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Biomedical subjects

Y Ohyama

Publications and source records attributed to Y Ohyama.

At least 145 records · Page 8Linked to original sources
PubMed1992

Stable expression of natriuretic peptide receptors: effects of HS-142-1, a non-peptide ANP antagonist.

We established clonal cell lines stably expressing each of two subtypes of membrane bound guanylate cyclases (GC-A and GC-B), which are known as natriuretic peptide receptors. Using these cell lines, we showed that GC-A is an ANP/BNP receptor, whereas GC-B is a specific receptor for CNP. Effects of HS-142-1, a novel non-peptide ANP antagonist, on GC-A and GC-B were examined by using these cells. In cells expressing either GC-A or GC-B, HS-142-1 inhibited cGMP production elicited by ANP or CNP with IC50 values of 1.8 micrograms/ml and 1.5 micrograms/ml, respectively, and also competitively blocked specific binding of the natriuretic peptides with IC50 values of 2.2 micrograms/ml and 3.3 micrograms/ml, respectively. These results indicate that HS-142-1 is a potent antagonist of CNP as well as ANP. We also showed that CNP suppressed the growth of cells expressing GC-B by 22% and that HS-142-1 blocked the antiproliferative action of CNP.

Animals↗
PubMed1992

Parathyroid hormone inhibits 25-hydroxyvitamin D3-24-hydroxylase mRNA expression stimulated by 1 alpha,25-dihydroxyvitamin D3 in rat kidney but not in intestine.

Using a cDNA probe for rat renal 24-hydroxylase, expression of its mRNA was compared in the rat kidney and intestine. Vitamin D-deficient rats received a single injection of 1 alpha,25-dihydroxyvitamin D3. Expression of 24-hydroxylase mRNA was first detected in the kidney at 3-h post-injection and increased thereafter. Similarly, 24-hydroxylase mRNA was expressed in the intestine after 1 alpha,25-dihydroxyvitamin D3 injection. However, the dose level of 1 alpha,25-dihydroxyvitamin D3 required to induce the intestinal 24-hydroxylase mRNA expression was only 1/100 the amount required to induce renal 24-hydroxylase mRNA. Induction of intestinal 24-hydroxylase mRNA expression by 1 alpha,25-dihydroxyvitamin D3 was far more rapid than that of renal 24-hydroxylase mRNA. Thyroparathyroidectomy shortened the time required to induce expression of renal, but not intestinal, 24-hydroxylase mRNA. Administration of either parathyroid hormone or cAMP to vitamin D-deficient rats greatly reduced the expression of 24-hydroxylase mRNA in the kidney but not in the intestine. When rats were fed a vitamin D-repleted diet containing 0.7% (adequate) or 0.03% (low) calcium for 2 weeks, intestinal expression of 24-hydroxylase mRNA could be induced only in the low calcium group. In contrast, renal mRNA expression was preferentially stimulated in the adequate calcium group. These results clearly demonstrate that the expression of 24-hydroxylase mRNA is down-regulated by parathyroid hormone in the kidney but not in the intestine.

Animals↗
PubMed1992

Cloning and characterization of two forms of C-type natriuretic peptide receptor in rat brain.

Two similar membrane bound guanylate cyclases (GC-A and GC-B) are known as natriuretic peptide receptors, but have not been well characterized yet. In this study, we have isolated two forms of GC-B cDNA clones along with GC-A cDNA clones from rat brain. The two forms of rat GC-B differ from each other only by 75bp deletion at 3'-flanking region of the putative transmembrane domain, the shorter form lacking the nucleotide binding site by the deletion. Expression of these cDNAs on mammalian cells revealed that (1) GC-B is a specific receptor for CNP whereas GC-A is stimulated effectively both by ANP and BNP, and (2) the two forms of GC-B possess practically the same high binding affinity for CNP while the shorter form could not induce cGMP production by the binding of CNP. These data indicate that in rat brain is present the non-functional receptor for CNP caused by the short deletion.

Amino Acid Sequence↗
PubMed1992

Analysis of the eicosapentaenoic acid (EPA) metabolite delta 17-6-keto-PGF1 alpha in human plasma by GC/SIM using [18O] delta 17-6-keto-PGF1 alpha.

A method of the microdetermination of delta 17-6-keto-PGF1 alpha, a hydrolyzed metabolite of PGI3, is described. An authentic delta 17-6-keto-PGF1 alpha (120 mg) was prepared from eicosapentaenoic acid (EPA) incubated with homogenate of bovine aortic intima. [18O]delta 17-6-Keto-PGF1 alpha was synthesized by repeating base-catalyzed hydrolysis of methyl ester derivatives in [18O]water, to obtain an internal standard in gas chromatography/selected ion monitoring (GC/SIM) of delta 17-6-keto-PGF1 alpha. Good linear response over the range of 10 pg-10ng was demonstrated. Chromatographic conditions using a MP-65HT column presented nearly baseline separation of delta 17-6-keto-PGF1 alpha and 6-keto-PGF1 alpha. We were able to detect delta 17-6-keto-PGF1 alpha in the range from 6 to 26 pg/ml of the human plasma. The present method can be applied to the determination of delta 17-6-keto-PGF1 alpha in the human urine and plasma.

6-Ketoprostaglandin F1 alpha↗
PubMed1992

Separation and concentration of delta 17-6-keto-PGF1 alpha using monoclonal antibody to omega 3-olefin structure of trienoic prostanoids.

A monoclonal antibody against cis-3-hexen-1-ol was prepared and used to separate and/or concentrate delta 17-6-keto-prostaglandin F1 alpha (PGF1 alpha) in the human sera. cis-3-Hexen-1-ol was conjugated with the human serum albumin (HSA) according to the N-succinimidylester method and hyperimmunized to BALB/c mouse. The monoclonal antibodies were obtained from hybridoma clones established by a fusion between SP2/0-Ag14-k13 mouse myeloma cells and splenocytes of a mouse. A monoclonal antibody, named 4G9-12B, recognized the epitope characteristic for omega 3-olefin structure. The 4G9-12B antibody became more specific for delta 17-6-keto-PGF1 alpha than 6-keto-PGF1 alpha by applying inhibition ELISA using amino-residue coating plates. Using the prepared immunoaffinity columns of this antibody, delta 17-6-keto-PGF1 alpha was clearly detected in 6 pg/ml of the human blood sera by GC/MS analysis. These results suggest that the monoclonal antibody to the partial structure of trienoic prostanoid, omega 3-olefin unit, and that its immunoaffinity columns are useful in separating and concentrating delta 17-6-keto-PGF1 alpha in the human blood or urine.

6-Ketoprostaglandin F1 alpha↗
PubMed1992

Purification and characterization of beta-galactoside-binding proteins from Caenorhabditis elegans.

Two carbohydrate-binding proteins (subunit molecular masses, 32 and 16 kDa, respectively) were isolated for the first time from a nematode, Caenorhabditis elegans. They were specifically extracted with lactose and adsorbed on asialofetuin-Sepharose in the absence of a metal ion. Although these two proteins were co-eluted from a gel filtration column at a position corresponding to an apparent molecular size of 30 kDa under non-denaturing conditions, they could be separated by reversed-phase chromatography. The 32 kDa protein, the main component, was further characterized. Together with its solubility, saccharide specificity and metal independence, some other structural properties, including its amino acid composition, UV spectrum, and partial amino acid sequence, strongly suggested that the 32 kDa protein is a member of a class of soluble beta-galactoside-binding lectins which had previously been only found in vertebrates.

Amino Acid Sequence↗
PubMed1992

Ontogenic expression of somatostatin-messenger RNA in the intestinal tract of neonatal rats.

Ontogenic expression of somatostatin (SRIF) -messenger RNA (mRNA) in the gastrointestinal tract was examined in neonatal rats aged from 1 day preterm to 60 days postpartum in comparison with that in the hypothalamus. SRIF-mRNA in the hypothalamus was already expressed in prenatal rats and its developmental change was relatively small. In contrast, a unique pattern of SRIF-mRNA expression was seen in the different intestinal regions, gastric antrum, duodenum, jejunum and colon. In the duodenum, SRIF-mRNA level was low at birth, markedly increased during the postnatal 3 days and declined to the previous level by day 21. Jejunal SRIF-mRNA was found in neonates but progressively decreased in a similar way to duodenum. On the contrary, gastric SRIF-mRNA level, which was low during early development, rose rapidly to a peak on day 21 and gradually declined to an adult level. In the colon age-related change was not conspicuous, remaining at a low level. These results indicate that (1) expression of SRIF gene in the intestinal tract is regulated by local factor(s) as well as developmental stage, and (2) shift of SRIF-mRNA pattern occurs during weaning from the duodenum-dominant infantile pattern to the gastric-dominant adult pattern.

Age Factors↗
PubMed1992

Usefulness and limitation of measurement of insulin-like growth factor binding protein-3 (IGFBP-3) for diagnosis of growth hormone deficiency.

To analyze the utility of insulin-like growth factor binding protein-3 (IGFBP-3) radioimmunoassay for diagnosis of growth hormone deficiency (GHD) we measured IGFBP-3 in sera from normal children, short children and patients with GHD. The sensitivity (true positive ratio) of IGFBP-3 for complete GHD (cGHD) was 93%, while the specificity (true negative ratio) for normal short children (NS) was 88%. In contrast, the sensitivity of IGFBP-3 for partial GHD (pGHD) was only 43%. The poor discrimination between patients with pGHD and NS may be the result of their relatively similar GH level, as compared to cGHD, or due to the limitations of GH stimulation tests. The specificity of IGFBP-3 for NS was excellent in children of all ages: less than 10 years old (87%) and older than 10 (88%). However, sensitivity for GHD was good for children less than 10 years old (84%) but poor for children older than 10 (64%). IGFBP-3 may be less sensitive for diagnosing GHD in older children because IGFBP-3 levels may also increase during puberty due to mechanisms independent of the GH-IGF-I axis.

Adolescent↗
PubMed1992

The influence of tetrodotoxin on the toxic effects of aconitine in vivo.

Both aconite toxins (aconitine, mesaconitine, and hypaconitine) and a pufferfish toxin (tetrodotoxin, TTX) were detected in the blood of a legal autopsy case. In order to elucidate the in vivo influence of TTX on the toxic effects of aconitine, a mixture of aconitine and TTX was administered to male ICR mice orally or intraperitoneally. The animal experiments revealed that the time of death due to aconitine was significantly delayed in proportion to the dose of TTX compared with the case for aconitine alone, and that the mortality of aconitine was lowered by TTX when the dose ratio of the two toxins was in a particular range. Accordingly, it is thought that the toxic effects of aconitine are attenuated by TTX in vivo.

Aconitine↗
PubMed1992

[Molecular biology of cytochrome P450 in kidney].

Cytochrome P450-catalyzed monooxygenation system is widespread in nature and many isoforms have been found in almost all the tissues of vertebrates. In the kidney, several isoforms of cytochrome P450 families, CYP1, 2, 3, 4, 24, 27 have been characterized by biological, immunochemical and molecular biological techniques. These cytochrome P450 isoforms are localized in the endoplasmic reticulum or mitochondria of kidney cortex and functional in the monooxygenation of a number of substrates, such as steroids, fatty acids, xenobiotics, and vitamin D. Particularly, hydroxylation of vitamin D at positions of 1 and 24 is known to be one of the most important functions of kidney specific P450 isoforms.

Animals↗
PubMed1991

Isolation and characterization of a cytochrome P-450 from rat kidney mitochondria that catalyzes the 24-hydroxylation of 25-hydroxyvitamin D3.

A cytochrome P-450 that catalyzes the 24-hydroxylation of 25-hydroxyvitamin D3 (P-450cc24: P-450cholecalciferol24) was purified to electrophoretic homogeneity from the kidney mitochondria of female rats treated with vitamin D3 (Ohyama, Y., Hayashi, S., and Okuda, K. (1989) FEBS Lett. 255, 405-408). The molecular weight was 53,000, and its absorption spectrum showed peaks characteristic of cytochrome P-450. The turnover number was 22 min-1 and the specific content was 2.8 nmol/mg protein. The N-terminal amino acid sequence, Arg-Ala-Pro-Lys-Glu-Val-Pro-Leu-, is different from the N-terminal sequence of any other cytochrome P-450s so far reported. Upon reconstitution with the electron-transferring system of the adrenal mitochondria, the enzyme showed a high activity in hydroxylating 25-hydroxyvitamin D3 as well as 1 alpha,25-dihydroxyvitamin D3 at position 24. However, the purified enzyme hydroxylated neither vitamin D3 nor 1 alpha-hydroxyvitamin D3. The enzyme was also inactive toward xenobiotics. The enzyme hydroxylated 25-hydroxyvitamin D3 at position 24 but not at 1 alpha, indicating that the enzyme is distinct from that catalyzing 1 alpha-hydroxylation. The reaction followed Michaelis-Menten kinetics, and the Km value for 25-hydroxyvitamin D3 was 2.8 microM. Both vitamin D3 and 1 alpha-hydroxyvitamin D3 inhibited the 24-hydroxylation of 25-hydroxyvitamin D3 in a competitive, concentration-dependent manner. 25-Hydroxyvitamin D3 24-hydroxylase activity was significantly inhibited by 7,8-benzoflavone, ketoconazole, and CO, whereas it was only slightly inhibited by aminoglutethimide, metyrapone, and SKF-525A. Mouse antibodies raised against the cytochrome P-450 inhibited the reaction about 70% and reacted with the P-450cc24 in immunoblotting but did not react with other kinds of cytochrome P-450 in rat liver microsomes and mitochondria.

Animals↗
PubMed1991

Cloning and expression of cDNA encoding 25-hydroxyvitamin D3 24-hydroxylase.

A cDNA encoding 25-hydroxyvitamin D3 24-hydroxylase (P450cc24) was isolated from a rat kidney cDNA library using specific antibodies to the enzyme. The isolated cDNA was 3.2 kbp long and contained a 1542-bp open reading frame encoding 514 amino acids. The deduced amino acid sequence contained a presequence typical of mitochondrial enzymes in the N-terminal region. The amino acid sequence shows less than 30% similarity to those of any other cytochrome P450s so far reported and, therefore, P450cc24 constitutes a novel family of P450. COS-7 cells transfected with the cDNA produced a protein that was reactive with the antibodies and catalyzed NADPH-dependent 24-hydroxylation of 25-hydroxyvitamin D3 in the presence of adrenodoxin and NADPH-adrenodoxin reductase. Using the cDNA as a probe we demonstrated that the increase of 24-hydroxylation activity caused by administration of vitamin D3 into rats was accompanied by an increase of the mRNA.

Amino Acid Sequence↗
PubMed1991

Novel derivatization and immunoextraction to improve microanalysis of 11-dehydrothromboxane B2 in human urine.

A new, highly selective procedure for the determination of 11-dehydrothromboxane B2 (11-dehydro-TXB2) in human urine is described. Following the addition of [19,19,20,20-2H4]11-dehydro-TXB2 as an internal standard, samples were extracted with an affinity column of anti-11-dehydro-TXB2. Conversion of the immunoextracted 11-dehydro-TXB2 into its 1-methyl ester-11-n-propylamide-9,12,15-tris-dimethylisopropylsilyl ether derivative was followed by gas chromatography-selected-ion monitoring. The mass spectrum of the 11-dehydro-TXB2 derivative was dominated by the base peak ion of [M-C3H7]+ at m/z 698, which accounted for more than 10% of the total ion current. A typical result showed that the immunoaffinity purification procedures provided an extremely clean alternative to more conventional methods of chromatographic fractionation, and that interfering substances from the urine matrix were almost entirely eliminated during the microanalysis.

Antibodies, Monoclonal↗
PubMed1991

Multi-functional property of rat liver mitochondrial cytochrome P-450.

To solve the problem of whether a common enzyme catalyzes both 5 beta-cholestane-3 alpha,7 alpha,12 alpha-triol 27-hydroxylation and 25-hydroxylation of 1 alpha-hydroxyvitamin D3 (a synthetic compound used therapeutically for vitamin D-deficient diseases) in rat liver mitochondria, enzymological and kinetic studies were performed. A cytochrome P-450 was purified from female rat liver mitochondria based on these catalytic activities and it was found that the two enzyme activities accompanied each other at all purification steps. The 5 beta-cholestane-3 alpha,7 alpha,12 alpha-triol 27-hydroxylation activity of the final preparation had a turnover number of 36 min-1, and the value of the corresponding 1 alpha-hydroxyvitamin D3 25-hydroxylation activity was 1.4 min-1. When the enzyme was partially denatured by heating at different temperatures, both enzyme activities declined in a parallel fashion. Treatment of the enzyme with N-bromosuccinimide decreased both enzyme activities in a similar manner. 5 beta-Cholestane-3 alpha,7 alpha,12 alpha-triol competitively inhibited 25-hydroxylation of 1 alpha-hydroxy-vitamin D3 and vice versa. From these results it was concluded that 5 beta-cholestane-3 alpha,7 alpha,12 alpha-triol 27-hydroxylation and 1 alpha-hydroxyvitamin D3 25-hydroxylation are catalyzed by a common enzyme in rat liver mitochondria.

Animals↗
PubMed1991

Triethylene-tetramine (trien) therapy for Wilson's disease.

Triethylene tetramine (trien), in increasing dose from 1.0-2.0 g/day to 2.5-3.0 g/day, was used for 4 Japanese patients with Wilson's disease who were intolerant of D-penicillamine (D-PC). Before the treatment, urinary copper excretion (UCE) was 70-96 micrograms/day. UCE increased to 1,512-2,352 micrograms/day on the day of initial administration, and remained at levels between 350-1,100 micrograms/day, thereafter. During 2 months of trien therapy, neurological deficits regressed in three patients, and only slightly in one patient. No adverse effects were observed. These results and the retrospective survey on 17 patients treated with D-PC confirmed that trien is less potent but a safer copper chelating agent than D-PC. The transient aggravation of neurological deficits seen in two patients during the early stage of the treatment suggested that trien, as D-PC, should be started in small doses and gradually increased.

Adolescent↗
PubMed1991

[A case of recurrent hepatocellular carcinoma after hepatic resection surviving over five years by hepatic arterial infusion of lipiodol-anticancer drug suspension].

A 63-year-old male with four intrahepatic recurrences of surgically resected hepatocellular carcinoma was admitted to our hospital in June 1985. He underwent lateral segmentectomy of the liver in November 1983. Pathologic finding of Edmondson II with liver cirrhosis had been confirmed by the operative specimen. Sizes of four recurrent tumors were assessed by CT as 3.5 x 2.2 cm, 2.6 x 2.2 cm, 2.2 x 2.2 cm and 2.2 x 2.2 cm, respectively. During five years until July 1990, the patient was treated with hepatic arterial infusion of Lipiodol-anticancer drug suspension eight times (total 5-FU 900 mg, ADM 77 mg, MMC 73 mg, and Lipiodol 36 ml) and hepatic arterial chemoembolization of MMC microcapsules one time. In addition, two hepatic arterial infusions of CDDP (total 70 mg) were given and 5-FU (total 10 g) was administered intravenously. Partial response (PR) was obtained for 19 months. Hepatic arterial infusion of Lipiodol-anticancer drug suspension was given only once every 6 months, and he maintained a good quality of life for over four and half years. The man died in July 1990. In general, multiple intrahepatic recurrence of surgical resected hepatocellular carcinoma has a poor prognosis. Therefore it was considered that hepatic arterial infusion of this drug brought about the relatively long survival of more than five years.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed1991

[A case of ventricular septal defect associated with active infective endocarditis which was successfully treated by triple valve replacement and ventricular septal defect patch closure].

A 47-year-old man was hospitalized in May, 1990, because of breathlessness and continuous fever which appeared about 4 weeks after he had had his periodontic tooth removed in December, 1989. He had been diagnosed as having ventricular septal defect (VSD) at the age of 6 years. When he was hospitalized, he was in a condition of class IV by NYHA classification, with a white blood cell count of 17,300/mm3, an increase in CRP, a red blood cell sedimentation rate of 108 mm/hr, and positive alpha-streptococcus in blood culture. His cardiothoracic ratio was 64% with signs of pulmonary congestion on a chest X-ray film. Echocardiography revealed the presence of VSD and huge vegetations on the tricuspid, mitral and aortic valves. He was considered to have active infective endocarditis (AIE) which had presumably been provoked by VSD and the tooth removal. Penicillin G at a daily dose of 20 million units and gentamicin at a daily dose of 80 mg were intravenously administered to treat the alpha-streptococcus infection for about 4 weeks. Furosemide was used for congestive heart failure. Since, although his cardiac function appeared to have been improved, the signs and symptoms of the infection persisted, triple valve replacement for the tricuspid, mitral and aortic valves and patch closure of the VSD were performed 4 weeks after the hospitalization. The operation revealed inflammatory lesions extending from the endocardium of the right ventricle to the mitral valves through the VSD, and huge vegetations on the tricuspid, mitral and aortic valves. The operation was successful and the inflammatory areas gradually disappeared.(ABSTRACT TRUNCATED AT 250 WORDS)

Aortic Valve↗