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Biomedical subjects

H Nishio

Publications and source records attributed to H Nishio.

At least 145 records · Page 8Linked to original sources
PubMed1995

Ca(2+)-independent fusion of secretory granules with phospholipase A2-treated plasma membranes in vitro.

The fusion of secretory granules with plasma membranes prepared from rat parotid gland was studied in vitro to clarify the mechanism of exocytosis. Fusion of the granules with plasma membranes was measured by a fluorescence-dequenching assay with octadecyl rhodamine B, and release of amylase was also measured to confirm the fusion as a final step of the secretory process. Plasma membranes that had been pretreated with porcine phospholipase A2 (PLA2) in the presence of 20 microM Ca2+ fused with the granules within 30 s, and induced amylase release by reacting with the membranes of granules, whereas without this pretreatment they had no significant effect. The fusion process accompanied by amylase release was induced in the presence of 10 mM EGTA, and therefore was apparently Ca(2+)-independent. On the other hand, the presence of EGTA or 100 microM quinacrine, an inhibitor of PLA2, during treatment of plasma membranes with PLA2 inhibited their fusogenic activity, suggesting the importance of activation of PLA2. Arachidonic acid and linoleic acid were released from the plasma membranes during the PLA2 treatment. The presence of albumin, an adsorbent of fatty acids, during the treatment also inhibited the activity. Pretreatment of the membranes with arachidonic acid or linoleic acid did not have any effect, but the presence of exogenously added arachidonic acid during PLA2 treatment enhanced the membrane-fusion-inducing effect of PLA2. Pretreatment of the membranes with lysophosphatidylcholine induced fusogenic activity. These findings suggest that the conformational change in the plasma-membrane phospholipids induced by PLA2 and the presence of arachidonic acid or linoleic acid produced by PLA2 are important in the process of fusion of secretory granules with the plasma membranes of rat parotid acinar cells and that the fusion process itself is independent of Ca2+.

5,8,11,14-Eicosatetraynoic Acid↗
PubMed1995

pp60c-src expression in rat spermatogenesis.

A fractionation of the membrane extract of rat testes revealed the existence of pp60c-src kinase activity. The expression of pp60c-src was examined in the developing rat testes. The immunecomplex kinase assay using a monoclonal antibody specific to pp60c-src (mAb327) showed that the expression of pp60c-src kinase activity increased during the development of rat testes and declined in the adult. The increase in pp60c-src kinase activity observed during the development of rat testes was accompanied with an increase in the amount of pp60c-src protein. The peak period in the increase of pp60c-src kinase activity well coincided with the timing, when the spermatogenesis by meiosis just began. The immunohistochemical staining of pp60c-src in rat testes demonstrated that pp60c-src is most abundantly expressed in the spermatids which are the spermatogenic cells in the post-meiotic phase of the spermatogenesis. These findings strongly suggest that pp60c-src is a developmentally regulated gene product which is involved in rat spermatogenesis.

Amino Acid Sequence↗
PubMed1995

Role of calcium ion in platelet serotonin uptake regulation.

It is generally accepted that intracellular Ca2+ is a key substance in the intracellular signal transducing mechanism of platelets. We investigated the possibility that extracellular and/or intracellular Ca2+ might regulate the transport activity of serotonin (5-HT) into platelets. We found that extracellular Ca2+ chelation with EGTA caused inhibition of 5HT uptake activity, which was recovered by extracellulary applied excess Ca2+. Intracellular Ca2+ chelation with acetoxymethyl bis(O-aminophenoxy)ethane-N,N,N'-tetraacetate (BAPTA-AM) did not, however, have any inhibitory effect on 5HT uptake activity in the presence of extracellular Ca2+. In the absence of extracellular Ca2+, BAPTA-AM significantly inhibited 5-HT uptake. The restorative effect of Ca2+ on 5-HT transport into EGTA-treated platelets was mimicked by Ba2+, but not by Sr2+. It was antagonised by inorganic Ca2+ channel antagonist including Ni2+, La3+ and Gd3+, but not by organic Ca2+ channel blockers including verapamil, nifedipine, diltiazem, omega-conotoxin GVIA and omega-agatoxin IVA. Furthermore, 3,4,5-trimethoxybenzoic acid 8-(diethylamino)octyl ester hydrochloride (TMB-8), an intracellular Ca2+ antagonist, was found to inhibit the restorative effect of Ca2+. These results have led to the suggestion that depletion of intracellular Ca2+ pool(s) by EGTA might result in a reduction of 5-HT uptake activity. Thus, the intracellular Ca2+ pool(s) susceptible to EGTA might have a regulatory role in maintaining 5-HT transport into blood platelets.

Animals↗
PubMed1995

Relation of angiographically defined coronary artery disease and plasma concentrations of insulin, lipid, and apolipoprotein in normolipidemic subjects with varying degrees of glucose tolerance.

We investigated the association between hyperinsulinemia and changes in lipid, lipoprotein, and apolipoprotein that would increase the risk of coronary artery disease (CAD) independent of glucose tolerance. A coronary angiogram was recorded in 127 male subjects, including 41 with normal glucose tolerance, 41 with impaired glucose tolerance, and 45 with non-insulin-dependent diabetes mellitus (NIDDM). Subjects were divided into 2 groups according to results: the group with CAD (n = 94) and the group with normal coronary arteries (n = 33). All subjects were normolipidemic (total cholesterol < 230 mg/dl and triglycerides < 150 mg/dl). The CAD group had a significantly lower plasma level of high-density lipoprotein (HDL) cholesterol and apolipoprotein A-I (apo A-I) and a higher level of apolipoprotein B (apo B) than the normal group with normal glucose tolerance. In considering subjects with impaired glucose tolerance or NIDDM, the CAD group had a significantly lower plasma level of HDL cholesterol and apo A-I and a significantly higher plasma level of total cholesterol, triglycerides, and apo B than the normal group. In each of the subjects with normal and impaired glucose tolerance, and NIDDM, the elevation of plasma insulin concentration during both the complete test period and the early phase of an oral glucose challenge was significantly higher in the CAD than in the normal group. In all subjects, graded reductions in HDL cholesterol and apo A-I and graded increases in plasma total cholesterol, triglycerides, and apo B were observed with increasing tertiles of the postglucose challenge measurements of insulinemia.(ABSTRACT TRUNCATED AT 250 WORDS)

Apolipoprotein A-I↗
PubMed1995

Modulation of in vitro splicing of the upstream intron by modifying an intra-exon sequence which is deleted from the dystrophin gene in dystrophin Kobe.

Molecular analysis of dystrophin Kobe showed that exon 19 of the dystrophin gene bearing 52-bp deletion was skipped during splicing, although the known consensus sequences at the 5' and 3' splice sites of exon 19 were maintained (Matsuo, M., T. Masumura, H. Nishio, T. Nakajima, Y. Kitoh, T. Takumi, J. Koga, and H. Nakamura. 1991. J. Clin. Invest. 87:2127-2131). These data suggest that the deleted sequence of exon 19 may function as a cis-acting element for exact splicing for the upstream and downstream introns. To investigate this potential role of exon 19, an in vitro splicing system using artificial dystrophin mRNA precursors (pre-mRNAs) was established. Pre-mRNA containing exon 18, truncated intron 18, and exon 19 was spliced precisely in vitro, whereas splicing of intron 18 was almost completely abolished when the wild-type exon 19 was replaced by the dystrophin Kobe exon 19. Splicing of intron 18 was not fully reactivated when dystrophin Kobe exon 19 was restored to nearly normal length by inserting other sequences into the deleted site. These results suggest that the presence of the exon 19 sequence which is lost in dystrophin Kobe is more critical for splicing of intron 18 than the length of the exon 19 sequence. Characteristically, the efficiency of splicing of this intron seemed to correlate with the presence of polypurine tracks within the downstream exon 19. Moreover, an antisense 31-mer 2'-O-methyl ribonucleotide complementary to the 5' half of the deleted sequence in dystrophin Kobe exon 19 inhibited splicing of wild-type pre-mRNA in a dose- and time-dependent manner. This first in vitro evidence that dystrophin pre-mRNA splicing can be modulated by an antisense oligonucleotide raises the possibility of a new therapeutic approach for Duchenne muscular dystrophy.

Base Sequence↗
PubMed1995

Synthesis of the esterified cerebroside in the epidermis of guinea pigs.

(2S,3R,4E,23'Z)-1-O-(beta-D-Glucopyranosyl)-N-(32'-linoleoyloxy -23'-dotriacontenoyl)-4-sphingenine (1), a component of the esterified cerebrosides in the epidermis of guinea pigs, was synthesized by selective N-acylation of (2S,3R,4E)-1-O-(beta-D-glucopyranosyl)-4-sphingenine (18) with activated omega-linoleoyloxy fatty acid ester 17.

Acylation↗
PubMed1995

Mutation in alpha 5(IV) collagen chain gene in nonfamilial hematuria.

Alport syndrome is an inherited disorder characterized by progressive nephritis with ultrastructural basket-weave changes of the glomerular basement membrane and neurosensory deafness. Mutations in the COL4A5 gene encoding the Type IV collagen alpha 5 chain have been reported to occur in patients with X-linked Alport syndrome. A girl with hematuric nephritis, characteristic basket-weave glomerular basement membrane changes, and abnormal expression of the Type IV collagen alpha 5 chain immunohistochemically, but no family history of nephritis, was identified. Mutation detection enhancement gel electrophoresis of the polymerase chain reaction-amplified exons of COL4A5 from this patient revealed a sequence variant in the exon 50 region. Sequence analysis of her polymerase chain reaction product demonstrated a single-base (C; nucleotide 4728 from the 5' end) deletion in exon 50. This novel mutation alters the reading frame and introduces a translation stop codon that would be expected to result in a noncollagenous domain with only 209, instead of the normal 229, amino acid residues. Gene tracking with restriction enzyme AfIIII demonstrated that her mother was normal. These findings represent a new mutation of the X-linked Alport syndrome in this patient and demonstrate that a COL4A5 gene mutation causes the abnormal expression of Type IV collagen alpha 5 chain protein.

Base Sequence↗
PubMed1995

Orientation of the toad, Bufo japonicus, toward the breeding pond.

A variety of orientation cues has been suggested for the migration to the breeding site in adult amphibians. We categorized the cues into the following 3 groups: 1) cues from the breeding pond such as male calling and pond odors, 2) celestial cues such as the sun light and the magnetic field of the earth and 3) cues from the area or route of the migration which compose a local map such as a visual and olfactory maps. To determine which of these is used by the toad, Bufo japonicus, we designed and conducted a displacement experiment in which migrating toads from one direction were transported to the ground in the opposite side of the pond. The displaced toads were completely disoriented and moved to random directions. We conclude that the toad uses a local map to orient to the breeding pond and cues from celestial bodies and the pond are not used. We also found that adult toads tracked the same route on both trips from and to the pond. This suggests that the local map was memorized by newly metamorphosed toads at their first terrestrial trip from the pond. The next step of our study was to determine what sense is used to receive the cue. We found blind toads, whose upper and lower eye-lids were stitched together, could reach the pond at a similar rate with the sham-operated and intact toads. However, anosmic toads, whose olfactory mucosa were damaged by the treatment with a 5% silver nitrate solution, rarely reached the pond.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

Cardiac involvement in a family with Becker muscular dystrophy.

We report a family with Becker muscular dystrophy (BMD) presenting with cardiac involvement. The proband was a 41-year-old Japanese man who was hospitalized with exertional dyspnea and muscle weakness. Cardiac examination showed findings consistent with dilated cardiomyopathy. Dystrophin immunohistochemical analysis showed a discontinuous patchy staining pattern in cardiac and skeletal muscles biopsied from the proband. His brothers had high creatine kinase (CK) activity and abnormal electrocardiogram. Dystrophin gene analysis revealed that the proband and his brothers had G-to-T transversion at the terminal nucleotide of exon13. We conclude that the mutated dystrophin gene may cause cardiac involvement as a symptom precedent to skeletal muscle involvement.

Adult↗
PubMed1995

Necessity of newly synthesized ATP by creatine kinase for contraction of permeabilized longitudinal muscle preparations of rat proximal colon.

Necessity of newly synthesized ATP by creatine kinase for synthesis of ATP as an energy source for smooth muscle contraction was studied in permeabilized longitudinal muscle preparations of rat proximal colon. In alpha-toxin-permeabilized preparations, Ca++ induced "phasic type" contraction in a normal bath solution containing 4 mM ATP and 5 mM phosphocreatine. Omission of phosphocreatine from the solution resulted in significant decrease in phasic contraction, and omission of ATP resulted in loss of the response to Ca++. When ADP, but not adenosine-5-O-(2-thiodiphosphate), with phosphocreatine was added as a substitute for ATP, Ca++ induced the same type of contraction as with ATP. The maximum tensions of the phasic and tonic phases of the contraction with ADP were approximately 60% of, and almost the same, respectively as those with ATP. A selective inhibitor of creatine kinase, 2,4-dinitrofluorobenzene, inhibited the phasic contraction induced by Ca++. After irreversible inhibition of endogenous creatine kinase by DNFB in beta-escin-permeabilized preparations, treatment of the preparations with exogenous creatine kinase restored Ca(++)-induced contraction. These findings suggest that ATP synthesized from ADP and phosphocreatine by creatine kinase was necessary for phasic contraction of permeabilized smooth muscle and that exogenous ATP was mainly used after its hydrolysis to ADP.

Adenosine Diphosphate↗
PubMed1995

Involvement of heterotrimeric GTP-binding protein and rho protein, but not protein kinase C, in agonist-induced Ca2+ sensitization of skinned muscle of guinea pig vas deferens.

We studied the involvement of protein kinase C (PKC) and a small GTP-binding protein (G-protein), rho, in receptor-mediated Ca2+ sensitization of the contractile apparatus of smooth muscle of guinea pig vas deferens. In beta-escin-permeabilized smooth muscle strips, norepinephrine (NE) in the presence of GTP caused further contraction of the preparations at a constant Ca2+ level (Ca2+ sensitization). Prazosin and GDP beta S, a nonhydrolyzable GDP analogue, inhibited NE-induced Ca2+ sensitization, indicating an alpha-1 adrenoceptor/G-protein mediated response. GTP alone (> 10 microM) and GTP gamma S, a non-hydrolyzable GTP analogue, also induced Ca2+ sensitization. Pretreatment of preparations with C3 exoenzyme of Clostridium botulinum, which is known to ADP-ribosylate rho family proteins, with NAD resulted in complete inhibition of NE- and GTP (GTP gamma S)-induced Ca2+ sensitization. AIF4-, which activates heterotrimeric G-, but not small G-protein also induced Ca2+ sensitization. Interestingly, AIF4(-)-induced Ca2+ sensitization was inhibited by not only GDP beta S but also C3-treatment, suggesting that activation of heterotrimeric GTP-binding protein precedes activation of rho protein. On the other hand, phorbol 12,13-dibutyrate, like NE, also induced Ca2+ sensitization. The sensitization was inhibited by PKC(19-31), a PKC inhibitor peptide. However, PKC(19-31) did not have any effect on NE- or AIF4(-)-induced Ca2+ sensitization.(ABSTRACT TRUNCATED AT 250 WORDS)

ADP Ribose Transferases↗
PubMed1995

Docosahexaenoic acid ethyl ester and its ultraviolet degradation products showed DNA-breaking activity in vitro and cytotoxic effects on the HSC-4 cell line.

The DNA-breaking activity and cytotoxicity of docosahexaenoic acid ethyl ester (DHAE) and its degradation products (dDHAE) induced by ultraviolet (UV) irradiation, were demonstrated in this study. The major component of dDHAE was identified as 4-hydroxyvaleric acid ethyl ester. DHAE and dHAE solutions at a concentration of 100 mg/ml induced single-strand breaks of plasmid DNA after an incubation at 37 degrees C and pH 7.6 for 15 hours. DHAE transformed a plasmid supercoiled DNA (Form I) into an open circular relaxed form (Form II), and dDHAE completely destroyed DNA molecules. HSC-4 cells, which were derived from human prickle cell carcinoma, were cultured with various concentrations (10, 30 and 50 micrograms/ml) of DHAE and dDHAE. Both inhibited the proliferation of HSC-4 cells. The potency of their cytotoxicity was dependent on their concentration. It is noteworthy that the lethal effects of DHAE and dDHAE on HSC-4 cells were quite similar, although the DNA-breaking activity of dDHAE was much greater than that of DHAE. These results suggested that DHAE becomes more cytotoxic after undergoing oxidization and metabolism in the cell.

Antineoplastic Agents↗
PubMed1994

[MR pyelography using fast spin echo technique in hydronephrosis].

MR pyelography using the fast spin echo technique and and a body coil were performed in ten patients with hydronephrosis. In all cases, pyelograms of the dilated urinary tract were obtained by this method, although they were obtained in only two cases by drip infusion pyelography. MR pyelography is an effective method with which to obtain images of dilated urinary tract non-invasively, and in a relatively short time. This method was also useful to evaluate intraureteral structure.

Adolescent↗
PubMed1994

[Usefulness of MR cholangiography with flex coil].

Fast SE MR cholangiography was performed in 10 healthy volunteers and 10 patients with biliary disease, using body, shoulder and flex coils. Surface coils provided better images than the body coil. Although high SNR images were obtained near surface coils, signal intensity decreased in areas distant from the shoulder coil. The flex coil greatly improved image quality, especially in intrahepatic bile ducts, compared with the shoulder coil. The application of a flex coil to MR cholangiography is a highly useful method to evaluate the biliary system.

Adult↗
PubMed1994

Differential effects of divalent cations on specific 3H-GR 65630 binding to 5-HT3 receptors in rat cortical membranes.

We have examined the ability of divalent cations to inhibit 3H-GR 65630 binding to 5-HT3 receptor recognition sites in rat brain cortical membranes. Among the divalent metal cations tested, Cd2+, Zn2+ and Cu2+ inhibited the specific 3H-GR 65630 binding activity to 5-HT3 receptors at a concentration of 0.1-1 mM. The other divalent metal cations tested (i.e. Co2+, Ni2+, Ba2+, Ca2+, Mg2+ and Mn2+) produced no inhibition of the specific 3H-GR 65630 binding. Cd2+, Zn2+ and Cu2+ did not change the Bmax value of the binding activity, but significantly increased the Kd value. It was suggested that these cations inhibited the binding activity by reducing affinity of the 5-HT3 receptor for the antagonist, resulting in apparent inhibition of the binding activity. As to the binding association rate, Cd2+, Cu2+ and Zn2+ were found to have an inhibitory effect. The binding dissociation rate, however, was shown to be decreased by Cu2+ but not by Cd2+ and Zn2+. Furthermore, the Zn(2+)-induced inhibition of 3H-GR 65630 binding was suggested to be antagonized by both concanavalin A and wheatgerm agglutinin. The Cu(2+)-induced inhibition, however, was not influenced by these lectins, indicating that Cu2+ has a different lectin sensitivity for its inhibitory effect. The different mechanism of action between Cu2+ and Zn2+ was suggested in their inhibitory effect on the specific 3H-GR 65630 binding activity.

Animals↗
PubMed1994

Modulation of substance P/neurokinin-1 receptor in human astrocytoma cells by antisense oligodeoxynucleotides.

1. An antisense oligodeoxynucleotide corresponding to the NH2-terminus of the substance P (SP)/neurokinin-1 (SP/NK1) receptor was constructed and added to cultures of human astrocytoma U-87 MG cells in vitro and rats in vivo. 2. Antisense oligodeoxynucleotide at a concentration of 30 microM progressively reduced the specific binding [3H][Sar9, Met(O2)11]-SP, selective SP/NK1 receptor agonist, in the U-87 MG astrocytoma cells by approx. 31% on the second day after treatment (control: 26.1 +/- 2.4 fmol/mg protein vs antisense oligodeoxynucleotide: 18.0 +/- 1.4 fmol/mg protein, P < 0.001). 3. Treatment with 30 microM antisense oligodeoxynucleotide for 2 days inhibited the SP/MK1 receptor-induced influx of 45Ca2+ into the U-87 MG cells by approx. 35%. 4. When the same antisense oligodeoxynucleotides were encapsulated in liposomes and injected into the lateral cerebral ventricle of rats, functional SP receptor was blocked.

Animals↗