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Takafumi Yoshikawa

Publications and source records attributed to Takafumi Yoshikawa.

At least 19 recordsLinked to original sources
PubMed2006

Suppressive effect of alpha2 Heremans-Schmid glycoprotein on in vitro calcification of osteogenesis.

BACKGROUND: alpha(2) Heremans-Schmid glycoprotein (alpha(2)HS glycoprotein) is predominantly found in bone. To date, we have investigated plasma alpha(2)HS levels in immature babies and neonates as well as the histological distribution in various neonatal tissues in order to clarify its physiological significance. In an effort to understand the physiological function of alpha(2)HS glycoprotein in bones, we studied the effects of alpha(2)HS glycoprotein in cultured osteogenesis model using rat marrow cells. METHODS: We added different concentrations of alpha(2)HS glycoprotein to cultured marrow cells, including osteoblasts in the presence of dexamethasone, in an attempt to elucidate the effects of alpha(2)HS glycoprotein on osteoblast growth and bone calcification in vitro. RESULTS: The results showed that total DNA content was significantly increased with 0.2-20 nM (f.c.) alpha(2)HS glycoprotein, but was neither suppressed nor increased with 200 nM (f.c.) alpha(2)HS glycoprotein. Although ALP activity increased with 0.2 or 2 nM (f.c.) alpha(2)HS glycoprotein, it decreased with 20 or 200 nM (f.c.) alpha(2)HS glycoprotein. While 0.2 nM (f.c.) alpha(2)HS glycoprotein had no effect on calcium or osteocalcin content, 2 nM (f.c.) alpha(2)HS glycoprotein decreased both calcium content and osteocalcin content by about half, and no calcium or osteocalcin was observed with 20 or 200 nM (f.c.). Calcium staining of cultured marrow cells revealed that the number of stained cell tubercles decreased in a concentration-dependent manner. CONCLUSION: These findings suggest that alpha(2)HS glycoprotein regulates the growth of osteoblasts and acts as an inhibitory factor in the regulation of bone calcification.

Animals↗
PubMed2006

Bone formation-promoting effect of genistein on marrow mesenchymal cell culture.

The effect of genistein, a soybean isoflavone, on new bone formation by bone marrow cells from mature rats and humans was examined. Bone marrow cells were collected from the femoral diaphysis of 7-week-old Fisher rats, cultured in MEM containing fetal calf serum and then cultured with or without the addition of dexamethasone to the bone-forming medium. Genistein was added at concentrations of 10(-5),10(-6),10(-7) or 10(-8) M. Bone formation was examined 2 weeks after culture. After informed consent was obtained from a 55-year-old woman with lumbar spondylosis deformans, bone marrow cells were collected from her ilium for culture by the same process, and bone formation investigated. In both rats and humans, when dexamethasone was added to the bone-forming medium, genistein (10(-7) M and 10(-8) M) caused a significant increase in the levels of calcium, alkaline phosphatase, and DNA compared with cells not cultured in genistein. In conclusion, genistein was found to promote bone formation at lower concentrations across species, and thus may be useful as a bone formation-promoting factor.

Animals↗
PubMed2005

Clinical and pathological features of non-alcoholic steatohepatitis.

Clinical and pathological features were reviewed in 76 Japanese patients with non-alcoholic steatohepatitis (NASH). Forty-one were male and 35 were female with the mean age of 49.7 years old (range 15-75 years old, males; 46.3, females; 53.7 years old). Fifty-four percent of patients were preobese with a body mass index (BMI) between 25 and 30, while 16% of the patients were non-obese, and only 30% of the cases were morbidly obese, indicating that Japanese have a greater tendency to develop insulin-resistance and fatty liver disease than Western people. Hyperlipidemia was found in 51%, diabetes mellitus in 38%, and hypertension in 33% of the patients. Abnormally elevated liver function tests were found in one-third to two-thirds of the patients and were characteristically mild with 2- to 3-fold elevation from the normal range in the majority of the cases. Histological features of the liver were similar or identical to those reported in English literature and were characterized by fatty change, perivenular and pericellular fibrosis in zone 3, hepatocyte ballooning and necrosis with occasional Mallory's body formation and polymorphonuclear leukocyte infiltration. Mallory's bodies were found in 39% of patients and were characteristically small and poorly formed compared with those in alcoholic hepatitis. Eosinophilic granular or dirty foggy aggregated, not sufficient to be identified as Mallory's bodies, were a rather characteristic cytoplasmic expression in NASH patients. Portal inflammation and fibrosis were not found in the early stage of NASH, but were found as the disease progresses with formation of C-C and/or P-C bridging fibrosis, and eventually resulting in liver cirrhosis.

Journal Article↗
PubMed2005

Cervical laminoplasty combined with muscle release in patients with athetoid cerebral palsy.

STUDY DESIGN: A retrospective study comparing cervical laminoplasty with or without muscle release for the treatment of cervical myelopathy resulting from athetoid cerebral palsy. OBJECTIVE: To assess the effectiveness of muscle release in the treatment of athetoid cerebral palsy. SUMMARY OF BACKGROUND DATA: While anterior and/or posterior spinal fusion has been generally accepted as necessary in surgical treatment for cervical myelopathy due to athetoid cerebral palsy, several studies have shown relatively favorable results following laminoplasty. Better results can be obtained by combining muscle release. METHODS.: Study participants were 10 patients who underwent cervical laminoplasty combined with muscle release (mean age, 44.6 years) and 15 patients who underwent cervical laminoplasty alone (mean age, 48.2 years). Therapeutic outcomes 1 year after surgery, as assessed by Kurokawa's methods and JOA scores, were compared between groups. RESULTS: Recovery rate 1 year after surgery was significantly higher for the muscle release group than for the control group. In both groups, recovery rates were significantly better for patients who could walk before surgery. CONCLUSIONS: Cervical laminoplasty combined with muscle release for the treatment of cervical myelopathy due to athetoid cerebral palsy is effective in simplifying postoperative therapy and improving JOA scores.

Adult↗
PubMed2005

Bio-artificial periosteum for severe open fracture--an experimental study of osteogenic cell/collagen sponge composite as a bio-artificial periosteum.

In an attempt to reduce complications in cases of severe open fracture, we developed a bio-artificial periosteum composed of osteogenic cells and collagen sponge. In the present study, we evaluated the osteogenic potential of the bio-artificial periosteum in vivo and in vitro. After 4-week incubation in vitro, the bio-artificial periosteum had high alkaline phosphatase activity and osteocalcin content. Moreover, energy dispersive X-ray analysis revealed numerous crystal structures consisting of P and Ca on the surface of the bio-artificial periosteum. Using a rat model for severe bone injury, we examined the bone formation process in defect sites covered with the bio-artificial periosteum. New bone formation occurred in the central part of the bone defect as well as at the bone edge. We conclude that by using the bio-artificial periosteum, the fracture site benefited from an improved osteogenic environment. These results indicate that a clinical trial to further evaluate this technique should be conducted.

Alkaline Phosphatase↗
PubMed2005

Stimulatory effects of statins on bone marrow-derived mesenchymal stem cells. Study of a new therapeutic agent for fracture.

Recent studies have reported that statins, inhibitors of hydroxymethylglutaryl-coenzyme A (HMG-CoA) reductase, increase bone formation in osteoblasts in vitro, suggesting that statins may have a new therapeutic application in the treatment of osteoporosis. During the reparative phase of healing of bone fractures, bone marrow-derived mesenchymal stem cells differentiate into osteoblasts or chondrocytes to form callus. If statins also stimulate bone formation in bone marrow-derived mesenchymal stem cells they may have beneficial effects in the treatment of bone fractures. In this study, we assessed the effect of statins on bone formation in rat bone marrow-derived mesenchymal stem cells in vitro. The statins fluvastatin, simvastatin and pravastatin did not significantly enhance mineralization, alkaline phosphatase (ALP) activity and bone gra protein (BGP, osteocalcin). These findings suggest that statins do not increase bone formation in bone marrow-derived mesenchymal stem cells.

Animals↗
PubMed2004

Alteration of sugar donor specificities of plant glycosyltransferases by a single point mutation.

In comparison with the amino acid sequences of seven species of glucosyltransferases and six species of galactosyltransferases, glutamine and histidine are highly conserved as the last amino acid residue of a glycosyltransferase-specific conserved region (UDPGT) in glucosyltransferases and galactosyltransferases, respectively. Consequently, the sugar donor specificities of glycosyltransferases are successfully altered by a single amino acid point mutation. UDP-galactose:anthocyanin galactosyltransferase (ACGaT), isolated from Aralia cordata, acquired glucosyltransferase activity in addition to the inherent galactosyltransferase activity by replacing histidine with glutamine. In contrast, UDP-glucose:flavonoid glucosyltransferase (UBGT), isolated from Scutellaria baicalensis, did not acquire galactosyltransferase activity by replacing glutamine with histidine, and exhibited a remarkable decrease in glucosyltransferase activity.

Amino Acid Sequence↗
PubMed2004

Effect of N- and C-terminal deletions on the RNA N-glycosidase activity and the antigenicity of karasurin-A, a ribosome-inactivating protein from Trichosanthes kirilowii var. japonica.

Karasurin-A, from root tubers of Trichosanthes kirilowii var. japonica, is a type I ribosome-inactivating protein (RIP) that displays activity of RNA N-glycosidase to remove an adenine in the conserved sarcin/ricin loop of the largest RNA in the ribosome. We expressed recombinant proteins of karasurin-A and its various mutants with N- or C-terminal deletions in Escherichia coli as fusion proteins with maltose-binding protein (MBP), and compared their enzymatic activities and antigenicities. Muteins of karasurin-A generated by deleting either the first 100 N-terminal or the last 30 C-terminal amino acid residues lost activity of RNA N-glycosidase. The mutant proteins whose 80 N-terminal or 20 C-terminal amino acids were deleted could depurinate rRNA although the activities were decreased drastically. The antigenicities of the recombinant proteins were considerably reduced by deleting 20 amino acid residues from either N- or C-terminal regions.

Carrier Proteins↗
PubMed2004

A citrus flavonoid hesperidin suppresses infection-induced endotoxin shock in mice.

Administration of a citrus flavonoid hesperidin (HES) to mice before LPS challenge significantly reduced tumor necrosis factor (TNF)-alpha production in a dose-dependent manner. Treatment of HES 3 h before intraperitoneal (i.p.) infection with 10(8) CFU Salmonella typhimurium aroA resulted in rescue from lethal shock as similar to LPS-nonresponder mice. Not only bacterial numbers in livers and spleens but also plasma LPS levels significantly decreased by pretreatment with HES. In addition, HES markedly suppressed plasma levels of TNF-alpha and high mobility group box chromosomal protein 1 (HMGB-1), decreased the number of apoptotic cells in livers and normalized the activated states of blood coagulation factors such as prothrombin time and platelet numbers caused by infection. Pretreatment of LPS with HES suppressed the chromogenic Limulus reaction.

Animals↗
PubMed2004

An apoptotic inducer, aralin, is a novel type II ribosome-inactivating protein from Aralia elata.

We recently found that aralin, a novel cytotoxic protein consisting of two subunits, from Aralia elata selectively induces apoptosis in transformed cells as compared to normal cells. Here we report that aralin is a lectin specific for galactose (Gal) and its derivatives, and possesses RNA N-glycosidase activity as a new type II ribosome-inactivating protein (RIP). The RNA N-glycosidase activity of aralin was detected in cell-free and whole cell systems by the generation of an R-fragment from 28S rRNA. Coinciding with appearance of the R-fragment in aralin-treated cells, significant inhibition of protein synthesis was observed prior to the onset of apoptosis. Aralin-evoked cell death was efficiently repressed by the addition of Gal and its derivatives. Interestingly, melibiose preferentially protected normal cells from apoptosis as compared with transformed cells. Using rhodamine-coupled aralin, the aralin receptor could be clearly detected around the cell surface of transformed cells, but to a lesser extent on normal cells. Receptor binding was suppressed by Gal. These results indicate that aralin is incorporated into cells via its Gal-containing cell surface receptor and induces apoptosis through its RIP activity. Moreover, the expression level and/or structural changes of the aralin receptor may affect the sensitivity toward aralin.

Animals↗
PubMed2004

Osteogenic potential of cultured bone/ceramic construct: comparison with marrow mesenchymal cell/ceramic composite.

Osteogenesis occurs in porous hydroxyapatite (HA) when porous HA blocks combined with marrow mesenchymal cells are grafted in vivo. In vitro bone formation occurs in HA pores when HA combined with marrow cells is cultured in osteogenic medium containing dexamethasone. This cultured bone/HA construct possesses higher osteogenic ability when it is grafted in vivo. In the present study, we compared the osteogenic potential of a cultured bone/HA construct with that of a marrow mesenchymal cell/HA composite. Marrow cells were obtained from the femoral bone shaft of 7-week-old, male Fischer 344 rats and were cultured in T-75 flasks. Cells were concentrated, then frozen and stored in liquid nitrogen for 6 months. The cryopreserved cells were then thawed and prepared for subculture in porous HA (5 x 5 x 5 mm, Interpore 500) and for implantation with porous HA. After 2 weeks of subculture, three cultured bone/HA constructs were separately implanted in the right side of the back of each syngeneic 7-week-old male Fischer rat, and three thawed cell/HA composites (without subculture) were separately implanted in the left side. These implants were harvested at 2 or 4 weeks postimplantation, and prepared for histological, biochemical, and genetic analysis. Alkaline phosphatase activity and osteocalcin content of cultured bone/HA constructs were much higher than those of the cell/HA composites at 2 and 4 weeks postimplantation. Histological examination and gene expression data agreed with these findings. The culture technique discussed herein should facilitate the development of biosynthetic bone implants with higher osteogenic capacity.

Alkaline Phosphatase↗
PubMed2003

Inhibitory effect of quercetin on carrageenan-induced inflammation in rats.

We examined the effect of quercetin on the inflammatory response induced by carrageenan in the rat. Air pouches were induced subcutaneously on the backs of rats and injected with carrageenan. The rats were treated with either vehicle or quercetin at a dose of 10 mg/kg one hour before carrageenan challenge. Fourty-eight hour after carrageenan challenge, the air pouches were removed and analyzed. The volume, protein amounts and cell counts in the exudation obtained from the quercetin-treated animals were significantly reduced compared to those from vehicle-treated animals. The contents of PGE(2), TNF-alpha, RANTES, MIP-2 and the mRNA for cyclooxygenase-2 were also suppressed in these rats. The histological examination displayed the suppression of the inflammatory response in the pouch tissues from quercetin-treated rats. As the anti-inflammatory effect of the flavonols was more or less at the similar level among the quercetin-, isoquercitrin- or rutin-treated rats, it appeared that the sugar parts did not influence on the anti-inflammatory effect. Our study indicated that the flavonols modulated the inflammatory response, at least in part, by modulating the prostanoid synthesis as well as cytokine production.

Animals↗
PubMed2003

Bone regeneration by grafting of an autogenous cultured bone/ceramic construct.

The in vivo osteogenic potential of autogenous cultured bone/ceramic constructs in large animals or humans is unknown, and thus we performed a preliminary study of this issue prior to clinical application. All autogenous cultured-bone/ceramic constructs at 3 weeks after implantation in dogs showed obvious histological bone formation within the ceramic pores. In many pores, the HE staining of decalcified specimens revealed thick lamellar bone formation on the pore surface of ceramic. On the surface of bone tissue, numerous active cuboidal osteoblasts were evident. Biochemically, high alkaline phosphatase activity was detected in all dogs. Histological examination of the constructs at 8 weeks postimplantation showed lamellar bone formation with vascular system invasion into the pores, and regenerated hematopoietic bone marrow was often detected in association with the new bone in grafting of human cultured bone/ceramic constructs. Trilineage hematopoietic cells (i.e., granulocytic, erythroblastic, and megakaryocytic cells) were identified in the ceramic pores. Biochemically, high alkaline phosphatase activity and significant human osteocalcin content was detected in the constructs. Based on these findings, in the near future, this technique (grafting of patient-derived cultured bone/HA constructs) will be able to be applied to various bone reconstruction surgical treatments.

Alkaline Phosphatase↗
PubMed2003

cDNA cloning and expression of isoflavonoid-specific glucosyltransferase from Glycyrrhiza echinata cell-suspension cultures.

A cDNA encoding UDP-glucose: formononetin 7- O-glucosyltransferase, designated UGT73F1, was cloned from yeast extract-treated Glycyrrhiza echinata L. cell-suspension cultures using probes from Scutellaria baicalensis UDP-glucose: flavonoid 7- O-glucosyltransferase. The open reading frame of the UGT73F1 cDNA encodes a 441-amino-acid protein with a predicted molecular mass of 48.7 kDa. The deduced amino acid sequence showed that the protein is related to the stress-inducible glucosyltransferases. UGT73F1 mRNA was not detected in untreated G. echinata cultures but was transiently induced by treatment with yeast extract. Recombinant UGT73F1 was expressed as a histidine-tag fusion protein in Escherichia coli and purified to near homogeneity by nickel chelate chromatography. The purified recombinant enzyme was selective for isoflavonoid, formononetin and daidzein as substrates, while flavonoids and various tested non-flavonoid compounds were poor substrates.

Cells, Cultured↗
PubMed2002

Upper thoracic spinal cord injury without vertebral bony lesion: a report of two cases.

STUDY DESIGN: Case report. OBJECTIVES: To describe a rarely reported type of upper thoracic spinal cord injury without vertebral bony lesion in two cases with multiple trauma. SUMMARY OF BACKGROUND DATA: Because it is supported by the stiffness of the rib cage, the upper thoracic spine has greater stability than the cervical and lumbar regions, and thus its fracture or fracture dislocation is less frequent. Nevertheless, when fracture or fracture dislocation of upper thoracic spine occurs, spinal cord involvement and severe concomitant injuries are frequently associated. METHODS: Two cases who were suspected to have thoracic spinal cord injuries were referred to our emergency center: a 19-year-old girl presented with paraparesis after her motorcycle collided with a truck, and a 63-year-old male involved in an industrial accident presented with paraplegia. RESULTS: Radiograph and computed tomography scan showed no abnormality or dislocation in the vertebral bodies in these two cases, although the upper thoracic spinal cord injuries were suspected by clinical features. Magnetic resonance images detected abnormal signals, suggesting spinal cord injuries, and these signals each emanated from levels that coincided with the observed clinical features. CONCLUSIONS: Transient subluxation or displacement might have caused the upper thoracic spinal cord injuries after the support of the rib cages was temporarily lost on application of excessive force, although such findings could not be confirmed by imaging procedures.

Accidents, Occupational↗
PubMed2002

Expression of c-Jun N-terminal kinases after axotomy in the dorsal motor nucleus of the vagus nerve and the hypoglossal nucleus.

c-Jun N-terminal kinases (JNKs)/stress-activated protein kinases (SAPKs) are a subgroup of mitogen-activated protein kinases (MAPKs), and important mediators of signal transduction from the cell surface to the nucleus. JNK phosphorylates the transcription factor c-Jun. c-Jun is one of the earliest and most consistent markers for neurons that respond to nerve-fiber transection. To elucidate the c-Jun metabolism after axotomy, we investigated the expression of JNKs mRNA and of JNKs and c-Jun proteins following vagus and hypoglossal nerve transection. We found that JNK 1, JNK 2 and JNK 3 mRNA were positive in the cytoplasm of neuronal and glial cells. JNK 1 and JNK 2 protein were distributed mainly in the cytoplasm of neurons and glial cells, while only JNK 3 immunoreactivity was observed intensely in the nuclei of neuronal cells. Activated JNK was also observed intensely in the nuclei of neuronal cells. In sham-operated animals, the cytoplasm of a few glial cells showed moderate immunoreactivity for activated JNK, while after axotomy the cytoplasm of all perineuronal microglial cells were stained intensely. Up-regulated c-Jun and phosphorylated c-Jun immunoreactivities were found in the nuclei of neuronal cells in the severed side of both the dorsal motor and hypoglossal nuclei. These results indicate that the principal activity of JNKs in neurons is contributed largely by JNK 3 under both normal and axotomized conditions, and that JNKs play an important role in signal transduction of perineuronal microglial cells after axotomy.

Animals↗
PubMed2002

Recombinant human bone morphogenetic protein-2 promotes osteogenesis within atelopeptide type I collagen solution by combination with rat cultured marrow cells.

We evaluated the combination effect of recombinant human bone morphogenetic protein-2 (rhBMP-2) and cultured rat bone marrow mesenchymal stem cells (MSCs) in atelopeptide type I collagen (AC) solution on osteogenesis in a diffusion chamber (DC) to develop a bone substitute having consistent osteogenic capability for clinical applications. The cultured MSCs were obtained by 10-day primary culture of fresh bone marrow cells of Fischer rats. We prepared three groups of DCs: AC solution with rhBMP-2, AC solution with cultured MSCs, and AC solution with rhBMP-2 and cultured MSCs. The prepared combined solutions were injected into DCs, which were subcutaneously implanted into the backs of syngeneic rats. DCs were harvested after 2, 4, or 8 weeks and analyzed for bone-forming capability by determining histological and osteoblastic biochemical markers. De novo bone formation was observed both inside and outside of the membrane filter of DCs in the group of AC solution with rhBMP-2 and cultured MSCs. The alkaline phosphatase activity and osteocalcin content in the group of AC solution with rhBMP-2 and cultured MSCs were significantly higher than those in the group of AC solution with cultured MSCs at any time. These findings indicate that AC aqueous solution is a useful material not only as a carrier of rhBMP-2 but also as a cell-anchorage for differentiation and proliferation of MSCs. Therefore, this study suggests that clinical repairs of bone defects are feasible using injectable AC solution with rhBMP-2 and cultured MSCs as a bone substitute.

Alkaline Phosphatase↗