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Biomedical subjects

H Teraoka

Publications and source records attributed to H Teraoka.

At least 19 recordsLinked to original sources
PubMed2005

Mechanism of cell death in inflamed superficial digital flexor tendon in the horse.

The aim of the present study was to clarify the presence and determine the role of apoptosis in the degenerative process of the superficial digital flexor tendon (SDFT) in the horse. Samples were obtained from normal and inflamed SDFTs of horses. To detect apoptosis and to identify apoptotic cells, the samples were subjected to immunohistochemical labelling and Western blot analysis. Although a large number of cells in degenerate areas showed positive reactions with caspase-3 and single stranded DNA antibodies, cells in normal tendon samples showed very weak reactions. Excessive apoptosis was confirmed by the results of Western blot analysis, which showed a significant increase in activated caspase-3 protein in the inflamed SDFTs, suggesting that apoptosis occurred in the tendinocytes via a caspase-3-dependent pathway. This is the first report of excessive apoptosis in inflamed SDFT of the horse. The results indicate that apoptosis may play an important role in the process of degeneration of the tendon as well as other tissues.

Animals↗
PubMed2004

Hedgehog and Fgf signaling pathways regulate the development of tphR-expressing serotonergic raphe neurons in zebrafish embryos.

Serotonin (5HT) plays major roles in the physiological regulation of many behavioral processes, including sleep, feeding, and mood, but the genetic mechanisms by which serotonergic neurons arise during development are poorly understood. In the present study, we have investigated the development of serotonergic neurons in the zebrafish. Neurons exhibiting 5HT-immunoreactivity (5HT-IR) are detected from 45 h postfertilization (hpf) in the ventral hindbrain raphe, the hypothalamus, pineal organ, and pretectal area. Tryptophan hydroxylases encode rate-limiting enzymes that function in the synthesis of 5HT. As part of this study, we cloned and analyzed a novel zebrafish tph gene named tphR. Unlike two other zebrafish tph genes (tphD1 and tphD2), tphR is expressed in serotonergic raphe neurons, similar to tph genes in mammalian species. tphR is also expressed in the pineal organ where it is likely to be involved in the pathway leading to synthesis of melatonin. To better understand the signaling pathways involved in the induction of the serotonergic phenotype, we analyzed tphR expression and 5HT-IR in embryos in which either Hh or Fgf signals are abrogated. Hindbrain 5HT neurons are severely reduced in mutants lacking activity of either Ace/Fgf8 or the transcription factor Noi/Pax2.1, which regulates expression of ace/fgf8, and probably other genes encoding signaling proteins. Similarly, serotonergic raphe neurons are absent in embryos lacking Hh activity confirming a conserved role for Hh signals in the induction of these cells. Conversely, over-activation of the Hh pathway increases the number of serotonergic neurons. As in mammals, our results are consistent with the transcription factors Nk2.2 and Gata3 acting downstream of Hh activity in the development of serotonergic raphe neurons. Our results show that the pathways involved in induction of hindbrain serotonergic neurons are likely to be conserved in all vertebrates and help establish the zebrafish as a model system to study this important neuronal class.

Animals↗
PubMed2003

The influence of the expanding HIV genetic diversity on molecular diagnosis in the Philippines.

Since the discovery in the Philippines of the first AIDS case in 1984, several subtypes of HIV-1 have been discovered. From the persons diagnosed in the early 1980s only subtype B was found and thereafter other subtypes, C, D, E, and F were also identified although HIV was not particularly prevalent at that time. In this paper, we determine whether the rapid expansion of genetic diversity will influence molecular diagnosis by polymerase chain reaction (PCR). First, we determine HIV-1 subtype on env (V3) and gag (p24) gene as a means of rapid genetic diversity. Secondly, we tried to analyse and identify homologous regions of gag (p24) gene of HIV genome for diagnostic purposes of designing primers. Out of 46 samples analysed, six subtypes were classified based on gag and env gene subtyping namely: 33 subtype B/B (71.2%), nine subtype A/E and one each subtype C/C, A/B and G/A (2.2% each). As a result, occurrence of non-subtype B and inter-subtype recombinant contributed to expanding genetic diversity. Based on inter- and intra-subtype gag alignment, oligonucleotides (>10 bases in length) could be easily selected as a universal primer to produce the PCR product composed of more than 100bp. This indicates that the PCR technology can be safely used with limited length of primers for the diagnosis of HIV infection in this country.

Base Sequence↗
PubMed2002

kdsA mutations affect FtsZ-ring formation in Escherichia coli K-12.

No one has, as yet, addressed the relationship between the nature of the outer membrane and cell division. kdsA encodes 3-deoxy-D-manno-octulosonic acid (KDO) 8-phosphate synthetase which catalyses the first step in the synthesis of KDO, the linker between lipid A and oligosaccharide of lipopolysaccharide (LPS). Seven temperature-sensitive mutants containing missense mutations in kdsA were affected in the production of KDO and all mutants stopped dividing at 41 degrees C and formed filaments with either one or no FtsZ ring. All observed defects were reversed by the plasmid-borne wild-type kdsA gene. Western blotting analysis, however, demonstrated that the amount of FtsZ protein was not affected by the mutation. The mutants were more susceptible to various hydrophobic materials, such as novobiocin, eosin Y and SDS at 36 degrees C. Methylene blue, however, restored kdsA mutant growth. Plasmid-borne wild-type msbA, encoding a lipid A transporter in the ABC family, partially suppressed kdsA mutation. A mutation of lpxA, functioning at the first stage in lipid A biosynthesis, inhibited both cell division and growth, producing short filaments. These results indicate that the instability of the outer membrane, caused by the defect in KDO biosynthesis, affects FtsZ-ring formation.

Aldehyde-Lyases↗
PubMed2001

5-HT(7) receptor and beta(2)-adrenoceptor share in the inhibition of porcine uterine contractility in a muscle layer-dependent manner.

To compare the inhibition of uterine contractility mediated by beta-adrenoceptors and 5-HT(7) receptors, the effects of catecholamines and 5-HT on spontaneous contractions were examined in longitudinal and circular muscles isolated from three different regions (cornu, corpus and cervix) of the non-pregnant proestrus porcine uterus. In addition, the distribution of beta-adrenoceptors between muscle layers was characterized by means of adenylate cyclase activity assay, cyclic AMP assay and [(3)H]dihydroalprenolol binding studies. In the cornu, isoprenaline, adrenaline and noradrenaline inhibited the spontaneous contraction of longitudinal and circular muscles but longitudinal muscle was more sensitive to catecholamines than was circular muscle. The inhibitory response to isoprenaline was antagonized by propranolol (300 nM) or (+/-)-1-[2,3-(dihydro-7-methyl-1H-inden-4-yl)oxy]-3-[(1-methylethyl)amino]-2-butanol (ICI 118,551; 100 nM). The rank order of potency was isoprenaline > or =adrenaline > noradrenaline. The beta(2)-adrenoceptor-selective agonist, clenbuterol, was more potent than xamoterol (beta(1)-selective) and (+/-)-4-[2-[(2-(3-chlorophenyl)-2-hydroxyethyl)amino]propyl]phenoxyacetic acid (BRL 37344; beta(3)-selective) to inhibit the spontaneous contraction of longitudinal muscles. Isoprenaline increased adenylate cyclase activity in both muscle layers, but the activity in the longitudinal muscle was greater than that in the circular muscle. Cyclic AMP production by isoprenaline was also more conspicuous in the longitudinal muscle than in the circular muscle. Although both muscle layers contained a single class of [3H]dihydroalprenolol binding sites with similar K(d) values (longitudinal muscle, 3.1+/-0.94 nM, n=4; circular muscle, 2.4+/-0.73 nM, n=4), B(max) in the longitudinal muscle (175.7+/-32.8 fmol/mg protein, n=4) was significantly higher than that in the circular muscle (53.1+/-5.1 fmol/mg protein, n=4). As previously reported [Br. J. Pharmacol. 130 (2000) 79], 5-HT also inhibited the spontaneous contraction of both muscle layers from the cornu and the 5-HT(7) receptor antagonist, 2a-[4-(4-phenyl-1,2,3,6-tetrahydropyridyl)butyl]-2a,3,4,5-tetrahydro-benzo[cd]indol-2(1H)-one (DR4004; 100 nM, n=4) blocked the 5-HT-induced inhibition of spontaneous contractions in the circular muscles, and reversed the less marked inhibition in the longitudinal muscles. In other regions of the uterus (corpus and cervix), 5-HT inhibited the spontaneous contraction of the circular muscles but contracted the longitudinal muscle strips. On the other hand, isoprenaline caused muscle layer-dependent inhibition (longitudinal muscle > circular muscle) in both regions, and the responsiveness tended to increase toward the cervix. In conclusion, beta(2)-adrenoceptors are present heterogeneously in the porcine uterus (longitudinal muscle > circular muscle) and share the inhibition of uterine contractility with 5-HT(7) receptors in a layer-dependent manner (longitudinal muscle: beta(2)-adrenoceptors, circular muscle: 5-HT(7) receptors).

Adenylyl Cyclases↗
PubMed2001

Enhanced VEGF production and decreased immunogenicity induced by TGF-beta 1 promote liver metastasis of pancreatic cancer.

TGF-betas are multifunctional polypeptides that regulate cell growth and differentiation, extracellular matrix deposition, cellular adhesion properties, angiogenesis and immune functions. In this study, we investigated the effect of TGF-beta1 on liver metastasis and its mechanism by using human pancreatic cancer cell lines Panc-1, Capan-2, and SW1990. Capan-2 and SW1990 cells demonstrated enhanced liver metastatic potential by in vivo splenic injection with TGF-beta1. Consequently, we examined the role of TGF-beta1 on in vitro angiogenesis and received cytotoxicity by peripheral blood mononuclear leukocytes (PBMLs). While TGF-beta1 slightly decreased cell proliferation, it also upregulated VEGF production in all cancer cells examined. The binding of PBMLs to cancer cells and cancer cell cytotoxicity during co-culture with PBMLs were remarkably decreased by treatment with TGF-beta1. Panc-1 cells revealed no liver metastasis despite their high immunogenetic and angiogenetic abilities, which was attributed to a lack of expression of the cell surface carbohydrates that induce attachment to endothelial cells. We concluded that the presence of TGF-beta1 in the microenvironment of tumour site might play an important role in enhancing liver metastasis of pancreatic cancer by modulating the capacity of angiogenesis and immunogenicity.

Animals↗
PubMed2001

2, 3, 7, 8-tetrachlorodibenzo-p-dioxin induces apoptosis in the dorsal midbrain of zebrafish embryos by activation of arylhydrocarbon receptor.

Neurotoxic effects of 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin (TCDD) has not been fully elucidated, despite the known potent agonist of arylhydrocarbon receptor (AhR), which activation induces cytochrome P450 1A and several representative toxicities of halogenated aromatic hydrocarbons. In the present study, the effects of TCDD on cell death in zebrafish embryos (Danio rerio) during the early stage of development were investigated. As shown by terminal transferase-mediated nick-end-labeling staining, TCDD exposure significantly increased the occurrence of pycnotic cell death (PCD), especially in the dorsal midbrain (optic tectum). The ultrastructures of these pycnotic cells showed apoptotic features such as condensation and cleavage of chromatin. TCDD-induced PCD was mimicked by beta-naphthoflavone (AhR agonist), and inhibited by alpha-naphthoflavone (AhR antagonist). These results suggest that AhR activation can induce apoptosis in the central nervous system during development.

Animals↗
PubMed2001

Degradation of human brain natriuretic peptide (BNP) by contact activation of blood coagulation system.

Brain natriuretic peptide (BNP) and atrial natriuretic peptide (ANP) were added to venous blood samples from healthy volunteers, and incubated in tubes made of various materials. The residual immunoreactivity was measured with radioimmunoassay for BNP and ANP. In blood samples stored in glass tubes, immunoreactivity of ANP was more stable than that of BNP. In siliconized glass or PET tubes, however, BNP immunoreactivity was more stable than ANP. The activation of blood coagulation factors was evaluated from the kallikrein activity in plasma. Kallikrein activity was increased in plasma stored in glass tube while it was negligible in plasma stored in siliconized glass or PET tubes. In kaolin-activated plasma, more rapid BNP degradation and higher kallikrein activity were observed. Our results indicated that the blood coagulation factors, especially kallikrein, played an important role in digestion of BNP.

Blood Coagulation↗
PubMed2001

Inhibitory effects of endogenous dopaminergic neurotoxin, norsalsolinol on dopamine secretion in PC12 rat pheochromocytoma cells.

Naturally occurring neurotoxins, 6,7-dihydroxy-1,2,3,4-tetrahydroisoquinolines (DHTIQs), thought to be the causative agents of Parkinsonism. DHTIQs including norsalsolinol have been found in the mammalian central nervous system. Norsalsolinol can be formed by a non-enzymatic Pictet-Spengler condensation reaction between dopamine and formaldehyde, and has been detected in the urine of Parkinsonian patients. However, the effects of DHTIQs on the secretion of dopamine, as well as other neurotransmitters, are not well understood. This study investigated the effects of norsalsolinol on dopamine secretion from nerve growth factor-differentiated PC12 cells. Norsalsolinol (1-100 microM) pretreatment suppressed both ATP (100 microM)- and K(+) (50 mM)-induced dopamine secretion from PC12 cells in a concentration-dependent fashion, but did not affect basal dopamine secretion. In beta-escin-permeabilized PC12 cells, norsalsolinol pretreatment suppressed Ca(2+) (pCa=4-8)-induced dopamine secretion, but did not inhibit the secretagogue-induced change in intracellular Ca(2+) concentration. These results suggest that norsalsolinol causes the inhibition of secretagogue-induced dopamine secretion from PC12 cells without altering intracellular Ca(2+) concentration. Inhibition of dopamine secretion by norsalsolinol may also be involved in postural abnormality in Parkinson's disease.

Animals↗
PubMed2001

Ca(2+) signaling in porcine duodenal glands by muscarinic receptor activation.

The duodenal glands have been thought to play an important role in defense against proximal duodenal ulcer; however, the secretory mechanisms of these glands remain to be determined. In isolated duodenal acinar cells of the pig, we investigated the effects of ACh on intracellular Ca(2+) concentration ([Ca(2+)](i)) and on membrane currents with fura 2 fluorometry and the patch clamp technique. ACh caused a transient increase in [Ca(2+)](i), and the increase was markedly inhibited by atropine or 4-diphenylacetoxy-N-methylpiperidine methiodide but not by hexamethonium, pirenzepine, or methoctramine. The expression of mRNA for the M(3) subtype far exceeded that for either M(1) or M(2) as revealed by real-time quantitative PCR and in situ hybridization. The rise in [Ca(2+)](i) evoked by ACh was largely inhibited by thapsigargin but slightly affected by extracellular Ca(2+) deprivation. Caffeine had no effect on [Ca(2+)](i). ACh elicited Ca(2+)-dependent K(+) currents, a finding similar to the response to inositol 1,4,5,-trisphosphate applied intracellularly. These results suggest the presence of M(3) receptors linked to Ca(2+) release in porcine duodenal glands.

Adenosine Triphosphatases↗
PubMed2001

TGF-beta1 promotes liver metastasis of pancreatic cancer by modulating the capacity of cellular invasion.

We investigated the effect of TGF-beta1 on liver metastasis of pancreatic cancer using surgical specimens of pancreatic cancer and human pancreatic cancer cell lines Capan-2 and SW1990. Immunostaining of TGF-beta1 showed that TGF-beta1 positivity was significantly related to venous invasion and tumor staging, and also relatively associated with liver metastasis. Cellular invasion and protease production of MMP-2 and u-PA, and in vivo liver metastasis were significantly enhanced after treatment of cells with TGF-beta1. These findings suggest that TGF-beta1 might play an important role in enhancing liver metastasis of pancreatic cancer.

Adenocarcinoma↗
PubMed2000

Hemin pretreatment ameliorates aspects of the nephropathy induced by mercuric chloride in the rat.

Mercuric chloride (HgCl(2)) is known to be a nephrotoxicant. When HgCl(2) is administered into rats, acute renal failure (ARF) is induced. Heme oxygenase-1 (HO-1) is antioxidative enzyme and is known to play a protective role against the oxidative injury. To elucidate the cytoprotective role of HO-1 against the nephrotoxicant-induced ARF, we examined the effect of hemin, HO-1 inducer, on HgCl(2)-induced ARF. Subcutaneous administration of hemin (30 mg/kg body weight) into rats once a day for two successive days obviously induced HO-1 protein in the kidneys at 24 h after the last injection. Under this situation, when HgCl(2) (1 mg/kg body weight) was intraperitoneally injected into rats at 24 h after the last injection of hemin improved the serum creatinine (SCr) and blood urea nitrogen (BUN) levels, markers for renal injury, at 24 h after the HgCl(2) injection as compared with the control rats without hemin pretreatment (HgCl(2) treatment alone). This result was further confirmed by histopathological analysis. These findings strongly suggest that the preinduction of HO-1 ameliorates the subsequent HgCl(2)-induced acute renal injury.

Acute Kidney Injury↗
PubMed2000

Cell death and cell-cycle arrest induced by incorporation of [3H]thymidine into human haemopoietic cell lines.

PURPOSE: To investigate the influence of [methyl-3H]thymidine ([3H]Tdr) incorporated into human haemopoietic cell lines. MATERIALS AND METHODS: HL-60, Molt-4, Jurkat, Raji and SKW6-CL4 cells were incubated in the presence of [3H]Tdr. Cell proliferation, cell viability, DNA fragmentation and expression of caspase-3 and Bcl-2 families were examined. The cell-cycle of HL-60 was analysed using flow cytometry. RESULTS: In HL-60, Molt-4 and Jurkat, cell death was accompanied by DNA nucleosomal fragmentation and activation of caspase-3. In Raji and SKW6-CL4, it was accompanied by neither. Protein levels of Bcl-2 and Bad in HL-60 and Molt-4 did not significantly change, and that of Bax was decreased after a 3-day incubation. HL-60 incubated in the presence of 74 or 185 kBq/ml [3H]Tdr arrested at G2/M phase, and then underwent apoptosis. In 7.4 kBq/ml, the cell-cycle progressed after the delay in S-phase. CONCLUSIONS: Two different modes of cell death were observed when [3H]Tdr was incorporated into the human haemopoietic cell lines. Incorporation into HL-60 cells resulted in delay of S-phase progression, arrest at G2/M and apoptosis.

Blotting, Western↗
PubMed2000

Genetic risk factors for coronary artery spasm: significance of endothelial nitric oxide synthase gene T-786-->C and missense Glu298Asp variants.

BACKGROUND: We recently identified two endothelial nitric oxide synthase (eNOS) gene polymorphisms, Glu298Asp and T-786-->C, which are independently associated with coronary spasm. eNOS gene intron 4b/a polymorphism is also reported to be involved in smoking-dependent coronary artery disease. The genetic linkage among these polymorphisms remains unknown. Also, it is unclear which variant is most responsible for coronary spasm. In the present study, we first examined the genetic linkage among these three variants. Next, we studied the risk factors of coronary spasm by using all significant genetic and conventional risk factors in a large-scale study. METHODS: The genotype and allele frequencies for the T-786-->C, intron 4b/a, and Glu298Asp variants were assessed in 423 randomly selected DNA samples to examine their genetic linkages. The relative capacities of all risk factors to predict coronary spasm were then analyzed using multiple logistic regression in 201 patients with coronary spasm and 345 volunteers. RESULTS: Comparison of allele frequencies revealed that the eNOS intron 4a allele was significantly linked to the T-786-->C mutation (P < 0.00001), whereas there was not a linkage between the intron 4a allele and the Glu298Asp variant (P = 0.1437) or between the Glu298Asp variant and the T-786-->C mutation (P = 0.1996). Multiple logistic regression revealed that the most predictive independent risk factor for coronary spasm was the T-786-->C mutation (P < 0.001), followed by cigarette smoking (P < 0.001), hypertension (P = 0.004), and the Glu298Asp variant (P = 0.028). CONCLUSIONS: We found that the T-786-->C mutation and the intron 4a allele are in linkage disequilibrium. We previously showed that the T-786-->C mutation reduced eNOS gene promoter activity. In that context, our results strongly suggest that the T-786-->C mutation underlies the functional characteristics of the intron 4a allele. Further, multiple logistic regression analysis revealed that the T-786-->C mutation is the most predictive risk factor for coronary spasm, followed by cigarette smoking. Given that those effects are potentially additive, patients carrying the eNOS gene variants should be strongly cautioned against smoking.

Adult↗
PubMed1999

Development of a sensitive enzyme-linked immunosorbent assay for eotaxin and measurement of its levels in human blood.

The CC chemokine eotaxin is potent eosinophil-selective chemoattractant, and it is thought that the function of eotaxin is closely related to the recruitment of eosinophils in certain inflammatory reactions. In order to learn more about the biological role of this molecule, we have developed a new sandwich ELISA method to measure human eotaxin using two monoclonal anitibodies and purified recombinant eotaxin as a standard. The minimal detectable concentration of eotaxin in this assay was 1.5 pg/ml, and the working range was 3.1--200 pg/ml with low CVs (< 10%). Both within- and between-run precision levels were less than 6.7% of the CVs. The dilution curves of two serum and two spiked plasma samples showed good linearity and the recovery range was 92.8--103.3%. No cross-reactivity was found with other similar chemokines. MCP-1, MCP-2, MCP-3, MCP-4, eotaxin-2 and RANTES. This assay was sensitive enough to measure the circulating eotaxin levels of healthy volunteers. However, the eotaxin levels in serum samples (mean+/-SD; 68.6+/-13.4 pg/ml, n=15) were significantly higher than those in matched plasma samples (19.2+/-5.4 pg/ml) separated from blood collected in tubes containing EDTA. Kinetic studies revealed that the eotaxin levels in serum markedly increased depending on the elapsed time before separation from blood cells, but such changes in EDTA-plasma were negligible up to 4 h at 25 degrees C. Our new ELISA is an accurate and useful method for quantifying human eotaxin in blood and demonstrates that the process of preparing blood samples affects the measurement of the eotaxin levels.

Chemokine CCL11↗
PubMed1999

A murC gene from coryneform bacteria.

The upstream flanking region of the ftsQ and ftsZ genes of Brevibacterium flavum MJ233, which belongs to the coryneform bacteria, was amplified by the inverse polymerase chain reaction method and cloned in Escherichia coli. Complementation analysis of E. coli mutant with a defective cell-wall synthesis mechanism with the cloned fragment and its DNA sequencing indicated the presence of the murC gene, encoding UDP-N-acetylmuramate:L-alanine ligase involved in peptidoglycan synthesis, just upstream from the ftsQ gene. The B. flavum murC gene could encode a protein of 486 amino acid residues with a calculated molecular mass of 51 198 Da. A 50-kDa protein was synthesized by the B. flavum murC gene in an in vitro transcription/translation system using E. coli S30 lysate. These results indicate that the genes responsible for cell-wall synthesis and cell division are located as a cluster in B. flavum similar to the E. coli mra region.

Amino Acid Sequence↗
PubMed1999

Stability of brain natriuretic peptide (BNP) in human blood samples.

Stability of immunoreactivity of human brain natriuretic peptide (BNP) in blood samples was investigated. After storage of the whole blood samples in the blood collecting tubes made of glass or polyethylene terephthalate (PET), residual immunoreactivity of BNP in the plasma was measured by sandwich radioimmunoassay for human BNP. BNP in the blood samples collected in the PET tubes were kept more stable than that in the glass tubes. The results suggested that commercially available PET tubes would enable more accurate BNP values and this would also help to simplify the sample preparation.

Humans↗