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Biomedical subjects

Y Inagaki

Publications and source records attributed to Y Inagaki.

At least 109 records · Page 6Linked to original sources
PubMed1996

Characterization of the interaction between RhoA and the amino-terminal region of PKN.

The yeast two-hybrid system and in vitro binding assay were carried out to characterize the interaction between PKN and a small GTP-binding protein, RhoA. It was revealed that the region corresponding to the amino acid residues 33-111 in the amino-terminal region of PKN was sufficient to confer the ability to associate with RhoA. Each synthetic peptide fragment corresponding to the amino acid residues 74-93 and 94-113 of PKN inhibited the interaction between PKN and RhoA in the in vitro binding assay, suggesting that this region is important in the association with RhoA. The endogenous and the GAP-stimulated GTPase activity of RhoA was inhibited by the interaction with PKN, suggesting the presence of a regulatory mechanism that sustains the GTP-bound active form of RhoA.

Binding Sites↗
PubMed1996

Cloning of the Mycoplasma capricolum gene encoding peptide-chain release factor.

In Mycoplasma capricolum (Mc), a relative of Gram+ eubacteria with a high genomic A + T-content, the UGA codon is assigned to Trp instead of being a stop codon. We previously showed the lack of peptide-chain release factor (RF) activity in vitro responding to the UGA codon in this bacterium [Inagaki et al., Nucleic Acids Res. 21 (1993) 1335-1338]. To obtain more information on the translation termination mechanism of Mc, we isolated and sequenced the gene encoding RF. The deduced amino-acid sequence has no RF-2-specific + 1 frameshift site and shows 50 and 36% identity to Escherichia coli RF-1 and RF-2, respectively. We conclude that this gene encodes the putative RF-1 which would possess the conserved 'five-domain' structure of RF family found in various organisms.

Amino Acid Sequence↗
PubMed1996

Histamine release induced by human natriuretic peptide from rat peritoneal mast cells.

We have been interested in the effects of some popular peptides on tracheal smooth muscle. Previously, we reported that atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP) and C-type natriuretic peptide (CNP) had dose-dependent relaxant effects on guinea pig tracheal smooth muscle. In this study, we compared the effects of ANP, BNP and CNP on histamine release from rat peritoneal mast cells. ANP and BNP were more potent than CNP, dose-dependently increasing histamine release at a concentration of 10(-7) M or higher. CNP induced histamine release at a concentration of 10(-6) M or higher. Extracellular calcium inhibited the histamine release induced by all 3 peptides. In conclusion, the effects of these 3 peptides in rat peritoneal mast cells demonstrated adverse reactions for respiratory diseases, although our previous results showed that these peptides caused relaxation of guinea pig tracheal smooth muscle. We should note that the drugs have different actions in each organ.

Amino Acid Sequence↗
PubMed1996

Identification of the first major allergen of a squid (Todarodes pacificus).

BACKGROUND: In Japan, squid is an important seafood, and some patients with food allergies are sensitive to squid. There has been no report, however, describing the major allergens of squid. OBJECTIVE: To characterize squid allergens, we isolated a major allergen from the Pacific flying squid (Todarodes pacificus) and compared it with a major allergen from a shrimp (Penaeus orientalis). METHODS: The major squid and shrimp allergens were isolated by column chromatography on diethylaminoethyl-Sepharose (Pharmacia, Uppsala, Sweden), hydroxylapatite, and Sephacryl S-300 (Pharmacia). The IgE reactivity of the isolated allergens was assessed by immunoblotting. The cross-reactivity between the squid and shrimp allergens was examined by use of mouse polyclonal and monoclonal antibodies to the major allergens. Amino acid sequence analyses of the isolated allergens were done. RESULTS: The isolated squid allergen is a 38 kd, heat-stable protein. IgE antibody binding to the purified squid allergen was demonstrated by immunoblotting. Cross-reactivity between major squid and shrimp allergens was demonstrated with sera from patients allergic to squid or shrimp or with allergen-specific monoclonal antibodies. The amino acid sequence analysis of the major squid allergen showed a marked homology with tropomyosin from blood fluke planorbid (Biomphalaria glabrata), which is a common vector snail of Schistosoma mansoni. CONCLUSION: This 38 kd protein is a major allergen of the squid, Todarodes pacificus, and is believed to be squid muscle protein tropomyosin. We named it Tod p 1 according to International Union of Immunological Societies allergen nomenclature regulation.

Allergens↗
PubMed1996

Cloning of variable regions of an antibody that reacts with the soluble fraction of human liver cells and its possible value in chronic liver disease.

A gene encoding the variable regions of the heavy and light chains of a mouse monoclonal antibody designated H2, which specifically reacts with human liver cells, was cloned into a phagemid vector. The clone of the variable region was designed to be expressed as a separate protein, the structure of which is the same as that of the mouse antibody. The cloned phage protein specifically reacted with anti-idiotypic antibodies produced in rabbits against the original mouse antibody, and this reaction was specifically blocked by the original antibody. The soluble protein, expressed as a fusion protein, was detected as a single 30-kd band on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and specifically bound to an anti-H2 idiotypic antibody as determined by Western blot analysis. Sera of patients with various diseases were assayed for antibodies to anti-H2 by sandwich enzyme-linked immunosorbent assay (ELISA). Only sera from patients with chronic liver disease reacted strongly. This binding was specifically blocked by the cloned soluble protein. The nucleotide sequences of the variable regions were determined by the dideoxy chain-termination method, and the sequences were approximately 95% identical to those of other mouse immunoglobulins. These findings suggest that a human antibody with the same idiotype as a mouse monoclonal antibody that reacts with human liver cells, can be detected in patients with chronic liver disease, suggesting that autoimmunity may be partly responsible for these diseases.

Adult↗
PubMed1996

Mode and site of analgesic action of epidural buprenorphine in humans.

This study was designed to clarify the site of analgesic action of epidural buprenorphine and its spinal segmental analgesia. Fifty patients undergoing gastrectomy were randomly assigned to five groups according to the dose of buprenorphine and route of administration: epidural saline group, epidural buprenorphine 2- and 4-micrograms/kg groups, and intravenous buprenorphine 2- and 4-micrograms/kg groups. The changes in pressure pain threshold (PPT) and visual analog scale (VAS) were compared within groups of patients receiving either buprenorphine, 2 or 4 micrograms/kg epidurally, and between groups of patients receiving buprenorphine 2 or 4 micrograms/kg either epidurally or intravenously. The PPT near the surgical incision was significantly greater (P < 0.05) in the epidural buprenorphine group compared to the intravenous group during the middle period (between 2 and 6 h after administration) of analgesia obtained after administration of buprenorphine. VAS response significantly (P < 0.05) decreased in the epidural group compared to the intravenous group at the same dose. In the smaller dose buprenorphine epidural group, PPT at the forehead (an index of systemic analgesia) exceeded the preoperative value during the 12 h after administration of buprenorphine. PPT near the surgical incision decreased by 30% from the preoperative value and did not exceed the preoperative value throughout the study. PPT at the forehead significantly exceeded the PPT changes measured near the surgical incision (an index of regional analgesia) (P < 0.05). In the larger dose buprenorphine epidural group, during the early period (up to 2 h after administration), PPT at the forehead increased by 30%-40%, while PPT near the surgical incision decreased by 10%-25% compared to the preoperative value (P < 0.01). However, in the middle period, there were no significant differences in the changes between PPT at the forehead and near the surgical incision. In the late period (more than 6 h after administration), PPT at the forehead maintained the level of preoperative value, whereas PPT near the surgical incision decreased by more than 25% from the preoperative value (P < 0.01). VAS value significantly (P < 0.01) decreased in the larger dose buprenorphine epidural group compared with that in the smaller dose group during the middle period. The larger dose of epidural buprenorphine provided better analgesia than the smaller dose. We conclude that epidural buprenorphine acts predominantly at the supraspinal region and produces spinal segmental analgesia in a dose-related manner.

Adult↗
PubMed1996

Microvascular disturbances in the colonic mucosa in antibiotic-associated haemorrhagic colitis: involvement of platelet aggregation.

An ultrastructural study of the colonic mucosa was performed in four patients with antibiotic-associated haemorrhagic colitis and new findings are reported. Colonoscopy was performed and biopsy specimens were obtained within 24 h of the onset of bloody diarrhoea. Colonoscopy demonstrated diffuse oedematous and haemorrhagic mucosa with erosions and white coat. Light microscopy revealed mucosal haemorrhage and inflammatory cell infiltration. Ultrastructurally, platelet aggregation was frequently present in the lumina of colonic mucosal capillaries, causing engorgement of red blood cells in adjacent microvessels. Mild to severe damage was observed in capillary endothelial cells, including discontinuity of basement membranes, gaps between endothelial cells and the destruction of capillaries. There was no evidence of microvascular spasm. In conclusion, our findings suggest that antibiotics directly or indirectly cause microcirculatory disturbances, which result in tissue damage and haemorrhage, in the colonic mucosa.

Adult↗
PubMed1996

[Pharmacokinetics of propofol during liver transplantation in small pigs].

A study using 14C propofol showed that the liver is the main eliminating organ for the agent. The current study was designed to clarify pharmacokinetics of propofol during liver transplantation in pigs. Five small pigs weighing 25.4 +/- 2.5 kg were anesthetized with isoflurane (0.5-1.5%) and mechanically ventilated under muscle paralysis with pancuronium. In the anhepatic phase, veno-veno bypass was placed from the inferior vena cava and portal vein to the superior vena cava. We studied pharmacokinetic parameters following an intravenous bolus injection of propofol at 2 mg.kg-1 in each phase, i.e. the pre-anhepatic, anhepatic and post-anhepatic phase during liver transplantation. Pharmacokinetic analysis showed that total plasma clearance of propofol in the anhepatic phase was significantly lower than that in the post-anhepatic phase. The results suggest that propofol may be metabolized extrahepatically and can be used at reduced doses in the anhepatic phase during liver transplantation.

Anesthetics, Intravenous↗
PubMed1996

[Cognitive function and MRI findings in very low birth weight infants].

Twenty-two very low birth weight infants at preschool ages of 5-6 years were studied to clarify the correlation between cognitive function and MRI findings. Cognitive function was evaluated by the Wechsler Intelligence Scale for Children-Revised (WISC-R) and the Frostig developmental test of visual perception. Ventricular enlargement, assessed by the bioccipital index (B. I.) measured on MRI, was correlated to cognitive disorders. Children with periventricular high intensity areas (T2-weighted images) extending from the posterior periventricular region to the parietal lobe tend to highly suffer from cerebral palsy and visuo-perceptual impairment. These results indicate that the disorders of cognitive function in very low birth weight infants were caused by a damage of association fibers in periventricular areas which was detectable by MRI.

Brain↗
PubMed1996

Effects of the new calcium antagonist efonidipine hydrochloride on resting and exercise hemodynamics in patients with stable effort angina.

The action of efonidipine hydrochloride ((+/-)-2-[benzyl(phenyl)-amino]ethyl 1,4-dihydro-2,6-dimethyl-5-(5,5-dimethyl-2-oxo-1, 3,2-dioxaphosphorinan-2-yl)-4-(3-nitrophenyl)-3-pyridinecarboxy late hydrochloride ethanol, CAS 1110011-76-8, NZ-105) a new dihydropyridine calcium antagonist, on cardiac hemodynamics at rest and during exercise as well as plasma concentration and pharmacokinetic parameters were studied in 9 patients with angina pectoris. NZ-105 was administered 40 mg once daily for a week and cardiac hemodynamics parameters were measured at rest and during exercise using a bicycle ergometer before and after treatment. All patients showed anginal symptoms during exercise before treatment, while only 4 showed anginal symptoms during exercise after treatment. Improvement on electrocardiograms (ECG) (> 0.1 treatment mV) was detected in 4 out of 9, and NZ-105 was recognized to have an anti-anginal action. The mean plasma concentration of NZ-105 at the time was 14.5 ng/ml. At rest, reduction in blood pressure and decrease in total peripheral vascular resistance were observed, however, NZ-105 showed no effect on heart rate, cardiac index, pulmonary arterial pressure and central venous pressure. During maximum exercise, a decrease in total peripheral vascular resistance, reduction in pulmonary arterial pressure and central venous pressure, increasing tendency of left ventricular ejection fraction, and increase in cardiac index were observed. However, NZ-105 showed no effect on heart rate and blood pressure. Based on the results mentioned above, cardiac hemodynamics of NZ-105 during exercise, featured primarily, reduction of afterload and improvement of cardiac functional deterioration due to exercise. In conclusion, NZ-105 is useful in patients with ischemic cardiac diseases by improving hemodynamics and ECG findings during exercise in patients with effort angina.

Angina Pectoris↗
PubMed1995

Second-site suppressor mutations for the Asp-66-->Cys mutant of the transposon Tn10-encoded metal-tetracycline/H+ antiporter of Escherichia coli.

Asp66 is the only essential acidic residue in the putative hydrophilic loop2-3 region of the transposon Tn10-encoded metal-tetracycline/H+ antiporter [TetA(B)] [Yamaguchi, A., Nakatani, M., & Sawai, T. (1992a) Biochemistry 31, 8344-8348]. Escherichia coli cells producing a D66C mutant of TetA(B) showed no tetracycline resistance. A spontaneous second-site revertant was isolated from the cells carrying the D66C mutant gene, which showed moderate resistance to tetracycline [minimum inhibitory concentration (MIC), 50 micrograms/mL]. The entire sequencing of the revertant genes revealed two secondary mutations, i.e., the codon 40 of GCT (Ala)-->GAT (Asp) and T-->G at 17 bases upstream from the initiation codon of the tetA gene. There was a T-->G mutation at position -17, which was a mutation in the tet promoter/operator region, which caused a decrease in TetA(B) production. The full expression of the A40D/D66C double and the A40D single mutants, which were constructed by site-directed mutagenesis, was deleterious for cell growth. The -17T-->G mutation mitigated the deleterious effect of these mutants through reduction of expression. The -17T-->G single mutation introduced into the wild-type tet gene did not affect the level of resistance, although the expression was significantly reduced. Intact cells carrying the A40D/D66C and -17T-->G/A40D/D66C mutant plasmids showed a reduced level of tetracycline accumulation due to active efflux, whereas no significant tetracycline uptake was observed in inverted membrane vesicles prepared from these mutant-producing cells. The A40D mutation is located at opposite side of the membrane to the D66C mutation in the putative secondary structure of TetA(B). The second-site mutation might mediate its effects through a structural perturbation propagated along the polypeptide backbone.

Amino Acid Sequence↗
PubMed1995

Translation of synonymous codons in family boxes by Mycoplasma capricolum tRNAs with unmodified uridine or adenosine at the first anticodon position.

In Myocoplasma capricolum, codon family boxes except for arginine and threonine (CUN leucine, GUN valine, UCN serine, CCN proline, GCN alanine, and GGN glycine) have only a single tRNA species with the anticodon sequence UNN (N is U, C, A or G), the first nucleoside U being unmodified. Incorporation of the [3H]amino acid into the peptide fraction was examined with the M. capricolum cell-free translation system. Synthetic mRNA containing each of the respective amino acid codons in the coding frame was subjected to translation. The tRNAUNN translated all the family box codons with similar, if not equal, efficiencies. In M. capricolum, there are two species of threonine tRNA, tRNA(UGUThr) and tRNA(AGUThr), the first nucleoside U or A being unmodified. The tRNA(UGUThr) species translates codons ACA, ACG and ACU efficiently, and ACC only poorly. In contrast, the tRNA(AGUThr) species translates codons ACU, ACC and ACG efficiently and ACA poorly.

Adenosine↗
PubMed1995

Chemical modification and inactivation of phospholipases A2 by a manoalide analogue.

Chemical modification and inactivation of bovine pancreatic, porcine pancreatic, Naja naja atra and Pseudechis australis phospholipases A2 (PLA2s), belonging to Group I, and of Trimeresurus flavoviridis, Vipera russelli russelli and Agkistrodon halys blomhoffii PLA2s, belonging to Group II, were investigated by the use of a manoalide (MLD)-analogue, 1-(2,5-dihydro-hydroxy-5-oxo-3-furanyl)-8,12-dimethyl-4-formyl-3,7, 11-tridecatrienol. At appropriate time intervals, residual PLA2 activities towards monodispersed, anionic mixed micellar and non-ionic mixed micellar substrates were measured. We tested the protective effect of micellar n-dodecylphosphocholine (n-C12PC) on enzyme inactivation. Inactivation of pancreatic PLA2s (Group I) was only observed towards anionic mixed micellar substrates. This inactivation was completely prevented by the presence of micellar n-C12PC. From a fragmentation study of modified bovine pancreatic PLA2 using lysyl endopeptidase, we speculated that Lys-56 of this enzyme was modified by MLD-analogue and that this modification was responsible for enzyme inactivation. Inactivation of non-pancreatic PLA2s was observed towards all types of substrate, except that no significant inactivation of N. naja atra PLA2 (Group I) towards monodispersed substrate was noted. Micellar n-C12PC protected N. naja atra PLA2 (Group I) completely from inactivation by MLD-analogue, but had lesser protective effects on P. australis PLA2 (Group I), T. flavoviridis and V. russelli russelli PLA2s (Group II). However, no significant protection of A. halys blomhoffii PLA2s (Group II) activity was observed. These results indicate that the inactivation of pancreatic and N. naja atra PLA2s originates from the modification of Lys residues at the interfacial recognition site, and that inactivation of P. australis, T. flavoviridis and V. russelli PLA2s arises from the modification of Lys residues at the catalytic site, interfacial recognition site and regions outside both sites. The inactivation of A. halys blomhoffii PLA2 was assumed to be due to the modification of Lys residues outside the two sites described above.

Amino Acid Sequence↗
PubMed1995

Structural analysis of Tpn1, a transposable element isolated from Japanese morning glory bearing variegated flowers.

The 6.4 kb transposable element Tpn1 belonging to the En/Spm family was found within one of the DFR (dihydroflavonol-4-reductase) genes for anthocyanin biosynthesis in a line of Japanese morning glory (Pharbitis nil) bearing variegated flowers. Sequencing of the Tpn1 element revealed that it is 6412 bp long and carries 28-bp perfect terminal inverted repeats. Its subterminal repetitive regions, believed to be the cis-acting sequences for transposition, show striking structural features. Twenty-two copies of the 10-bp sequence motif GACAACGGTT can be found as direct or inverted repeats within 650 bp of the 5' end of the element, and 33 copies of the sequence motif lie within 800 bp of the 3' terminus. All these 22 copies of the sequence motif near the 5' terminus and 30 copies in the 3' terminal region are arranged as inverted repeats and 3-8 bp AT-rich sequences are detected between these inverted repeats. In addition, four copies of 122-bp tandem repeats and six copies of 104-bp tandem repeats are present in the 5' and 3' subterminal repetitive regions, respectively. No large open reading frame characteristic of autonomous elements of the En/Spm family can be detected within the element. The results are discussed with respect to heritable changes in flower variegation in this line of Japanese morning glory.

Alcohol Oxidoreductases↗