Biomedical subjects

A Togari

Publications and source records attributed to A Togari.

At least 37 records · Page 2Linked to original sources

PubMed2000

Osteoclast differentiation factor in human osteosarcoma cell line.

A sensitive sandwich enzyme-linked immunosorbent assay (ELISA) for human osteoclast differentiation factor (ODF/RANKL/ OPGL/TRANCE) utilizing a polyclonal antibody that recognizes both human soluble ODF and mouse ODF in combination with a osteoclasogenesis inhibitory factor (OCIF/OPG) was developed. We can quantify the ODF level in not only human ODF (detection limit: 0.05 ng/ml), but also mouse ODF by virtue of cross-reactivity. Employing this assay system, we demonstrated that ODF is constitutively present as a membrane-bound form in both the human osteosarcoma cell lines, MG-63, HOS and SaOS-2, and the mouse osteoblastic cell line MC3T3-E1.

Animals↗

PubMed2000

Cytokines in Parkinson's disease.

We found that in Parkinson's disease (PD) the levels of various cytokines [tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta, IL-2, IL-4, IL-6, epidermal growth factor (EGF), transforming growth factor (TGF)-alpha, TGF-beta1] were significantly increased in the striatum (caudate and putamen) of the postmortem brain and in ventricular or spinal cerebrospinal fluid (VCSF, LCSF). Furthermore, the levels of the apoptosis-related proteins such as bcl-2 and soluble Fas (sFas) in the striatum were also elevated in PD. In 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated parkinsonism mice, the levels of IL-1beta in the striatum were significantly increased, but those of nerve growth factor (NGF) were significantly decreased, compared with control mice. In hemiparkinsonism rats produced by injection of 6-hydroxydopamine (6-OHDA) into one side of the median forebrain bundle, the levels of TNF-alpha in the 6-OHDA-treated side were increased in the striatum and substantia nigra, but not in the cerebral cortex, compared with those in the control side. Repeated administration of L-DOPA in the 6-OHDA-treated rats did not change the TNF-alpha levels in the control side and in the 6-OHDA-treated side in the substantia nigra, striatum, and cerebral cortex. Our results suggest that the changes in the levels of cytokines, neurotrophins, and apoptosis-related proteins in the nigrostriatal regions of PD may be involved in apoptosis and degeneration of the nigrostriatal DA neurons.

Animals↗

PubMed1999

Involvement of nitric oxide and biopterin in proinflammatory cytokine-induced apoptotic cell death in mouse osteoblastic cell line MC3T3-E1.

We previously demonstrated that the addition of proinflammatory cytokines (tumor necrosis factor-alpha, interleukin-1beta, and interferon-gamma) caused induction of mRNAs for inducible nitric oxide (NO) synthase and GTP cyclohydrolase I, a rate-limiting enzyme for 5,6,7,8-tetrahydrobiopterin (BH4) biosynthesis, and produced their end-products, NO and BH4, in osteoblastic cells. In the present study, we examined whether NO and BH4, biologically active substances produced in response to proinflammatory cytokines, are involved in the effect of these cytokines on cell viability and apoptotic cell death involving DNA fragmentation. Cytokines as well as S-nitroso-N-acetyl-d,l-penicillamine, an NO generator, decreased cell viability, whereas sepiapterin, which was converted intracellularly to BH4, increased it. The examination of cytotoxicity measured in terms of lactate dehydrogenase release and apoptotic cell death assessed by flow cytometric analysis showed that cytokine-induced reduction of cell viability may be based upon cell death by apoptosis, but not lytic death as in necrosis. In the presence of sepiapterin, cytokine treatment resulted in a statistically pronounced reduction in the amount of DNA fragmentation. Furthermore, this fragmentation could be blocked by 2-(4-carboxy-phenyl)-4,4,5,5-tetramethylimidazole-1-oxyl 3-oxide, an NO scavenger. These results suggest that cytokine-induced apoptotic cell death is attributed to NO and is protected by BH4, and that osteoblastic cells in response to proinflammatory cytokines operate both a stimulatory process resulting in NO production and an inhibitory one resulting in BH4 production for apoptotic cell death. Cytokine-induced apoptotic cell death may be a consequence of the predominance of the stimulatory process over the inhibitory process.

3T3 Cells↗

PubMed1999

Brain-derived growth factor and nerve growth factor concentrations are decreased in the substantia nigra in Parkinson's disease.

Using highly sensitive sandwich enzyme-linked immunosorbent assays (ELISA), we measured for the first time the concentrations of brain-derived growth factor (BDNF) in the brain (substantia nigra, caudate nucleus, putamen, cerebellum, and frontal cortex) from control and parkinsonian patients. BDNF in the human brain (the order of ng/mg protein) was significantly lower specifically in the nigrostriatal dopamine (DA) regions from parkinsonian patients than in those from control patients. The concentration of nerve growth factor (NGF) was also significantly decreased in the substantia nigra of parkinsonian patients in comparison with that in the controls. Since BDNF and NGF may play important roles in survival and differentiation of neuronal cells, the present data indicate that the lack of neurotrophins, especially BDNF, may be involved in the pathogenesis of PD during progress of neurodegeneration of the nigrostriatal DA neurons.

Adult↗

PubMed1999

Increase in level of tumor necrosis factor (TNF)-alpha in 6-hydroxydopamine-lesioned striatum in rats without influence of systemic L-DOPA on the TNF-alpha induction.

We previously reported that the levels of proinflammatory cytokines such as tumor necrosis factor (TNF)-alpha are increased in the striatum and cerebrospinal fluid from patients with Parkinson's disease (PD) and in the striatum from 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated mice, a murine model of PD. Presently we examined the changes in cytokine levels in the nigrostriatal dopaminergic regions in rats treated with an intrastriatal injection of 6-hydroxydopamine (6-OHDA) as a model of slowly progressive neurodegeneration similar to that seen in PD. We compared the content of TNF-alpha in the nigrostriatal dopaminergic regions of the control side with that of the 6-OHDA-injected experimental side, and also explored the effects of 6-OHDA injection combined with the L-DOPA treatment on the TNF-alpha level in the dopaminergic regions of rats. TNF-alpha was measured by a highly sensitive sandwich enzyme-linked immunosorbent assay (ELISA). The concentrations of TNF-alpha in the dopaminergic regions (striatum and substantia nigra) on the 6-OHDA injection side (right side: R) were significantly higher than those in the regions on the control side (left side: L) (Wilcoxon's test, P < 0.05). The ratio of the concentration of TNF-alpha on the injection side to that on the control side (TNF-alpha (R/L)) of each rat was not significantly different in the striatum and substantia nigra between the control group and the group treated with 25 or 50 mg/kg L-DOPA (Mann-Whitney Utests). These results show that TNF-alpha is increased in the striatum and substantia nigra in 6-OHDA-injected dopaminergic regions in rats, which finding is similar to the increase in the striatal dopaminergic regions in patients with PD. The results also indicate that L-DOPA alone or together with 6-OHDA does not increase the level of TNF-alpha in the brain in vivo.

Animals↗

PubMed1999

Electron microscopic observation on the parathyroid gland of the cardiomyopathic hamster (UM-X7.1 Syrian hamster).

The ultrastructure of the parathyroid gland (PTG) of the UM-X7.1 cardiomyopathic hamster (the UM-X7.1) was qualitatively and quantitatively compared with that of the normal hamster. In the UM-X7.1, the Golgi apparatus and rER were well developed compared with those of the control hamster. Morphometric analysis revealed that the mean values of the Golgi apparatus, rER and large vacuolar bodies were significantly greater in the UM-X7.1 than in the normal hamster and the mean value of lipid droplets was significantly less in the UM-X7.1 than in the normal hamster. Ultrastructurally, we consider that in the UM-X7.1, the synthesis and release of the parathyroid hormone may be activated by an excessive amount of circulating catecholamine, and the functional activity of the PTG may be higher than that in the normal hamster.

Animals↗

PubMed1999

Anti-proliferative capsular-like polysaccharide antigen from Actinobacillus actinomycetemcomitans induces apoptotic cell death in mouse osteoblastic MC3T3-E1 cells.

Actinobacillus actinomycetemcomitans (A. actinomycetemcomitans) has been implicated in the etiology of localized juvenile periodontitis (LJP), and produces a multiplicity of tissue-damaging products. Among those products, the capsular-like polysaccharide antigen (CPA) from A. actinomycetemcomitans is a potent mediator of bone resorption. In fact, this CPA (serotype b) is known to promote osteoclast-like cell formation via interleukin (IL)-1alpha production in mouse marrow cultures. Although osteoblasts complete bone formation, there are few reports focusing on the effect of CPA in bone-forming activity of osteoblasts in inflammatory disease sites. We hypothesized that CPA plays a mediating role in osteoblastic cells. Therefore, the purpose of this study was to examine the effect of CPA from A. actinomycetemcomitans on the mouse osteoblastic cell line MC3T3-E1 and human osteosarcoma SaOS-2 cells. A. actinomycetemcomitans serotype c resulted in a potent dose-dependent inhibition of cell proliferation of both cell lines. Characterization of the antiproliferative activity in the CPA demonstrated that it was not cytotoxic for MC3T3-E1. A 20-hour incubation with CPA-c resulted in a significant increase in apoptotic cell death in the cells, as evaluated by both cellular DNA fragmentation ELISA and FACS analysis. In contrast to the results obtained with a cytokine mixture (tumor necrosis factor-alpha, IL-1beta, and interferon-gamma), no inducible nitric oxide (NO) synthase gene expression or NO release could be detected in MC3T3-E1 after incubation with CPA-c. Further, both CPA-b and -c caused potent induction of apoptosis-related modifiers, e.g., Fas mRNA, whereas bcl-2 mRNA levels were unchanged. Therefore, this study has shown that CPA from A. actinomycetemcomitans contains a potent antiproliferative polysaccharide whose activity is associated with apoptotic cell death in MC3T3-E1, and that CPA per se is an inducer of apoptosis mediated by the Fas system but not by NO.

Aggregatibacter actinomycetemcomitans↗

PubMed1998

Effects of repeated systemic administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) to mice on interleukin-1beta and nerve growth factor in the striatum.

Interleukin (IL)-1beta and nerve growth factor (NGF) were measured for the first time in the brain (caudate nucleus and putamen, and frontal cortex) from control mice and mice treated with a parkinsonism-inducing neurotoxin, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), by highly-sensitive sandwich enzyme-linked immunosorbent assays (ELISAs) The concentrations of interleukin (IL)-1beta in the striatal regions were significantly higher in MPTP-treated mice than those in control mice treated with saline (P < 0.005), whereas those in the frontal cortex did not show significant differences between MPTP-treated and control mice. The present results agreed with our previous data on increased IL-1beta in the postmortem striatum from patients with Parkinson's disease (PD). In contrast, the concentrations of nerve growth factor (NGF) in the striatal regions were significantly lower in MPTP-treated mice, down to a 54% level of control mice (P < 0.05), but those in the frontal cortex did not show significant differences between MPTP-treated and control mice. Since NGF may play important roles as neurotrophic factors in the brain, the present results suggest that both the elevation of pro-inflammatory cytokine IL-1beta and the decrease of NGF in the dopaminergic striatal region of MPTP- treated mice may be related to neuronal cell death.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine↗

PubMed1998

Coexpression of GTP cyclohydrolase I and inducible nitric oxide synthase mRNAs in mouse osteoblastic cells activated by proinflammatory cytokines.

Proinflammatory cytokines, a combination of IL-1beta, TNF-alpha, and IFN-gamma, caused mRNA expression of GTP cyclohydrolase I (GTP-CH), the rate-limiting enzyme in tetrahydrobiopterin (BH4) biosynthesis, and of inducible nitric oxide synthase (iNOS) in a well-characterized osteoblastic clone MC3T3-E1 cell line. We found the expression of the GTP-CH gene in osteoblasts for the first time. The expression of GTP-CH and iNOS mRNAs was found to be maximal at 3 and 9 h, respectively. The expression of both genes elicited increases in BH4 and NO levels. Pharmacological studies using 2,4-diamino-6-hydroxypyrimidine, an inhibitor of GTP-CH activity, showed that BH4 is involved in the activity of iNOS, but not in the induction of iNOS mRNA. The results using an inhibitor of nuclear factor (NF)-kappaB and activating protein-1 (AP-1) activation suggested that coinduction of the two genes in response to cytokines occurred via activation of NF-kappaB and AP-1. In MC3T3-E1 cells BH4 and sepiapterin, producing BH4, could protect against apoptosis, i.e. the degradation of nuclear DNA in the cells, induced by NO derived from S-nitroso-N-acetyl-D-L-penicillamine. These results suggest that the induction of BH4 together with NO by proinflammatory cytokines could protect against NO-induced apoptosis in MC3T3-E1 cells.

3T3 Cells↗

PubMed1998

Comparison of the effects of carboxyl-terminal parathyroid hormone peptide[53-84] and aminoterminal peptide[1-34] on mouse tooth germ in vitro.

The biological activity of carboxyl-terminal parathyroid hormone peptide[53-84] (PTH[53-84]) and aminoterminal peptide [1-34] (PTH[1-34]) during cultivation of the lower molar tooth germ from mouse embryos was examined. PTH[1-34] increased alkaline phosphatase activity in the tooth germ at an early developmental stage (early to advanced bell stage), and decreased it at a late developmental stage (advanced to late bell stage). On the other hand, PTH[53-84] decreased alkaline phosphatase activity at the early stage and increased it at the late stage. Thus, both PTH fragments had distinct effects on tooth development in vitro, which were confirmed by histological observation of their marked effects on the formation of dentine and enamel. These results show that PTH[53-84] has biological activity in the murine tooth germ, opposite to the action of PTH[1-34].

Alkaline Phosphatase↗

PubMed1998

Involvement of vitamin D in lead-induced cutaneous calcification in rodents.

Experimental cutaneous calcification was induced by a subcutaneous injection of lead acetate in parathyroidectomized (PTX) rats and vitamin D-deficient rats whose plasma calcium concentrations were controlled by diets containing various amounts of calcium. Although, in PTX rats, the calcium content of the calcified plaque induced by lead acetate was correlated with their plasma calcium concentration, calcium content of the calcified plaque in vitamin D-deficient rats with normocalcemia was significantly lower than those in control and PTX rats with normocalcemia. A subcutaneous administration of 1 alpha,25-dihydroxyvitamin D3 (1 alpha,25(OH)2D3) following the lead acetate injection increased calcium content of the calcified plaque in vitamin D-deficient mice, suggesting the involvement of vitamin D in the cutaneous calcification. As the synthesis of osteocalcin, osteopontin and interleukin-1 (IL-1) receptor is well known to be increased by 1 alpha,25(OH)2D3, gene expressions of mRNAs of these proteins were examined in the skin with calcified plaque by reverse transcription polymerase chain reaction (RT-PCR) analysis. These mRNA expressions in the skin with calcified plaque were increased by a subcutaneous administration of 1 alpha,25(OH)2D3 following the lead acetate injection. On the contrary, they decreased in vitamin D-deficient mice. These findings suggest that vitamin D is directly involved in the process of experimental calcification.

Animals↗

PubMed1997

Expression of mRNAs for neuropeptide receptors and beta-adrenergic receptors in human osteoblasts and human osteogenic sarcoma cells.

In human periosteum-derived osteoblastic cells (SaM-1) and human osteosarcoma-derived cells (SaOS-2, HOS, MG-63), the mRNA expressions of calcitonin gene-related peptide receptor (CGRP-R), substance P receptor (SP-R), neuropeptide Y receptor (NPY-R), beta-adrenergic receptors (beta1-R, beta2-R, beta3-R), vasoactive intestinal polypeptide type 1 and type 2 receptors (VIP-1R, VIP-2R) and pituitary adenylate cyclase activating polypeptide receptor (PACAP-R) were examined by reverse transcription-polymerase chain reaction (RT-PCR). According to the magnitude of the mRNA expression of alkaline phosphatase (ALP), the relative state of commitment of these osteoblastic cell lines to the osteoblast lineage was SaM-1 > SaOS-2 > HOS > MG-63. CGRP-R, NPY-R, VIP-1R and beta2-R, but not SP-R, VIP-2R, PACAP-R, beta1-R and beta3-R, were expressed in osteoblasts as well as osteosarcoma cells. Expression of these receptors seems to be a common feature in osteoblastic cells, but the magnitude of expression was not dependent upon the relative state of commitment of the osteoblastic cells to the osteoblast lineage. In addition, VIP mRNA was not expressed in osteoblastic cells, suggesting the absence of an autocrine system of VIP in osteoblasts. These observations suggest that these neuropeptides and norepinephrine are involved in local regulation of human bone metabolism.

Adult↗

PubMed1997

Fluoride profiles in the cementum and root dentine of human permanent anterior teeth extracted from adult residents in a naturally fluoridated and a non-fluoridated area.

OBJECTIVES: To determine the effect of water fluoride concentration on the fluoride profile across the entire thickness of the cementum and root dentine of human permanent anterior teeth in adults. SUBJECTS: Twenty-eight human permanent anterior teeth from individuals aged from 30 to over 60 years were studied. SETTING: Teeth were obtained from a natural high-fluoride area (West Hartle-pool, UK; 1.0-1.3 ppm F in drinking water, WHP) and the other from a non-fluoridated naturally low fluoride area (Leeds, UK; 0.1 ppm F in drinking water, LDS). DESIGN: Cementum and root dentine were sampled using an abrasive micro-sampling technique from the cementum surface to the pulpal surface of root dentine. RESULTS: Fluoride concentration was higher in tooth roots (the cementum and dentine) taken from the naturally fluoridated area (WHP) than from the non-fluoridated area (LDS). Age and average fluoride concentration showed a positive correlation in WHP dentine, middle region of the root (r = 0.78, P < 0.001) and in the apical region of the root (r = 0.61, P < 0.05). WHP cementum had the strongest fluoride concentration correlation with age in the cervical region of the root (r = 0.67, P < 0.01). An analysis of variance (ANOVA) showed that the area (water fluoride content), age and number of years lived in the area combined with total age were significant. CONCLUSIONS: The fluoride content of cementum and root dentine in adult residents is related to fluoride content in drinking water.

Adult↗

PubMed1995

Morphological differences in the skull of ascorbic acid-deficient ODS rats.

The physiological importance of ascorbic acid (AsA) in bone formation has been thought to be due mainly to its effects on collagen production and, consequently, matrix formation. However, effects of AsA deficiency on growing animals are poorly understood. The purpose of this experiment was to find how AsA deficiency affects craniofacial growth cephalometrically, and the periodontal ligament and the bone density of alveolar septum histologically. Five-week-old growing male rats with hereditary defects in AsA synthesis (ODS rats) placed on an AsA-deficient or minimally supplemented diet for 4 weeks showed a marked reduction in plasma AsA level and a negative or slight increment in body weight. Cephalometric evaluation revealed that craniofacial growth was influenced by AsA deficiency. Histological observations of periodontal structure and bone density in AsA-deficient animals showed no difference from controls, whereas the periodontal ligament of the upper first molar in the deficient animals was narrower than that of controls. These observations indicate that AsA deficiency reduces craniofacial size and the width of the periodontal ligament significantly in growing rats, whereas bone density is not affected. Thus, AsA, through collagen production, may be important to the increase in size of the craniofacial structures in growing animals.

Alveolar Bone Loss↗

PubMed1995

Alteration of catecholamine phenotype in transgenic mice influences expression of adrenergic receptor subtypes.

Agonist-induced regulation of adrenergic receptors (ARs) has an important role in controlling physiological functions in response to changes in catecholamine stimulation. We previously generated transgenic mice expressing phenylethanolamine N-methyltransferase (PNMT) under the control of a human dopamine beta-hydroxylase gene promoter to switch catecholamine specificity from the norepinephrine phenotype to the epinephrine phenotype. In the present study, we first examined changes in catecholamine metabolism in peripheral tissues innervated by sympathetic neurons of the transgenic mice. In the transgenic target tissues, a high-level expression of PNMT led to a dramatic increase in the epinephrine levels, whereas the norepinephrine levels were decreased to 48.6-87.9% of the nontransgenic control levels. Analysis of plasma catecholamines in adrenalectomized mice showed large amounts of epinephrine derived from sympathetic neurons in the transgenic mice. Subsequently, we performed radioligand binding assays with (-)-[125I]iodocyanopindolol to determine changes in binding sites of beta-AR subtypes. In transgenic mice, the number of beta 2-AR binding sites was 56.4-74.9% of their nontransgenic values in the lung, spleen, submaxillary gland, and kidney, whereas the beta 1-AR binding sites were regulated in a different fashion among these tissues. Moreover, northern blot analysis of total RNA from the lung tissues showed that down-regulation of beta 2 binding sites was accompanied by a significant decrease in steady-state levels of the receptor mRNA. These results strongly suggest that alteration of catecholamine specificity in the transgenic sympathetic neurons leads to regulated expression of the beta-AR subtypes in their target tissues.

Animals↗

PubMed1995

[Pharmacological study of tooth formation and mineralization using a tooth germ cultivation system].

In this study, we introduce a cultivation method for mandibular first molars from mouse embryos. We also investigate the in vitro effects of several drugs on tooth formation. Histological observations demonstrated that the tooth germ dissected from 17-day-old embryonic mice and cultivated for 7 days under the conditions described here showed clear mineralization in the dentin and enamel. Alkaline phosphatase (ALP) activity and calcium content in the tooth germ increased developmentally during 7-day cultivation. The increase of calcium was preceded by that of ALP in the germ. The removal of ALP attached covalently to the external surface of plasma membranes by phosphatidylinositol specific phospholipase C disturbed in vitro mineralization in the tooth germ, suggesting that ALP functioning as an ectoenzyme is involved in the physiological process of tooth formation. To elucidate the effects of calcium regulating hormones and several drugs on tooth mineralization, ALP and calcium content were measured and also the tooth germ was observed histologically. The results obtained from the present study suggest that this in vitro system provides a useful tool for investigating both the direct action of drugs on tooth formation and the mechanisms of drug action.

Alkaline Phosphatase↗

PubMed1995

Alteration of bone status with ascorbic acid deficiency in ODS (osteogenic disorder Shionogi) rats.

Rats with hereditary defects in ascorbic acid (AsA) synthesis (ODS rats) subjected to AsA-deficiency for 3 weeks showed reductions of plasma alkaline phosphatase and dry and ash weights of the tibia, but no body weight alteration. In accordance with the chemical changes, bone loss and decrease of bone formation by AsA deficiency but not by malnutrition were observed in contact microradiographs of the tibia and by a tetracycline double labeling technique, respectively. The mechanical properties of femora measured by a three point-bending procedure were also altered by AsA deficiency for 3 weeks and showed decreases of 59% in toughness, 32% in strength, 32% in ductility and 22% in stiffness. The biomechanical changes by AsA deficiency were greater than the chemical changes in bone, indicating the usefulness of measuring mechanical properties as a sensitive method for the evaluation of the bone status. The second moment of the area of the femur was not changed by AsA deficiency. These results suggest that AsA deficiency in ODS rats causes marked bone loss and reduction in bone formation, which is accompanied by a greater reduction in biomechanics of the femur without causing macroarchitectural changes.

Animals↗

PubMed1993

Inhibition of in vitro mineralization in osteoblastic cells and in mouse tooth germ by phosphatidylinositol-specific phospholipase C.

We have examined the effect of phosphatidylinositol-specific phospholipase C (PIPLC) on the in vitro mineralization during cultivation of both osteoblast-like MC3T3-E1 cells and the lower molar tooth germ from mouse embryo. PIPLC not only caused the release of alkaline phosphatase (ALP) from cell membranes into the culture medium but also produced a disturbance of in vitro mineralization in MC3T3-E1 cells and in mouse embryonic tooth germ. These findings strongly suggest that ALP, functioning as an ectoenzyme, is involved in the physiological mineralization process of both bone and teeth.

Alkaline Phosphatase↗