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Biomedical subjects

M Kanashiro

Publications and source records attributed to M Kanashiro.

At least 37 records · Page 2Linked to original sources
PubMed1990

Non-invasive 31P NMR study on the development of brain membrane of gerbils and jimpy mice, a myelin-deficient mutant.

The broad background resonance observed in the in vivo 31P NMR spectra of adult murine heads was investigated in terms of phosphorus atoms in bone and membrane phospholipids. The broad background resonance was found to be weak in a juvenile and increase with advance of age. Fractionation of adult gerbil heads showed that the broad signal was derived from bone and membrane, of which myelin is the major component. The two origins of the broad background resonance exhibited considerably different line shapes in spectra, which enabled us to extract the membrane component from an intact murine head spectrum (sequential subtraction method). By the use of this method, the development of membrane in gerbil brain at various age grades could be estimated. The membrane component was shown to be suppressed in a mutant mouse, jimpy, which has a deficiency in myelin formation ability. Furthermore, the value of T1 of the membrane component was estimated to be 0.9 sec, which was in good agreement with previously reported values for excised brain.

Animals↗
PubMed1990

Effect of bifemelane on the intracellular pH and energy state of the ischemic brain.

4-(o-Benzylphenoxy)-N-methylbutylamine hydrochloride (bifemelane, CAS 90293-01-9, Celeport) has been reported to exert a protective effect on the brain against ischemic insults. However, the underlying mechanism of this action has not yet been fully elucidated. The effects of bifemelane on the intracellular pH (pHi) and energy metabolism of the ischemic brain were examined in Mongolian gerbils using in vivo 31P nuclear magnetic resonance spectroscopy. Transient global ischemia was produced by clipping both common carotid arteries for 45 min, and the brain was reperfused by releasing the clips. Bifemelane (10 or 20 mg/kg) or normal saline was administered intraperitoneally 30 min prior to the ischemia. During the ischemia, adenosine triphosphate (ATP) and phosphocreatine (PCr) were markedly reduced in association with an increase in inorganic phosphate (Pi) and decrease in pHi in both the control and bifemelane groups. The extents of energy disturbance and intracellular acidosis in the three groups were identical. After reperfusion, ATP, PCr, Pi and pHi recovered towards the pre-ischemic levels in all the groups. In the bifemelane groups, the recovery of pHi was significantly faster than in the control group. Of the two bifemelane groups, the 20 mg/kg group showed more excellent pHi recovery as compared to the 10 mg/kg group. The energy recovery in the three groups were almost identical, although the 20 mg/kg group showed some tendency towards faster recovery as compared to the control group. The present results suggest that bifemelane may accelerate recovery of the pHi after cerebral ischemia. Such an action may contribute to the cerebral protective effects of this drug against ischemic insults.

Adenosine Triphosphate↗
PubMed1989

Decrease in the fluidity of brush-border membrane vesicles induced by gentamicin. A spin-labeling study.

In our previous paper (Horio et al., Biochim Biophys Acta 858: 153-160, 1986), we reported that the addition of gentamicin in vitro to rabbit renal brush-border membrane vesicles decreases the apparent Vmax of Na+-dependent D-glucose transport without affecting the apparent Km. In the present study, we investigated the effects of gentamicin on the physical state of spin-labeled rabbit renal brush-border membranes, using electron spin resonance spectrometry. Brush-border membrane vesicles were prepared from outer cortex (mainly contains early proximal tubule) and outer medulla (containing primarily late proximal tubule), and the gentamicin toxicities in both preparations were compared. Significant decreases were observed in the membrane fluidity of 5 mM gentamicin-treated brush-border membranes. The fluidity of outer cortical brush-border membranes was affected at both 25 degrees and 35 degrees, whereas that of outer medullary membranes was affected only at 35 degrees. Two different stearic acid spin labels revealed that gentamicin affected the fluidity only in the superficial region of the membranes. We also demonstrated that the gentamicin-induced decreases in Na+-dependent D-glucose transport and in the membrane fluidity were recovered by washing gentamicin-treated brush-border membranes. We suggest that gentamicin binds to the superficial region of brush-border membranes and inhibits Na+-dependent D-glucose transport across brush-border membranes through the decrease in the membrane fluidity.

Animals↗
PubMed1989

Clinical application of sodium-23 nuclear magnetic resonance for measurement of red cell sodium concentrations.

Red cell sodium (RBC-Na+) concentrations were measured using 23Na nuclear magnetic resonance (NMR), without the destruction of erythrocyte membranes. Subjects were categorized into four groups: 20 normotensive subjects (NT group), 20 age-matched essential hypertensive patients (EHT group), 10 patients with primary aldosteronism (PA group), and 18 patients treated with digoxin (DIG group). Although RBC-Na+ concentrations were similar between the NT group (6.14 +/- 0.80 (Mean +/- SD) mmol/l) and the EHT group (5.92 +/- 0.99), they were significantly higher in both the PA group (7.55 +/- 0.88, p less than 0.001) and the DIG group (8.43 +/- 3.81, p less than 0.02). In the PA group, RBC-Na+ concentrations decreased significantly after resection of the adenoma, and there was an inverse relationship between serum potassium and RBC-Na+ concentrations (r = -0.65, p less than 0.01). In the DIG group, RBC-Na+ concentrations tended to increase in proportion to serum digoxin levels (r = 0.53, p less than 0.05). These results support the view that RBC-Na+ concentrations are determined primarily by Na+/K+-pump activity of red cell membranes. This study showed also that Na+ NMR is an useful method determining intracellular Na+ concentrations.

Digoxin↗
PubMed1989

Effects of propentofylline on energy metabolism of the ischemic brain studied by in vivo 31P nuclear magnetic resonance spectroscopy.

The effects of 3-methyl-1-(5'-oxohexyl)-7-propylxanthine (propentofylline, HWA 285) on transient cerebral ischemia were studied in Mongolian gerbils by measuring the in vivo 31P nuclear magnetic resonance (NMR) spectra and cerebral water content. Transient ischemia was produced by bilateral common carotid artery occlusion for 30 min, which was followed by 60 min of reperfusion. Propentofylline (1, 2.5 or 30 mg/kg) or normal saline was administered intravenously at 2 min after the reperfusion. The 31P spectra during the occlusion showed a marked reduction in adenosine triphosphate (ATP) and phosphocreatine (PCr) with elevation of inorganic phosphate (Pi) in all groups. The intracellular pH (pHi) calculated from the chemical shift of Pi was markedly reduced in all groups. After the reperfusion, ATP, PCr, Pi and pHi gradually recovered towards the normal levels in the control group. In the 2.5 mg/kg propentofylline group, the energy recovery was significantly faster than in the controls. The cerebral water content measured at the end of reperfusion was significantly lower in the 2.5 mg/kg propentofylline group than in the controls. However, such cerebral protective effects were not observed in the 1 mg/kg and 30 mg/kg groups. The present results suggest that propentofylline may accelerate the energy recovery of the transiently ischemic brain and suppress the development of post-ischemic cerebral edema. The effects, however, were not dose-dependent in manner. The detailed mechanism of the effects requires further investigation.

Adenosine Triphosphate↗
PubMed1988

Flow thresholds for cerebral energy disturbance and Na+ pump failure as studied by in vivo 31P and 23Na nuclear magnetic resonance spectroscopy.

The relationships among CBF, cerebral energy metabolism, Na+ pump activity, and electrocorticograms (ECoG) following graded hypotension were studied in 48 gerbils. Energy metabolism and Na+ pump activity were estimated by in vivo 31P and 23Na nuclear magnetic resonance (NMR) spectroscopy, and CBF was determined by [14C]iodoantipyrine methods at the end of the experiments. The CBF measured in normotensive animals was 0.51 +/- 0.07 ml/g brain/min. Following graded hypotension, no 31P spectral change was observed until CBF fell to 0.21-0.27 ml/g brain/min, at which level the intracellular pH began to decrease in association with ECoG voltage reduction. At a CBF level of 0.18-0.23 ml/g brain/min, phosphocreatine (PCr) began to decrease in association with inorganic phosphate (Pi) elevation. At this level, ECoG became isoelectric, although no adenosine triphosphate (ATP) change yet resulted. At a flow level of 0.12-0.14 ml/g brain/min, ATP began to decrease gradually. At 0.04-0.05 ml/g brain/min, PCr and ATP virtually disappeared, and the 23Na signal intensity suddenly changed. The present study demonstrated flow thresholds for the development of tissue acidosis, PCr-Pi changes, and ATP reduction. It appears that functional suppression occurs prior to ATP changes, whereas Na+ pump failure results after ATP depletion.

Animals↗
PubMed1988

Therapeutic effect of 15-deoxyspergualin on acute graft rejection detected by 31P nuclear magnetic resonance spectrography, and its effect on rat heart transplantation.

We investigated the effect of 15-deoxyspergualin (DSG) on graft rejection, starting administration at the onset of rejection and on the induction of immunologic unresponsiveness. Hearts from WKAH rats were transplanted into the neck of ACI rats. The energy metabolism of the grafted hearts was followed by 31P nuclear magnetic resonance spectroscopy. The day that energy metabolism started to fall was defined as the onset of rejection, and intraperitoneal administration of DSG was initiated at 5 mg/kg/day for 15 days from this day. The grafted heart arrested in 2 of 10 rats 9 and 11 days after transplantation, respectively, but the remaining 8 recovered from rejection and 5 of them showed evidence of immunologic unresponsiveness. Of 10 rats treated with DSG from the day of transplantation, only 1 rat showed evidence of unresponsiveness. The initiation of DSG treatment from the onset of rejection resulted in a higher percentage of induction of unresponsiveness. Therefore, DSG was considered to specifically inhibit lymphocyte clone expansion at the onset of rejection. Spleen cells obtained from recipients 7-10 days after the end of DSG treatment were administered to syngeneic ACI rats grafted with WKAH hearts. Graft survival was significantly prolonged, but long-term unresponsiveness could not be transferred. However, immunologic unresponsiveness could be adoptively transferred in 3 of 5 rats receiving spleen cells from syngeneic rats that had recovered from rejection after DSG treatment and had acquired long-term unresponsiveness. These results suggest that suppressor cells are resistant to DSG and are spared and participate in the maintenance of immunologic unresponsiveness.

Animals↗
PubMed1988

Isolation, structure, and properties of the beta-carboline formed from 5-hydroxytryptamine by the superoxide anion-generating system.

A yellow substance was isolated by Sephadex LH-20 gel chromatography, silica gel TLC, and reversed-phase HPLC after incubation of 5-hydroxytryptamine (5-HT) with the superoxide anion (O2-)-generating system, i.e., the xanthine-xanthine oxidase system, in the presence of the Fe-EDTA complex and glycine in alkaline medium. The product gave a blue color with Ehrlich's reagent very slowly but no color with xanthydrol and Gibbs' reagent. Its reduced form, however, gave an immediate blue-violet color with all three reagents. No color was developed with ninhydrin, but the reduced form was orange-red. The chemical structure of the yellow substance was identified by 1H-nuclear magnetic resonance and field desorption-mass spectrometry as 4,9-dihydro-3H-pyrido[3,4-b]indol-6-ol (6-hydroxy-3,4-dihydro-beta-carboline, 5-hydroxy-2,3-dihydrotryptoline). The one carbon unit inserted into 5-HT came from glycine, with its 14C-2 being incorporated into C-1 of the yellow substance. The mechanism for the formation of the yellow substance from 5-HT is discussed. This compound inhibited 5-HT uptake into rat brain cortical synaptosomes with an IC50 of 1.5 X 10(-4) M and a Ki value of 1.2 X 10(-5) M.

Animals↗
PubMed1987

In vivo measurements of intra- and extracellular Na+ and water in the brain and muscle by nuclear magnetic resonance spectroscopy with shift reagent.

The introduction of new paramagnetic shift reagents in the nuclear magnetic resonance (NMR) method has made it possible to distinguish intra- and extracellular ions in tissues or organs in vitro. We measured the intra- and extracellular 23Na and 1H in vivo in the gerbil brain and skeletal muscle by NMR spectroscopy employing the shift reagent, dysprosium triethylenetetraminehexaacetate (Dy[TTHA]3-). Without Dy(TTHA)3-, the 23Na and 1H signals were seen only as single peaks, but gradual intravenous infusion of Dy(TTHA)3- separated these signals into two peaks, respectively. The unshifted peaks reflected the intracellular 23Na and 1H signals, while the shifted peaks reflected the extracellular signals. In the brain spectra, an additional small peak, which represented intravascular signals, was detected and its intensity increased after injection of papaverine hydrochloride. The present method is advantageous over the microelectrode technique because of its nondestructiveness and its capability for obtaining intra- and extracellular volume information from measurements of the 1H spectra, the peaks of which reflect the intra- and extracellular water amounts. The intracellular Na+ increase associating with increased cellular volume after ouabain in the muscle was clearly visualized by this method. The technique is clearly of use for physiological and pathophysiological studies of organs.

Aging↗
PubMed1987

Effect of a new immunosuppressant, 15-deoxyspergualin, on heterotopic rat heart transplantation, in comparison with cyclosporine.

In this study, 15-deoxyspergualin (DSG) or cyclosporine (CsA) was administered to heterotopically heart-grafted rats for 15 days, commencing on the day of transplantation. In addition, a 31P nuclear magnetic resonance (NMR) technique was applied to investigate the in vivo energy metabolism of the graft. A significant prolongation of graft survival was observed in groups treated with 2.5 mg/kg and 5 mg/kg of DSG, when compared with the control group not treated with an immunosuppressant. One graft in the DSG 2.5 mg/kg-treated group and one in the 5 mg/kg-treated group survived for more than 100 days after grafting. The 31P NMR study demonstrated that, although rejection occurred in the rats treated with 2.5 mg/kg of DSG during the early period after transplantation, 5 mg/kg of DSG inhibited rejection completely. As for CsA, while 2 mg/kg of the drug did not affect graft survival, 5 mg/kg and 14 mg/kg significantly prolonged survival. It was revealed by 31P NMR, however, that CsA 5 mg/kg did not quite inhibit rejection by itself, and 14 mg/kg of CsA, which was the tolerogenic dose, exerted a cardiotoxic effect. In consequence, DSG seems to be a powerful immunosuppressant with a low toxic effect.

Animals↗
PubMed1985

A spin-label study on human high density lipoprotein.

Human plasma high density lipoproteins (HDL) have been labeled with N-(1-oxyl-2,2,6,6-tetramethyl-4-piperidinyl)maleimide (NEM-TEMPO). The spin-labeled HDL exhibited an ESR spectrum containing signals of both strongly immobilized and weakly immobilized components by the reaction with a high concentration of NEM-TEMPO, while an ESR spectrum containing only signals of a strongly immobilized component range between 4 degrees C and 37 degrees C, the signal height of the strongly immobilized component exhibited reversible temperature-dependent changes, whereas that of the weakly immobilized component changed irreversibly at temperatures above 25 degrees C. The activation energy of the irreversible change was estimated to be 26 kcal per mol. The strongly immobilized component was derived from NEM-TEMPO which modified apolipoprotein A-I covalently, while the weakly immobilized component was derived from NEM-TEMPO noncovalently bound to HDL. The rate of binding of NEM-TEMPO to either the strongly binding or weakly binding sites and the number of the strongly binding sites in apolipoprotein A-I were estimated to be 125 M-1.day-1 and 1.78, respectively. The binding of NEM-TEMPO to the strongly binding sites was suppressed greatly by pretreatment of HDL with 2,4,6-trinitrobenzene sulfonic acid (TNBS). The slow reaction and suppression with TNBS suggest that NEM-TEMPO binds to some amino acid residue, probably a lysine residue, in apoprotein A-I. The strongly immobilized and weakly immobilized components were reduced almost completely by ascorbate at the same rate, 0.048 min-1 at pH 7.4 and at 4 degrees C.

Ascorbic Acid↗
PubMed1983

Electron spin resonance studies on the selective labeling of N-(1-oxyl-2,2,5,5-tetramethyl-3-pyrrolidinyl)maleimide of rat liver mitochondria.

N-(1-Oxyl-2,2,5,5-tetramethyl-3-pyrrolidinyl)maleimide (MSL) was incorporated into rat liver mitochondria and the nitroxide radical incorporated was found to decay considerably. The incorporation was blocked by a high concentration of NEM, but not by pCMB. Spin labeled fatty acid derivatives, 2-(3-carboxypropyl)-2-tridecyl-4,4-dimethyl-3-oxazolidinyloxyl (FSL1) and 2-(14-carboxytetradecyl)-2-ethyl-4,4-dimethyl-3-oxazolidinyloxyl (FSL2), were also incorporated and the nitroxide radical decayed. However, incorporation of FSL1 or FSL2 was not blocked by NEM or pCMB. The ESR spectrum of 3-carboxyl-2,2,5,5-tetramethyl-pyrroline-1-oxyl (CSL) did not change on reaction with the mitochondria. The labeled MSL exhibited an ESR spectrum composed of both strongly immobilized and weakly immobilized components. A similar reaction with FSL1 gave an ESR spectrum mainly composed of a strongly immobilized component, the weakly immobilized component was negligibly small, while FSL2 exhibited an ESR spectrum in which free-like signals of the nitroxide radical were predominant. The results suggest that MSL is labeled selectively in the mitochondrial membrane through those SH groups that are not reactive to pCMB, and the labeled nitroxide radical is reduced in situ. The mode of incorporation into the mitochondria differs between MSL and the other spin labeled reagents, and labeling of MSL at the binding site may precede reduction of the nitroxide radical. The incorporation of MSL was dependent on the concentration of MSL used. ADP-acceleration of mitochondrial oxygen uptake with succinate was inhibited by labeling the mitochondria with MSL without loss of the electron transferring activity.

Animals↗
PubMed1982

A spin trap study on anaerobic dehalogenation of halothane by a reconstituted liver microsomal cytochrome P-450 enzyme system.

In the presence of a spin trapping reagent, N-tert-butyl-alpha-phenylnitrone, anaerobic incubation of halothane (2-bromo-2-chloro-1,1,1-trifluoroethane) with a reconstituted cytochrome P-450 enzyme system of rabbit liver microsomes exhibited an electron spin resonance spectrum containing signals of a nitroxide radical, and all components of the reconstituted system were necessary to produce the nitroxide radical. Formation of 2-chloro-1,1,1-trifluoroethane, the product of anaerobic dehalogenation of halothane by the reconstituted system, was inhibited by N-tert-butyl-alpha-phenylnitrone. These results indicate that a radical intermediate of halothane is produced during the anaerobic dehalogenation reaction, and that the radical is trapped by N-tert-butyl-alpha-phenylnitrone to form a N-tert-butyl-alpha-phenylnitrone radical adduct.

Anaerobiosis↗
PubMed1982

A spin label study on human low density lipoprotein.

Selective labeling with N-(1-oxyl-2,2,6,6-tetramethyl-4-piperidinyl)-maleimide of human serum LDL has been performed. The spin-labeled LDL exhibited an ESR spectrum containing signals of a strongly immobilized component only. The signals were completely reversible between 4 degrees C and 37 degrees C and fairly stable at each temperature. The spin-labeled LDL which was prepared by the usual method exhibited an ESR spectrum containing signals of both strongly immobilized and weakly immobilized components (5, 6). The latter was unstable above 25 degrees C and changed irreversibly. The strongly binding site showed higher affinity for the nitroxide radical than the weakly binding site, and two kinds of the strongly binding site were demonstrated kinetically. The rate of binding of the nitroxide radical to the two kinds of strongly binding site were estimated to be 4.7 x 10(4) M-1 . day-1 and 0.16 x 10(4) M-1 . day-1 at pH 7.4 and 4 degrees C, respectively. Both the strongly immobilized and weakly immobilized radicals were reduced with ascorbate at the same rate. It was also shown on gel filtration of the SDS-treated LDL derivatives that the strongly immobilized component was on the apoprotein B moiety, whereas either noncovalent binding to LDL or binding to some small molecular species other than protein was suggested for the weakly immobilized component.

Binding Sites↗