Biomedical subjects

Junzo Tanaka

Publications and source records attributed to Junzo Tanaka.

At least 19 recordsLinked to original sources

PubMed2007

Histological changes and apoptosis of cartilage layer in human anterior cruciate ligament tibial insertion after rupture.

The purpose of this study is to investigate the histological changes and apoptosis of cartilaginous layers in human anterior cruciate ligament (ACL) tibial insertion at different time periods after rupture. By using a core reamer, 35 tibial insertions of ruptured ACLs were obtained during primary ACL reconstructions (number of days after injury: 19-206 days). A histological examination was performed and a terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end labeling (TUNEL) staining assay was carried out to detect apoptosis. The average thickness of the cartilage layer, the glycosaminoglycan-stained area and the number of chondrocytes per millimeter decreased with time. The percentage average of TUNEL-positive chondrocytes was 42.0 +/- 16.2. The histological degenerative changes of the cartilage layer in the ruptured ACL tibial insertion progressed with time, especially in the first 2 months. Moreover, chondrocyte apoptosis continued from 19 to 206 days after rupture. The results may help elucidate the etiology of the histological changes of the insertion, and may help in devising optimal treatment protocols for ACL injuries if apoptosis is controlled. Moreover, we consider that using a surviving ligament and minimizing a debridement of ACL remnant during ACL reconstruction may be important for ACL reconstruction to maintain cartilage layers in ACL insertions.

Adolescent↗

PubMed2007

Fabrication and mechanical and tissue ingrowth properties of unidirectionally porous hydroxyapatite/collagen composite.

This study investigated the effects of the three-dimensional (3-D) pore structure of a porous hydroxyapatite/collagen (HAp/Col) composite on their mechanical properties and in vivo tissue ingrowth. The unique 3-D pore structure, comprising unidirectionally interconnected pores, was fabricated by the unidirectional growth of ice crystals by using a cooling stage and a subsequent freeze-drying process. The unidirectional pores had a spindle-shaped cross section, and their size gradually increased from the bottom to the upper face. The porous composite showed an elastic property and anisotropic compressive strength for the pore directions. While the strength and modulus parallel to the pore axis were 1.3- and twofold higher than those of the porous composite with spherical pores formed randomly, the strength and modulus perpendicular to the pore axis showed the lowest values. The subcutaneous implantations revealed that when compared with the random pores, the unidirectional pores promote the ingrowth of the surrounding tissues into the pores.

Absorbable Implants↗

PubMed2007

Amniotic membrane immobilized poly(vinyl alcohol) hybrid polymer as an artificial cornea scaffold that supports a stratified and differentiated corneal epithelium.

Poly(vinyl alcohol) (PVA) is a biocompatible, transparent hydrogel with physical strength that makes it promising as a material for an artificial cornea. In our previous study, type I collagen was immobilized onto PVA (PVA-COL) as a possible artificial cornea scaffold that can sustain a functional corneal epithelium. The cellular adhesiveness of PVA in vitro was improved by collagen immobilization; however, stable epithelialization was not achieved in vivo. To improve epithelialization in vivo, we created an amniotic membrane (AM)-immobilized polyvinyl alcohol hydrogel (PVA-AM) for use as an artificial cornea material. AM was attached to PVA-COL using a tissue adhesive consisting of collagen and citric acid derivative (CAD) as a crosslinker. Rabbit corneal epithelial cells were air-lift cultured with 3T3 feeder fibroblasts to form a stratified epithelial layer on PVA-AM. The rabbit corneal epithelial cells formed 3-5 layers of keratin-3-positive epithelium on PVA-AM. Occludin-positive cells were observed lining the superficial epithelium, the gap-junctional protein connexin43-positive cells was localized to the cell membrane of the basal epithelium, while both collagen IV were observed in the basement membrane. Epithelialization over implanted PVA-AM was complete within 2 weeks, with little inflammation or opacification of the hydrogel. Corneal epithelialization on PVA-AM in rabbit corneas improved over PVA-COL, suggesting the possibility of using PVA-AM as a biocompatible hybrid material for keratoprosthesis.

3T3 Cells↗

PubMed2006

Cartilaginous tissue formation from bone marrow cells using rotating wall vessel (RWV) bioreactor.

This is the first successful report of the rapid regeneration of three-dimensional large and homogeneous cartilaginous tissue from rabbit bone marrow cells without a scaffold using a rotating wall vessel (RWV) bioreactor, which simulates a microgravity environment for cells. Bone marrow cells cultured for 3 weeks in DMEM were resuspended and cultured for 4 weeks in the chondrogenic medium within the vessel. Large cylindrical cartilaginous tissue with dimensions of (1.25 +/- 0.06) x (0.60 +/- 0.08) cm (height x diameter) formed. Their cartilage marker expression was confirmed by mRNA expressions of aggrecan, collagen type I and II, and glycosaminoglycan (GAG)/DNA ratio. Their cartilaginous properties were demonstrated by toluidine blue, safranin-O staining, and polarization.

Animals↗

PubMed2006

pH-responsive swelling behavior of collagen gels prepared by novel crosslinkers based on naturally derived di- or tricarboxylic acids.

The aim of this study was to compare the physicochemical properties of alkali-treated collagen (AlCol) gels prepared using two kinds of naturally derived crosslinkers made from citric and malic acids (CAD and MAD, respectively) that we have developed. From the crosslinking reaction between active ester groups and amino groups of AlCol, we successfully obtained AlCol gels, named AlCol-CAD and AlCol-MAD, prepared using CAD and MAD, respectively. The gelation time of the AlCol solution containing CAD initially decreased with increasing CAD concentration up to 70 mM, and then increased as the CAD concentration increased further. The gelation time reached its minimum and began to increase. On the other hand, for AlCol-MAD solution, gelation occurred within 40s at any MAD concentration. Moreover, the residual amino groups in AlCol-CAD and AlCol-MAD were found to decrease with increasing CAD or MAD concentrations, whereas increased residual carboxyl groups were detected only in the case of AlCol-CAD. The swelling ratio of AlCol-CAD significantly increased at CAD concentrations above 50mM. On the other hand, AlCol-MAD showed little increase in swelling ratio with increasing MAD concentration. Also, AlCol-CAD was swollen when the gels were immersed in a solution with high pH. On the other hand, no significant increase in swelling ratio was observed when AlCol-MAD was immersed in a similar solution. These results suggest that the different amounts of carboxyl groups in AlCol-CAD affected the swelling behavior of gels and that this pH-responsive AlCol-CAD has potential for drug delivery systems and tissue engineering.

Collagen↗

PubMed2006

Fabrication of hydroxyapatite ultra-thin layer on gold surface and its application for quartz crystal microbalance technique.

We present a method for coating gold quartz crystal microbalance with dissipation (QCM-D) sensor with ultra-thin layer of hydroxyapatite nanocrystals evenly covering and tightly bound to the surface. The hydroxyapatite layer shows a plate-like morphology and less than 20 nm in thickness. The hydroxyapatite sensor operated in liquid with high stability and sensitivity. The in-situ adsorption mechanism and conformational change of fibrinogen on gold, titanium and hydroxyapatite surfaces were investigated by QCM-D technique and Fourier-transform infrared spectroscopy. The change of secondary structures of fibrinogen adsorbed on the surfaces depended on the adsorbed amounts of protein. The secondary structure of fibrinogen adsorbed on the surfaces changes with increasing coverage. This is explained by repulsion among fibrinogens, affecting water structure and thus the strength of fibrinogen interactions on the surface. The study indicates that the hydroxyapatite sensor is applicable for qualitative and conformational analysis of protein adsorption.

Adsorption↗

PubMed2006

Pre-adsorbed type-I collagen structure-dependent changes in osteoblastic phenotype.

Type-I collagen is the most abundant extracellular matrix in bones and modulates various functions of osteoblasts. We prepared two different structures of type-I collagen on tissue culture grade polystylene (TCPS) surfaces, one is feltwork structure of filamentous molecules from acid solutions (ACs) and the other is network structure of fibrils from neutral solutions (NCs), to examine effects of the structures on the maturation process of osteoblast-like cells. No significant differences of cell proliferation were observed between TCPS and ACs, but NCs delayed the proliferation. In initial cell attachment, the cells on ACs had tense lamellipodia with sharp tips, while those on NCs had loose lamellipodia. No detectable differences in levels of expressed integrin alpha2- and alpha5-subunits were observed between the structures. Although the matrix mineralization in NCs was also delayed in comparison with TCPS and ACs, fully mineralized levels in NCs were the same as those of TCPS and ACs. In addition, although we examined the effects of densities of pre-adsorbed collagen molecules on osteoblast maturation, the effects were less serious than those of the structures. This study suggests that the structures of collagen affect proliferation and mineralization of osteoblast-like cells.

Animals↗

PubMed2006

Collagen-immobilized poly(vinyl alcohol) as an artificial cornea scaffold that supports a stratified corneal epithelium.

The cornea is a transparent tissue of the eye, which is responsible for the refraction of incoming light. Both biological corneal equivalents and synthetic keratoprostheses have been developed to replace donor tissue as a means to restore vision. However, both designs have drawbacks in terms of stability and biocompatibility. Clinically available synthetic devices do not support an intact epithelium, which poses a risk of microbial infection or protrusion of the prosthesis. In the present study, type I collagen was immobilized onto poly(vinyl alcohol) (PVA-COL) as a possible artificial cornea scaffold that can sustain a functional corneal epithelium. Human and rabbit corneal epithelial cells were air-lift cultured with 3T3 feeder fibroblasts to form a stratified epithelial layer on PVA-COL. The epithelial sheet expressed keratin 3/12 differentiation markers, the tight junction protein occludin, and had characteristic microvilli structures on transmission electron microscopy. Functionally, the stratified epithelium contained normal glycogen levels, and an apical tight-junction network was observed to exclude the diffusion of horseradish peroxidase. Furthermore, the epithelium-PVA-COL composite was suturable in the rabbit cornea, suggesting the possibility of using PVA-COL as a biocompatible material for keratoprosthesis.

Animals↗

PubMed2006

Organic-inorganic interaction and the growth mechanism of hydroxyapatite crystals in gelatin matrices between 37 and 80 degrees C.

The crystal development of hydroxyapatite[HAp] phase in gelatin[GEL] matrices was investigated in the temperature range 37 to 80 degrees C by using X-ray diffraction, scanning electron microscopy(SEM), thermoanalytical measurement(DT/TGA), Fourier-Transformed Infra-Red(FT-IR) spectroscopy, and transmission electron microscopy(TEM) with electron diffraction(ED). It was found that during the coprecipitation of apatite phase in GEL matrices and the next aging process the crystallites were formed and developed through the two reaction mechanisms of organic-inorganic interaction between apatite phase and GEL molecules, and thermodynamic reaction for the crystal growing. The analytical evidences showed that there was a definite competition between these two mechanisms with the reaction temperature. Below 50 degrees C the crystal development of HAp was greatly suppressed by the existence of the GEL molecules, indicating the heterogeneous nucleation by the supposed number of carboxyl groups in GEL. Above 50 degrees C the effective organic components as a template for the heterogeneous nucleation of apatite crystallites were greatly degraded and so more amount of inorganic ions could be favorably accredited on the preexisting crystallites in virtue of the limited nucleation chance, finally resulting in the crystal growth. At higher temperature pretty big HAp crystals were developed with the depletion of the organics to be bound with crystallites in the slurry solution. Presumably it is believed that the poisoning of the functional groups in GEL molecules was vigorously occurred in the phosphoric acid environment above approximately 50 degrees C.

Biocompatible Materials↗

PubMed2006

Synergistic effect of silanol group and calcium ion in chitosan membrane on apatite forming ability in simulated body fluid.

A chitosan membrane modified with silanol groups and calcium ions on its surface and in its structure, respectively, was newly developed and evaluated for the potential application as a bioactive-guided bone-regeneration membrane. The chitosan membrane, which contained calcium nitrate tetrahydrate, was prepared and further subjected to surface modification with 3-isocyanatopropyl triethoxysilane (IPTS) following hydrolysis with HCl solution. As control, chitosan membranes which contained only calcium nitrate tetrahydrate and modified with only silanol groups were prepared, respectively. Three membranes were exposed to simulated body fluid (SBF) for a period ranging from 3 h to 7 days. The SBF exposure led to the deposition of a layer of apatite crystals on the surface of the chitosan membrane modified with silanol groups and calcium ions, while those modified with only calcium ions or silanol groups did not show the apatite-forming ability. It implies that the silanol groups and calcium ion acted together in a synergistic fashion in the formation of apatite crystals; the silanol groups and calcium ions acted as the nucleation sites and accelerator for the formation of apatite crystals, respectively. Therefore, this new chitosan membrane is likely to have a potential for the application as a bioactive guided bone regeneration membrane because of its apatite-forming ability in the SBF.

Apatites↗

PubMed2006

Synthesis of polypyridine-graft-PEG copolymer for protein repellent and stable interface.

Polypyridine grafted with poly(ethylene glycol) (Py-g-PEG) have been synthesized. Radical copolymerization of methyl-terminated PEG macromonomer with 4-pyridylmethyl methacrylate homogeneously proceeded and the obtained copolymer spontaneously adsorbs from aqueous solution onto gold surfaces, where the pyridine parts act as the multipoint anchor to the surface and the PEG parts provide the strong steric repulsion between the chains. As a result, the highly protein repellent and stable surface was constructed through multipoint pyridine attachment as compared with singlepoint pyridine attachment. Py-g-PEGs thus synthesized are promising material to functionalize metal and semiconductor material and to self-assemble into micelle in biotechnological and biomedical field.

Adsorption↗

PubMed2006

Guided bone regeneration with beta-tricalcium phosphate and poly L-lactide-co-glycolide-co-epsilon-caprolactone membrane in partial defects of canine humerus.

This study was performed to evaluate the effect of betatricalcium phosphate and poly L-lactide-co-glycolide-coepsilon- caprolactone (TCP/PLGC) membrane in the repair of partial bone defects in canine proximal humerus. Three adult mixed-breed dogs were used during the experimental period. The length of the defect was quarter of the full length of humerus, and width of the defect was quarter of middle diameter of the lateral aspect of humerus. The humeri of each dog were divided into treatment (TCP/ PLGC) and control groups. The defect was covered with TCP/PLGC membrane in treatment group. To evaluate regeneration of the bone, computerized tomography (CT) and histopathologic examination were performed. The radiopaque lines were appeared at the original defect sites in TCP/PLGC group but below the original site in control at 4th week. Radiopacity and thickness of the defect sites, and radiopaque lines were more increased at 8th week than those of 4th week. Histopathologic findings revealed fibrous connective tissue migration into the defect and the migration inhibited the structure of new cortex to be placed in the original level in control whereas new cortex growth was found in the level of original line in TCP/ PLGC group. However, the new cortical bone in the TCP/ PLGC group was thinner and less organized than the adjacent intact cortex, and the amount of new cancellous bones were also scanty. The result suggested that TCP/ PLGC membrane is a good guided bone regeneration material to restore the original morphology of humerus in partial defect.

Absorbable Implants↗

PubMed2005

Reuptake of extracellular amelogenin by dental epithelial cells results in increased levels of amelogenin mRNA through enhanced mRNA stabilization.

Amelogenin is an extracellular matrix protein secreted by ameloblasts and is a major component of enamel matrix. Recently, in addition to their role in enamel formation, the biological activity of enamel proteins in the process of cell differentiation has recently become widely appreciated. In this study, we examined the biological activity of amelogenin on ameloblast differentiation. Recombinant mouse amelogenin (rm-amelogenin) enhanced the expression of endogenous amelogenin mRNA in a cultured dental epithelial cell line (HAT-7), despite a lack of increased amelogenin promoter activity. To solve this discrepancy, we analyzed the effects of rm-amelogenin on the stability of amelogenin mRNA. The half-life of amelogenin mRNA is extremely short, but in the presence of rm-amelogenin its half-life was extended three times longer than the control. Furthermore, we showed the entry of exogenous fluorescein isothiocyanate-conjugated rm-amelogenin into the cytoplasm of HAT-7 cells. It follows from our results that exogenous amelogenin increases amelogenin mRNA levels through stabilization of mRNA in the cytoplasm of HAT-7 cells. Here we speculated that during differentiation, dental epithelial cells utilize a unique mechanism for increasing the production of amelogenin, the reuptake of secreted amelogenin.

3' Untranslated Regions↗

PubMed2005

Long-term effects of muscle-derived protein with molecular mass of 77 kDa (MDP77) on nerve regeneration.

The long-term effects of the 77-kDa muscle-derived protein (MDP77) on motor and sensory nerve regeneration were examined in vivo. Fourteen-millimeter bridge grafts of the right sciatic nerve of SD rats were carried out with silicone tubes containing a solution of type I collagen together with 0, 5, 10, or 20 microg/ml recombinant human MDP77 (N = 10 in each group). Recovery of motor and sensory function was evaluated monthly by the maximal toe-spread index (TSI) and hot-plate test, respectively, for 6 months after the operation. Electrophysiology (nerve conduction velocity), histology (diameter and total number of the regenerated myelinated axons in the tube), and immunohistochemistry (total number of Schwann cells in the tube), as well as measurement of soleus muscle weight, were also performed at this time. Motor, but not sensory, function recovered rapidly in the MDP77-treated groups in a dose-dependent manner. Electrophysiological measurements and the ratio of soleus muscle weight corroborated the positive effects of MDP77 on motor nerve regeneration, but no facilitation of sensory nerve recovery was observed. Furthermore, histological and immunohistochemical evaluations suggested that MDP77 treatment accelerates Schwann cell migration, followed by enhanced maturation of regenerating axons, resulting in functional recovery of both the nerves and the atrophied, denervated muscle.

Animals↗

PubMed2005

Synthesis of high surface area hydroxyapatite nanoparticles by mixed surfactant-mediated approach.

A new surfactant-mediated approach was developed to synthesize hydroxyapatite (HAp) nanoparticles with high surface areas by calcination of their precursors encapsulated with calcium stearate using mixed surfactant-containing reaction mixtures. Acidic aqueous solution of calcium phosphate was mixed with both or either nonaoxyethylene dodecyl ether (C12EO9) and polyoxyethylene(20) sorbitan monostearate (Tween 60) and then was treated with aqueous ammonium at 25 degrees C. The C12EO9-based single surfactant system yielded an aggregate of platy HAp nanoparticles 20-40 nm in size, whereas the Tween 60-based single and mixed systems led to lath-shaped HAp nanoparticles 2-8 nm wide and encapsulated with calcium stearate. On calcination at 500 degrees C, the stearate-encapsulated HAp nanoparticles in the latter two systems were deorganized into high surface area HAp nanoparticles. Particularly, the HAp nanoparticles in the mixed system exhibited a specific surface area as high as 364 m2 g(-1) that is roughly 3 times larger than 160 m2 g(-1) for those in the single system. The significantly high surface area for the former is attributed to much less adhesion of decapsulated HAp nanoparticles, which originated from the particle-separating effect of the C12EO9 molecules adsorbed on the outer surface of the stearate-encapsulated HAp nanoparticles to inhibit their agglomeration or interfacial coordination. The present results demonstrate that the mixed use of two different surfactants as a source of encapsulating and templating agent and a particle-separating agent is specifically effective for the synthesis of high surface area HAp nanoparticles.

Journal Article↗

PubMed2005

Effects of a laminin peptide (YIGSR) immobilized on crab-tendon chitosan tubes on nerve regeneration.

Thiolated and nonthiolated hydroxyapatite-coated crab-tendon chitosan (t-chitosan/HAp-SH and t-chitosan/HAp, respectively) tubes, both alone and conjugated with CDPGYIGSR (YIGSR) peptide, were compared, in order to determine their biocompatibility and efficacy as nerve conduits. YIGSR peptide was adsorbed on the t-chitosan/HAp (HAp) tubes, and covalently bound on the t-chitosan/HAp-SH (HAp-SH) tubes (Y/HAp and Y/HAp-SH tubes, respectively). HAp, HAp-SH, Y/HAp, or Y/HAp-SH tubes measuring 15 mm were bridge grafted into the sciatic nerve of SD rats. Grafting of 15-mm-long Type I atelocollagen tubes and isografting of sciatic nerves were also carried out (N = 6 in each group). After 12 weeks, evoked muscle action potentials were recorded to calculate the terminal latency quotient. Histological observation and analysis of myelinated axons were also carried out. Nerve-tissue regeneration did not occur directly on the tubes' surfaces in the YIGSR peptide-unconjugated groups. Transplantation of YIGSR-conjugated tubes, however, gave rise to regenerated nerve tissue attached to thin layers of epineurium-like structure formed on the inner-tube surface. Histological and electrophysiological analyses suggested that although thiolation retards nerve-tissue regeneration, adsorbed YIGSR, and, to a lesser extent, peptide that had been covalently bound onto the tube surfaces, enhance nerve regeneration, promoting sprouting from the proximal nerve stump and bridging of regenerated axons throughout the tube.

Animals↗

PubMed2005

Nano-scaled hydroxyapatite/polymer composite III. Coating of sintered hydroxyapatite particles on poly(4-methacryloyloxyethyl trimellitate anhydride)-grafted silk fibroin fibers.

A novel composite coupling between nano-scaled hydroxyapatite (HAp) particles and poly[4-methacryloyloxyethyl trimellitate anhydride (4-META)]-grafted silk fibroin (SF) through ionic interaction was synthesized. The weight gain of poly(4-META) by graft-polymerization increased with increasing the reaction time, eventually reaching a plateau value of about 20 wt%. The HAp nano-particles were adsorbed equally and dispersively on the treated SF fiber surface. The HAp content in the composite was 4.554 wt% +/- 0.098 (n = 4), confirmed by thermogravimetry (TG). This synthetic system requires no heat to connect HAp to SF and is useful when applying to non-heat-resistant polymers. The L-929 cell-adhesion test shows that the HAp/SF composite improves bioactivity compared to the original SF.

Coated Materials, Biocompatible↗

PubMed2005

The interaction between osteoclast-like cells and osteoblasts mediated by nanophase calcium phosphate-hybridized tendons.

We developed a novel technique of hybridizing calcium phosphate (CaP) with bioorganic soft tissue using an alternating soaking process. By this technique, we hybridized CaP with a grafted tendon tissue to bond with a bone tunnel. Tendons were soaked in Ca and NaHPO(4) solutions alternately for 10 min. Needle-like CaP crystals 30-50 nm in length including low-crystalline apatite were deposited on and between collagen fibrils from the surface to 200 microm deep in the tendon. In light and transmission electron microscopic images, osteoclast-like cells and osteoblasts appeared on the implanted tendon and osteoid was observed on the tendon surface at 1 week postoperatively. At 2 weeks postoperatively, osteoclast-like cells resolved the tendon by forming Howship's lacuna-like spaces on the surfaces and osteoblasts formed osteoid in these spaces. Direct bonding between the implanted tendon and the newly formed bone was observed. At 3 weeks postoperatively, thick newly formed bone firmly bonded to tendon surface. From these results, we conclude that the tendons prepared by an accelerated CaP hybridization method efficiently enhance osteoclast-like cells and osteoblasts to bond the implanted tendons to newly formed bone.

Animals↗