Biomedical subjects

M Ohno

Publications and source records attributed to M Ohno.

At least 289 records · Page 16Linked to original sources

PubMed1994

Topical application of AT1 receptor antagonists prevents medial and neointimal proliferation after balloon injury.

Angiotensin II plays an important role in neointima formation after vascular injury. A rat model of carotid artery injury was used i) to investigate the effect of single topical application of angiotensin II subtype 1 (AT1) receptor antagonists (CV11974, DuP753) in suppressing medial proliferation at day 2 and neointimal proliferation at day 14, and ii) to investigate the antiproliferative effects of additional application of L-arginine (a nitric oxide precursor). Drugs mixed in 25% (W/W) solutions of F127 pluronic gel were applied topically to injured vessels. Early medial proliferation of smooth muscle cells, assessed by the S-bromo-2'-deoxyuridine labelling index, was significantly suppressed by application of CV11974 (5 mg/kg), 7.5% +/- 2.2% vs. 19% +/- 3.9% in the control group. The intima/media ratio following CV11974 (10 mg/kg) or DuP753 (12.5 mg/kg) at day 14 was significantly lower than that in the control group (42% +/- 7%, 43% +/- 14%, and 123% +/- 11%, respectively). Additional application of L-arginine seemed to increase effectiveness, but was not statistically significant. In conclusion, single topical application of AT1 receptor antagonists was effective in suppressing early medial proliferation and neointima formation after balloon injury, suggesting that they may be clinically useful after angioplasty or vascular surgery.

Administration, Oral↗

PubMed1994

[A comparative study of a bioelectrical impedance method and dual energy X-ray absorptiometry for body composition analysis].

The accuracy of a bioelectrical impedance analysis (BIA) was examined by comparison with a dual energy X-ray absorptiometry (DXA) to measure the body composition. The subjects consisted of 52 patients (21-78y.) with chronic renal failure (32 cases), osteoporosis (6 cases) and diabetes mellitus with obesity (14 cases). Bioelectrical impedance between hand and leg was measured by a body composition analyzer (Model BIA-BC-3, RJL) with a four-electrode system. One pair of current supply and detector electrode was attached to the right hand and another pair was to the leg. The total fat mass measured by BIA was well correlated with the value by DXA (r = 0.959, p < 0.01). The total fat mass by BIA was also correlated with the fat mass in the arms (r = 0.863), fat mass in the legs (r = 0.911), and fat mass in the trunk (r = 0.920) by DXA. The correlation rates were p < 0.01. These correlation coefficients were lower than the correlation rate of the total fat mass measured by DXA. The total lean mass measured by BIA was well correlated with the value by DXA (r = 0.963, p < 0.01). It was correlated with the lean mass in the arms (r = 0.899), lean mass in the legs (r = 0.929), and lean mass in the trunk (r = 0.906) by DXA, but these values were lower than the correlation rate of the total lean mass obtained by DXA. These findings suggest that BIA is a safe and simple method for analyzing the body composition of the total fat mass and the total lean mass, and total body composition is reflected more clearly than regional body composition.

Absorptiometry, Photon↗

PubMed1994

Assessment of production of endothelium-derived relaxing factor (EDRF) by cultured human vascular endothelial cells based on its anti-aggregatory effect on human platelets.

In order to establish an experimental system to assess the production of endothelium-derived relaxing factor (EDRF) by cultured human umbilical vascular endothelial cells (HUVECs), the effect of endothelial cells on thrombin-induced platelet aggregation was examined. Cultured HUVECs were harvested from umbilical veins by collagenase treatment. The platelet aggregation experiments were performed using cuvettes lined with HUVECs. The cuvettes were prepared by seeding HUVECs in gelatin-coated cuvettes at a cell density of 2 x 10(5) cells/ml and culturing for 48 hours. Thrombin-induced platelet aggregation was inhibited in the presence of HUVECs. This HUVEC-dependent anti-platelet aggregatory effect was enhanced by the addition of bradykinin, which stimulates the production of EDRF, and thrombin-induced platelet aggregation was completely inhibited. Indomethacin reduced the HUVEC-dependent anti-platelet aggregatory effect. These findings suggest that this simple, new experimental system is useful in assessing the production of EDRF by HUVECs and in examining the effects of various chemicals (or agents) on EDRF production.

Biological Assay↗

PubMed1994

Molecular cloning of a novel non-receptor tyrosine kinase, HYL (hematopoietic consensus tyrosine-lacking kinase).

We identified a novel non-receptor tyrosine kinase from a human megakaryoblastic cell line, UT-7, by means of a PCR-based cloning method. The HYL gene contained a SH2 and SH3 domain and a tyrosine kinase catalytic domain. The deduced amino acid sequence of the protein encoded by this gene was most homologous to CSK (c-src kinase). This gene and CSK shared some unique structural properties such as the absence of a myristylation signal and phosphorylation sites of tyrosine residues corresponding to tyrosines 416 and 527 of chicken p60c-src. Unlike CSK, the SH3 domain of HYL was unique since the ALYDY motif was absent. Northern blot analysis revealed a 2.2 kb transcript in various myeloid cell lines but not in adult tissues except for the brain and the lung, whereas CSK mRNA was ubiquitously expressed. The expression of HYL was upregulated when these myeloid cells were differentiated by induction with phorbol myristate acetate. We named this gene, hematopoietic consensus tyrosine-lacking kinase, HYL. The HYL gene was assigned to chromosome 19 at band p13. It is suggested that HYL plays a significant role in the signal transduction of hematopoietic cells.

Amino Acid Sequence↗

PubMed1994

Comparison of effects of dihydropyridine calcium antagonists on left ventricular systolic and diastolic performance.

We compared the effects of three dihydropyridine calcium antagonists (felodipine, nifedipine and amlodipine) on left ventricular (LV) contractile performance and diastolic filling dynamics in eight conscious animals. After administering metoprolol and atropine, felodipine (25 nmol/kg i.v.) produced significant decreases in LV end-systolic pressure (PES) (109 +/- 15 vs. 88 +/- 12 mmHg, P < .05) and arterial elastance (Ea) (12.6 +/- 4.5 vs. 8.5 +/- 3.4 mmHg/ml, P < .05), whereas the heart rate was unchanged. Felodipine increased the slopes of the end-systolic P-V relation (7.4 +/- 0.9 vs. 9.9 +/- 1.0 mmHg/ml, P < .05), the dP/dtmax-end diastolic volume (VED) relation (68.1 +/- 11.2 vs. 94.9 +/- 14.3 mmHg/sec/ml, P < .05), and the stroke work (SW)-VED relation (72.1 +/- 3.1 vs. 82.8 +/- 5.2 mmHg, P < .05), and shifted all three relations to the left, indicating enhanced contractile performance. In contrast, at doses that produced equivalent reductions of PES, nifedipine (375 nmol/kg i.v.) and amlodipine (780 nmol/kg i.v.), significantly decreased the slopes of the end-systolic P-V relation, the dP/dtmax-VED relation and the SW-VED relation and shifted all three relations to the right, indicating depressed LV contractile performance. Felodipine decreased the time constant (T) of LV relaxation (32.2 +/- 5.2 to 28.8 +/- 5.2 msec, P < .05) and increased the maximum rate of early diastolic LV filling (dV/dtmax) (167 +/- 22 to 207 +/- 26 ml/sec, P < .05). Amlodipine had the opposite effect, slowing T (31.0 +/- 4.9 to 33.9 +/- 5.4 msec, P < .05) and decreasing dV/dtmax (173 +/- 39 to 154 +/- 30 ml/sec, P < .05), whereas nifedipine had no significant effects on T, PGmax or dV/dtmax. Thus, we conclude that in conscious dogs after autonomic blockade, at dosages that produced equivalent arterial vasodilation, felodipine augmented, whereas amlodipine depressed, LV contractile performance, LV relaxation and early LV filling. Nifedipine decreased LV contractile performance but had no significant effect on LV relaxation and early LV filling.

Amlodipine↗

PubMed1993

Deficits in working memory following inhibition of hippocampal nitric oxide synthesis in the rat.

In order to elucidate the roles of hippocampal nitric oxide (NO) synthesis in working and reference memory performance of rats, the effects of intrahippocampal injections of the NO synthase inhibitor, NG-nitro-L-arginine methyl ester (L-NAME), on this behavior were examined with a three-panel runway task. In the working memory task, L-NAME, injected bilaterally at 10 and 32 micrograms/side into the dorsal hippocampus, significantly increased the number of errors (attempts to pass through two incorrect panels of the three panel-gates at four choice points). This effect of intrahippocampal L-NAME (32 micrograms/side) on working memory was attenuated by concurrent injection of 100 micrograms/side L-arginine, the precursor of NO. Intrahippocampal injection of the inactive isomer D-NAME at doses up to 32 micrograms/side had no effect on the number of working memory errors. In the reference memory task, neither L-NAME nor D-NAME affected the number of errors when injected into the hippocampus at doses up to 32 micrograms/side. These results suggest that processes mediated by NO synthesis in the hippocampus are involved in working memory, but not in reference memory.

Amino Acid Oxidoreductases↗

PubMed1993

Recombinant thyrotropin stimulates cAMP formation in CHO-K1 cells expressing recombinant chorionic gonadotropin receptor.

Using CHO-K1 cells expressing rat thyrotropin-receptor (CHO-rTSH-R cells) or rat chorionic gonadotropin-receptor (CHO-rCG-R cells), we have examined their reactivity or cross-reactivity to recombinant human thyrotropin (TSH) or human chorionic gonadotropin (hCG). TSH stimulated cAMP formation in CHO-rTSH-R cells dose-dependently, and the maximum response was obtained at 1 mIU/ml. hCG also increased cAMP in the cells from 10(4) mIU/ml, and the maximum stimulation was observed at 10(6) mIU/ml. 10(7) mIU of hCG had the same potency with 1 mIU of TSH to rTSH-R. On the other hand, 10(-1) mIU/ml of hCG increased cAMP content in CHO-rCG-R cells and reached the maximum level at 10(2) mIU/ml. TSH also showed the stimulatory activity to CHO-rCG-R cells. It increased cAMP formation in CHO-rCG-R cells dose-dependently from 10 mIU/ml to 10(3) mIU/ml. Thirty mIU of TSH was calculated to be equivalent to 1 mIU of hCG to CG-R. These results indicate that TSH and hCG cross-react each other with rTSH-R or rCG-R.

Animals↗

PubMed1993

Angiotensin converting enzyme inhibitors or DuP753 prevent neointimal formation following balloon injury with single topical or multiple systemic application.

Angiotensin II plays an important role in neointimal formation after vascular injury. Our objectives were 1) to investigate the difference between angiotensin converting enzyme inhibitors (captopril, delapril) and an angiotensin II subtype 1 (AT1) receptor antagonist (DuP753) in suppressing neointimal proliferation; and 2) to investigate the antiproliferative effects of these drugs given topically to the injured vessels. All these treatments effectively prevented neointimal formation (p < 0.01). Even a single topical application of either type of drug with F127 pluronic gel to be injured vessel after ballooning is found to be significantly effective probably due to the inhibition of smooth muscle cell migration (p < 0.01). Multiple systemic application of angiotensin converting enzyme inhibitors was more effective than that of DuP753 at the same blood pressure level. The effectiveness of topical application of these drugs suggests clinical usefulness after angioplasty or vascular surgery.

Administration, Topical↗

PubMed1993

MHC class II antigen expression and T-cell infiltration in the demyelinating CNS and PNS of the twitcher mouse.

The expression of the major histocompatibility complex class II antigens (Ia) was investigated in the central and peripheral nervous systems of the twitcher mouse, an authentic murine model of globoid cell leukodystrophy (Krabbe disease) in humans. In this mutant, demyelination is noted as early as postnatal day 10 in the peripheral nerve and day 20 in the spinal cord. Emergence of Ia antigen expressing cells (Ia+ cells) was largely coincident with the initiation of demyelination, suggesting the importance of local factors for the induction of Ia antigens. Ia+ cells gradually increased in number with the progression of demyelination, but reached a plateau between postnatal days 30 and 40, then rapidly decreased despite continuous demyelination in both central and peripheral nervous systems. The only exception was in the spinal cord where Ia+ cells were numerous even at postnatal day 50. The cells expressing L3T4 antigen (L3T4+ cells), which is a mouse CD4 antigen, and the interleukin-2 receptor antigen expressing cells (IL-2R+ cells), also appeared in the regions where Ia+ cells were observed. L3T4+ cells were still abundant after P45, when Ia+ and IL-2R+ cells decreased. Combined autoradiographic and immunocytochemical studies demonstrated mitotic activity in a few Ia+ cells. However, low labeling indices of Ia+ cells in both cerebrum and sciatic nerve suggest that the increase of Ia+ cells was largely due to either induction of Ia antigens on local cells or new recruitment of Ia+ cells from the circulation. Expression of Ia antigens associated with an emergence of L3T4+ or IL-2R+ cells in the demyelinating lesions may indicate involvement of immunological responses in the pathogenesis of this metabolic demyelinating disorder. Alternatively, these apparent immunological phenomena may be tentative and non-specific local reactions to degenerating tissue components such as myelin. The mechanism(s) regulating the expression of Ia antigens in this genetic metabolic demyelinating disease is yet to be determined.

Animals↗

PubMed1993

Effect of salts on conformational change of basic amphipathic peptides from beta-structure to alpha-helix in the presence of phospholipid liposomes and their channel-forming ability.

A synthetic model peptide, H-(Leu-Al alpha-Arg-Leu)3-(Leu-Arg-Al alpha-Leu)3-OH (4(6)) can form ion channels in planar lipid bilayers by taking an amphipathic alpha-helix (Agawa, Y., Lee, S., Ono, S., Aoyagi, H., Ohno, M., Taniguchi, T., Anzai, K. and Kirino, Y. (1991) J. Biol. Chem. 266, 20218-20222). For further study of ion channels formed by this type of peptides, we planned to synthesize [Trp1]-4(6)(Ser) and [Trp12]-4(6)(Ser) in which a hydrophilic amino acid, Ser, was introduced in several positions of 4(6) instead of hydrophobic ones. This modification was expected to decrease the ability of membrane perturbation and to simplify various current levels of the channel observed for 4(6). Furthermore, additional Trp was introduced to the N-terminus or position 12 to monitor the lipid-peptide interaction. CD study showed that both peptides formed a random structure in buffer, but an alpha-helix in the presence of egg PC and a beta-structure in egg PC/egg PG (3:1). Moreover, addition of NaCl to the acidic liposomes induced the conformational transition in the peptide from beta-structure to alpha-helix. Salt-induced conformational transition in the presence of acidic liposomes was discussed in terms of membrane binding and ion-channel formation in planar lipid bilayer. Despite introduction of hydrophilic residues instead of hydrophobic residues in 4(6), the peptide showed nearly the same dye-release ability from egg PC- egg PG liposomes as 4(6). [Trp12]-4(6)(Ser) was able to form cation-selective ion channels with two levels of conductance (mainly 250 and occasionally 125 pS) in asolectin planar lipid bilayer, suggesting that appropriate orientation of hydrophobic and hydrophilic residues in amphipathic peptide can simplify channel current levels.

Amino Acid Sequence↗

PubMed1993

Impairment of working memory induced by scopolamine in rats with noradrenergic DSP-4 lesions.

In a working memory task with a three-panel runway set-up, DSP-4 (N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine), a noradrenergic neurotoxin, at 50 mg/kg i.p. given 14 days before testing, had no effect on the number of errors (attempts to pass through two incorrect panels of the three-panel gates at four choice points). Working memory errors were significantly increased by scopolamine, a muscarinic receptor antagonist, at 0.32 mg/kg i.p. given 20 min before testing, whereas errors were not affected by the 0.1 mg/kg dose. In rats with noradrenergic DSP-4 lesions, 0.1 mg/kg scopolamine significantly increased the number of working memory errors. However, DSP-4 at 50 mg/kg and scopolamine at 0.1 mg/kg whether they were administered alone or in combination had no effect on reference memory errors. These results suggest that noradrenergic deficits aggravate the working memory impairment induced by blockade of muscarinic receptors.

Animals↗

PubMed1993

Accelerated evolution of Trimeresurus flavoviridis venom gland phospholipase A2 isozymes.

Six Trimeresurus flavoviridis (Habu snake) venom gland phospholipase A2 (PLA2) isozyme genes were found to consist of four exons and three introns and to encode proteins of 138 amino acid residues, including the signal sequence of 16 amino acid residues. Comparison of the nucleotide sequences showed that the introns are much more homologous than the protein-coding regions of exons except for the signal peptide-coding region of the first exon. The numbers of nucleotide substitutions per site (KN) for introns are approximately one-fourth of the numbers of nucleotide substitutions per synonymous site (KS) for the protein-coding regions, indicating that the introns are unusually conserved. The absence of an apparent functional role for the introns suggests that the protein-coding regions, except for the signal peptide-coding domains, have evolved at greater substitution rates than introns. The fact that the numbers of nucleotide substitutions per nonsynonymous site (KA) are close to or larger than KS values for relevant pairs of genes revealed that Darwinian-type accelerated substitutions have occurred in the protein-coding regions or exons. This is compatible with the presence of PLA2 species with diverse physiological activities in the venom.

Amino Acid Sequence↗

PubMed1993

Receptor-mediated specific biological activity of a beta-amyloid protein fragment for NK-1 substance P receptors.

A beta-amyloid protein fragment AP21-35 (tetradeapeptide with amino acid residues 21-35) was found to be a highly selective agonist of substance P receptors (NK-1) among three tachykinin receptor subtypes. This peptide fragment contracted the smooth muscle preparations of guinea pig ileum in a dose dependent manner (EC50 = 22 microM). This activity was completely reversed by CP-96,345-1, a specific antagonist of NK-1 receptors, whereas atropine for NK-3 had no effect. The peptide was inactive in rat vas deferens which contains predominantly NK-2 receptors. These results indicated that AP21-35 is a highly selective agonist for NK-1 receptors. In the radio-ligand receptor binding assay using [125I]-Bolton-Hunter substance P to membranes from guinea pig ileum, the fragment exhibited a distinct dose-response curve (IC50 = 11 microM). All the present data strongly suggest that beta-amyloid protein function as a peptide ligand of substance P receptors with some pathophysiologic activities.

Amino Acid Sequence↗

PubMed1993

Enhancement of thrombin receptor activation by thrombin receptor-derived heptapeptide with para-fluorophenylalanine in place of phenylalanine.

Thrombin receptor-derived peptide SFLLRNP (one-letter amino acid code) which corresponds to the N-terminal heptapeptide of tethered ligand is able to activate thrombin receptor and to stimulate the phosphoinositide (PI) turnover. The replacement of Phe-2 by Ala eliminated this activity completely, showing the crucial role of the Phe-phenyl group in receptor activation. It was found that substitution of para-fluorophenylalanine ((p-F)Phe) for Phe-2 enhanced several times the PI-turnover activity of SFLLRNP. This is the first example to date of a substitution with one order of magnitude greater increase in receptor activation. The Phe-2/Tyr substitution diminished the activity drastically (almost 2% of SFLLRNP), indicating the importance of hydrophobicity of Phe2-phenyl. The Phe-2/Leu substitution, however, diminished also the activity (less than 2% of SFLLRNP). These results suggested that highly specific hydrophobic interaction exists between Phe-2 of the tethered ligand and its binding site in thrombin receptor.

Amino Acid Sequence↗

PubMed1993

Water-soluble chymotrypsin specific inhibitors containing arginine.

In an attempt to develop water-soluble, potent inhibitors for chymotrypsin, structurally rigid dipeptides containing arginine were designed and synthesized. The dipeptide H-D-Arg-Phe-NHBzl inhibited chymotrypsin very strongly (Ki = 5.9 microM). The dipeptide with the inverse sequence, H-D-Phe-Arg-NHBzl, was also a moderate inhibitor for chymotrypsin with Ki of 240 microM. In spite of the presence of arginine in these dipeptides, they inhibited trypsin only weakly, indicating that they are highly specific for chymotrypsin. High resolution 1H-NMR (400-MHz) indicated that these dipeptides can make a strong intramolecular hydrophobic interaction between Arg-beta, gamma, delta-methylenes and Phe-phenyl, producing a rigid hydrophobic core which interacts with the chymotrypsin S2 site. Since these dipeptides are easily soluble in water, they are regarded as the sophisticated and effective inhibitors for chymotrypsin.

Amino Acid Sequence↗

PubMed1993

Working memory deficits following muscarinic blockade combined with depletion of brain somatostatin in rats.

In a working memory task with three-panel runway paradigm, cysteamine, a depletor of somatostatin, at 100 or 200 mg/kg i.p. given 24 h before testing, had no effect on the number of errors (attempts to pass through two incorrect panels of the three panel-gates at four choice points). Cysteamine at 100 mg/kg caused a significant reduction in somatostatin-like immunoreactivity in the rat brain, including the hippocampus and cerebral cortex. Working memory errors were significantly increased by scopolamine, a muscarinic receptor antagonist, at 0.32 mg/kg i.p. given 20 min before testing, whereas errors were not affected by the 0.1 mg/kg dose. Combined administration of 100 mg/kg cysteamine and 0.1 mg/kg scopolamine significantly increased the number of working memory errors. However, cysteamine at 100 mg/kg and scopolamine at 0.1 mg/kg had no effect on reference memory errors, whether they were administered alone or in combination. These results suggest that depletion of brain somatostatin aggravates working memory deficits induced by blockade of muscarinic receptors.

Animals↗

PubMed1993

Working memory deficits induced by intrahippocampal administration of 8-OH-DPAT, a 5-HT1A receptor agonist, in the rat.

In a test of working memory using a three-panel runway task, the 5-HT1A receptor agonist, 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT), at 1.0 mg/kg i.p. significantly increased the number of errors (attempts to pass through two incorrect panels of the three panel gates at four choice points), but it had no effect on errors in a test of reference memory. Intrahippocampal injection of 8-OH-DPAT at 10 micrograms/side also significantly increased the number of working memory errors, without affecting that of reference memory errors. The effect of intrahippocampal 8-OH-DPAT (10 micrograms/side) on working memory was blocked by the 5-HT1A receptor antagonist, (-)-propranolol, at 5 mg/kg i.p. These results suggest that activation of hippocampal 5-HT1A receptors impairs working memory, but does not affect retention of reference memory.

8-Hydroxy-2-(di-n-propylamino)tetralin↗

PubMed1993

Thyrotropin receptor in non-thyroid tissues.

Thyrotropin receptor (TSH-R), the main target for the autoantibody in Graves' disease, has been thought to be a thyroid-specific protein. However, we successfully obtained the cDNA fragments of TSH-R from rat retro-orbital tissues and adipose tissues by using polymerase chain reaction methods. Sequencing has revealed that the nucleotide sequence of the cDNAs from these non-thyroid tissues was identical to that from the thyroid. TSH-R peptide antibody, which recognizes rat TSH-R, stained a 104 kDa protein from the retro-orbital tissues and the adipose tissues. The band was not detected with the antibody preabsorbed with the peptide. These results suggest that the message is translated to make a TSH-R protein even in these non-thyroid tissues.

Animals↗