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Biomedical subjects

H Usui

Publications and source records attributed to H Usui.

At least 55 records · Page 3Linked to original sources
PubMed1997

Inhibitory effects of various spasmolytics on the vagal afferent gastric excitatory response in cats.

The inhibitory effects of atropine, cimetropium, pirenzepine and N-butylscopolamine on the vagal afferent gastric excitatory response in cats under anesthesia with pentobarbital sodium and infusion of gallamine were examined. Electrical stimulation of vagal trunk in left side (10 Hz in frequency, 3 msec in duration, 15 V in intensity and for 10 sec) caused an initial gastric excitatory response during the period of stimulation followed by a late excitatory gastric response after stimulation in normal cats. The initial response was inhibited by atropine (100 microg/kg, i.v.) and hexamethonium (10 mg/kg, i.v.), while the late response was inhibited by atropine but not by hexamethonium (10 mg/kg, i.v.). In chronic supranodose vagotomized cats 11-15 days after the operation, stimulation of the vagal trunk caused a late gastric excitatory response after the stimulation period, which was inhibited by atropine (100 microg/kg, i.v.) but not by hexamethonium (10 mg/kg, i.v.). The two types of gastric responses in normal cats have been defined as follows: the initial gastric excitatory response (atropine- and hexamethonium- sensitive) is due to activation of vagal efferent fibers and the late gastric excitatory response (atropine-sensitive and hexamethonium-resistant) is due to activation of vagal afferent fibers. ED50 values of atropine, cimetropium, pirenzepine and N-butylscopolamine in inhibiting the vagal afferent gastric response were 7.2 microg/kg (n=4), 2.4 microg/kg (n=6), 82.6 microg/kg (n=3) and 93.0 microg/kg (n=4), respectively. The inhibitory effects of atropine and cimetropium on the vagal afferent gastric excitatory response (hexamethonium-resistant) were more potent than those of pirenzepine and N-butylscopolamine. These results suggested that the potent inhibitory effects of cimetropium and atropine on the vagal afferent gastric response may involve a potent spasmolytic effect.

Animals↗
PubMed1997

Identification of casoxin C, an ileum-contracting peptide derived from bovine kappa-casein, as an agonist for C3a receptors.

Casoxin C (Tyr-Ile-Pro-Ile-Gln-Tyr-Val-Leu-Ser-Arg) is a bioactive peptide that was isolated from a tryptic digest of bovine kappa-casein as an anti-opioid peptide in longitudinal strips of guinea pig ileum. Casoxin C also evokes contraction of the ileal strips, and we found that this process was biphasic with rapid and slow components. The contractile profile was very similar to that of human complement C3a(70-77), which is the COOH-terminal octapeptide of C3a and has, although less potent, qualitatively the same biological activities as C3a. Casoxin C also has homology with C3a(70-77). The rapid contraction was mediated by histamine release and the slow contraction was mediated by a prostaglandin E2-like substance, judging from the effects of various pharmacological inhibitors and antagonists on the ileal contraction. Casoxin C had affinity for C3a receptors (IC50 = 40 microM) in the radioreceptor assay. In addition, casoxin C showed phagocyte-stimulating activities. Casoxin C is therefore the first milk-derived peptide identified, that acts through complement C3a receptors.

Amino Acid Sequence↗
PubMed1997

Identification of autophosphorylation sites in c-Yes purified from rat liver plasma membranes.

c-Yes was purified 322-fold from a rat liver plasma membrane fraction to a single 60-kDa band on SDS-PAGE. The purified protein contained essentially no phosphotyrosine residues and was autophosphorylated with Mg2+. ATP exclusively at tyrosine residues with a concomitant increase in the protein-tyrosine kinase activity. The autophosphorylated c-Yes was extensively digested by trypsin and the resultant two major phosphopeptides, peptides I and II, were purified by HPLC on a reversed-phase C-18 column. The amino acid sequence of peptide I was determined to be LIEDNEYTAR, which is identical with the sequence from Leu-418 through Arg-427 of mouse c-Yes, indicating that one of the autophosphorylation sites corresponds to Tyr-424 of the mouse c-Yes. After partial determination of the N-terminal sequence of 10 amino acid residues of peptide II, the 230 bp sequence of rat cDNA that encodes the N-terminal 76 amino acid residues of c-Yes covering peptide II, was determined. From the predicted amino acid sequence, the sequence of peptide II was assumed to be from Tyr-16 through Lys-46, YTPENPTEPVNTSAGHYGVEHATAATTSSTK. The purified c-Yes phosphorylated the tyrosine residue of synthetic peptides covering Tyr-32 and its surrounding sequence but did not phosphorylate peptides covering Tyr-16 and its surrounding sequence, suggesting that the other autophosphorylation site is Tyr-32.

Amino Acid Sequence↗
PubMed1997

Tissue and subcellular distributions, and characterization of rat brain protein phosphatase 2A containing a 72-kDa delta/B" subunit.

A 74-kDa delta/B" subunit was isolated by heparin-Sepharose column chromatography from human erythrocyte protein phosphatase 2A (PP2A) consisting of a 34-kDa catalytic subunit (alpha/C) and 63- and 74-kDa regulatory subunits (beta/A and delta/B") in a ratio of 1:1:1. The purified delta/B" was used as an immunogen in mice, to prepare specific antisera against delta/B". Immunoblot analyses with the antisera detected an immunoreactive 72-kDa protein in the cytosol from various rat tissues including erythrocytes, brain, lung, testis, adrenal gland, heart, spleen, kidney, and liver. The 72-kDa protein was highly abundant in brain and was distributed evenly in cerebral cortex, cerebellum, and brain stem. The 72-kDa protein was also detected in mitochondria and microsome fractions. An immunoreactive 68-kDa protein was detected mainly in nuclear and microsome fractions. The 72-kDa protein from rat brain cytosol copurified with phosphorylated H2B histone phosphatase activity during successive chromatographies on DEAE-Toyopearl, AH-Sepharose, Sephadex G-150, H1 histone-Toyopearl, TSK DEAE-5PW, protamine-Toyopearl, and TSK G3000SW columns. The purified enzyme migrated as a single protein band on nondenaturing PAGE and as three protein bands of 34, 63, and 72 kDa in a ratio of 1:1:1 on SDS-PAGE. The molecular weight of the enzyme was estimated to be 170,000 from the s20,W value of 7.2 +/- 0.3 S and the Stokes radius of 5.5 +/- 0.1 nm. The rat brain enzyme was classified as PP2A, based on the following properties; (1) an IC50 for okadaic acid of 10(-9) M; (2) its preferential dephosphorylation of the a subunit of phosphorylase kinase; (3) its insensitivity to protein inhibitor 2; and (4) its heterotrimeric subunit structure. The Km value and the molecular activity of the enzyme for phosphorylated H2B histone were 72.3 +/- 0.3 microM and 192 +/- 2 mol Pi released/min/mol enzyme, respectively, and were comparable to those of human erythrocyte PP2A (alpha1 beta1 delta1/ CAB"). The 72-kDa subunit in the purified rat brain PP2A was phosphorylated in vitro by cAMP-dependent protein kinase.

Animals↗
PubMed1997

Hyaluronidase activity in human pus from which Streptococcus intermedius was isolated.

Hyaluronidase (HAase) activity was detected in both a human pus sample and the culture supernatant of the only bacterial isolate from the pus, Streptococcus intermedius, using a zymographic technique. The optimum pH range for HAase activity was similar for both samples. Although the bands showing the strongest HAase activity of these samples differed from each other with respect to molecular size, both samples were equally inhibited by an antiserum raised against HAase of S. intermedius. These results suggest that S. intermedius may produce HAase in vivo as well as in vitro, and that this enzyme and/or its fragments may play an important role in host tissue degradation.

Adult↗
PubMed1997

[Tachykinin receptor subtypes involved in endothelium-dependent and -independent responses in rabbit intrapulmonary arteries].

In the present study, it was demonstrated that SP, neurokinin A (NKA), neurokinin B (NKB), SP methyl ester (SPME), [Ala5, beta-Ala8]-alpha-neurokinin fragment 4-10 (AANF) at 10(-8) M all caused contraction in non-contracted endothelium-intact arteries. SP- and SPME-induced contraction were reduced by removal of endothelium. All the peptides with the exception of AANF induced transient relaxation in the precontracted arteries. The relaxation were attenuated by removal of endothelium. The potency orders for endothelium-dependent contraction (EDC), -dependent relaxation (EDR) and -independent contraction (EIC) were SP > SPME >> NKA [symbol: see text] NKB [symbol: see text] AANF, SP > SPME > NKA > NKB >> AANF and NKA > AANF > NKB >> SP [symbol: see text] SPME, respectively. SP-induced EDC and EDR were attenuated by an NK1 antagonist but not by an NK2 antagonist. The SP-induced EIC was reduced by an NK2 antagonist. SP-induced EDC was attenuated by aspirin, OKY-046, and S-1452. The EDR was attenuated by L-NAME and methylene blue. The EDC induced by SPME was non-competitively attenuated by CP-99994, an NK1 antagonist. EDR was competitively inhibited by CP-99994. In conclusion, SP and related peptides caused EDC via NK1 receptors and TXA2 production, EDR via NK1 receptors and NO release and EIC via NK2 receptors in rabbit intrapulmonary arteries.

Animals↗
PubMed1997

[The chronological changes in the size of maxillary defect].

Use of a maxillary prosthesis is now considered one of the good methods to ameliorate the crippling caused by the defect in the maxillary palate area after an ablative procedure for a maxillary tumor and it consumes no physical resources from the patient. There has been debate on whether the prosthesis should be made in the very early postoperative days or after the wound is stabilized. In this study, changes in the area of the defect in the palate after surgery were investigated by using plaster models from clinical cases we have experienced. Twenty-one cases were investigated according to the following features: "maxillary sinus cancer "+" more than 1.5 years of follow-up for maxillofacial prosthetics "+" presence of more than two casts of the case". These casts were photographed from a distance of 1.5 m, and the photographs were enlarged to the original size of the cast, then the defect area was measured with a Planimeter (Uchida Co. Ltd). The area of the defect had shrunk in 16 cases. It took over 1 year to get stabilized in 80% of the cases. It is believed to take around 3.5 years for all cases to become stable. This tendency had no relationship with the pattern of the defect, the stroke of the mouth opening, the pattern of the rest of the teeth, or the defect area itself. These findings suggest that it may be advisable to start the prosthetic work as early as possible rather than waiting for stability of the defect wound, to improve the patients's QOL.

Adult↗
PubMed1996

Isolation of cDNA clones of the rat mRNAs expressed preferentially in the prenatal stages of brain development.

For better understanding of the molecular mechanisms underlying the developmental processes of the mammalian brain, we isolated rat fetal brain-enriched (FBE) cDNA clones, whose corresponding mRNAs were expressed at least 5-fold more in the fetal brain than in the adult brain. Our modified differential screening procedure, which utilized a two-vector (pT7T3D and pBluescript) system and showed low background levels of colony hybridization for screening, efficiently identified 64 candidate FBE clones from a small number (475) of colonies in the fetal brain cDNA library. After subsequent second screening of the candidate FBE clones by Northern blot analysis, we successfully isolated 22 distinct FBE clones. The nucleotide sequence analysis of the 22 FBE clones revealed that 13 of them had no significant matches to the sequences reported in the databases, whereas 9 of them matched previously reported sequences (alpha-tubulin M alpha 1, beta-tubulin M beta 5, thymosin-beta 10, stathmin, beta-tubulin M beta 2, alpha-internexin, ferritin Lg chain, neuronatin and amphoterin), most of which have been shown to be down-regulated during brain development. We also found that the Northern blot analysis in the second screening could be replaced by cDNA library DNA-Southern blot analysis, in most clones corresponding to relatively abundantly expressed mRNAs. Thus, once the cDNA library is constructed, clone selection will be possible in such clones without the use of additional RNA or Northern blot in screening, allowing the analysis of small brain regions of interest.

Animals↗
PubMed1996

Molecular cloning of a 74-kDa regulatory subunit (B" or delta) of human protein phosphatase 2A.

Based on amino acid sequence data of a 74-kDa regulatory subunit (B" or delta) of a human heterotrimeric protein phosphatase 2A, a cDNA encoding the subunit was isolated from a human cerebral cortex library. The cDNA had an open reading frame encoding an M(r) 66,138 protein of 570 amino acids. Bacterial expression of the cDNA yielded a protein immunoreactive with antisera specific to the 74-kDa subunit. The predicted primary structure of the subunit had no similarity to already reported sequences of PP2A regulatory subunits including A, B, and PR72. Potential phosphorylation sites for protein kinases A and C, a bipartite motif of putative nuclear localization signal, and SH3 accessible proline-rich domain, and a unique PQ repeat were found in the sequence. The subunit mRNA of about 2.9 kb was ubiquitously expressed in rat tissues.

Amino Acid Sequence↗
PubMed1996

Possible involvement of endothelial leukotrienes in acetylcholine-induced contraction in rabbit coronary artery.

In endothelium-intact preparations of rabbit coronary artery, acetylcholine (3 x 10(-8)-10(-6) M) caused a contraction in the presence of N omega-nitro-L-arginine methyl ester (10(-4) M). Removal of endothelium significantly attenuated the contraction. The present experiments were undertaken to elucidate whether the acetylcholine-induced contraction in endothelium-intact preparations was due to arachidonic acid metabolites, endothelin or superoxide. The acetylcholine-induced contraction in endothelium-intact preparations was attenuated by manoalide (10(-6) M and 3 x 10(-6) M) or oleyloxyethyl phosphorylcholine (10(-5) M) (phospholipase A2 inhibitors), BAY x 1005 (3 x 10(-6) and 10(-5) M) or L 663,536 (10(-6) and 10(-5) M) (leukotriene synthesis inhibitors) and ONO-1078 (10(-6) and 3 x 10(-6) M) or SK&F 104353 (10(-6) and 3 x 10(-6) M) (leukotriene antagonists). The contraction was not affected by aspirin (10(-4) M) or indomethacin (10(-6) M) (cyclooxygenase inhibitors), S-1452 (10(-8) M) or ONO-3708 (10(-7) M) (thromboxane A2 antagonists), FR139317 (10(-6) M) (endothelin receptor antagonist) or superoxide dismutase (150 u/ml) combined with catalase (1000 u/ml), or allopurinol (10(-5) M) (antioxidants). In contrast, the endothelium-independent contraction induced by acetylcholine was unaffected by any of these inhibitors and antagonists listed above. The present experiments demonstrate that in the presence of N omega-nitro-L-arginine methyl ester, acetylcholine partly causes an endothelium-dependent contraction in rabbit coronary artery which is probably due to leukotriene C4 and D4, but does not involve production of thromboxane A2, endothelin or superoxide.

Animals↗
PubMed1996

Studies on the ileum-contracting mechanisms and identification as a complement C3a receptor agonist of oryzatensin, a bioactive peptide derived from rice albumin.

Oryzatensin (Gly-Tyr-Pro-Met-Tyr-Pro-Leu-Pro-Arg) is an ileum-contracting and immunostimulating peptide derived from rice albumin. The mechanisms for the ileal contraction that it induces, consisting of rapid and slow components, were examined. The rapid contraction was mediated by histamine release and the slow contraction by a prostaglandin E2-like substance, judging from the effects of various pharmacological inhibitors and antagonists on ileal contraction and titration of histamine release. The contractile profile was very similar to that of human complement C3a(70-77), which is the COOH-terminal octapeptide of C3a and has, although less potent, qualitatively the same biological activities as C3a. Oryzatensin showed homology with C3a(70-77) and affinity for C3a receptors (IC50 = 44 microM) by radioreceptor assay. This is the first report of a food-derived bioactive peptide acting through complement C3a receptors.

Amino Acid Sequence↗
PubMed1996

Vasorelaxation by casomokinin L, a derivative of beta-casomorphin and casoxin D, is mediated by NK1 receptor.

Casomokinin L (Tyr-Pro-Phe-Pro-Pro-Leu), a derivative of beta-casomorphin and casoxin D, showed endothelium-dependent vasorelaxing activity on the canine mesenteric artery (EC50 = 7 x 10(-8) M). This relaxing activity was partly blocked by 10(-5) M NAME (nitric oxide synthase inhibitor), and the inhibition by NAME was restored by addition of 10(-4) M arginine, suggesting the involvement of nitric oxide as an endothelium-dependent relaxing factor. The relaxing activity was blocked by 10(-8) M CP-99994 and 10(-7) M FK888 (NK1 antagonists), but not by 10(-6) M SR-48968 (NK2 antagonist). Casomokinin L binds to neurokinin NK1 receptors (Ki = 5.8 x 10(-5) M), and lowered blood pressure in SHR (0.1 mg/kg, i.v. and 3 mg/kg, PO). Thus, despite its only three residues in common with substance P, casomokinin L binds to NK1 receptors, relaxes the artery, and exerts an antihypertensive effect.

Amino Acid Sequence↗
PubMed1996

Pharmacological nature of TP receptor mediated contraction in human intrapulmonary artery.

The present experiments were undertaken to elucidate the pharmacological nature of thromboxane A2/prostaglandin H2 receptor (TP)-mediated contraction in human intrapulmonary arteries. 9,11-epithio-11, 12-methano-thromboxane A2 (STA2) and (15S)-hydroxy-9 alpha, 11 alpha-(epoxymethano) prosta-5Z, 13E-dienoic acid (U46619) (TXA2 agonists) caused contractions in a concentration-dependent manner with EC50 values of 1.4 x 10(-9) M and 3.1 x 10(-9) M, respectively. S-1452 and ONO-3708 (TP receptor antagonists) concentration-dependently attenuated the STA2 (10(-8) M)-induced contraction with IC50 values of 5.8 x 10(-9) M and 4.2 x 10(-8) M, respectively. U-73122 (3 x 10(-6) M) and 2-nitro-4-carboxyphenyl-N,N-diphenylcarbamate (3 x 10(-5) M) (phospholipase C inhibitors) significantly attenuated the STA2-induced contraction. Ca(+2)-induced contraction in the presence of STA2 (10(-8) M) in Ca(+2)-free medium was attenuated by nifedipine (10(-6) M) by 40%. The remaining nifedipine-resistant Ca(+2)-induced contraction was not attenuated by nitroglycerin (10(-5) M), but forskolin (10(-5) M) (adenylate cyclase stimulant) significantly decreased it by 75%. The results clearly indicate that in human intrapulmonary artery, there are TP receptors coupled with phospholipase C activation and that TP receptor-mediated Ca(+2)-mobilization is in part nifedipine- and nitroglycerin-resistant, but forskolin-sensitive.

Adult↗
PubMed1995

Functional couplings of the delta- and the kappa-opioid receptors with the G-protein-activated K+ channel.

The delta- and the kappa-opioid receptors were individually co-expressed with the G-protein-activated K+ (GIRK1) channel in Xenopus oocytes. Stimulation of these opioid receptors with selective agonists induced currents which were blocked by the opioid receptor antagonist naloxone. The current responses showed inward rectification and were blocked by 300 microM Ba2+, indicating that the effect of activated opioid receptors was mainly mediated by the GIRK1 channel. The EC50 value obtained from these responses was 45 nM for the agonist DPDPE in the oocytes injected with the delta-opioid receptor mRNA and the GIRK1 mRNA and 15 nM for the agonist U50488H in the oocytes injected with the kappa-opioid receptor mRNA and the GIRK1 mRNA. The Hill coefficient was 0.92 in the former and 0.93 in the latter. These results suggest that each of the delta- and the kappa-opioid receptors functionally couples with the GIRK1 channel.

3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cycloh↗
PubMed1995

Endothelium-dependent contraction induced by acetylcholine in isolated rat renal arteries.

We investigated whether or not acetylcholine elicited an endothelium-dependent contraction and whether an arachidonic acid metabolite was involved in the acetylcholine-induced contraction in ring preparations of rat renal arteries. Acetylcholine (0.1-100 microM) caused a transient contraction in endothelium-intact arteries in a concentration-dependent manner. The contraction induced by acetylcholine (10 microM) was enhanced by pretreatment with NG-nitro-L-arginine (100 microM), a nitric oxide synthase inhibitor, and was abolished by mechanical removal of the endothelium. Atropine (0.1 microM), quinacrine (1 and 3 microM), manoalide (0.1 and 1 microM), aspirin (1 and 10 microM), indomethacin (30 and 300 nM), ONO-3708 (9,11-dimethyl-methane-11,12-methano-13,14-dihydro-13-aza-14- oxo-15(beta)-cyclophenyl-omega-pentenor-thromboxane A2 L-arginine salt) (10 nM), S-1452 (calcium (5Z)-1R,2S,3S,4S-7-[3-phenylsulphonyl-aminobicyclo[2.2.1]hep t-2yl]-5- heptenoate hydrate) (3 nM) and SQ29,548 ([1S- [1 alpha,2 beta(5Z),3 beta,4 alpha]]-7-[3-[[2-[(phenylamino) carbonyl]hydrazino]methyl]-7-oxabicyclo[2.2.1]hept-2-yl]-5-heptenoi++ + c acid) (3 and 10 nM), but not hexamethonium (1 microM), OKY-046 (sodium (E)-3-[4-(1- imidazolylmethyl)phenyl]-2-propenoic acid hydrochloride monohydrate) (100 microM) and CS-518 (sodium 2-(1-imidazolylmethyl)-4,5- dihydrobenzo[b]thiophene-6-carboxylate) (10 microM) significantly attenuated the acetylcholine (10 microM)-induced endothelium-dependent contractions in renal arteries pretreated with NG-nitro-L-arginine. These findings suggest that acetylcholine causes endothelium-dependent contraction by stimulation of muscarinic receptors in rat renal arteries, and that an arachidonic acid metabolite(s) of the cyclooxygenase pathway is involved in this endothelium-dependent contraction.

Acetylcholine↗