Fabrication of Gentamicin-Loaded Hydroxyapatite/Collagen Bone-Like Nanocomposite for Anti-Infection Bone Void Fillers.

A gentamicin-loaded hydroxyapatite/collagen bone-like nanocomposite (GNT-HAp/Col) was fabricated and evaluated for its absorption-desorption properties, antibacterial efficacy, and cytotoxicity. The hydroxyapatite/collagen bone-like nanocomposite (HAp/Col) powder was mixed with gentamicin sulfate (GNT) in phosphate-buffered saline (PBS) at room temperature. After 6 h mixing, the GNT adsorption in all conditions reached plateau by Langmuir's isotherm, and maximum GNT adsorption amount was 34 ± 7 µg in 250 µg/mL GNT solution. Saturated GNT-loaded HAp/Col powder of 100 mg was soaked in 10 mL of PBS at 37 °C and released all GNT in 3 days. A shaking culture method for a GNT extraction from the GNT-HAp/Col and an inhibition zone assay for the GNT-HAp/Col compact showed antibacterial efficacy to Escherichia coli (E. coli) at least for 2 days. From the release profile of the GNT from the GNT-HAp/Col powder, antibacterial efficacy would affect E. coli at least for 3 days. Further, no cytotoxicities were observed on MG-63 cells. Thus, the GNT-HAp/Col is a good candidate of bioresorbable anti-infection bone void fillers by prevention initial infections, which is the primary cause of implant-associated infection even for rapid bioresorbable materials.

Formation of osteon-like structures in unidirectional porous hydroxyapatite substitute.

Unidirectional porous hydroxyapatite (UDPHAp) bone substitute comprises a microstructure of cross-sectionally oval pores with diameters ranging from 30 to 300 µm. Bone remodeling within the UDPHAp is expected upon implantation into bone; however, the mechanism and factors influencing this bone growth remain unclear. The objectives of the present study were to assess the vasculature and microstructure of newly formed bone and to determine how bone formation is affected by load transfer and UDPHAp pore size. Formation of osteon-like structures, defined by the presence of lacunae, canaliculi and a central lumen containing capillaries, was observed within the implanted UDPHAp material in all animals after six weeks. The number of osteocytes and osteon-like structures in areas adjacent to the cortex of recipient bone was significantly higher than in areas next to the medullary cavity throughout the recovery period. Notably, osteon-like structures tended to form in smaller diameter pores. Continuous bone remodeling might be promoted by the rapid formation of unidirectional capillaries and the osteocyte lacunae-canalicular system. Load transfer and smaller pore size could positively affect cortical bone regeneration. © 2018 The Authors Journal of Biomedical Materials Research Part B: Applied Biomaterials Published by Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 2665-2672, 2018.

Bone Regeneration and Remodeling within a Unidirectional Porous Hydroxyapatite Bone Substitute at a Cortical Bone Defect Site: Histological Analysis at One and Two Years after Implantation.

Unidirectional porous hydroxyapatite (UDPHAp) is an artificial bone substitute with a unique microstructure consisting of 100-300-µm oval pores that present the material unidirectionally. UDPHAp has a compression strength of 14 MPa and a porosity of 75%, which promotes cell migration and capillary formation within the material. Despite these advantageous properties, bone remodeling and bone formation with UDPHAp remain unclear. To examine long-term remodeling and differences in bone formation based on the defect site, trapezoidal prism-shaped UDPHAp blocks were implanted into rectangular-shaped cortical bone defects in the proximal tibia of Japanese white rabbits. Histological analysis performed at 52 and 104 weeks after implantation revealed that bone and capillaries had formed within the implanted UDPHAp material. Bone formed within the UDPHAp implanted in the cortical defect of rabbit tibia and remodel up to two years. The percentage of new bone area within UDPHAp was larger in cortical lesions than that in medullary lesions. These findings suggest that UDPHAp is a promising material for the repair of non-critical-sized cortical bone defects.

Novel long-term immobilization method for radioactive iodine-129 using a zeolite/apatite composite sintered body.

The amount of radioactive iodine generated from nuclear power plants is expected to increase with the proliferation of nuclear energy production, and long-term immobilization methods for such radioactive elements need to be developed to make nuclear energy sustainable. The standard immobilization method of radioactive elements, vitrification, is not very effective for radioactive iodine-129 because of the low solubility of iodine in silicate melts, its very high volatility at standard vitrification process temperatures, and its instability in the alkaline environment of deep geological layers below 300 m. We have developed a novel three-phase ceramic composite produced by a sintering process. Iodine adsorbed onto Ca-type zeolite A was covered with a hydroxyapatite nanolayer through the exchange reaction of ammonium with calcium. Clusters of iodine of 30 nm within the zeolite structure were found to be thermally stable up to 1253 K because of the partial blockage of the alpha-cage apertures by ammonium ions and the partial change from a crystalline phase to an amorphous phase at 473 K. No gasification of iodine molecules was found to occur during the sintering process. The outer phase was highly crystalline hydroxyfluorapatite in which the hydroxyapatite nanolayer plays an important role for successful sintering. The elution of iodine in low-dioxygen water, similar to that found within the Earth's crust, was investigated and was found to occur only in the surface layer of the sintered body.

DNA microarray analysis of human gingival fibroblasts from healthy and inflammatory gingival tissues.

In the inflammatory gingival tissues of patients with periodontitis, cytokines such as interleukin (IL)-1 alpha, IL-1 beta, IL-6, IL-8, and tumor necrosis factor (TNF)-alpha have been detected. Gingival fibroblasts are the major constituents of gingival tissue. We recently demonstrated that lipopolysaccharide (LPS) from periodontopathic bacteria induces inflammatory reactions in various tissues via CD14 and/or Toll-like receptors (TLRs) in gingival tissues [Biochem. Biophys. Res. Commun. 273 (2000) 1161]. To confirm this, we examined the expression of IL-1 alpha, IL-1 beta, IL-6, IL-8, TNF-alpha, CD14, TLR2, and TLR4 in human gingival fibroblasts (HGFs) obtained from patients with healthy or inflammatory gingiva using DNA microarray analysis. We also studied the expression levels of these proteins by flow cytometric analysis (FACS). The expression levels of all eight genes in the HGFs of the Inflammatory group were significantly higher than those in the Healthy group on DNA microarray analysis. FACS revealed that the expression levels of all eight proteins on the HGFs of the Inflammatory group were higher than those on the Healthy group. Our data indicated that these eight proteins in HGFs are involved in inflammatory conditions in the gingiva, including periodontal disease. Our results suggested that these eight proteins, in turn, act directly or indirectly on the immune response by activating host cells involved in inflammatory processes.

Effect of anti-CD14 antibody on experimental periodontitis induced by Porphyromonas gingivalis lipopolysaccharide.

The lipopolysaccharide (LPS) released by Porphyromonas gingivalis, a Gram-negative bacterium found in the periodontal pockets of patients with periodontitis, induces bone resorbing activity in vivo. We previously showed that a receptor for LPS on human gingival fibroblasts and gingival epithelial cells is CD14. In this study, we established a mouse model of experimental periodontitis by applying a P. gingivalis LPS solution to the buccal region of mice. P. gingivalis LPS-induced bone resorption and interleukin-6 production in the gingival tissues were significantly inhibited by pretreatment with anti-CD14 antibody for 5 weeks prior to LPS treatment. This result suggests that anti-CD14 antibody may be usable as a prototype for the development of drugs for the treatment of periodontal disease.

Crystal growth and structure analysis of twin-free monoclinic hydroxyapatite.

Single crystals of hydroxyapatite were grown by a flux method using Ca(3)(PO(4))(2) and Ca(OH)(2) under 100 MPa of Ar gas. The crystals obtained had stoichiometric composition of Ca(10)(PO(4))(6)(OH)(2) and some of them were twin-free single crystals. From X-ray diffraction analyses, the space group was confirmed to be monoclinic P2(1)/b with cell parameters a=0.9419(3) nm, b=1.8848(6) nm, c=0.6884(2) nm, and gamma=119.98(2) degrees. The detailed crystal structure was determined with a reliability factor R(w)=0.033; the O atoms of OH were located just on the 2(1) axis while the H atoms of OH occupied the positions a little deviated from the 2(1) axis. The origin of this structure was ascribed to the formation of hydrogen bonds between the H atoms of the OH ions and the specific O atoms of the PO(4) ions.