Search PubMed⌕ Search

Biomedical subjects

Junzo Tanaka

Publications and source records attributed to Junzo Tanaka.

At least 37 records · Page 2Linked to original sources
PubMed2005

Encapsulation of chondrocytes in injectable alkali-treated collagen gels prepared using poly(ethylene glycol)-based 4-armed star polymer.

An in situ gel system was developed to encapsulate chondrocytes under physiological conditions using an alkali-treated collagen (AlCol) and pentaerythritol poly(ethylene glycol) ether tetrasuccinimidyl glutarate (4S-PEG) as a crosslinker. AlCol gels were obtained at crosslinker concentrations from 0.1 to 3.0 mM. Chondrocytes were encapsulated and dispersed homogeneously in AlCol gels. Results of MTT staining showed that cells survived after encapsulation in AlCol gels. Biochemical analysis demonstrated that DNA content in AlCol gels was constant after 3 weeks. Glycosaminoglycan content and mRNA expression of type II collagen and aggrecan increased with culture time. These results suggest that this in situ gel system is useful for regenerating cartilage in vitro and for minimally invasive therapy for cartilage defects.

Alkalies↗
PubMed2005

Immobilization of laminin peptide in molecularly aligned chitosan by covalent bonding.

We developed a new biomaterial effective for nerve regeneration consisting of molecularly aligned chitosan with laminin peptides bonded covalently. Molecularly aligned chitosan was prepared from crab (Macrocheira kaempferi) tendons by ethanol treatment and 4 wt%-NaOH aqueous solutions to remove proteins and calcium phosphate, followed by deacetyl treatment using a 50 wt%-NaOH aqueous solution at 100 degrees C. Molecularly aligned tendon chitosan was chemically thiolated by reacting 4-thiobutyrolactone with the chitosan amino group. The introduction of thiol groups and their distribution to tendon chitosan and chitosan cast film were confirmed using ATR FT-IR, (1)H-NMR, and EDS. The 1.24 micromol/g of thiol groups introduced on the surface of tendon chitosan and the chitosan cast film was confirmed using ultraviolet (UV) spectra. Thiol groups of cysteine located at the end of synthetic laminin peptides were then reacted chemically with thiolated chitosan to form chitosan-S-S-laminin peptide. YIGSR estimated at 0.92 micromol/g and IKVAV estimated at 0.28 micromol/g on thiolated tendon chitosan were confirmed using UV spectra. YIGSR was estimated at 0.85 micromol/g and IKVAV was estimated at 0.34 micromol/g on the thiolated chitosan cast film.

Animals↗
PubMed2005

Culturing of skin fibroblasts in a thin PLGA-collagen hybrid mesh.

A thin biodegradable hybrid mesh of synthetic poly(DL-lactic-co-glycolic acid) (PLGA) and naturally derived collagen was used for three-dimensional culture of human skin fibroblasts. The hybrid mesh was constructed by forming web-like collagen microsponges in the openings of a PLGA knitted mesh. The behaviors of the fibroblasts on the hybrid mesh and PLGA knitted mesh were compared. The efficiency of cell seeding was much higher and the cells grew more quickly in the hybrid mesh than in the PLGA mesh. The fibroblasts in the PLGA mesh grew from the peripheral PLGA fibers toward the centers of the openings, while those in the hybrid mesh also grew from the collagen microsponges in the openings of the mesh resulting in a more homogenous growth. The proliferated cells and secreted extracellular matrices were more uniformly distributed in the hybrid mesh than in the PLGA mesh. Histological staining of in vitro cultured fibroblast/mesh implants indicated that the fibroblasts were distributed throughout the hybrid mesh and formed a uniform layer of dermal tissue having almost the same thickness as that of the hybrid mesh. However, the tissue formed in the PLGA mesh was thick adjacent to the PLGA fibers and thin in the center of the openings. Fibroblasts cultured in the hybrid mesh were implanted in the back of nude mouse. Dermal tissues were formed after 2 weeks and became epithelialized after 4 weeks. The results indicate that the web-like collagen microsponges formed in the openings of the PLGA knitted mesh increased the efficiency of cell seeding, improved cell distribution, and therefore facilitated rapid formation of dermal tissue having a uniform thickness. PLGA-collagen hybrid mesh may be useful for skin tissue engineering.

Absorbable Implants↗
PubMed2005

The effect of calcium ion concentration on osteoblast viability, proliferation and differentiation in monolayer and 3D culture.

Our research group aims to develop an osteochondral composite using type II collagen gel with hydroxyapatite (HAp) deposited on one side. Soaking gels in Ca2+ and phosphate solution is indispensable to HAp deposition, so relationships between cell behavior and Ca2+ concentration were examined in two- and three-dimensional cultures. The present results indicate that 2-4 mM Ca2+ is suitable for proliferation and survival of osteoblasts, whereas slightly higher concentrations (6-8 mM) favor osteoblast differentiation and matrix mineralization in both 2- and 3-dimensional cultures. Higher concentrations (>10 mM) are cytotoxic. Purely from the perspective of calcium deposition, higher concentrations lead to increased accumulation of Ca2+. Culturing cells in phosphate-containing gel in media with Ca2+ also leads to time-dependent formation of HAp in the gel. Considering the viability of embedded cells, culturing scaffolds in media with Ca2+ concentrations around 5mM is useful for both HAp deposition and osteoblast behavior.

Animals↗
PubMed2005

A new biotechnology for articular cartilage repair: subchondral implantation of a composite of interconnected porous hydroxyapatite, synthetic polymer (PLA-PEG), and bone morphogenetic protein-2 (rhBMP-2).

OBJECTIVE: Articular cartilage repair remains a major obstacle in tissue engineering. We recently developed a novel tool for articular cartilage repair, consisting of a triple composite of an interconnected porous hydroxyapatite (IP-CHA), recombinant human bone morphogenetic protein-2 (rhBMP-2), and a synthetic biodegradable polymer [poly-d,l-lactic acid/polyethylene glycol (PLA-PEG)] as a carrier for rhBMP-2. In the present study, we evaluated the capacity of the triple composite to induce the regeneration of articular cartilage. METHODS: Full-thickness cartilage defects were created in the trochlear groove of 52 New Zealand White rabbits. Sixteen defects were filled with the bone morphogenetic protein (BMP)/PLA-PEG/IP-CHA composite (group I), 12 with PLA-PEG/IP-CHA (group II), 12 with IP-CHA alone (group III), and 12 were left empty (group IV). The animals were killed 1, 3, and 6 weeks after surgery, and the gross appearance of the defect sites was assessed. The harvested tissues were examined radiographically and histologically. RESULTS: One week after implantation with the BMP/PLA-PEG/IP-CHA composite (group I), vigorous repair had occurred in the subchondral defect. It contained an agglomeration of mesenchymal cells which had migrated from the surrounding bone marrow either directly, or indirectly via the interconnecting pores of the IP-CHA scaffold. At 6 weeks, these defects were completely repaired. The regenerated cartilage manifested a hyaline-like appearance, with a mature matrix and a columnar organization of chondrocytes. CONCLUSIONS: The triple composite of rhBMP-2, PLA-PEG, and IP-CHA promotes the repair of full-thickness articular cartilage defects within as short a period as 3 weeks in the rabbit model. Hence, this novel cell-free implant biotechnology could mark a new development in the field of articular cartilage repair.

Animals↗
PubMed2005

Bone repair analysis in a novel biodegradable hydroxyapatite/collagen composite implanted in bone.

The purpose of this study was to evaluate a biodegradable hydroxyapatite/collagen composite and to examine the use of the calcium ion contained for bone formation and growth. Surgical holes were prepared in the femora and tibiae of beagle dogs, and were filled with the hydroxyapatite/collagen composite labeled with alizarin red. After 4 weeks, calcein was administered to the experimental dogs. After 1 additional week, the femora and tibiae were removed surgically and fixed in formalin. Light microscopy and confocal laser scanning microscopy were used to examine the surgical holes with their implanted materials and the surrounding bone. There were only a few inflammatory cells adjacent to the hydroxyapatite/collagen composite. The newly formed bone in the cortical bone was stained with calcein, which binds to serum calcium, and new bone near the hydroxyapatite/collagen composite in the holes was stained positive for alizarin red, which binds to the calcium in the hydroxyapatite/collagen composite. In addition, osteoblasts near the hydroxyapatite/collagen composite as well as newly formed bone adjacent to the osteoblasts showed alizarin red staining, but the new bone at a distance from the hydroxyapatite/collagen implant reacted only to calcein staining. These results, using the tissue labeling method with calcein and alizarin red, suggested that the calcium bound to the alizarin red released from the hydroxyapatite/collagen composite materials might have been translocated to sites of new bone formation. The present experiment showed that the novel hydroxyapatite/collagen composite is a useful implant material for bone augmentation and that the calcium in the newly formed bone might have been released from the implant.

Absorbable Implants↗
PubMed2005

Development of a novel biomaterial, hydroxyapatite/collagen (HAp/Col) composite for medical use.

A hydroxyapatite/type I collagen (HAp/Col) composite, aligning hydroxyapatite nano-crystals along collagen molecules, has been synthesized. The biocompatibility, osteoconductivity and efficacy as an rhBMP-2 carrier of this novel biomaterial implanted in the weight-bearing site have been examined. The HAp/Col implants adsorbing 0 or 400 microg/ml of rhBMP-2 were implanted into bone defects of tibiae in 3 beagle dogs and fixed according to the Ilizarov method. As a control, bone defects of 20 mm remaining in 2 beagle dogs and the dogs were allowed to walk using a Ilizarov external skeletal fixator. The radiological and histological findings suggest that the implants induce bone remodeling units and are a superior carrier of rhBMP-2 due to the stimulation of early callus and new bone formation. As a next step, anterior fusion was carried out on 6 beagle dogs with the implants adsorbing 400 microg/ml of rhBMP-2, and 9 dogs with the implants without rhBMP-2. In 3 dogs of the rhBMP-treated group, as well as 6 dogs of the non-rhBMP-treated group, the implant was fixed with a poly-L-lactide plate. Histological and radiographical analysis suggest that enhancement of callus formation and bone bridging by rhBMP-treatment is effective to prevent collapse of the implant.

Animals↗
PubMed2004

Chondrogenic differentiation of human mesenchymal stem cells cultured in a cobweb-like biodegradable scaffold.

Human mesenchymal stem cells (MSCs) were cultured in vitro in a cobweb-like biodegradable polymer scaffold: a poly(dl-lactic-co-glycolic acid)-collagen hybrid mesh in serum-free DMEM containing TGF-beta3 for 1-10 weeks. The cells adhered to the hybrid mesh, distributed evenly, and proliferated to fill the spaces in the scaffold. The ability of the cells to express gene encoding type I collagen decreased, whereas its ability to express type II collagen and aggrecan increased. Histological examination by HE staining indicated that the cells showed fibroblast morphology at the early stage and became round after culture for 4 weeks. The cartilaginous matrices were positively stained by safranin O and toluidine blue. Immunostaining with anti-type II collagen and anti-cartilage proteoglycan showed that type II collagen and cartilage proteoglycan were detected around the cells. In addition, a homogeneous distribution of cartilaginous extracellular matrices was detected around the cells. These results suggest the chondrogenic differentiation of the mesenchymal stem cells in the hybrid mesh. The PLGA-collagen hybrid mesh enabled the aggregation of mesenchymal stem cells and provided a promotive microenvironment for the chondrogenic differentiation of the MSCs.

Absorbable Implants↗
PubMed2004

Calcium-phosphate-hybridized tendon directly promotes regeneration of tendon-bone insertion.

We developed a novel technique to improve tendon-bone attachment by hybridizing calcium phosphate (CaP) with tendons using an alternate soaking process. We characterized the deposited CaP on or in tendons and determined the healing process of anterior cruciate ligament (ACL) grafts by implanting CaP-hybridized free tendons in bone tunnels intra-articularly. Tendons to be implanted were alternately soaked 10 times in a Ca-containing solution and a PO(4)-containing solution for 30 s each. Treated tendons had ash contents threefold that of untreated tendons. Low-crystallinity apatite was found on or in treated tendons. In animal experiments, the CaP-hybridized tendon exhibited osteoclasts at the tendon-bone interface at 5 days after operation. At 2 weeks after operation, there were more osteoclasts and osteoblasts around the tendon than at 5 days after operation. Directly bonded areas were partially found between the implanted tendon and newly formed bone. The formation of a cartilage layer was partially apparent at 3 weeks after operation. The newly formed bone was observed almost around the tendon. We conclude that CaP-hybridized tendons clearly enhance the healing process of ACL grafts at the tendon-bone interface and regenerate a direct insertion-like formation of tendons similar to a normal healthy ACL insertion within 3 weeks after operation.

Animals↗
PubMed2004

Growth factor combination for chondrogenic induction from human mesenchymal stem cell.

During the last decade, many strategies for cartilage engineering have been emerging. Stem cell induction is one of the possible approaches for cartilage engineering. The mesenchymal stem cells (MSCs) with their pluripotency and availability have been demonstrated to be an attractive cell source. It needs the stimulation with cell growth factors to make the multipluripotent MSCs differentiate into chondrogenic lineage. We have shown particular patterns of in vitro chondrogenesis induction on human bone marrow MSCs (hBMSCs) by cycling the growth factors. The pellet cultures of hBMSCs were prepared for chondrogenic induction. Growth factors: TGF-beta3, BMP-6, and IGF-1 were used in combination for cell induction. Gene expression, histology, immunohistology, and real-time PCR methods were measured on days 21 after cell induction. As shown by histology and immunohistology, the induced cells have shown the feature of chondrocytes in their morphology and extracellular matrix in both inducing patterns of combination and cycling induction. Moreover, the real-time PCR assay has shown the expression of gene markers of chondrogenesis, collagen type II and aggrecan. This study has demonstrated that cartilage tissue can be created from bone marrow mesenchymal stem cells. Interestingly, the combined growth factors TGF-beta3 and BMP-6 or TGF-beta3 and IGF-1 were more effective for chondrogenesis induction as shown by the real-time PCR assay. The combination of these growth factors may be the important key for in vitro chondrogenesis induction.

Cartilage↗
PubMed2004

Muscle-specific protein MDP77 specifically promotes motor nerve regeneration in rats.

This study has examined the effects of recombinant human MDP77 (rhMDP77) on sciatic motor nerve regeneration in vivo. We carried out bridge grafting (14 mm) into the sciatic nerve of Sprague-Dawley rats using silicone tubes containing a mixture of type-I collagen and 0, 5, 10, or 20 microg/ml of rhMDP77, or containing phosphate-buffered solution (N = 6 in each group). Electrophysiological and histological evaluations carried out 12 weeks after implantation suggest that rhMDP77 has a positive effect on terminal and collateral sprouting of regenerating nerves and thereby promotes motor nerve regeneration in a dose-dependent manner.

Action Potentials↗
PubMed2004

Nano-scaled hydroxyapatite/polymer composite IV. Fabrication and cell adhesion properties of a three-dimensional scaffold made of composite material with a silk fibroin substrate to develop a percutaneous device.

Nano-scaled sintered hydroxyapatite (HAp) particles with an a-axis length of 87 +/- 23 nm, a c-axis length of 236 +/- 81 nm, and a mean aspect ratio ( c/ a) of 2.72 were covalently linked onto a silk fibroin (SF) substrate chemically modified by graft polymerization with gamma-methacryloxypropyl trimethoxysilane (MPTS). Graft polymerization with poly(MPTS) on SF was conducted by free-radical initiation in a water solvent with pentaethylene glycol dodecyl ether as a nonionic surfactant. The alkoxysilyl groups of the graft polymers avoided hydrolysis and maintained their activity in coupling with the hydroxyl groups on the HAp surface despite the use of water as the reaction solvent. The weight gain of poly(MPTS) on SF increased with increasing the reaction time, eventually reaching a plateau value of about 15 wt% after 50 min of reaction time. After HAp covalent coating, the particles separated or aggregated into several crystals, as shown by scanning electron microscopic observation. L929 fibroblast cells adhered more plentifully on HAp-coated SF compared to untreated SF and hydrolyzed poly(MPTS)-grafted SF during 24 h or 48 h of incubation. The cells adhered only on the HAp surface but not at all on the dehydrated grafted surface of SF without HAp. A button-shaped prototype for a percutaneous device was manufactured by transplantation of HAp-coated SF fibers of about 100 microm in length onto silicone moldings using an adhesive, and the device showed good cell adhesiveness.

Biocompatible Materials↗
PubMed2004

Development of guided bone regeneration membrane composed of beta-tricalcium phosphate and poly (L-lactide-co-glycolide-co-epsilon-caprolactone) composites.

To create biodegradable and thermoplastic materials for guided bone regeneration, GBR, and guided tissue regeneration, GTR, membranes, composites of beta-tricalcium phosphate, TCP, and biodegradable polyesters, poly (L-lactide-co-glycolide-co-epsilon-caprolactone), PLGC, and poly (L-lactide-co-epsilon-caprolactone), PLCL, were prepared by a heat-kneading method. The composites maintained thermoplasticity and mechanical strength by formation of a chemical interaction between Ca on TCP and C=O on the lactide segment of PLGC or PLCL. The composites also indicated composite effects in pH auto-regulation property and elongation of biodegradation period, e.g., the composites maintained their mechanical strength up to 12 weeks after soaking in both physiological and phosphate-buffered saline, and the period was sufficient time to use for GBR and GTR membranes. Animal tests for GBR indicated that the present composite membrane successfully regenerated beagles' mandible defects 10 x 10 x 10 mm3 in size. These results suggested that the TCP/PLGC bioresorbable composites could be utilized for GBR and GTR therapy.

Animals↗
PubMed2004

Relationship between bone mineral density of the distal radius and ulna and fracture characteristics.

PURPOSE: This study was designed to elucidate the different decrease patterns of bone mineral density (BMD) in the distal tenth and third of the radius and ulna that influence the incidence of fractures, the fracture type, and redisplacement after closed reduction and casting. METHODS: The BMD of 1024 healthy women and 86 women treated for distal radius and ulna fracture was measured in the distal tenth and distal third zones of the radius and ulna (termed R10, R3, U10, and U3, respectively) using dual-energy x-ray absorptiometry. The BMD in the fracture group was compared with age-matched BMD in the nonfracture group. The relationship between BMD, type of fracture, and radiographic parameters (radial length, radial inclination, and palmar tilt) determined after closed reduction and at bone union were examined. RESULTS: The BMD of the distal radius in zones R10 and R3 in the fracture group were significantly lower than those in the nonfracture group among subjects older than 80 years, whereas the BMD in the U10 and U3 in the fracture group were lower than those in the nonfracture group among subjects aged 50 to 59 years. No significant difference in the mean BMD in any of the fracture group subjects was found, irrespective of the fracture pattern. A significant correlation was found between the BMD in the R10 and radial length measured at bone union and between radial length measured at bone union and RL measured after closed reduction. The radial length, radial inclination, and palmar tilt measured at bone union correlated significantly with each other. CONCLUSIONS: The BMD in the R10 and R3 are potential indicators of a distal radius fracture in women over 80 years old, whereas the U10 and U3 are indicators of an ulnar styloid fracture in women in their fifties. In addition the distal radius (R10) is an important prognostic parameter of possible deformity of the radius after closed reduction and casting.

Absorptiometry, Photon↗
PubMed2004

Glutaraldehyde cross-linked hydroxyapatite/collagen self-organized nanocomposites.

To control the mechanical properties and biodegradability of self-organized hydroxyapatite/collagen (HAp/Col) nanocomposites, cross-linkage was introduced into the composites with glutaraldehyde (GA). The HAp/Col composite suspensions, prepared by a simultaneous titration method and aged for 3h, were cross-linked with the reagents for 10min under vigorous stirring. The precipitates obtained were filtrated and compacted by dehydration under a uniaxial pressure. The particle size distribution, 3-point bending strength, contained water amount and swelling ratio of the composites were examined as a function of cross-linkage amount; the biodegradability was estimated by animal tests using rabbits. As regards the cross-linked composites, no long-rage alignment of HAp crystals along collagen molecules was found with a transmission electron microscope, suggesting that the cross-linking reagents suppressed their long-range self-organization mechanism. The 3-point bending strength increased with the GA content and took a maximal value at 1.35mmol/g(col). The animal tests indicated no toxicity and osteoclastic resorption with good osteoconductivity. The resorption rate was decreased with increasing GA concentration. These results suggest that GA cross-linkage controls mechanical properties and resorption rate without reducing high biocompatibility of the composite.

Animals↗
PubMed2004

Development of a hydroxyapatite/collagen nanocomposite as a medical device.

The effect of cross-linking of a hydroxyapatite/collagen (HA/Col) nanocomposite, in which HA nanocrystals and collagen fibers are aligned like natural bone by a self-organization mechanism between HA and collagen in vitro, on mechanical properties was examined. The influence of degree of cross-linking, as well as rhBMP-2 preadsorption to the composite on the substitution pattern and rate with bone, was examined. In Experiment 1, anterior fusion was carried out at the C3-C4 vertebrae on 10 dogs and they were implanted as follows: without cross-linking and without adsorbed rhBMP-2 (three dogs), with cross-linking and without adsorbed rhBMP-2 (three dogs), without cross-linking and with adsorbed rhBMP-2 (two dogs), and with cross-linking and adsorbed rhBMP-2 (two dogs). Implants were removed from each dog for histology determinations after 12, 16, and 24 weeks in the non-rhBMP-treated groups, and after 16 and 24 weeks in the rhBMP-treated groups. In Experiment 2, the HA/Col composites with cross-linking and both with and without rhBMP-2 pretreatment were implanted into a bone defect of 20 mm made in the central part of tibiae in dogs (N = 3 in each group). As a control, bone defects of 20 mm remained without implantation (N = 3). The dogs were allowed to walk using an Ilizarov extra skeletal fixator. The implants were removed after 12, 16, and 24 weeks from one dog in each group. The cross-linking of the HA/Col composite was effective in controlling both the mechanical strength and bioresorbability. A "self-organization process" on the HA/Col implant surface resulted in the formation of bone remodeling units in and around the implant. Radiographic and histological findings suggest that a combined treatment of cross-linking of the HA/Col composite with preadsorption of rhBMP-2 molecules may be a very suitable replacement of existing ceramic systems in the anterior fusion of the cervical spine, as well as inlay grafting of bone defects in weight-bearing sites.

Animals↗