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M Homma

Publications and source records attributed to M Homma.

At least 19 recordsLinked to original sources
PubMed2001

MRNA and enzyme activity of hepatic 11beta-hydroxysteroid dehydrogenase type 1 are elevated in C57BL/KsJ-db/db mice.

To evaluate the importance of 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1) in insulin resistant diabetic C57BL/KsJ-db/db mice, we measured the activity and mRNA level of 11beta-HSD1 in the liver of db/db mice and their heterozygote litter mates, db/+m mice. The blood glucose, plasma insulin, and corticosterone levels of db/db mice were significantly higher than those of db/+m mice. Despite hyperinsulinemia, the activity level of this enzyme was significantly higher in db/db mice, and the mRNA level of hepatic 11beta-HSD1 was also significantly higher in db/db mice. Since hepatic 11beta-HSD1 in vivo mainly functions as 11-keto-reductase and does not work as 11beta-oxidase, these results suggest that the rate of hepatic conversion of 11-dehydrocorticosterone to corticosterone is increased in db/db mice, resulting in higher glucocorticoid activity in the liver. The increased hepatic corticosterone concentration due to the elevation of 11beta-HSD1 and high plasma corticosterone concentration may antagonize the action of insulin and cause insulin resistance. These findings have a potentially important implication for relationships between increased hepatic 11beta-HSD1 and insulin resistance in db/db mice. The present paper is the first to demonstrate the increased activities and mRNA level of hepatic 11beta-HSD1 in db/db mice.

11-beta-Hydroxysteroid Dehydrogenase Type 1↗
PubMed2001

Na(+)-driven flagellar motor of Vibrio.

Bacterial flagellar motors are molecular machines powered by the electrochemical potential gradient of specific ions across the membrane. Bacteria move using rotating helical flagellar filaments. The flagellar motor is located at the base of the filament and is buried in the cytoplasmic membrane. Flagellar motors are classified into two types according to the coupling ion: namely the H(+)-driven motor and the Na(+)-driven motor. Analysis of the flagellar motor at the molecular level is far more advanced in the H(+)-driven motor than in the Na(+)-driven motor. Recently, the genes of the Na(+)-driven motor have been cloned from a marine bacterium of Vibrio sp. and some of the motor proteins have been purified and characterized. In this review, we summarize recent studies of the Na(+)-driven flagellar motor.

Amino Acid Sequence↗
PubMed2001

Role of K(+) efflux in apoptosis induced by AMPA and kainate in mouse cortical neurons.

Activation of ionotropic glutamate receptors can induce neuronal apoptosis in vitro and in vivo. We showed previously that activation of the N-methyl-D-aspartic acid (NMDA) subtype of glutamate receptors in a low Ca(2+) and low Na(+) condition induced apoptotic neuronal death, and that the K(+) efflux via NMDA receptor channels was likely a key event in NMDA-induced apoptosis. Since non-NMDA receptors, alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionate (AMPA) and kainate receptors, are also permeable to K(+), we tested the hypothesis that stimulating K(+) efflux via non-NMDA receptor channels could induce apoptosis in cultured cortical neurons. Using a Ca(2+)-free and Na(+)-free external solution, application of kainate revealed outward membrane currents carried by K(+) efflux. In a low Ca(2+)/low Na(+) medium, a 5-h exposure to 50-500 microM AMPA in the presence of the NMDA receptor antagonist MK801 induced dose-dependent neuronal death 24 h after the onset of the insult, accompanied by intracellular K(+) reduction and caspase-3 activation. The AMPA-induced cell death was attenuated by the caspase inhibitor Z-Val-Ala-Asp(OMe)-fluoromethyl ketone (Z-VAD-FMK) and by the protein synthesis inhibitor cycloheximide. Reducing K(+) efflux by raising extracellular K(+) concentration from 5 to 25 mM attenuated AMPA-triggered cell death, the Ca(2+) channel antagonist nifedipine showed no effect on the AMPA toxicity. Kainate induced similar neuronal death sensitive to attenuation by Z-VAD-FMK or elevated extracellular K(+).We suggest that the non-NMDA receptor-mediated K(+) efflux may participate in apoptotic process and that blocking excessive K(+) efflux mediated by NMDA and non-NMDA receptors may selectively prevent neuronal apoptosis under certain pathological conditions.

Animals↗
PubMed2001

Assessing systemic 11beta-hydroxysteroid dehydrogenase with serum cortisone/cortisol ratios in healthy subjects and patients with diabetes mellitus and chronic renal failure.

11beta-hydroxysteroid dehydrogenase (11beta-HSD), an enzyme regulating mineralocorticoid like action of glucocorticoid, oxidizes active cortisol to inactive cortisone. Impaired activity of this enzyme is associated with apparent mineralocorticoid excess (AME) syndrome and is characterized by hypertension and hypokalemia. Recent investigations suggest the presence of hypertensive subjects with low activity of 11beta-HSD. The blood concentration ratio of cortisone/cortisol reflects the overall conversion of cortisol to cortisone and may be an index to assess the systemic activity of 11beta-HSD. We evaluated the peripheral blood concentration ratio of cortisone/cortisol as a possible marker to identify subjects with hypertension thought to represent impaired 11beta-HSD activity. We compared this ratio in healthy subjects and patients with diabetes mellitus (DM) or chronic renal failure (CRF). Peripheral blood samples were collected from 69 healthy subjects, 44 DM, and 36 CRF patients in the morning (9:00 to 11:00 AM). Twenty-six DM patients (59%) and 32 CRF patients (89%) met the criteria for having hypertension. Serum cortisol and cortisone concentrations were determined by high performance liquid chromatography (HPLC). All values for serum cortisone and cortisol levels were within the normal range. Serum cortisone/cortisol ratio in the healthy subjects was distributed with a range of 0.113 to 0.494 (median, 0.243). Compared with healthy subjects, DM and CRF patients had significantly low (P <.01) serum cortisone/cortisol levels (median, 0.188 [range, 0.092 to 0.313] in DM and 0.088 [range, 0.031 to 0.140] in CRF). Bimodal distribution of cortisone/cortisol, found in DM patients with hypertension, represented high- and low-ratio groups around the border of the ratio 0.2. Kidney function, DM duration, and complications varied between the high- and low-ratio groups. The low ratio group (<0.2), whose 11beta-HSD activity was considered low, had an increase in blood urea nitrogen (BUN) levels and experienced nephropathy, neuropathy, retinopathy, and prolonged DM duration when compared with the group with a ratio greater than 0.2. The data suggest that the serum cortisone/cortisol ratio reflects the change in 11beta-HSD activity and is dependent kidney function. This is a possible marker to evaluate glucocorticoid excess hypertension observed in DM and CRF patients.

11-beta-Hydroxysteroid Dehydrogenases↗
PubMed2001

Magnolol from Magnolia officinalis inhibits 11beta-hydroxysteroid dehydrogenase without increases of corticosterone and thymocyte apoptosis in mice.

Magnolol is an 11beta-hydroxysteroid dehydrogenase (11beta-HSD) inhibitor contained in Magnolia officinalis which is used in Chinese remedies. We have reported that glycyrrhetinic acid, a strong 11beta-HSD inhibitor isolated from licorice, induces apoptosis of murine thymocytes via accumulation of corticosterone. In this paper, we report that magnolol inhibited 11beta-HSD without increases in the blood concentration of corticosterone and in thymocyte apoptosis in mice. Oxidative activities of the enzyme (from corticosterone to 11-dehydrocorticosterone) in liver, kidney and thymus in vitro were examined 24 h after a single administration of magnolol. Magnolol inhibited the enzyme activity in kidney (P < 0.0001) and thymus (P < 0.002), while the activity in liver was not affected. Blood concentrations of corticosterone in the magnolol-treated mice were unexpectedly lower than those in the control animals (P < 0.002). This means that the inhibition of 11beta-HSD by magnolol did not increase the systemic level of corticosterone which is relevant to thymocyte apoptosis. Accordingly, our flow cytometric analysis of thymocytes after magnolol treatment showed no change in the number of apoptotic cells. We concluded that unlike glycyrrhetinic acid, magnolol selectively inhibited 11beta-HSD in kidney and thymus but not in liver, so that the blood concentrations of corticosterone could not exceed the control level.

11-beta-Hydroxysteroid Dehydrogenase Type 1↗
PubMed2001

Cloning and characterization of motX, a Vibrio alginolyticus sodium-driven flagellar motor gene.

Four motor proteins, MotX, MotY, PomA, and PomB, have been identified as constituents of the Na(+)-driven flagellum of Vibrio species. In this study, the complete motX gene was cloned from Vibrio alginolyticus and shown to complement three mot mutations, motX94, motX115, and motX119, as well as a V. parahaemolyticus motX mutant. The motX94 mutant contains a frameshift at Val86 of MotX, while the motX115 and motX119 mutations comprise substitutions of Ala146 to Val and Gln 194 to amber, respectively. When MotX was overexpressed in Vibrio cells, the amount of MotY detected in the membrane fraction increased, and vice versa, suggesting that MotX and MotY mutually stabilize each other by interacting at the membrane level. When a plasmid containing the motX gene was introduced into motY mutants NMB117 (motY117) and VIO542 (motY542), the mutations were suppressed. In contrast, motY could not cause the recovery of any swarm-defective motX mutants studied. Considering the above evidence, we propose that MotX is more directly involved than MotY in the mechanical functioning of the Na(+)-type flagellar motor, and that MotY may stabilize MotX to support its interaction with other Mot proteins.

Amino Acid Sequence↗
PubMed2001

Effect of ascites on tacrolimus disposition in a liver transplant recipient.

To examine the effects of ascites on tacrolimus disposition, the authors measured tacrolimus concentration in blood and ascitic fluid from a patient with a living related liver transplant recipient who required removal of 500 to 2400 mL ascitic fluid daily. Tacrolimus levels in ascitic fluid ranged from 0.07 to 0.29 ng/mL and in whole blood from 7.5 to 20.3 ng/mL. The tacrolimus concentration in ascitic fluid positively correlated with that in whole blood ( r = 0.878, P <0.0001). Because the amounts of tacrolimus excreted into the ascitic fluid corresponded to only 0.01% to 0.09% of the dose administered, the authors concluded that the effects of ascites on tacrolimus disposition were negligible even though large amounts of ascitic fluid were drained regularly.

Ascites↗
PubMed2001

Glycyrrhetinic acid induced apoptosis in murine splenocytes.

Glycyrrhetinic acid (GA) is known to inhibit glucocorticoid metabolism inhibiting 11beta-hydroxysteroid dehydrogenase (11beta-HSD). Moreover, GA administration to mice has been shown to affect the lymphoid organs through elevation of endogenous corticosterone concentration. The effect of GA administration on thymus has been demonstrated to show that considerable amounts of thymocytes undergo apoptosis by elevated levels of corticosterone in systemic circulation. However, the effect of GA administration on peripheral lymphocytes has remained unknown. In our current study, we demonstrated that a significant involution of spleen as well as thymus occurred within 24 h of a single administration of GA in mice. In addition, a flow cytometric analysis of the splenocytes taken from mice treated with GA showed a significant increase in the number of apoptotic cells which exhibited translocated phosphatidylserine outside the plasma membrane. Furthermore, considerable inhibition of 11beta-HSD activity in GA-treated mice was observed in liver and spleen, resulting in a significant increase in concentration of corticosterone in the blood. These facts showed that the apoptosis of splenocytes was the result of indirect effect of GA through elevated levels of corticosterone. We confirmed this using cultured splenocytes in vitro where no apoptotic effect of GA was observed. We concluded that GA administration induces cell death of not only thymocytes that are naive to corticosterone, but also splenocytes that are usually stable to its physiological concentrations.

11-beta-Hydroxysteroid Dehydrogenases↗
PubMed2001

Effects of absorbed components of saiboku-to on the release of leukotrienes from polymorphonuclear leukocytes of patients with bronchial asthma.

Previous studies confirmed the efficacy of the Chinese herbal remedy Saiboku-to in patients with steroid-dependent asthma. We studied 8 phenolic compounds, isolated from the urine of patients receiving Saiboku-to, with respect to their effects on leukotriene (LT) release by polymorphonuclear leukocytes (PMN) obtained from 6 patients with atopic asthma and 8 healthy subjects. The compounds (0.01-10 micrograms/ml) were incubated with Ca2+ ionophore (A23187)-stimulated PMN, and concentrations of leukotriene B4 (LTB4) and leukotriene C4 (LTC4) in the supernatant were measured by EIA. Each compound suppressed the release of LTB4 and LTC4 by PMN obtained from healthy and asthmatic subjects. In particular, baicalein and magnolol were 5-10 times more potent than azelastine, an antiallergic drug, and significantly suppressed LTC4 release by PMN obtained from asthmatic patients, as compared with healthy subjects. Suppression of LTC4 release by these compounds may play an important role in the clinical efficacy of Saiboku-to.

Adult↗
PubMed2000

Intermolecular cross-linking between the periplasmic Loop3-4 regions of PomA, a component of the Na+-driven flagellar motor of Vibrio alginolyticus.

PomA and PomB form a complex that conducts sodium ions and generates the torque for the Na(+)-driven polar flagellar motor of Vibrio alginolyticus. PomA has four transmembrane segments. One periplasmic loop (loop(1-2)) connects segments 1 and 2, and another (loop(3-4)), in which cysteine-scanning mutagenesis had been carried out, connects segments 3 and 4. When PomA with an introduced Cys residue (Cys-PomA) in the C-terminal periplasmic loop (loop(3-4)) was examined without exposure to a reducing reagent, a 43-kDa band was observed, whereas only a 25-kDa band, which corresponds to monomeric PomA, was observed under reducing conditions. The intensity of the 43-kDa band was enhanced in most mutants by the oxidizing reagent CuCl(2). The 43-kDa band was strongest in the P172C mutant. The motility of the P172C mutant was severely reduced, and P172C showed a dominant-negative effect, whereas substitution of Pro with Ala, Ile, or Ser at this position did not affect motility. In the presence of DTT, the ability to swim was partially restored, and the amount of 43-kDa protein was reduced. These results suggest that the disulfide cross-link disturbs the function of PomA. When the mutated Cys residue was modified with N-ethylmaleimide, only the 25-kDa PomA band was labeled, demonstrating that the 43-kDa form is a cross-linked homodimer and suggesting that the loops(3-4) of adjacent subunits of PomA are close to each other in the assembled motor. We propose that this loop region is important for dimer formation and motor function.

Anti-Bacterial Agents↗
PubMed2000

Coupling ion specificity of chimeras between H(+)- and Na(+)-driven motor proteins, MotB and PomB, in Vibrio polar flagella.

We have shown that a hybrid motor consisting of proton-type Rhodobacter sphaeroides MotA and sodium-type VIBRIO: alginolyticus PomB, MotX and MotY, can work as a sodium-driven motor in VIBRIO: cells. In this study, we tried to substitute the B subunits, which contain a putative ion-binding site in the transmembrane region. Rhodobacter sphaeroides MotB did not work with either MotA or PomA in Vibrio cells. Therefore, we constructed chimeric proteins (MomB), which had N-terminal MotB and C-terminal PomB. MomB proteins, with the entire transmembrane region derived from the H(+)-type MotB, gave rise to an Na(+) motor with MotA. The other two MomB proteins, in which the junction sites were within the transmembrane region, also formed Na(+) motors with PomA, but were changed for Na(+) or Li(+) specificity. These results show that the channel part consisting of the transmembrane regions from the A and B subunits can interchange Na(+)- and H(+)-type subunits and this can affect the ion specificity. This is the first report to have changed the specificity of the coupling ions in a bacterial flagellar motor.

Amino Acid Sequence↗
PubMed2000

Multimeric structure of PomA, a component of the Na+-driven polar flagellar motor of vibrio alginolyticus.

Four integral membrane proteins, PomA, PomB, MotX, and MotY, are thought to be directly involved in torque generation of the Na(+)-driven polar flagellar motor of Vibrio alginolyticus. Our previous study showed that PomA and PomB form a complex, which catalyzes sodium influx in response to a potassium diffusion potential. PomA forms a stable dimer when expressed in a PomB null mutant. To explore the possible functional dependence of PomA domains in adjacent subunits, we prepared a series of PomA dimer fusions containing different combinations of wild-type or mutant subunits. Introduction of the mutation P199L, which completely inactivates flagellar rotation, into either the first or the second half of the dimer abolished motility. The P199L mutation in monomeric PomA also altered the PomA-PomB interaction. PomA dimer with the P199L mutation even in one subunit also had no ability to interact with PomB, indicating that the both subunits in the dimer are required for the functional interaction between PomA and PomB. Flagellar rotation by wild-type PomA dimer was completely inactivated by phenamil, a sodium channel blocker. However, activity was retained in the presence of phenamil when either half of the dimer was replaced with a phenamil-resistant subunit, indicating that both subunits must bind phenamil for motility to be fully inhibited. These observations demonstrate that both halves of the PomA dimer function together to generate the torque for flagellar rotation.

Amiloride↗
PubMed2000

Functional reconstitution of the Na(+)-driven polar flagellar motor component of Vibrio alginolyticus.

The bacterial flagellar motor is a molecular machine that couples the influx of specific ions to the generation of the force necessary to drive rotation of the flagellar filament. Four integral membrane proteins, PomA, PomB, MotX, and MotY, have been suggested to be directly involved in torque generation of the Na(+)-driven polar flagellar motor of Vibrio alginolyticus. In the present study, we report the isolation of the functional component of the torque-generating unit. The purified protein complex appears to consist of PomA and PomB and contains neither MotX nor MotY. The PomA/B protein, reconstituted into proteoliposomes, catalyzed (22)Na(+) influx in response to a potassium diffusion potential. Sodium uptake was abolished by the presence of Li(+) ions and phenamil, a sodium channel blocker. This is the first demonstration of a purification and functional reconstitution of the bacterial flagellar motor component involved in torque generation. In addition, this study demonstrates that the Na(+)-driven motor component, PomA and PomB, forms the Na(+)-conducting channel.

Bacterial Outer Membrane Proteins↗
PubMed2000

Total energy expenditure of elite synchronized swimmers measured by the doubly labeled water method.

To determine the daily energy requirement of elite synchronized swimmers during moderate-intensity training, the average daily energy expenditure measured by the doubly labeled water method, was calculated for nine female Japanese national team synchronized swimmers [four senior; mean (SD) 22.5 (1.0) years old, 52.2 (3.6) kg, and five junior; 17.6 (1.1) years old, 52.8 (2.3) kg]. Their total energy expenditure (TEE) was 11.5 (2.8) MJ x day(-1) [2738 (672) kcal day(-1)]. When compared with estimated energy requirements derived from "Recommended Dietary Allowances for the Japanese", 12.1 (0.6) MJ day(-1) [2897 (139) kcal day(-1)], there was no difference between mean actual and estimated energy requirements. However, there were considerable differences observed on an individual basis. Their energy intake, estimated from 7- day self-reported dietary records, was 8.9 (1.7) MJ day(-1) [2128 (395) kcal x day(-1)], which was significantly lower than their TEE (P<0.05). Resting energy expenditure (REE), as determined by indirect calorimetry, was 5.2 (0.3) MJ x day(-1) [1247 (75) kcal x day(-1)]. Their physical activity level (TEE/REE) was 2.18 (0.43). These results demonstrate that the TEE values of elite female synchronized swimmers are not dissimilar to those reported for athletes participating in other sports, especially competitive swimmers during moderate-intensity training.

Adolescent↗
PubMed2000

Comparative study of lymphocyte-suppressive potency between prednisolone and methylprednisolone in rheumatoid arthritis.

We compared lymphocyte-suppressive potencies of prednisolone and methylprednisolone in rheumatoid arthritis (RA). IC(50)s of the glucocorticoids (GCs) on concanavalin A-induced blastogenesis of peripheral-blood mononuclear cells (PBMCs) from 44 RA patients and 30 healthy subjects were estimated in vitro, and differences in the IC(50)s of the two GCs were evaluated. The mean (+/-SD) IC(50)s for prednisolone and methylprednisolone on PBMC-blastogenesis of RA were 17.2+/-17.1 and 12.6+/-18.4 ng/ml, respectively, and no significant differences were observed between prednisolone-IC(50) and methylprednisolone-IC(50). In contrast, the mean IC(50)s of prednisolone and methylprednisolone on healthy PBMCs were 19.4+/-22. 4 and 3.7+/-3.9 ng/ml, respectively, and thus methylprednisolone potency was significantly higher than prednisolone potency (p<0.01). Methylprednisolone potency against PBMCs in RA patients exhibiting a high level of rheumatoid factor (RF) (>20 IU/ml) and the rheumatoid arthritis particle-agglutination value (RAPA) (>80) was significantly higher than that of patients exhibiting a lower level of RF or RAPA (p<0.05). In prednisolone-IC(50), however, such differences between the two patient-subgroups were not observed. Unlike reported cases of renal transplantation and healthy subjects, there was no difference in the lymphocyte-suppressive potencies for both prednisolone and methylprednisolone on RA-PBMCs. However, PBMCs from RA patients exhibiting high levels of RF or RAPA are more sensitive to methylprednisolone rather than prednisolone.

Aged↗
PubMed2000

Liquid chromatographic determination of urinary 6beta-hydroxycortisol to assess cytochrome p-450 3A activity in HIV positive pregnant women.

Assessing the activity of CYP3A4 is important for predicting the pharmacokinetic behavior of protease inhibitors in HIV positive patients, especially in pregnant women. The endogenous hormonal ratio of 6beta-hydroxycortisol (beta-OHF) to cortisol (F) in the urine is an index for metabolic enzyme activity of cytochrome p-450 (CYP) 3A4. Because the ratio is a unique way to assess the enzyme activity without using any exogenous probes for this isozyme, it is practical for use in pregnant women. In this paper, we describe a method using high performance liquid chromatography (HPLC) for 6beta-OHF in urine from pregnant women to estimate the ratio of 6beta-OHF/F. Urinary 6beta-OHF was measured by using C18-cartridge solid phase extraction and isocratic HPLC. Aliquots (1 ml) of urine samples spiked with internal standard, 6beta-hydroxyprednisolone (6beta-OHPSL), were alkalinized with NaOH, then applied to C18-cartridges, which were washed with water and hexane and eluted with ethyl acetate. After the effluents were dried and reconstituted in 10% acetonitrile, the samples were analyzed by HPLC using an isocratic mobile phase (acetic acid/acetonitrile/50 mM potassium dihydrogenphosphate: 0.2/9/90.8; v/v) and ultraviolet detection at 245 nm. The recoveries of 6beta-OHF from C18 cartridges were 93.2 and 93.9% when the authentic 6beta-OHF was added to the urine sample at the concentration of 50 and 300 ng/ml, respectively. Intra- and inter-day variations estimated at concentrations of 113-674 ng/ml were 2.9-5.6 and 4.9-8.1%, respectively. The method was applied to morning urine samples collected from HIV-positive pregnant women managed with protease inhibitor containing anti-retroviral regimens.

Adult↗
PubMed2000

The aspartate chemoreceptor Tar is effectively methylated by binding to the methyltransferase mainly through hydrophobic interaction.

In the chemotaxis of Escherichia coli, adaptation requires the methylation and demethylation of transmembrane receptors, which are catalysed by the methyltransferase CheR and the methylesterase CheB respectively. CheR binds to major chemoreceptors through their C-terminal motif NWETF, which is distinct from the methylation sites. In this study, we carried out a systematic mutagenesis of the pentapeptide sequence of Tar. Receptor methylation and adaptation were severely impaired by the alanine substitution of residue W550 and, to a lesser extent, by that of F553. Substitution of residues N549, E551 and T552 had only a slight or little effect. The defects of the W550A and F553A mutations were suppressed by high- and low-level overproduction of CheR respectively. Expression of a fusion protein containing the NWETF sequence, but not its W550A and F553A versions, inhibited chemotaxis of the Che+ strain. In an in vitro assay, CheR bound to the wild-type version but not to the mutant versions. These results and further mutagenesis suggest that the hydrophobicity and the size of residues W550 and F553 are critical in the interaction with CheR, a conclusion that is consistent with the crystal structure of a CheR-NWETF complex. On the other hand, the negatively charged side chain of E551 and the polar side chains of N549 and T552 may not be strictly required, although the presence of a salt bridge and hydrogen bonds between these residues and residues from CheR has been noted in the co-crystal.

Aspartic Acid↗