Abstract: The risks of nanomaterials for future generations should be elucidated. Thus, it is important to establish an experimental method to accurately examine embryotoxicity. We have conducted an in vitro embryotoxicity test with mouse ES cells to examine the embryotoxicities of various nanomaterials. In this study, the C60 fullerene did not influence the differentiation of ES-D3 cells and "non embryotoxicity". In the future, the biological safety should be comprehensively examined by improving dispersion in medium.
Abstract: The aim of the present investigation was to examine Si release from the silicon-containing apatite fiber scaffold (Si-AFS) and the biocompatibility of the Si-AFS. We have successfully synthesized silicon-containing apatite fibers (Si-AF) by a homogenous precipitation method. Three-dimensional Si-AFS were fabricated using these Si-AFs. The concentrations of Si in the starting solution were 0 (AF) and 0.8 (0.8Si-AF) mass%. The 0.8Si-AFS1000 were fabricated by firing Si-AF slurry compacts (carbon/Si-AF [w/ ratio: 10/1) at 1300 °C for 5 h. Solubility experiments were carried out in 0.05 mol/dm3 Tris-HCl buffer solutions at pH 7.30 using 0.8Si-AFS1000 (porosity: ~98%), together with Si-free AFS1000 (~98%) for 21 days. The Ca2+, PO43- and SiO44- concentrations in the solution were determined by inductively-coupled plasma atomic emission spectrometry (ICP-AES). The biocompatibility of the Si-AFS was examined in vitro using osteoblastic cell, MC3T3-E1 for 21 days. The results of the ICP-AES analysis indicated that the amount of SiO44- ions released from 0.8Si-AFS1000 rapidly increased at 1 day, and then the released SiO44- ions remained constant over a period for 21 days. The cells seeded on/in the 0.8Si-AFS1000 well-proliferated as compared to those on/in the AFS1000. Consequently, we can conclude that the 0.8Si-AFS offers as a potential novel scaffold material, creating a three-dimensional cell culture environment.
Abstract: We have successfully developed porous apatite-fiber scaffolds (AFSs) which have three-dimensional (3D) inter-connected pores; subsequently, we have clarified that the AFSs have an excellent bioactivity on the basis of both in vitro and in vivo evaluations. In addition, we have reconstructed the tissue-engineered bone with 3D structure through 3D-cell culture of mesenchymal stem cells derived from rat bone marrow (RBMC) using the AFS settled into the radial-flow bioreactor (RFB), and examined effect of flow rate of medium in the RFB on the differentiation of osteoblasts in tissue-engineered bone. Aim in the present work is to establish of the optimal conditions of flow rate in this construction method of 3D tissue-engineered bone. The flow rates were set to 0.4, 1.3, 6.3, 11.5 and 16.5 cm3min-1; tissue-engineered bones cultured by the individual flow rates are defined as bones#1~#5. The level of differentiation of osteoblasts in all the bones#1~#5 was examined by determining the content of two kinds of differentiation maker into osteoblast, alkaline phosphatase (ALP) for initial/middle stage and osteocalcin (OC) for late stage. The ALP activity normalized for DNA content of bone#3 showed the highest value among all of them. Moreover, the OC amount normalized for DNA content of bone#3 also indicated the highest among all the examined samples. These results demonstarate that the flow rate of 6.3 cm3min-1 may promote the differentiation into osteoblast. In conclusion, we determined that this flow rate was the optimal conditions for the bone regeneratrion in RFB.
Abstract: We have successfully developed the apatite-fiber scaffold (AFS) with enhanced mechanical porosity for tissue engineering of bone and liver via two routes: i) use of two type of carbon beads with diameter of ~150 μm and ~20 μm and following ii) uniaxial pressing of the green compacts. Our Aim is to add vascular formation ability into the above AFS in order to maintain the regenerated tissues for a long time. In the present study, the AFSs with various porosities (68±2.4, 85±1.5, 89±0.6, 92±1.0%) were fabricated, and then loaded with vascular endothelial growth factor (VEGF). Drug release from VEGF-loaded AFSs with various porosities was examined by immersing them into phosphate buffer. The AFSs with the highest porosity (92%) could be released with the most VEGF among examined AFSs. In addition, we carried out preliminary study for the compatibility of vascular endothelial cells, M1 cells established by Matsuura et al. to the VEGF-loaded AFS (porosity: 92%), in order to account for the vascular formation into the pore of the AFS. The numbers of M1 cells cultured in/on the VEGF-loaded AFS were about 1.5 times that of VEGF-free AFS over a period of cell culture. These results demonstrate that the VEGF-loaded AFS with enhanced mechanical property have a good compatibility to the M1 cells as a model of vascular endothelial cells.
Abstract: Integrin, a component of the hemidesmosome, plays a role for epithelial cell migration and adhesion. This study investigated the process of peri-implant epithelium (PIE) formation after implantation, and compared it to the process of oral mucosa healing after tooth extraction. At the healing site of extraction socket without implant, the original junctional epithelium (JE) had disappeared at week 2, and the oral epithelium (OE) with integrin-α3 positive basal cells extending from the sides of the wound, then joined in the middle of the extraction socket. On the other hand in implant group, newly formed epithelium with integrin-α3 positive cells from the OE extended apically 1 week after implantation. After 3 weeks, basal cells of the new epithelium consisted of those with integrin-α3 positive but β4 negative. Finally, after 4 weeks, integrin-β4 was expressed at the implant-PIE interface. These findings suggest that integrin α3β1 plays a role in cell migration during PIE formation from OE. Furthermore, after the completion of PIE constitution, integrin α6β4 contributes to the attachment to titanium.
Abstract: Bone is a composite of approximately 65% inorganic phase (carbonate apatite, CHA) and a 35% organic phase (mostly collagen). To date, several commercial composites consisting of natural or synthetic polymers and calcium phosphates ( hydroxyapatite, tricalcium phosphate, biphasic calcium phosphates) are recommended for use in bone repair. Objective: The aim of this study was to compare the physico-chemical properties of gelatin/carbonate apatite composites with that of bovine bone. Native (Gel) or cross-linked (Gel*) was used. Methods: The CHA was prepared by hydrolysis method. The gelatin (denatured collagen) was cross-linked using Genipin. The gelatin/CHA composite were prepared by mixing of 35% gelatin and 65% CHA and freeze-drying. The composites were characterized using x-ray diffraction (XRD), FT-IR spectroscopy, scanning electron microscopy (SEM) and thermogravimetry (TGA). Dissolution properties were determined in acidic buffer (0.1M KAc, pH 6, 37°C). Mechanical strength was determined using 3-point bend test. Bovine bone was similarly characterized for comparison. Results: The composition and crystallite size of the CHA were similar to that of the bone mineral. The Gel/CHA and Gel*/CHA composites showed several physico-chemical properties (crystallinity, composition, thermal stability, mechanical strength, dissolution rate) similar to that of bone. Gel*/CHA compared to Gel/CHA composites showed lower elastic modulus, flexural strength, dissolution rate, swelling and higher porosity. Conclusion: The Gel*/CHA composites presented several properties similar to those of bovine bone and may have potential as bone substitute materials.
Abstract: Carbonate apatite (CO3Ap) foam with interconnecting porous structure is a potential candidate as bone substitute material owing to its similarity to the cancellous bone with respect to composition, morphology and osteoclastic degradation. However, it is brittle and difficult to handle. This is thought to be caused by no organic material in the CO3Ap foam. The aim of this study is to reinforce the CO3Ap foam with poly (DL-lactide-co-glycolide) (PLGA). Immersion and vacuum infiltration methods were compared as reinforcing methods. Compressive strength of unreinforced CO3Ap foam, (12.0 ± 4.9 kPa) increased after PLGA reinforcement by immersion (187.6 ± 57.6 kPa) or by vacuum infiltration (407 ± 111.4 kPa). Scanning electron microscopy (SEM) showed the preservation of full interconnecting porous structure of CO3Ap foam after PLGA reinforcement using immersion or vacuum infiltration. Interface between the PLGA and CO3Ap foam, however revealed that no gap was found between the PLGA and CO3Ap foam interface when vacuum was used to reinforce the PLGA whereas a gap was found when simple immersion was used. Strong interface between PLGA and CO3Ap foam is therefore thought to be the key for higher compressive strength. In conclusion, vacuum infiltration is a more efficient method to reinforce the CO3Ap foam with PLGA for improving the mechanical strength without sacrificing the cancellous bone-type morphology.
Abstract: Dermal fillers are injectable implants made of biological materials (collagen, autologous fat and hyaluronic acid animal) or synthetic (PMMA microparticles of hydroxyapatite and non-animal hyaluronic acid), biodegradable or not, that include features such as ideal biocompatibility, durability, non-profile migration and ability to promote a smooth, natural-looking correction. Its main indication is intended to treat contour defects caused by aging, photo damage, disease, trauma or scarification. The fact of biodegradable fillers are absorbed within a year after application resulted in the emergence of products permanent and semi-permanent to offer patients long-lasting effects. Currently, one of the most effective strategies has been the development of scaffolds formed by combining two or more biomaterials seeking the restoration of tissue function. The bioceramic associated with water-soluble polymers have been developed as substitutes for the repair of soft tissues with optimal biological response. The objective of this study was to process and characterize a composite hydrogel in the form of hyaluronic acid (HA) microspheres and biphasic calcium phosphate (BCP) in order injectable applications for repair of soft tissue. The powders of HA and BCP were characterized by Infrared Spectroscopy Fourier Transform (FTIR) and X-ray diffraction (XRD) and Scanning Electron Microscopy (SEM). The characterization of the hydrogel injectability pure and the composite with different ratios of HA and BCP was performed. The components were characterized compatible for use as dermal fillers. The composite of hyaluronic acid (HA) and biphasic calcium phosphate (BCP) had adequate characterization and injetabilidade proving to be a potential candidate for restoration of soft tissue.
Abstract: Chemically synthesized collagen with triple helix structure similar to natural collagen has been developed as a safe biomaterial. If the chemically synthesized collagen is deposited with apatite, they are expected for novel bone substitutes having bioactivity and bioresorbability. Although apatite formation on the chemically synthesized collagen has been examined, highly supersaturated condition such as 1.5SBF with ion concentration 1.5 times those of simulated body fluid (SBF) is needed to achieve apatite formation. In the present study, we intended acceleration on the apatite formation on the chemically synthesized collagen by immobilization with polyglutamic acid (PGA). PGA is known as biodegradable and biocompatible polypeptide having excellent apatite-forming ability. We examined effects of the immobilization procedure on mineralization behavior in SBF. At first, PGA was immobilized on porous sponges of chemically synthesized collagen in aqueous solutions containing PGA and CaCl2. As a result, not only apatite but also calcite-type CaCO3 was deposited on the specimens in SBF. The calcite formation was occurred during the treatment with PGA solution. pH of the solution was adjusted to 7 by NaOH solution in order to avoid dissolution of the collagen. During this procedure, Ca (OH)2 would be precipitated by locally increase in pH of the solution and converted into the calcite. When the PGA solution treatment was shortened so as to prevent the calcite formation, single phase of the apatite was formed in SBF. The present results indicate that crystalline phase deposited on the chemically synthesized collagen can be controlled by fabrication procedure, and provide fundamental design of composites containing apatite and chemically synthesized collagen useful for bone regeneration.
Abstract: Biomimetic composites of hydroxyapatite (HAp) and collagen with fast bio-absorption and good biocompatibility were designed utilizing salmon bone and skin at 283-293 K and pH 7.5-7.9 by a dissolution-precipitation method. Simultaneously, dissolved-precipitated HAp (dp-HA) was prepared at pH 9-10 using the calcined bone. The HAp/collagen composites (HA-C) were constituted by Ca2+-deficient HAp, I type-collagen and gelatin. At the synthetic temperature of 283 K, collagen fiber and HAp microcrystals were seen, while at 293 K, frock like-agglomerated particles or fiber like-oriented columnar ones were observed depending on the composition ratio (H/C) of HAp to collagen. Specific surface areas and total pore volumes for the HA-C synthesized at 293 K clearly increased with increasing the H/C, although there were micro-and-meso-pores in the pore diameters of 3-30 nm. Concerning water vapor-adsorption isotherms at 298 K for the HA-C powders, hysteresis-curves of the amounts of water vapor adsorbed (V) were recognized in the adsorption-desorption processes. The V values increased with increasing the H/C under the low relative pressures (P/PS) of 0-0.3 that mean monolayer-adsorption of water molecule. However, for H/C=2.2, the V values were the highest under the high P/PS of 0.70-0.90 that mean multilayer-adsorption and the biggest hysteresis-curve was found under the P/PS of 0.45-0.60, suggesting that the HA-C (H/C=2.2) powder not only adsorb water molecule on the surfaces but also absorb one into the crystal structure. Accordingly, the biomimetic HA-C powders will be applied as water-absorbable adsorption materials for cosmetic products or bone-regeneration therapy.