porous three-dimensional constructs with complex shapes. In this study, the use of another silicate-based bioactive glass, referred to as 13-93, was investigated for the preparation of porous constructs. Particles of 13-93 glass (255-325 microm) were consolidated and sintered to form cylindrical constructs. Characterization of these constructs was performed using mercury porosimetry, scanning electron microscopy (SEM), and mechanical testing. Constructs with porosities of 40-45% and pore sizes in the range 100-300 microm were found to have a compressive strength of 22 +/- 1 MPa. The bioactivity of the 13-93 glass was studied by immersing disks in a simulated body fluid at 37 degrees C and characterizing the reaction products. X-ray diffraction, Fourier transform infrared (FTIR) spectroscopy, and SEM showed the formation of a crystalline hydroxyapatite layer on the glass surface after approximately 7 days. The ability to fabricate the complex geometrical shape of the articulating surface of a human tibia from 13-93 glass particles was demonstrated.
Assuntos
Materiais Biocompatíveis , Vidro , Tíbia/anatomia & histologia , Tíbia/cirurgia , Engenharia Tecidual/métodos , Fenômenos Biomecânicos , Durapatita/química , Humanos , Teste de Materiais , Microscopia Eletrônica de Varredura , Próteses e Implantes , Espectroscopia de Infravermelho com Transformada de Fourier , Propriedades de Superfície , Difração de Raios XRESUMO
Biomaterials with nanostructured surfaces influence cellular response in a significantly different, and often beneficial, manner compared to materials with coarser features. Hydroxyapatite [HA, Ca10(PO4)6(OH)2] and strontium-apatite [Sr10(PO4)6(OH)2] microspheres that present nanotopographies similar to biological apatites were incubated in albumin solutions, at physiological conditions (40 mg ml-1; 37 °C), for up to 72 h. Electronic and vibrational circular dichroism spectroscopies revealed spectral signatures characteristic of stacked ß-sheet regions in higher ordered structures (e.g., fibrils). The presence of stacked ß-sheets was further evidenced by thioflavin T staining. The sequestration of interfacial Ca atoms by pyrophosphate ions (P2O74-), prior to albumin adsorption, prevented stacked ß-sheet formation on hydroxyapatite. These results suggest that the charge and/or spatial arrangement of Ca atoms direct stacked ß-sheet formation during bovine serum albumin adsorption. Stacked ß-sheet spectral features were also observed after incubating HA in fetal bovine serum, highlighting that this phenomena could direct cellular response to these biomaterials in vivo.
Assuntos
Durapatita/química , Nanoestruturas/química , Soroalbumina Bovina/química , Animais , Bovinos , Estrutura Secundária de ProteínaRESUMO
A major limitation of using synthetic scaffolds in tissue engineering applications is insufficient angiogenesis in scaffold interior. Bioactive borate glasses have been shown to promote angiogenesis. There is a need to investigate the biofabrication of polymer composites by incorporating borate glass to increase the angiogenic capacity of the fabricated scaffolds. In this study, we investigated the bioprinting of human adipose stem cells (ASCs) with a polycaprolactone (PCL)/bioactive borate glass composite. Borate glass at the concentration of 10 to 50 weight %, was added to a mixture of PCL and organic solvent to make an extrudable paste. ASCs suspended in Matrigel were ejected as droplets using a second syringe. Scaffolds measuring 10 x 10 x 1 mm3 in overall dimensions with pore sizes ranging from 100 - 300 µm were fabricated. Degradation of the scaffolds in cell culture medium showed a controlled release of bioactive glass for up to two weeks. The viability of ASCs printed on the scaffold was investigated during the same time period. This 3D bioprinting method shows a high potential to create a bioactive, highly angiogenic three-dimensional environment required for complex and dynamic interactions that govern the cell's behavior in vivo.