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Y Nonaka

Publications and source records attributed to Y Nonaka.

At least 73 records · Page 4Linked to original sources
PubMed1992

Functional expression of cDNAs for bovine 11 beta-hydroxylase-aldosterone synthases, P450(11 beta)-2 and -3 and their chimeras.

Two molecular species of bovine P450(11 beta), P450(11 beta)-2 and P450(11 beta)-3 have been identified, in which the amino acid differences were found at the 6th, 36th and 82nd positions from the NH2-termini of the mature proteins. They catalyzed the 11 beta-, 18- and 19-hydroxylation and aldosterone formation from 11-deoxycorticosterone, and the rate of production of 18-hydroxycorticosterone and aldosterone by P450(11 beta)-3 was greater than that by P450(11 beta)-2 [Morohashi et al., J. Biochem. 107 (1990) 635-640]. In this study, chimeric clones were constructed whose 6th, 36th and 82nd amino acid residues were exchanged with each other. Two original clones and six chimeric clones were expressed in COS-7 cells, and their steroidogenic activities studied. The ratio of aldosterone or 18-hydroxycorticosterone production to corticosterone production by one clone was compared with that of the other. The ratios for the four clones having Gly36 [P450(11 beta)-3 type] were 0.08-0.22, whereas those for the clones having Ser36 [P450(11 beta)-2 type] were 0.03-0.05, suggesting that the Gly36 structure is important for aldosterone production.

Animals↗
PubMed1992

The blood vascular bed of the human pancreas, with special reference to the insulo-acinar portal system. Scanning electron microscopy of corrosion casts.

Microdissection and scanning electron microscopy of corrosion casts showed the structures of the vascular bed of the human pancreas to consist mainly of the capillary plexuses of the exocrine lobules, extralobular ducts and endocrine islets. A considerable number of the exocrine lobules were found to contain one to four marked endocrine islets larger than 30 microns in diameter. These intralobular islets received one or more arterioles (afferent vessels) and emitted conspicuous insulo-acinar portal vessels which continued into the lobular capillaries, suggesting insular control over the functions of the exocrine acini of the pancreas. Direct drainage of the intralobular islets into the veins was never reproduced. Not excluding the possibility that some lobules might contain smaller, unidentifiable islets, there nonetheless were many lobules which directly received arterioles. These lobules are free of control by an islet. Rarely, an islet was located in the interlobular tissue space or along an extralobular duct. Such an extralobular islet issued no portal vessels, and drained into the interlobular or periductal veins. The surface of this type of islet comprised a thin network of fine capillaries. A possibility was suggested that this cortical network might be homologous with the lobular capillaries. No portal route was observed between the islets and extralobular ducts. Few connections were noted between the capillary plexuses of the lobules and ducts.

Adult↗
PubMed1991

Functional expression of the cDNAs encoding rat 11 beta-hydroxylase [cytochrome P450(11 beta)] and aldosterone synthase [cytochrome P450(11 beta, aldo)].

Expression plasmids containing two cDNAs of a rat cytochrome P450(11 beta) family, pcP450(11 beta)-62 [Nonaka, Y., Matsukawa, N., Morohashi, K., Omura, T., Ogihara, T., Teraoka, H. & Okamoto, M. (1989) FEBS Lett. 255, 21-26] and pcP450(11 beta, aldo)-46 [Matsukawa, N., Nonaka, Y., Ying, Z., Higaki, J., Ogihara, T. & Okamoto, M. (1990) Biochem. Biophys. Res. Commun. 169, 245-252], were constructed and introduced into COS-7 cells by electroporation. Enzymatic activities of the expressed cytochromes P450(11 beta) and P450(11 beta, aldo) were determined by using 11-deoxycorticosterone, corticosterone, 18-hydroxy-11-deoxycorticosterone, 18-hydroxycorticosterone, or 19-hydroxy-11-deoxycorticosterone as a substrate. Cytochrome P450(11 beta) catalyzed 11 beta-, 18- and 19-hydroxylations of 11-deoxycorticosterone and 19-oxidation or 19-hydroxy-11-deoxycorticosterone at substantial rates, 18-hydroxylation of corticosterone at a very low rate, but no aldosterone production. Cytochrome P450(11 beta, aldo) catalyzed 11 beta- and 18-hydroxylations of 11-deoxycorticosterone, 18-hydroxylation of corticosterone and aldosterone production from 11-deoxycorticosterone or corticosterone. But neither 19-hydroxylation of 11-deoxycorticosterone nor 19-oxidation of 19-hydroxy-11-deoxycorticosterone was catalyzed by cytochrome P450(11 beta, aldo).

Adrenodoxin↗
PubMed1991

Localization of the gene transcripts of 11 beta-hydroxylase and aldosterone synthase in the rat adrenal cortex by in situ hybridization.

Using in situ hybridization, localization of the gene transcripts of 11 beta-hydroxylase and aldosterone synthase was investigated in order to clarify the sites for the synthesis of corticosterone (glucocorticoid) and aldosterone (mineralocorticoid) in the rat adrenal cortex. The gene transcript of 11 beta-hydroxylase was localized in all the endocrine cells of the entire adrenal cortex, while that of aldosterone synthase was exclusively confined in zona glomerulosa cells. These results represent that every endocrine cell of all the cortical zones synthesizes 11 beta-hydroxylase which converts 11-deoxycorticosterone to corticosterone, and only glomerulosa cells synthesize aldosterone synthase which produces aldosterone from corticosterone. Thus it is clearly shown that zona glomerulosa cells synthesize mineralocorticoid, while zona fasciculata as well as reticularis cells produce glucocorticoid.

Adrenal Cortex↗
PubMed1991

Bovine adrenal cytochrome P-450(11 beta)-mediated conversion of 11-deoxycortisol to 18- and 19-hydroxy derivatives; structural analysis by 1H-NMR.

Incubation of 11-deoxycortisol with a cytochrome P-450(11 beta)-reconstituted system yielded, in addition to cortisol, several new steroid products. In this study, structures of the three steroid products were elucidated. Retention time of the first product (Peak 2 substance) coincided with that of authentic 18-hydroxycortisol on reverse phase HPLC. To further confirm the chemical identity of this product, the purified sample was subjected to 1H-NMR analysis. The spectrum was essentially identical to that of 18-hydroxycortisol. The retention time of the second product (Peak 3 substance) did not coincide with those of commonly occurring steroids. The one- and two-dimension 1H-NMR spectra provided strong evidence for its structure of 19-hydroxy-11-deoxycortisol. The retention time of the third product (Peak 4 substance) did not coincide with those of commonly occurring steroids. The 1H-NMR spectrum showed the presence of signals of 19-CH3 and 18-CH2 protons. There was also evidence that this product is not hydroxylated at the 11-position. Further analysis of the COSY spectra identified its structure as 18-hydroxy-11-deoxycortisol. From these results, we conclude that bovine P-450(11 beta) can catalyze the hydroxylation of 11-deoxycortisol at 11 beta-, 18- and 19-positions and produce cortisol, 18-hydroxy-11-deoxycortisol, 18-hydroxycortisol and 19-hydroxy-11-deoxycortisol.

Adrenal Cortex↗
PubMed1991

Molecular nature of aldosterone synthase, a member of cytochrome P-450(11 beta) family.

The molecular nature of the aldosterone synthesizing enzymes of cattle and rat is discussed. In bovine adrenal cortex, one molecular species of cytochrome P-450(11 beta) catalyzes aldosterone synthesis as well as 11 beta-hydroxylation. The intactness of the mitochondrial membrane surrounding P-450(11 beta) in the zonae fasciculata-reticularis is essential to keep the aldosterone synthesizing activity of the cytochrome in these zones latent. In rat adrenal cortex, two distinct molecules belonging to a P-450(11 beta) family exist. One is 11 beta-hydroxylase, and the other aldosterone synthase.

Adrenal Cortex↗
PubMed1991

On the ligand-protein and ligand-flavin interactions in NADPH-adrenodoxin reductase as studied by 31P- and 13C-NMR. Use of 13C-enriched FAD as a probe.

The interaction between 2',5'-ADP and NADPH-adrenodoxin reductase from bovine adrenocortical mitochondria was examined by titrating the enzyme with 2',5'-ADP, while the 31P-signals of 2',5'-ADP were being monitored by 31P-NMR. From the titration profile, the dissociation constant for the complex of the enzyme with 2',5'-ADP was estimated to be 0.22 +/- 0.05 mM. Adrenodoxin reductase was reconstituted with 13C-enriched FADs. The 13C-enriched FADs used were [2-13C]-, [4,10 alpha-13C2]-, and [4 alpha-13C]FAD. The 13C-NMR spectra of these reconstituted enzyme preparations showed 13C-resonance peaks corresponding to the enriched carbon atoms at 160.6 , 165.1, 136.6, and 152.4 ppm (2-, 4-, 4 alpha-, and 10 alpha-13C atoms, respectively). When 2',5'-ADP was bound to the reconstituted enzyme, these 13C-resonance peaks did not shift appreciably from those of the unbound enzyme, whereas in the complex of the reconstituted enzyme with NADP+, the signals for 4- and 10 alpha-13C shifted to higher fields by 2.1 and 0.7 ppm, respectively and the 4 alpha-13C signal shifted to a lower field by 1.4 ppm. These results suggest that in the complex of the enzyme with NADP+ the pyridine moiety is located in the vicinity of C(4 alpha)-C(4) region and that the pi-electron density of the 4 alpha-position of flavin is decreased in the enzyme-NADP+ complex. This argues in favor of the electron transfer from the dihydropyridine moiety of NADPH to the electron-deficient N(5) = C(4 alpha) region of flavin.

Adenosine Diphosphate↗
PubMed1991

The possible involvement of free radical scavenging properties in the actions of cytokines.

Among the cytokines which show an antiproliferative effect on certain tumor cells, we investigated the free radical scavenging properties of tumor necrosis factor-alpha, interferon-alpha, and interferon-gamma. The scavenging properties were determined in vitro by the ability to reduce DPPH free radicals. All of these cytokines exhibited marked DPPH free radical scavenging ability.

Bepridil↗
PubMed1990

Molecular cloning and expression of cDNAS encoding rat aldosterone synthase: variants of cytochrome P-450(11 beta).

Two distinct forms of cDNA encoding rat aldosterone synthase were cloned from an adrenal capsular tissue cDNA library. The deduced amino acid sequences showed that one of the enzymes (P-450(11 beta),aldo-1) had a long extension peptide composed of 34 amino acid residues while the other (P-450(11 beta),aldo-2) had an extension peptide identical to that of rat P-450(11 beta). Glu at the 320th position of P-450(11 beta),aldo-1 was replaced with Lys in P-450(11 beta),aldo-2. The amino acid sequence of the aldosterone synthase was highly homologous (81%) to rat P-450(11 beta). Constructed expression vector containing the cDNA for extension peptide of P-450(11 beta) and the mature protein of P-450(11 beta),aldo-1 was transfected into COS-7 cells. The cells converted 11-deoxycorticosterone into corticosterone, 18-hydroxycorticosterone, and aldosterone.

Amino Acid Sequence↗
PubMed1990

Rate enhancement of the electron transfer of the adrenodoxin-adrenodoxin reductase system by inorganic and nucleotide phosphates.

Phosphate and pyrophosphate increased the rate of reduction of adrenodoxin by NADPH-adrenodoxin reductase and NADPH, pyrophosphate being one order more effective than the former. However, the cytochrome c reduction by the electron transport system was inhibited in the presence of inorganic (pyro)phosphate. On the other hand, ADP and ATP enhanced the rates of reduction of both adrenodoxin and cytochrome c through adrenodoxin by the electron transport system. GTP also enhanced the rate of reduction of cytochrome c by this system, whereas AMP showed no appreciable enhancement. These inorganic and nucleotide phosphates did not affect the rate of ferricyanide reduction by the reductase.

Adenosine Triphosphate↗
PubMed1990

Enzymatic activities of P-450(11 beta)s expressed by two cDNAs in COS-7 cells.

Expression plasmids were constructed using two cDNA clones of P-450(11 beta), pcP-450-(11 beta)-2, and pcP-450(11 beta)-3 (Morohashi et al. (1987) J. Biochem. 102, 559-568 and Kirita et al. (1988) J. Biochem. 104, 683-686), and introduced into COS-7 cells by electroporation. The expression of P-450(11 beta) proteins and their localization in the mitochondria were demonstrated by immunoblotting, immunofluorescence microscopy, and immunoelectron microscopy. The enzymatic activities of the expressed P-450(11 beta)s were determined using deoxycorticosterone (DOC), deoxycortisol, and corticosterone as substrates. Though the activities of the two P-450(11 beta)s for 11-, 18-, and 19-hydroxylation of DOC were almost equal, the production of 18-hydroxycorticosterone and aldosterone from corticosterone by P-450(11 beta)-3 was greater than that by P-450(11 beta)-2.

Animals↗
PubMed1989

Molecular cloning and sequence analysis of cDNA encoding rat adrenal cytochrome P-450(11)beta.

A cDNA clone encoding cytochrome P-450(11)beta of rat adrenal has been cloned and sequenced using a bovine P-450(11)beta cDNA insert (pcP-450(11 beta)-2; (1987) J. Biochem. 102, 559-568) as a probe. The nucleotide sequence contains an open reading frame sufficient to encode the entire amino acid sequence of a P-450(11)beta precursor protein consisting of 499 amino acids including an extension peptide of 24 amino acids at the NH2-terminus. The cDNA contains 1247 nucleotides at the 3'-noncoding region including 51 nucleotides of poly A, but lacks the 5'-noncoding region. The deduced amino acid sequence shows 61% similarity to that of bovine P-450(11)beta. Putative binding sites for heme and steroid are highly conserved among steroidogenic P-450s of known structure.

Adrenal Glands↗
PubMed1989

Rate enhancement of the electron transfer the adrenodoxin-adrenodoxin reductase system by dicarboxylic acids.

The rate of electron transport in the cytochrome P-450 system in adrenocortical mitochondria was studied with purified adrenodoxin reductase, adrenodoxin and cytochrome c. Oxaloacetate enhanced the rate at concentrations of less than 1 mM; malate, succinate and fumarate enhanced the rate to a lesser extent; and pyruvate and alpha-ketoglutarate had no appreciable effect. The rate enhancement was observed when the reagents were preincubated with adrenodoxin, but not with adrenodoxin reductase. Rate enhancement was also evident when the rate limiting step was at adrenodoxin in the electron transport system.

Adrenodoxin↗
PubMed1989

Immunohistochemical demonstration of adrenodoxin reductase in bovine and human adrenals.

Adrenodoxin reductase (ADR) was purified from bovine adrenocortical mitochondria and specific antibody was raised in rabbits. Immunohistochemical analysis of ADR was performed in the bovine and human adrenals. ADR was present in all of the zones in both bovine and non-pathological human adrenal cortex. In non-pathological human adrenals, the immunoreactivity was particularly prominent in the zona glomerulosa (ZG) and reticularis (ZR). Intensive immunoreactivity was observed in the ZG and some cells of the outer fasciculata and the ZR in the adrenal glands with idiopathic hyperaldosteronism. In adrenal glands with Cushing's disease, immunoreactivity was present in the compact cells of cortical micronodules. In all cases, sites of immunoreactivity correspond to sites of increased steroidogenesis. In aldosteronoma and cortical adenoma with Cushing's syndrome, the immunoreactivity was generally marked in compact cells but not in large cells with clear cytoplasm, ADR was present in the ZG and the ZR, and the ZG in the non-neoplastic adrenal glands attached to aldosteronoma and Cushing's adenoma, respectively. ADR was present in the compact cells in adrenocortical carcinoma clinically manifesting Cushing's syndrome.

Adrenal Gland Diseases↗
PubMed1989

Neurosis from the viewpoint of DIS (Diagnostic Interview Schedule).

We examined the relationship between clinical and DIS-Lifetime diagnoses given independently on 106 psychiatric patients clinically diagnosed as suffering from neurosis. They had many coexisting DIS diagnoses, and some of them had no DIS diagnosis. The key to the coexistence relationships in DIS diagnosis was a major depressive episode, and the subjects were classified into four types by the DIS coexistence relationships; Type I: 28 cases (26.4%) had coexisting diagnoses belonging to anxiety disorders or somatoform disorders, in addition to a major depressive episode. They were suffering from clinically severe neurosis accompanied by borderline personality disorder. Type II: 30 cases (28.3%) belonged to anxiety disorders or somatoform disorders without a major depressive episode without anxiety disorders or somatoform disorders, and had clinically depressive neurosis or depressive episode with less distortion of the personality. Type IV: 30 cases (28.3%) were other than Type I-III, and were clinically similar to symptomatic neurosis.

Adolescent↗
PubMed1989

Changes in levels of mucosal glycoprotein on cysteamine-induced duodenal ulcers in rats.

Duodenal ulcers were induced in rats by cysteamine administration. Time-course changes in duodenal hexosamine, blood flow and intragastric pH were measured, as well as changes in the susceptibility of Brunner's glands to concanavalin A (Con A) staining. Hexosamine contents in the duodenum decreased significantly at 5 and 24 h. In the control group, Brunner's glands were stained brown using Con A staining. One day after cysteamine administration, the extent of Con A staining markedly decreased. At 3 and 7 days, gland susceptibility to staining was essentially the same as that of the control group.

Animals↗
PubMed1988

Inhibition of bovine adrenocortical mitochondrial cytochrome P-450(11)beta-mediated reactions by imidazole derivatives and mineralocorticoid analogs.

The effects of several imidazole antimycotic agents, an imidazole and several mineralocorticoid analogs on the cytochrome P-450(11)beta-catalyzed 11 beta-hydroxylation of 11-deoxycorticosterone and aldosterone synthesis were examined. Ketoconazole, clotrimazole, miconazole and etomidate were found to be potent inhibitors of the reactions, causing 50% inhibition of the 11 beta-hydroxylase activity at concentrations between 10(-8) and 10(-7) M. The potency of etomidate as to the inhibition of aldosterone- and 18-hydroxycorticosterone-production was found to be almost equal to that in the case of 11 beta-hydroxylation. Spironolactone and other newly synthesized mineralocorticoid analogs were also found to inhibit the cytochrome P-450(11)beta-mediated reactions. The ID50 values of these drugs for inhibition of the 11 beta-hydroxylase activity were almost equal to those in the case of the aldosterone- and 18-hydroxycorticosterone-biosynthetic activities. The results of kinetical studies indicated that one of the mineralocorticoid analogs, Compound 23-0586, acts as a competitive inhibitor for the cytochrome P-450(11)beta-mediated reactions.

Adrenal Cortex↗