Eri Hirata

Development of a 3D Collagen Scaffold Coated With Multiwalled Carbon Nanotubes

Carbon nanotubes (CNTs) have attractive biochemical properties such as strong cell adhesion and protein absorption, which are very useful for a cell cultivation scaffold. In this study, we prepared a multiwalled carbon nanotube-coated collagen sponge (MWCNT-coated sponge) to improve the surface properties of the collagen sponge, and its cell culturing properties were examined. The suface of the collagen sponge was homogeneously coated with MWCNTs by dispersion. MC3T3-E1 cells were cultured on and inside the MWCNT-coated sponge. The DNA content on the MWCNT-coated sponge after 1 week of culture was significantly higher than on an uncoated collagen sponge (p < 0.05). There was no significant difference between the estimated ALP activity normalized by DNA quantity on the MWCNT-coated sponge and that on the uncoated collagen sponge which is well known as one of the best scaffolds for cell cultivation. In addition, the MWCNT-coated surface shows strong cell adhesion. Therefore, the MWCNT-coated collagen sponge is expected to be a useful 3D scaffold for cell cultivation. (c) 2009 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2009.

3D collagen scaffolds coated with multiwalled carbon nanotubes : Initial cell attachment to internal surface

The cell adhesion in a multiwalled carbon nanotube-coated collagen sponge (MWCNT-coated sponge) was investigated. Immediately after seeding, the cells adhered to the inner surface of the MWCNT-coated sponge and a significantly larger number of cells were observed there than for a pure collagen sponge used as control. On the MWCNT-coated sponge, the cells appeared favorable adhesion and spread in the early stages in the center part of the sponge which cells rarely attached without MWCNT-coating. It was suggested that the physical structure of MWCNTs was effective for initial adhesion of cells from the result of serum-free culture. MWCNT-coating makes the material a suitable three-dimensional scaffold for cell culturing, as opposed to other scaffold systems where such an effect is not seen.

Carbon nanotubes functionalized with fibroblast growth factor accelerate proliferation of bone marrow-derived stromal cells and bone formation

Multi-walled carbon nanotubes (MWCNTs) were functionalized with fibroblast growth factor (FGF) and the advantages of their use as scaffolds for bone augmentation were evaluated in vitro and in vivo. The activity of FGF was assessed by measuring the effect on the proliferation of rat bone marrow stromal cells (RBMSCs). The presence of FGF enhanced the proliferation of RBMSCs and the FGF covalently conjugated to the nanotubes (FGF-CNT) showed the same effect as FGF alone. In addition, FGF-CNT coated sponges were implanted between the parietal bone and the periosteum of rats and the formation of new bone was investigated. At day 14 after implantation, a larger amount of newly formed bone was clearly observed in most pores of FGF-CNT coated sponges. These findings indicated that MWCNTs accelerated new bone formation in response to FGF, as well as the integration of particles into new bone during its formation. Scaffolds coated with FGF-CNT could be considered as promising novel substituting materials for bone regeneration in future tissue engineering applications.

Carbon nanohorns allow acceleration of osteoblast differentiation via macrophage activation

Carbon nanohorns (CNHs), formed by a rolled graphene structure and terminating in a cone, are promising nanomaterials for the development of a variety of biological applications. Here we demonstrate that alkaline phosphatase activity is dramatically increased by coculture of human monocyte derived macrophages (hMDMs) and human mesenchymal stem cells (hMSCs) in the presence of CNHs. CNHs were mainly localized in the lysosome of macrophages more than in hMSCs during coculturing. At the same time, the amount of Oncostatin M (OSM) in the supernatant was also increased during incubation with CNHs. Oncostatin M (OSM) from activated macrophage has been reported to induce osteoblast differentiation and matrix mineralization through STAT3. These results suggest that the macrophages engulfed CNHs and accelerated the differentiation of mesenchymal stem cells into the osteoblast via OSM release. We expect that the proof-of-concept on the osteoblast differentiation capacity by CNHs will allow future studies focused on CNHs as ideal therapeutic materials for bone regeneration.

Transmission electron microscopic observation of cells cultured on multiwalled carbon nanotube-coated sponges

The cell structure and interface between cultured cells and a multiwalled carbon nanotube (MWCNT)-coated sponge (MWCNT-coated sponge) were observed by transmission electron microscopy (TEM). Moreover, the atomic structure of MWCNTs that entered the cells was also examined by means of high-resolution TEM (HRTEM). MWCNTs were observed in the cytoplasm, and a few MWCNTs were recognized in the cell nuclei. Those MWCNTs maintained their structure there. Subcellular organelles did not appear to be different from those on the collagen sponge despite the cellular uptake of MWCNTs.

Various Nanotube Scaffolds for Cell Proliferation

Carbon nanotubes (CNT) and their derivatives with different structure and compositions have unique features. In the present study, cell proliferation was performed on various nanotubes such as single walled CNTs, multiwalled CNTs and imogolite which is nanotubes of aluminosilicate. SEM observation of the growth of osteoblast-like cells cultured on CNTs showed the morphology fully developed for the whole direction, which was different from that extended to the one direction on the usual scaffold. Numerous filopodia were grown from cell edge, extended far long and combined with CNT meshwork. Apatite precipitation in simulated body fluid, affinity for proteins and saccharides, and nanosize meshwork structure with large porosity would be the properties responsible for these cell adhesion and growth. Imogolite showed the similar properties to CNTs. Nanotubes could be the favorable materials for biomedical applications.

Visualization of Invasion into the Body and Internal Diffusion of Nanoparticles

Nanoparticles may invade directly into the internal body through the respiratory or digestive system and diffuse inside body. The behavior of nanoparticles in the internal body is also essential to comprehend for the realization of DDS. Thus it is necessary to reveal the internal dynamics for the proper treatments and biomedical applications of nanoparticles. In the present study the plural methods with different principles such as X-ray scanning analytical microscope (XSAM), MRI and Fluorescent microscopy were applied to enable the observation of the internal diffusion of micro/nanoparticles in the (1) whole body level, (2) inner organ level and (3) tissue and intracellular level. Chemical analysis was also done by ICP-AES for organs and compared with the results of XSAM mapping.

The Effects of the Coating of Anodized Titanium with Multi-Walled Carbon Nanotubes on Bone Formation

Carbon nanotubes (CNTs) have excellent chemical, physical, and biological properties such as strong cell adhesion, protein adsorption and cell proliferation in vitro. Excellent osteocompatibility for the CNT monolith was also reported in vivo. The purpose of this study was to evaluate the effects of anodized titanium coated with multiwalled CNTs (MWCNTs) on human osteosarcoma Saos2 cells and bone tissue. Saos2 cells on CNT-Ti showed excellent proliferation with extension of cell morphology in all directions. CNT-Ti wire was implanted in the bone marrow of femurs of rats. At 2 weeks after surgery, histological investigations revealed that bone tissue attached to the surface of the CNT-Ti directly. Thus the surface modification of anodized Ti by MWCNTs can be effective for bone formation.