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Biomedical subjects

H Miyata

Publications and source records attributed to H Miyata.

At least 181 records · Page 10Linked to original sources
PubMed1997

Tranilast inhibits the proliferation, chemotaxis and tube formation of human microvascular endothelial cells in vitro and angiogenesis in vivo.

1. First developed as an antiallergic drug, tranilast inhibits chemical mediator release from mast cells. In the present study, we examine the effects of tranilast on angiogenesis in vitro and in vivo and discuss the application of tranilast for angiogenic diseases. 2. Tranilast inhibited significantly the proliferation (IC50: 136 microM, 95% confidence limits: 134-137 microM) and vascular endothelium growth factor (VEGF)-induced chemotaxis (IC50: 135 microM, 95% confidence limits: 124-147 microM) of human dermal microvascular endothelial cells (HDMECs) at concentrations greater than 25 micrograms ml-1. No toxicity to HDMECs measuring by LDH release and no inhibitory effects on metalloproteinase (MMP)-2 and MMP-9 activity were observed even at 100 micrograms ml-1 (306 microM). 3. Tube formation of HDMECs cultured on the matrigel as an in vitro angiogenesis model was inhibited by tranilast in a concentration-dependent manner. The IC50 value and 95% confidence limits were 175 microM and 151-204 microM, respectively. 4. In vivo angiogenesis was induced in mice by the subcutaneous injection of matrigel containing 30 ng ml-1 VEGF and 64 micrograms ml-1 heparin. Tranilast was administered orally twice a day for 3 days. Tranilast dose-dependently suppressed angiogenesis in the matrigel and a significant change was observed at a dose of 300 mg kg-1. 5. These results indicate that tranilast is an angiogenesis inhibitor which may be beneficial for the improvement of angiogenic diseases such as proliferative diabetic retinopathy, age-related macular degeneration, tumour invasion and rheumatoid arthritis.

Animals↗
PubMed1997

Cooperative association of actin protomers and crosslinked actin oligomers in filaments at low ionic strength.

F-Actin treated with a half molar equivalent of the heterobifunctional cross-linker, 4-bromomethyl-3-nitrobenzoic acid succinimide ester (BNBA-SE), produced a population of actin oligomers containing 2-14 or more protomers and a significant amount of uncrosslinked actin protomers. The crosslinking reaction is presumed to occur between Cys 374 of one actin protomer with Lys 191 of an adjacent protomer on the genetic helix of F-actin, in a similar manner to N-maleimidobenzoic acid succinimidyl ester, which shares similar reactive groups and crosslinking dimensions. Crosslinked oligomers and uncrosslinked protomers were found to form filaments that sediment after high-speed centrifugation, even in a buffer of low ionic strength [G-buffer: 2 mM tris-hydroxymethylaminomethane (pH 8.0), 0.2 mM CaCl2, 0.2 mM ATP, 0.3 mM NaN3, 5 mM 2-mercaptoethanol] where affinity between actin protomers usually becomes weak, leading to the depolymerization of native F-actin. By performing the crosslinking reaction in the presence of an environment-sensitive fluorescent label, 6-acryloyl-2-(dimethylamino)naphthalene (acrylodan), which competes with BNBA-SE for Cys 374 of actin protomers, fluorescent, crosslinked F-actin filaments were obtained which were also stable in G-buffer. Fluorometric analysis of actin labeled with acrylodan (prodan-actin) in these depolymerization-resistant filaments suggested that the molecular environment around Cys 374 of actin protomers is similar to that of actin protomers in native F-actin in F-buffer (G-buffer plus 0.1 M KCl and 1 mM MgCl2). When G-actin labeled with acrylodan was co-polymerized with non-fluorescent crosslinked F-actin, some of the labeled actin was incorporated into filaments that were resistant to depolymerization in G-buffer. The emission spectrum of the prodan-actin in these filaments measured in G-buffer was almost identical to that of prodan-actin in native F-actin in F-buffer. Our interpretation of this result is that actin protomers are locked into an F-actin-like conformation in the depolymerization-resistant filament by the subunit interactions they make with crosslinked oligomers. We also present evidence which suggests that the depolymerization rate in G-buffer of filaments made with crosslinked oligomers is much slower than that of native actin because the ends of the depolymerization-resistant filaments are capped with crosslinked oligomers.

Actins↗
PubMed1997

Inhibitory effects of dietary protocatechuic acid and costunolide on 7,12-dimethylbenz[a]anthracene-induced hamster cheek pouch carcinogenesis.

The modifying effects of dietary exposure to two natural products, protocatechuic acid (PCA) and costunolide during the development of neoplasms in oral carcinogenesis initiated with 7,12-dimethylbenz[a]anthracene (DMBA) were investigated in male Syrian golden hamsters. All hamsters except those in the test chemical alone and control groups received DMBA (0.5%) in mineral oil to the right buccal pouch 3 times per week for 4 or 6 weeks. At 13 weeks of age, the groups exposed to DMBA were fed diet containing PCA or costunolide at a dose of 0.2 g/kg diet (200 ppm) for 17 weeks. The other groups consisted of hamsters given mineral oil alone for 6 weeks, or given 200 ppm PCA or costunolide alone, or untreated. All animals were necropsied at the termination of the experiment (week 24). PCA or costunolide significantly decreased the tumor burden (P < 0.001-P < 0.05) and the extent of dysplastic areas (%) (P < 0.001-P < 0.05). PCA significantly decreased the mean number of AgNORs/nucleus (P < 0.05). The BrdUrd-labeling index was reduced by dietary administration of test compounds, though not significantly. These results suggest that PCA and costunolide inhibited hamster buccal pouch carcinogenesis and such inhibition may be related to suppression of cell proliferation in the buccal mucosa. It was also found that telomerase activity expressed in neoplastic and preneoplastic lesions of hamster buccal pouch epithelium after DMBA treatment correlated with the histopathological degree of malignancy.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1997

Metabolic and phenotypic adaptations of diaphragm muscle fibers with inactivation.

We hypothesized that metabolic adaptations to muscle inactivity are most pronounced when neurotrophic influence is disrupted. In rat diaphragm muscle (Dia(m)), 2 wk of unilateral denervation or tetrodotoxin nerve blockade resulted in a reduction in succinate dehydrogenase (SDH) activity of type I, IIa, and IIx fibers (approximately 50, 70, and 24%, respectively) and a decrease in SDH variability among fibers (approximately 63%). In contrast, inactivity induced by spinal cord hemisection at C2 (ST) resulted in much less change in SDH activity of type I and IIa fibers (approximately 27 and 24%, respectively) and only an approximately 30% reduction in SDH variability among fibers. Actomyosin adenosinetriphosphatase (ATPase) activities of type I, IIx, and IIb fibers in denervated and tetrodotoxin-treated Dia(m) were reduced by approximately 20, 45, and 60%, respectively, and actomyosin ATPase variability among fibers was approximately 60% lower. In contrast, only actomyosin ATPase activity of type IIb fibers was reduced (approximately 20%) in ST Dia(m). These results suggest that disruption of neurotrophic influence has a greater impact on muscle fiber metabolic properties than inactivity per se.

Adaptation, Physiological↗
PubMed1997

Two types of identified ascending interneurons with distinct GABA receptors in the crayfish terminal abdominal ganglion.

More than half of the identified ascending interneurons originating in the terminal abdominal ganglion of the crayfish received inhibitory sensory inputs from hair afferents innervating the tailfan on the side contralateral to their main branches. Biochemical aspects of this transverse lateral inhibition of ascending interneurons were examined by the use of neurophysiological and pharmacological techniques. Local application of gamma-aminobutyric acid (GABA) and its agonist muscimol into the neuropil induced membrane hyperpolarization of identified ascending interneurons with an increase in membrane conductance. Because the reversal potential of inhibitory postsynaptic potential (IPSPs) in ascending interneurons elicited by the sensory stimulation and GABA injection was similar, and the sensory-stimulated IPSPs of the interneurons were blocked by GABA and muscimol application, this study strongly suggests a GABAergic nature for transverse lateral inhibition of ascending interneurons. According to the response to the GABAA antagonists bicuculline and picrotoxin, ascending interneurons were classified into two types, picrotoxin-sensitive and picrotoxin-insensitive interneurons. Identified ascending interneurons VE-1 and RO-4 showed a pharmacological profile similar to that of the classical GABAA receptor of the vertebrates. Bath application of both bicuculline and picrotoxin reversibly reduced the amplitudes of IPSPs. The other identified ascending interneurons CA-1, RO-1, and RO-2 were not affected significantly by the bath application of GABAA and GABAB antagonists, although bath application of low-chloride saline reversed the sensory-stimulated IPSPs. IPSPs of the picrotoxin-sensitive interneurons had a rather faster time course and shorter duration in comparison with those of the picrotoxin-insensitive interneurons.

Animals↗
PubMed1997

[Effects of ulinastatin on experimental ulcerative colitis in rats].

We examined the effect of ulinastatin in comparison with prednisolone (PSL) and salazosulfapyridine (SASP), well-known drugs for ulcerative colitis (UC), on two experimental UC models induced by dextran sulfate sodium (DSS) or trinitrobenzene sulfonic acid (TNB) in rats. Ulinastatin at the doses of 3000 approximately 10,000 units/kg/day (i.v.) significantly ameliorated the formation of erosion and infiltration of inflammatory cells in colonic mucosa in DSS-induced rat UC models. Moreover, ulinastatin at the dose of 10,000 units/kg/day (i.v.) significantly suppressed inflammation with ulcer in the colonic mucosa in TNB-induced rat UC models. PSL at the dose of 1 mg/kg/day (p.o.) also was as effective as ulinastatin on the above two UC models, while SASP at the dose of 100 mg/kg/day (p.o.) was less effective than ulinastatin and PSL. In addition, ulinastatin inhibited the activities of elastase and cathepsin G from human leukocytes, and it suppressed TNF alpha, IL-8 and superoxide production by rat macrophages and rabbit leukocytes in vitro. These results suggest that the suppression of inflammatory mediators produced by leukocytes is involved in the mechanism of the anti-UC action of ulinastatin.

Animals↗
PubMed1997

[The volume of white matter in cerebral palsy with MRI findings of periventricular leukomalacia].

The volume of the white matter in 5 cases of cerebral palsy (CP) of term delivery without hypoxic episodes with MRI findings of periventricular leukomalacia (PVL), was compared with that of 12 CP cases in preterm delivery manifesting typical MRI findings of PVL. It was significantly decreased in the latter. The volume for the full-term CP cases was the same as that for children born at term and developing normally. These findings suggest that the grade of perinatal brain damage is different between the full-term and pre-term CP cases, and that the intrauterine timing of fetal circulatory disturbance is different.

Brain↗
PubMed1997

Antiplatelet effect of FK633, a platelet glycoprotein IIb/IIIa antagonist, on thrombus formation and vascular patency after thrombolysis in the injured hamster carotid artery.

The antithrombotic and restenosis-preventing effects of FK633, an inhibitor of platelet aggregation via binding to the glycoprotein (GP) IIb/IIIa receptor, were studied, IC50 value of FK633 against platelet aggregation ex vivo induced by 2.5 microM adenosine diphosphate (ADP) was 5.4 x 10(-7) M as determined using hamster platelet rich plasma. The inhibitory effect was also investigated in vivo on thrombus formation at the carotid arterial wall injured by a modified catheter. As a control, the left carotid artery was injured and the time required to develop a thrombotic occlusion (3.9 +/- 1.1 min, mean +/- S.E.M., n = 18) was determined. Then, the right carotid artery of the same animal was injured while a continuous intravenous (i.v.) infusion of FK633 was administered at doses of 0 (saline), 0.1, 0.3 or 1.0 mg/kg/h. The time to occlusion was dose-dependently prolonged. In a separate experiment, 10% of the total tPA dose (0.52 mg/kg) was injected into the injured artery as a bolus and the remaining was infused i.v. at a constant rate for 30 min. When FK633 (0.3 or 1.0 mg/kg/h) was infused together with tPA, late patency of the reperfused artery was much improved as compared with that of treatment with tPA alone. Bleeding time, measured at the end of the tPA infusion, was markedly prolonged when the higher dose of FK633 (1.0 mg/kg/h) was coadministered, however coadministration of the lower dose of FK633 (0.3 mg/kg/h) was almost without prolongation on the bleeding time, despite a significant effect on the vascular patency after thrombolysis. Next, neointima formation was evaluated 2 weeks after the vascular injury. When FK633 (0.3 mg/kg/h) was continuously infused i.v. by an implanted osmotic pump for 3, 7 or 14 days after the vascular injury, the neointimal area formation was significantly suppressed in the treatment groups for 7 or 14 days. These findings suggest that FK633 inhibits platelet activation in the injured artery and improves vascular patency after thrombolysis with tPA with a concomitant suppression of neointima formation.

Animals↗
PubMed1997

Metabolic properties of the sensory neurons in the rat dorsal root ganglion.

The dorsal root ganglion (DRG) neurons were classified in the rat on the basis of their metabolic enzyme properties as determined by quantitative analysis in histochemical staining. In particular, nicotinamide adenine dinucleotide-diaphorase (NADH-d) and alpha-glycerophosphate dehydrogenase (alpha-GPD) activities were examined on two serial sections from the same neurons in the lumbar (L4) DRG. The DRG neurons were classified into three groups based on the soma diameter distribution; small, intermediate and large size DRG neurons. The NADH-d activity showed a unimodal distribution in all size groups, while the alpha-GPD activity clearly showed a bimodal distribution in the intermediate and large size neurons, but not in the small size neurons.

Animals↗
PubMed1997

Primary prevention of HTLV-1 in Japan.

The Nagasaki Prefecture, Japan (population: 1.5 million), is one of the hot endemic foci of Human T-lymphotropic virus type 1 (HTLV-1). Prevalence of HTLV-1 carriers are approximately 10% in the age group over 40 years old (40,000 individuals), approximately 10 times of the national average. Annual registry of adult T-cell leukemia (ATL) in the Prefecture is approximately 60 cases (estimated incidence: 100 cases), or a half percent of total deaths. A effective measure to control the endemic cycle of HTLV-1 has been imperative, since practical ways to prevent or control ATL are not available. A prefecture wide intervention at Nagasaki by refrain from breast-feeding blocked approximately 80% of mother-to-child transmission of HTLV-1.

Adult↗
PubMed1997

[Relationship between cholinergic symptoms caused by distigmine and the activities of serum AChE and BChE].

Distigmine is widely used for the treatment of dysuria, which is caused by various types of psychotropic medications. Distigmine, however, is also known to induce adverse cholinergic effects, such as diarrhea and salivation, with a decreased level of serum cholinesterase. We evaluated the possibility of using serum acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) as specific clinical markers for the adverse cholinergic effects of distigmine. Of the twelve patients treated with distigmine for dysuria caused by psychotropic drugs six patients presented both adverse cholinergic effects and decreased levels of serum AChE and BChE. The other six presented neither of these changes. This study suggests that the values of serum AChE and BChE may be useful markers for the manifestation of adverse cholinergic effects caused by distigmine.

Acetylcholinesterase↗
PubMed1997

[Influence for cultured renal cell growth ability by toxins of Escherichia coli].

There have been many reports indicating that Escherichia coli from pyelonephritis may exhibit several specific phenotypes. The purpose of the present study was to examine the effects of various toxins from Escherichia coli on cultured renal cell growth. alpha-hemolysin suppressed growth of Mardin Darby canine kidney (MDCK) cells in a dose-dependent manner. A low dose of O-antigen suppressed MDCK cell growth, while a high dose of the antigen increased the cell growth slightly. K:1-antigen had no effect on MDCK cell growth. Mesangial cell growth was not affected by alpha-hemolysin. A significant increase in mesangial cell growth was recognized by the O- or K:1-antigen. From these results it can be speculated that alpha-hemolysin from Escherichia coli may play a leading role, and O- or K-antigen may play a supporting role in the pathogenesis of pyelonephritis.

Animals↗
PubMed1996

Signaling from the small GTP-binding proteins Rac1 and Cdc42 to the c-Jun N-terminal kinase/stress-activated protein kinase pathway. A role for mixed lineage kinase 3/protein-tyrosine kinase 1, a novel member of the mixed lineage kinase family.

Certain small GTP-binding proteins control the enzymatic activity of a family of closely related serine-threonine kinases known as mitogen-activated protein kinases (MAPKs). In turn, these MAPKs, such as p44(mapk) and p42(mapk), referred to herein as MAPKs, and stress-activated protein kinases, also termed c-Jun N-terminal kinases (JNKs), phosphorylate and regulate the activity of key molecules that ultimately control the expression of genes essential for many cellular processes. Whereas Ras controls the activation of MAPK, we and others have recently observed that two members of the Rho family of small GTP-binding proteins, Rac1 and Cdc42, regulate the activity of JNKs. The identity of molecules communicating Rac1 and Cdc42 to JNK is still poorly understood. It has been suggested that Pak1 is the most upstream kinase connecting these GTPases to JNK; however, we have observed that coexpression of Pak1 with activated forms of Cdc42 or Rac1 diminishes rather than enhances JNK activation. This prompted us to explore the possibility that kinases other than Pak might participate in signaling from GTP-binding proteins to JNK. In this regard, a computer-assisted search for proteins containing areas of homology to that in Pak1 that is involved in binding to Rac1 and Cdc42 led to the identification of mixed lineage kinase 3 (MLK3), also known as protein-tyrosine kinase 1, as a potential candidate for this function. In this study, we found that MLK3 overexpression is sufficient to activate JNK potently without affecting the phosphorylating activity of MAPK or p38. Furthermore, we present evidence that MLK3 binds the GTP-binding proteins Cdc42 and Rac1 in vivo and that MLK3 mediates activation of MEKK-SEK-JNK kinase cascade by Rac1 and Cdc42. Taken together, these findings strongly suggest that members of the novel MLK family of highly related kinases link small GTP-binding proteins to the JNK signaling pathway.

Animals↗
PubMed1996

Direct measurement of the torsional rigidity of single actin filaments.

Flexural and torsional rigidities of actin filaments are important factors in cell motility and muscle contraction, where actin filaments serve as mechanical elements. The flexural rigidity has already been determined by directly observing the bending of individual filaments under a microscope, but measurement of the torsional rigidity has been relatively scarce and indirect, because torsion of an actin filament is difficult to visualize. This paper shows that the torsional rigidity can be measured directly by visualizing the torsional Brownian motion of a single actin filament with a novel methodology based on an optical trapping technique. Actin filaments (F-actin) were prepared by polymerizing actin monomers binding Ca2+ ion or Mg2+ ion at the high affinity site. The torsional rigidity of F-Ca(2+)-actin ((8.5(+/- 1.3)) x 10(-26) N m2) was about three times as large as that of F-Mg(2+)-actin ((2.8(+/- 0.3)) x 10(-26) N m2), whereas the flexural rigidity ((6.0(+/- 0.2)) x 10(-26) N m2) was almost independent of the kind of the bound cation. The dynamic structure of F-actin is regulated by the bound metal in an anisotropic manner. The torsional rigidities above, whether of F-Ca(2+)-actin or F-Mg(2+)-actin, are one to two orders of magnitude greater than previous experimental estimates.

Actins↗
PubMed1996

Effect of Clostridium difficile toxin B on IgE receptor-mediated signal transduction in rat basophilic leukemia cells: inhibition of phospholipase D activation.

Antigen (Ag)-stimulated phospholipase D (PLD) activation and secretion were almost abolished by pretreatment of rat basophilic leukemia (RBL)-2H3 cells for 4 h with 5 ng/ml Clostridium difficile Toxin B which is known to inhibit Rho family proteins (Rho, Cdc42, Rac). The concentration-dependent inhibition of PLD activation was well correlated with the level of glucosylation of Rho family proteins. In streptolysin O-permeabilized RBL cells, Toxin B suppressed [3H] phosphatidylbutanol (PBut) formation in response to guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S) and phorbol 12-myristate 13-acetate (PMA) by 67 and 43%, respectively. The synergistic PLD activation by GTP gamma S and PMA was also reduced by Toxin B by 67%. These results suggest that the IgE receptor-coupled PLD activation is largely mediated by Rho proteins.

Animals↗
PubMed1996

Strength and lifetime of the bond between actin and skeletal muscle alpha-actinin studied with an optical trapping technique.

The force required to break the bond between skeletal muscle actin and alpha-actinin (unbinding force) was measured at the level of individual molecules with an optical trapping technique. An actin filament, to the barbed-end of which was attached a gelsolin-coated polystyrene bead, was bound to alpha-actinin molecules adsorbed to a nitrocellulose-coated glass surface (approximately equal to 1 alpha-actinin molecule per 1 micron actin filament). The filament-bound bead was held by the optical trap and the force was applied to break the bond by pulling the bead. The unbinding force ranged from 1.4 to 44 pN. The average magnitude of the force was approximately equal to 18 pN. As the probability of the bond breakage has been suggested to be governed by the magnitude of the external force, the relationship was studied between the magnitude of the unbinding force and the time required to break the bond (unbinding time). The unbinding time ranged from approximately equal to 0.1 to approximately equal to 20 seconds, and tended to become shorter as the unbinding force became larger. The unbinding time seemed to be classifiable into two major groups: one group having a time value of 1 sec or less and the other having a time value ranging from several to 20 seconds. This suggests the existence of at least two classes of the actin-actinin bonds.

Actinin↗
PubMed1996

Ceramide inhibits IgE-mediated activation of phospholipase D, but not of phospholipase C, in rat basophilic leukemia (RBL-2H3) cells.

Ceramide, a product of sphingomyelin hydrolysis by sphingomyelinase, elicits various cellular functions and has recently been regarded as a second messenger. To investigate the role of ceramide in rat basophilic leukemia (RBL-2H3) cells, the effects of a cell-permeable analogue, N-acetylsphingosine (C2-ceramide), on Ag-mediated cellular responses were examined. C2-Ceramide inhibited Ag- or PMA-induced activation of phospholipase D (PLD), whereas Ca2+ ionophore A23187-induced PLD activation was not affected. C2-Ceramide failed to inhibit PLD activity in two different in vitro assay systems. Since PLD activity is known to be regulated by several factors, the effects of C2-ceramide on these factors were examined. We have previously reported the possible involvement of protein tyrosine kinase in Ag-mediated PLD activation. However, C2-ceramide had no effect on Ag-induced protein tyrosine phosphorylation, including mitogen-activated protein (MAP) kinases. In fura-2-loaded RBL-2H3 cells, C2-ceramide suppressed Ag-induced Ca2+ influx, leaving initial Ca2+ increase and inositol phosphate production unaffected. Western blot analysis revealed that Ag caused translocation of protein kinase C (PKC) alpha, beta 1, beta 2, delta and epsilon isozymes from cytosol to membrane fraction. Translocation of alpha, beta 1, and beta 2 isozymes was specifically prevented by C2-ceramide. Moreover, C2-ceramide suppressed Ag-induced serotonin release. In the absence of extracellular Ca2+, Ag-induced PLD activation and release reaction were greatly reduced. The inhibitory profile was nearly the same as that obtained in C2-ceramide-treated cells. These results suggest that C2-ceramide inhibits Ag-induced PLD activation and serotonin release, probably through the blockage of Ca2+ influx and translocation of Ca(2+)-dependent PKC isozymes in RBL-2H3 cells.

Animals↗