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N Ohnuma

Publications and source records attributed to N Ohnuma.

At least 109 records · Page 6Linked to original sources
PubMed1986

Difference in molecular size of receptors for alpha-rat atrial natriuretic polypeptide among the kidney, aorta, and adrenal gland as identified by direct UV-photoaffinity labeling.

In order to identify the molecular size of receptors for alpha-rat atrial natriuretic polypeptide (alpha-rANP), we utilized the direct UV irradiation method for photoaffinity labeling with the biologically active [125I] alpha-rANP. In the preparation of isolated glomerulus and the inner medullary collecting duct (IMCT)-rich fraction, the autoradiograms of the electrophoresed sodium dodecyl sulfate (SDS)-polyacrylamide gels showed a single radioactive band which is displaceable with unlabeled alpha-rANP. The dose-dependent displacement fit very well with a binding-inhibition curve representing the binding affinity of 6.5 X 10(-10) M. The molecular size of the ligand-receptor complex was about 65,000 daltons for both glomerulus and IMCT-rich fraction. In contrast, in homogenate of the aorta and adrenal gland, the ligand-receptor complex was 140,000 daltons.

Adrenal Glands↗
PubMed1986

Studies of the influence of chloro-substituent sites and conformational energy in polychlorinated biphenyls on uroporphyrin formation in chick-embryo liver cell cultures.

Treatment of cultured chick-embryo liver cells with polychlorinated biphenyls (PCB) results in decreased uroporphyrinogen decarboxylase activity and increased uroporphyrin accumulation. In the present study we examined the effect of the chloro- or bromo-substituent sites in biphenyls (BP) on uroporphyrin accumulation in cultured hepatocytes and the three-dimensional structure of these congeners determined by molecular orbital calculations using a MNDO ('modified neglect of diatomic overlap') method. Among 20 congeners examined, those which were effective in stimulating porphyrin accumulation contained at least two Cl or Br atoms at the lateral adjacent positions in each phenyl ring, e.g. 3,4,3',4'-tetrachloro-, 2,4,3',4'-tetrachloro-, 3,4,5,3',4',5'-hexachloro- and 3,4,5,3',4',5'-hexabromobiphenyl, whereas those which contained less than two halogen atoms or more than three halogen atoms in each phenyl ring or those which contained halogen atoms at 2,2'-positions were not effective. On the basis of the conformational energy (delta E, difference from the most stable conformational energy), which is calculated as a function of the dihedral angle (theta) between the two phenyl rings, biphenyl congeners can be classified into four groups with different conformations. The conformation of active PCB was relatively flexible, whereas inactive species had a rigidly angulated conformation. Furthermore, the calculated probability of the conformation distribution for each congener indicated that the probability of co-planarity was higher for active biphenyls than for inactive congeners. These structural characteristics suggest the significance of both the chloro-substituent sites and the conformational energy reflecting the phenyl-ring twist angles in determining the inhibitory effect of PCB on uroporphyrinogen decarboxylase activity.

Animals↗
PubMed1986

Localization of binding sites for alpha-rat atrial natriuretic polypeptide in rat kidney.

To determine the intrarenal localization of receptors for atrial natriuretic polypeptide (ANP) in the rat, we performed autoradiography and binding assay by using 125I-labeled alpha-rat ANP (alpha-rANP). Autoradiography at the slice and microscopic level in the kidney and binding assay in isolated glomeruli demonstrated that receptors in the renal cortex were distributed mainly in glomeruli. Although dense silver grains were distributed diffusely both in inner medulla and outer stripe of outer medulla, a marked displacement of the grains was observed only in the inner medulla. Autoradiography at the microscopic level also showed that silver grains were distributed in the renal artery, renal pelvis, and inner medullary collecting tubule (IMCT) prepared by the microdissection method, but not in the arcuate artery, interlobular artery, and afferent or efferent arterioles. Specific binding was demonstrated in the isolated glomeruli and the preparation was rich in fragments of IMCT. Apparent binding affinity (Kd) and receptor density (R) for 125I-labeled alpha-rANP in isolated glomeruli and IMCT were Kd = 3.2 and 21 X 10(-9) M, and R = 320 and 420 fmol/mg protein, respectively. These observations suggest that alpha-rANP has a physiological action on glomeruli and possibly on the inner medullary collecting tubules in addition to the renal artery.

Animals↗
PubMed1986

Intrarenal localization of receptors for alpha-rat atrial natriuretic polypeptide: an autoradiographic study with [125I]-labeled ligand injected in vivo into the rat aorta.

We examined intrarenal localization of receptors for alpha-rat atrial natriuretic polypeptide (alpha-rANP) by injecting [125I]-labeled ligand in vivo into the rat aorta. We found that the receptors for alpha-rANP are distributed also on the vasa recta of the outer and inner medulla in addition to the previously reported sites, i.e., the renal arteries, renal pelvis, glomeruli, and inner medullary tissues including collecting tubules. In the vascular bundle of the outer medulla, the majority of grains was preferentially localized on the arterial vasa recta. The electron microscopic autoradiography of the glomerulus showed that the binding sites were mainly localized on the foot process of the podocyte. Since alpha-rANP injected into the aorta under physiological conditions was bound to the glomerulus and vasa recta in the kidney, the effect of ANP on these binding sites may be important in the mechanism of natriuresis.

Animals↗
PubMed1985

[Bowel function of spina bifida].

Bowel function of the spina bifida was studied by clinical assessment and anorectal manometric studies. Anorectal pressure profiles were well correlated with clinical assessment but not with recto-anal inhibitory reflexes. Early training of the defecation is emphasized to obtain good bowel function.

Defecation↗
PubMed1983

Isolation and identification of 1,25-dihydroxy-24-oxo-vitamin D3 and 1,23,25-trihydroxy-24-oxo-vitamin D3. New metabolites of vitamin D3 produced by a C-24 oxidation pathway of metabolism for 1,25-dihydroxyvitamin D3 present in intestine and kidney.

Two new vitamin D metabolites were isolated in pure form from separate incubations of homogenates of chick small intestinal mucosa or rat kidney employing either 1 alpha,25-dihydroxyvitamin D3 (28 microM) or 1 alpha,24R,25-trihydroxyvitamin D3 as substrate (0.17-1.3 microM). The newly characterized compounds and the amounts isolated in pure form from separate isolations are, respectively: 1 alpha,25-dihydroxy-24-oxo-vitamin D3 (1,25(OH)2-24-oxo-D3), 147 micrograms from kidney and 4.2 and 40 micrograms from intestine; 1 alpha,23,25-trihydroxy-24-oxo-vitamin D3 (1,23,25(OH)3-24-oxo-D3), 155 micrograms from kidney and 5.9 and 34 micrograms from intestine. Their structures were identified after extensive high pressure liquid chromatography by means of ultraviolet absorption spectrometry, mass spectrometry of the free compounds and their trimethylsilylated derivatives, proton nuclear magnetic resonance spectrometry, specific chemical reduction of the 24-oxo functionality with sodium borohydride, as well as direct comparison with synthetic 1,25(OH)2-24-oxo-D3. These structural assignments for both compounds correct previous determinations which had been proposed (Ohnuma, N., Kruse, J. R., Popjak, G., and Norman, A. W. (1982) J. Biol. Chem. 257, 5097-5102). The activity of the C-24 oxidation pathway used for the production of the 1,25(OH)2-24-oxo-D3 and 1,23,25(OH)3-24-oxo-D3 can be enhanced 10-fold by prior priming of the chicks or rats with a single intravenous dose of 1,25(OH)2D3 (1-12 nmol/100 g body weight); the induction of the enzyme activity is maximal by 3-6 h and returns to basal levels within 12 h. Further, 1,25(OH)2D3, 1,24,25(OH)3D3, and 1,25(OH)2-24-oxo-D3 all were found to be capable of serving as a precursor with chick intestine and rat kidney homogenates of 1,23,25(OH)3-24-oxo-D3. Collectively these results suggest the existence of a C-24 oxidation pathway for metabolism of 1,25(OH)2D3 by the target intestinal mucosa and kidney to 1,23,25(OH)3-24-oxo-D3. The pathway may play an important role in controlling the tissue levels of this hormonally active form of vitamin D3.

Animals↗
PubMed1983

Biological activity assessment of the vitamin D metabolites 1,25-dihydroxy-24-oxo-vitamin D3 and 1,23,25-trihydroxy-24-oxo-vitamin D3.

Two new metabolites of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3], namely 1,25(OH)2-24-oxo-vitamin D3 and 1,23,25(OH)3-24-oxo-vitamin D3, have been prepared in vitro using chick intestinal mucosal homogenates. To investigate the binding of 1,25(OH)2-[23-3H]-24-oxo-D3 and 1,23,25(OH)3-[23-3H]-24-oxo-D3 to the chick intestinal receptor we have isolated both metabolites in radioactive form using an incubation system containing 1,25(OH)2-[23,24-3H))-D3 with a specific radioactivity of 5.6 Ci/mmol. Both metabolites were highly purified by using Sephadex LH-20 chromatography followed by high-pressure liquid chromatography (HPLC). Sucrose density gradient sedimentation analysis showed specific binding of both tritium-labeled metabolites to the chick intestinal cytosol receptor. Experiments were carried out to determine the relative effectiveness of binding to the chick intestinal mucosa receptor for 1,25(OH)2D3. The results are expressed as relative competitive index (RCI), where the RCI is defined as 100 for 1,25(OH)2D3. Whereas the RCI obtained for 1,25(OH)2-24-oxo-D3 was 98 +/- 2 (SE), the RCI for 1,23,25(OH)3-24-oxo-D3 was only 28 +/- 6 (SE). Also, the biological activity of both new metabolites was assessed in vivo in the chick. In our assay for intestinal calcium absorption, 1,25(OH)2-24-oxo-D3 was active at a dose level of 1.63 and 4.88 nmol/bird (at 14 h), whereas 1,23,25(OH)3-24-oxo-D3 showed only weak biological activity in this system. In our assay for bone calcium mobilization, administration of both new metabolites showed modest activity at the 4.88-nmol dose level, which was reduced at the 1.63-nmol dose level. The results indicate that biological activity declines as 1,25(OH)2D3 is metabolized to 1,24R,25(OH)3D3, 1,25(OH)2-24-oxo-D3, and then 1,23,25(OH)3-24-oxo-D3.

Animals↗
PubMed1982

Identification of a new C-23 oxidation pathway of metabolism for 1,25-dihydroxyvitamin D3 present in intestine and kidney.

Evidence is presented for the existence of a new C-23 oxidation pathway for the metabolism of the hormonally active form of vitamin D3, namely 1 alpha,25-dihydroxyvitamin D3 (1,25(OH)2D3). Homogenates of intestinal mucosa or kidney, but not liver, from rats and chicks convert 1,25(OH)2[26,27-3H]D3 into two new metabolites, 1 alpha,25-dihydroxy-23-oxo-vitamin D3 (1,25(OH)2-23-oxo-D3) and 1 alpha,25,26-trihydroxy-23-oxo-vitamin D3 (1,25,26(OH)3-23-oxo-D3), and an unknown metabolite(s) which has been possibly subjected to side chain modification/cleavage so that the tritium of the substrate has been converted into a form which is volatile. The isolation and chemical characterization of 1,25(OH)2-23-oxo-D3 and 1,25,26(OH)3-23-oxo-D3 from homogenates of chick intestinal mucosa has recently been described (Ohnuma, N., Kruse, J., Popjak, G., and Norman, A. W. (1982) J. Biol. Chem. 257, 5097-5102). Based on kinetic studies with chick intestinal homogenates, the proposed C-23 oxidation pathway is: 1,25(OH)2D3 leads to 1,25(OH)2-23-oxo-D3 leads to 1,25,26(OH)3-23-oxo-D3 leads to unknown metabolite with altered side chain. The relative enzyme activities of the C-23 pathway in tissues from vitamin D-replete chicks are: intestine, kidney liver, 18:3:less than 1. The activity of the C-23 pathway in homogenates of chick intestinal mucosa can be enhanced 10 x by prior priming of the birds with a single intravenous dose of 500 ng (1.3 nmol) of 1,25(OH)2D3; the induction of the enzyme activity is maximal by 3-6 h and returns to basal levels by 12 h. A comparison was made in chick and rat intestinal homogenates of this C-23 pathway and the previously known C-24 oxidation pathway, which converts 1,25(OH)2D3 into 1,24,25-trihydroxyvitamin D3 (1,24,25(OH)3D3); it was calculated that under these conditions 72% of the 1,25(OH)2D3 was metabolized by the C-23 oxidation pathway, 13% by the C-24 pathway, and only 14% by other as yet unspecified pathways. It is proposed that the newly discovered C-23 pathway for metabolism of 1,25(OH)2D3 by the target intestinal mucosa and kidney may play a prominent role under physiological conditions of controlling the tissue levels of this hormonally active form of vitamin D3.

Animals↗
PubMed1982

Isolation and chemical characterization of two new vitamin D metabolites produced by the intestine. 1,25-Dihydroxy-23-oxo-vitamin D3 and 1,25,26-trihydroxy-23-oxo-vitamin D3.

Two new vitamin D metabolites were isolated in pure form from incubations of 53 nM 1,25-dihydroxyvitamin D3 with homogenates of small intestinal mucosa of vitamin D-replete chicks. The birds were injected intravenously with 8 to 9 nmol of 1,25-dihydroxyvitamin D3/100 g body weight 5 to 8 h before death. The isolation involved methanol-chloroform extraction and four successive chromatographic procedures (Sephadex LH-20 and high performance liquid chromatography). Chemical structures of the metabolites are proposed on the basis of (a) their chromatographic behavior, (b) their mass spectra, and (c) ultraviolet absorption spectra. They are identified as 1 alpha,25-dihydroxy-23-oxo-vitamin D3 and 1 alpha,25,26-trihydroxy-23-oxo-vitamin D3. Neither of the two new metabolites is produced by the intestinal mucosa when 1,25S,26-trihydroxyvitamin D3 is used as a substrate.

Animals↗
PubMed1980

Studies on vitamin D metabolism: catabolism of 1 alpha, 25-dihydroxyvitamin D3 and 1 alpha-hydroxyvitamin D3 to a metabolite inactive on bone mineral mobilization.

A heretofore unknown metabolite of vitamin D3 was isolated from the 1 alpha, 24, 25-trihydroxyvitamin D3 fraction of lipid extracts obtained from plasma of rats which were given intravenous or oral doses of 100 pmol/100 g of either 1 alpha-hydroxyvitamin D3 or 1 alpha, 23-dihydroxyvitamin D3. Doses of 25-250 pmoles of the new metabolite when given to a vitamin D deficient rat were completely inactive in terms of stimulating the classic vitamin D response of bone calcium mobilization. The nature of the metabolism of 1 alpha-hydroxyvitamin D3 or 1 alpha, 25-dihydroxyvitamin D3 to the metabolite is not clear at the present time, but it is probable that neither of these steroids undergo side-chain cleavage to yield the new metabolite.

Animals↗
PubMed1980

Aspirin inhibition of hypercalcemic effect of 1 alpha-hydroxyvitamin D3 in rats.

Interaction of calcium regulating hormone, active form of vitamin D3, and aspirin (ASA) was studied in rats. One alpha-hydroxyvitamin D3 (1 alpha-OH-D3) produced a hypercalcemia in parathyroidectomized, thyroparathyroidectomized and thyroparathyroidectomized nephrectomized rats. The effect of 1 alpha-OH-D3 was inhibited by treatment with 200, 50 and 20 mg/kg p.o. of ASA. ASA did not stimulate the gut absorption of calcium in vitamin D deficient rats. These data suggest that ASA acts by inhibiting 1 alpha OH-D3 induced bone resorption. This is the first report demonstrating the inhibitory effect of ASA on bone resorption as induced by 1 alpha-OH-D3, although partial inhibition of bone resorption in vitro has been reported.

Animals↗