Papers by Author: Nobutaka Hanagata

Abstract: Silica-chitosan hybrid was synthesized from a mixture of chitosan and glycidyloxypropyltrimethoxysilane and then coated on a microgrooved polydimethylsiloxane (PDMS) mold to produce the microgrooved silica-chitosan hybrid membrane. After incubated with osteoblast-like MC3T3-E1 cells, the microgrooved hybrid membranes strongly directed the alignment and elongation of cells and exhibited a strong cell contact guidance ability, indicating a potential application as engineering bone tissue regenerative materials.

Abstract: Luminescent europium(III)-doped nanoporous silica spheres (Eu:NPS) were synthesized, and folic acid derivative (FA: folate N-hydroxysuccinimidyl ester) as targeting ligand for cancer cells was immobilized on the sphere through a mediation of 3-aminopropyltriethoxysilane (APTES). The ordered nanopores were preserved with the immobilization. The surface area decreased only with the APTES immobilization, suggesting that the FA was immobilized only on the outer surface of the nanopores. The photoluminescence of the spheres showed the characteristic peak due to interactions between the FA and Eu³⁺, and further the luminescence can be clearly detected by a fluorescent microscopy. The spheres were highly dispersed stability in cell culture medium to exhibit the nontoxic in the proliferation stage of Hela cancer cells and NIH3T3 fibroblasts, and specifically bindto the Hela cells. The binding and uptake spheres showed the intense luminescence. Thus, the luminescent FA-immobilized Eu:NPS spheres will exhibit the targeting and imaging abilities for cancer cells.

Abstract: Insulin delivery carriers using low crystalline porous zinc hydroxyapatite (ZnHAp) microparticles with different crystal sizes and novel formulation method of poly (lactic acid) (PLA) have been developed to achieve the sustained-release of insulin. The adsorption isotherm curves of insulin dissolved into acetic acid of pH4.5 onto ZnHAp nanocrystals show non-Langmuir type due to the dissolution of the ZnHAp microparticles; the adsorption amounts were increased with the increase of crystalline sizes. The PLA formulation, coating the outer layer of microparticles, showed the apparent reduction of initial burst for insulin. The amounts of initial release of insulin decreased with the increase of crystalline sizes of ZnHAp, which could be attributed to the different meso-porous structure depending on its crystal sizes.

Abstract: Hydroxyapatite (HAp) sensor, available for quartz crystal microbalance with dissipation (QCM-D) technique, has been fabricated by an electrophoretic deposition method. The method of re-usability of the sensor after adsorption of fibrinogen and the biological apatite (BAp) growth on the sensor with and without the adsorption of feral bovine serum (FBS) from 1.5 simulated body fluid were investigated. The re-usability of the sensor, cleaning with the combination of ammonia and hydrogen peroxide mixture and UV/ozone treatment, achieved ten times reuses. BAp was grown on the HAp surface but not on the gold surface at 37.5 oC for 40 hours. The viscoelastic property (DD/Df value) of the BAp layer on the HAp sensor showed harder than that of the protein adsorption films from FBS. The amount of the BAp grown on the HAp sensor adsorbed FBS is lower than that on the HAp sensor. The adsorption of FBS proteins on the HAp surface strongly inhibited the BAp growth.

Abstract: The adsorption of multiple proteins derived from fetal bovine serum (FBS) in phosphate buffer saline (PBS) and alpha minimum essential (aMEM) was in situ analyzed with a quartz crystal microbalance with dissipation technique on gold, titanium and HAp sensors. The adsorption behaviors of FBS proteins were varied depending on the sensors. The DD/Df value of the HAp sensor were clearly different in PBS and aMEM, and others were not changed. The viscoelastic properties of the protein films adsorbed on the HAp sensor in PBS were flexible in comparison with those on the gold and titanium sensors. The D-f plots incidated that the proteins adsorbed on HAp in PBS would lead to competitive adsorption and conformational change and those in aMEM could form a monolayer. The adsorption behavior on the HAp in carbonate buffer saline was found to be similar to that in aMEM. These differential adsorption behaviors on the HAp surface were attributed to the pre-adsorptive ion, such PO43- or CO32- in the solvent.

Abstract: Surfaces of sintered hydroxyapatite (HAp) were treated with four kinds of acids, hydrochloric acid (HCl), nitric acid (HNO3), sulfuric acid (H2SO4) and hydrogen peroxide (H2O2). On these acid-treated HAp surfaces, osteoblast-like MC3T3-E1 cells were cultured for 20 days. The matrix mineralization was observed in HCl-treated HAp and HNO3-treated HAp, but not in non-treated HAp, H2SO4-treated HAp and H2O2-treated HAp. Gene expressions of Bglap2 encoding osteocalcin and Akp2 encoding alkaline phosphatase were higher in HCl-treated HAp and HNO3-treated HAp than those in the others. These results indicate that not only hydrochloric acid but also nitric acid improves performance of HAp to mature osteoblast.

Abstract: Rigid hydroxyapatite (HAp)-alginate beads were prepared as drug delivery carriers for an anti-cancer drug, paclitaxel (Taxol). Paclitaxel was loaded into the HAp microparticle in process of a spray-drying technique. The HAp-alginate beads including paclitaxel were obtained by a droplet method into barium solution as ionic cross-linkage and dehydration. Cross-sectional analyses indicated the homogeneity of HAp microparticles and barium ions inside the bead. The ratio of alginate to HAp in the beads dominated both mechanical and swelling properties. Drug-release experiment demonstrated the sustained release of paclitaxel from the beads cross-linked with barium ion for 7 days.

Abstract: Real time adsorption behaviors of six proteins with different isoelectric points on hydroxyapatite (HAp) nanocrystal surfaces have been investigated by using HAp sensors for quartz crystal microbalance with dissipation technique (QCM-D). The dissipation (D)–frequency (f) plots clearly showed that the different types of protein adsorption behaviors; the D-f plots of acidic proteins lie on one straight line with a constant slope under all initial protein concentrations, while those of neutral and basic proteins lie on two straight lines with different slopes. The acidic proteins formed a monolayer, while the neutral and basic proteins could cause conformational changes with the adsorbed amount of proteins. The QCM-D technique with novel HAp nanocrystal sensor is useful for the liquid phase changes of proteins on the surface.

Abstract: The adsorption properties of metal ions containing hydroxyapatite (Ca10(PO4)6(OH)2, HAp) were elucidated for the development of protein drug carrier. As-prepared metal ion containing HAp nanocrystal showed plate-like morphology with 10-20nm in length and 5-10nm in width. The metal ion containing HAp microparticles had higher specific surface area than the HAp microparticle. The adsorption amount per unit area of HAp showed higher than those of metal ion containing HAp. The adsorption behavior followed the Langmuir curves for each protein, indicating the monolayer adsorption. The loaded amount of proteins could be one of the most important properties for the application of drug delivery carrier.

Abstract: Gene expression profile of osteoblast-like cells cultured on dense disk materials and porous materials of calcium phosphate ceramics was constructed from DNA microarray analyses. The profile revealed that gene expression patterns of porous materials were significantly different from those of dense disk materials. The porous materials had a capacity to induce expressions of genes involved in osteoblast differentiation, while dense disk materials regulated gene expressions related to osteoclastogenesis.