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Biomedical subjects

N Fujitsuka

Publications and source records attributed to N Fujitsuka.

At least 37 records · Page 2Linked to original sources

Glycogen depletion of the intrafusal fibers in a mouse muscle spindle during prolonged swimming.

This study investigated the recruitment of different types of intrafusal fibers during prolonged swimming at 60-75% of VO2max. We used 56 male adult mice and examined depletion of glycogen in soleus (Sol) and extensor digitorum longus (EDL) muscle spindles by visual inspection and a newly developed optical scanning method. More than 80% of all spindles from six mice consisted of four fibers: one type I nuclear bag (bag1) fiber, one type II nuclear bag (bag2) fiber, and two nuclear chain fibers. Glycogen content was estimated in muscle fibers from groups of six mice that had rested or swum for either 0.5, 1, 2, 4, or 8 h. The optical scanning intensity of periodic acid Schiff (PAS)-stained sections was correlated with their biochemically determined glycogen content (r = 0.93). Both methods showed fundamentally the same result: each type of intrafusal fiber has its own typical recruitment pattern during exercise. In the initial phase (0-0.5 h), glycogen depletion was largest in nuclear bag1 fibers and insignificant in the bag2 and chain fibers. With the bag1 fibers having become fatigued, nuclear bag2 fibers mainly took over during the middle phase (2-4 h). During the last phase (4-8 h), only the glycogen content of chain fibers decreased significantly (4-8 h). There were significant correlations between the recruitment pattern of bag1 and extrafusal type I fibers in both Sol and EDL, between nuclear bag2 and type IIa fibers in Sol, and between nuclear chain and type IIb fibers in EDL. This suggests that, during moderately intense exercise, glycogen depletion occurs first in the slow, then the intermediate, and, finally, the fast intrafusal fibers.

Animals↗

Exercise training prevents maturation-induced decrease in insulin sensitivity.

We examined the effects of exercise training initiated before maturation or after maturation on insulin sensitivity and glucose transporter GLUT-4 content in membrane fractions of skeletal muscle. Female Wistar rats (4 wk of age) were divided into sedentary and exercise-trained groups. At 12 wk of age, a subset of the trained animals (Tr) was killed along with a subset of sedentary controls (Sed). One-half of the remaining sedentary animals remained sedentary (Sed-Sed) while the other half began exercise training (Sed-Tr). The remaining rats in the original trained group continued to train (Tr-Tr). Euglycemic clamp (insulin infusion rate at 6 mU.kg body wt-1. min-1) was performed at 4, 12, and 27 wk. After euglycemic clamp in all animals except the 4-wk-old, hindlimb (gastrocnemius and part of quadriceps) muscles were removed for preparation of membrane fractions. In sedentary rats, glucose infusion rate (GIR) during euglycemic clamp was decreased from 15.9 mg.kg-1.min-1 at 4 wk of age to 9.8 mg.kg-1.min-1 at 12 wk of age and 9.1 mg.kg-1.min-1 at 27 wk of age. In exercise-trained rats, the GIR was not significantly decreased by maturation (at 12 wk) and further aging (at 27 wk). Initiation of exercise after maturation restored the GIR at 27 wk of age to the same levels as these for the corresponding exercise-trained rats. GLUT-4 content in plasma and intracellular membrane fractions of hindlimb muscle obtained just after euglycemic clamp showed the same trend as the results of GIR. These results suggest that exercise training prevented the maturation-induced decrease in insulin sensitivity. Improvement of insulin sensitivity caused by exercise training was attributed, at least in part, to the increase in insulin-sensitive GLUT-4 on the plasma membrane in skeletal muscle.

Age Factors↗

Effect of acteoside on mesangial proliferation in rat anti-Thy 1 nephritis.

We investigated whether acteoside can inhibit mesangial matrix expansion or mesangial cell proliferation in mesangioproliferative anti-Thy 1 nephritis. Untreated control rats were compared to the rats treated with acteoside either during early (day 1 to 8) or late (day 4 to 12) period after the induction of anti-Thy 1 nephritis. The result showed that acteoside and prednisolone treatments (in either early or late period) significantly reduced proteinuria, mesangial matrix expansion (the index of matrix expansion) and mesangial proliferation as determined by the number of proliferating nuclear cell antigen (PCNA)-positive cells. Acteoside also reduced glomerular macrophage infiltration and ICAM-1 expression in glomeruli of anti-Thy 1 nephritic rats. Furthermore, acteoside treatment markedly increased the activities of matrix metaloproteinases (MMP) in glomeruli. These results suggest that acteoside can inhibit mesangial cell proliferation and extracellular matrix overproduction by either inhibiting ICAM-1 expression or increasing activities of MMP.

Animals↗

[Effect of Onpi-to (TJ-8117) on TGF-beta 1 in rats with 5/6 nephrectomized chronic renal failure].

We and others have already reported that Onpi-to (TJ-8117), a Kampo medicine composed mainly of Rhei Rhizoma, has a beneficial effect on an adenine-induced renal failure model and 5/6 nephrectomized renal failure. However, little is known about the detailed mechanism of this medicine when used for renal failure. The present study was designed to clarify whether or not TJ-8117 affects TGF-beta 1 production or activation in glomeruli of 5/6 nephrectomized rats. TJ-8117 (400 mg/kg/ day) and captopril (50 mg/kg/day) were administered as drinking water from the day immediately after the operation and continued throughout the experiment. All rats were sacrificed at 4 weeks and renal cortical tissue was removed to quantify the protein and activity of TGF-beta 1 and activities of metalloproteinase. TIMP expression and extracellular matrix (collagen type I, IV) in the glomeruli were analysed histologically. TJ-8117 inhibited proteinuria, and the accumulation of collagen type I and IV in glomeruli of nephrectomized rats. In addition, TJ-8117 inhibited the TGF-beta 1 positive area in the glomeruli and the elevation of mature TGF-beta 1 level in the renal cortex of nephrectomized rats. In the TJ-8117 treated group, activities of metalloproteinase 1, 2 or 9 in the renal cortex were elevated compared with the control group. Captopril failed to affect the TGF-beta 1 level. We also found that the constitutive herbs in TJ-8117, Rhei Rhizoma and Ginseng radix, inhibited the process from inactive TGF-beta 1 to mature TGF-beta 1.

Animals↗

[Effects of neutralizing antibodies on cytokine treatment for anti-GBM nephritis in mouse].

The process of glomerular injury in nephrotoxic serum nephritis (NTN) is dependent on proinflammatory cytokines. In the present investigation, we assessed the actions of neutralizing antibody against IL-1 beta, TNF-alpha, IL-6 and TGF-beta 1 on glomerular injury. Marked increase in IL-1 beta and IL-6 was detected in cultured glomeruli of NTN in mice throughout the experiments from disease induction. Protein of TNF-alpha and TGF-beta 1 also increased in NTN mice 1 day after disease induction. Treatment with either IL-1 beta or TNF-alpha neutralizing antibody reduced proteinuria from 71 +/- 11.2 m g/24 hr to 32.2 +/- 6.0 (P < 0.01). 34.3 +/- 6.8 mg/24 hr (P < 0.01), respectively. Although the effect of IL-6 neutralizing antibody on proteinuria was not remarkable, the decreased creatinine clearance was improved more than that of IL-1 beta or TNF-alpha. Antibody against TGF-beta 1 had no effect on proteinuria and creatinine clearance. Treatments with IL-1 beta, TNF-alpha and IL-6 neutralizing antibodies inhibited glomerular hypercellularity in NTN mice. TGF-beta 1 neutralizing antibody suppressed the index of mesangial matrix expansion. IL-1 beta and TNF-alpha neutralizing antibodies prevented the increase in the number of macrophages in the glomeruli. The number of PCNA positive cells and alpha-smooth muscle actin expression in glomeruli was significantly reduced in the IL-6 neutralizing antibody-treated group. These results confirm the direct involvement of IL-1 beta, TNF-alpha and IL-6 in mouse NTN. We speculate that TGF-beta 1 may inhibit excessive proliferation in glomerular cells.

Animals↗

Regulation of valine catabolism in canine tissues: tissue distributions of branched-chain aminotransferase and 2-oxo acid dehydrogenase complex, methacrylyl-CoA hydratase and 3-hydroxyisobutyryl-CoA hydrolase.

To clarify the valine catabolism, the activities of principal enzymes in its catabolic pathway, branched-chain aminotransferase, branched-chain 2-oxo acid dehydrogenase complex, methacrylyl-CoA hydratase and 3-hydroxyisobutyryl-CoA hydrolase, were measured using canine tissues. After killing of beagle dogs, tissues (liver, pancreas, kidney, heart, skeletal muscle and mucosae of digestive organs such as stomach, small intestine and colon) were removed and immediately frozen. Branched-chain aminotransferase activity in liver was the lowest among the tissues measured. In contrast, the activities of branched-chain 2-oxo acid dehydrogenase complex in liver as well as in kidney were relatively high and the enzyme complex activities were markedly low in small intestine and skeletal muscle. The activities of methacrylyl-CoA hydratase and 3-hydroxyisobutyryl-CoA hydrolase were relatively high in all tissues, suggesting that a cytotoxic intermediate, methacrylyl-CoA, is immediately degraded to non-toxic compounds, 3-hydroxyisobutyrate and free CoA. These findings suggest that the consumption of branched-chain amino acids in the absorption site (small intestine) is suppressed in order to supply them to the whole body, in particular to skeletal muscle and that skeletal muscle might act as a storage of gluconeogenic amino acids. The high capacity to dispose methacrylyl-CoA produced in the valine catabolism is suggested to play an important role in protecting cells against the toxic effects of methacrylyl-CoA.

3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)↗

Branched-chain alpha-keto acid dehydrogenase complex in rat skeletal muscle: regulation of the activity and gene expression by nutrition and physical exercise.

Branched-chain alpha-keto acid dehydrogenase complex is the rate-limiting enzyme in the catabolism of branched-chain amino acids in skeletal muscle. It is suggested that activation of this enzyme in the muscle during exercise plays an important role in the increased oxidation of branched-chain amino acids in the muscle. Evidence suggests that branched-chain alpha-keto acids, the substrates for the enzyme, regulate the activity state of the enzyme in the muscle during exercise through phosphorylation/dephosphorylation cycle of the enzyme protein. We propose a model for the mechanism of enzyme activation by exercise. In addition to this acute effect of exercise, we present evidence suggesting that exercise training modulates the enzyme activity and gene expression for the enzyme. Increases in the total activity as well as enzyme proteins by exercise training are suggested to be associated with mitochondrial biogenesis in the muscle.

3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)↗

[Effects of Onpi-to (TJ-8117) on mesangial injury induced by anti Thy-1 antibody].

We investigated the effect of Onpi-to (TJ-8117) on the mesangial injury induced by anti-Thy-1 antibody. TJ-8117 (400 mg/kg/day, p.o.) given from the 1st day (from the day of injection of the anti-thymocyte serum) or 4th day (after mesangial proliferation), markedly inhibited the mesangial proliferation and hypercellularity in glomeruli. TJ-8117 prevented the increase in the number of PCNA or ED-1 positive cells in glomeruli. These results suggest that TJ-8117 is effective against glomerular disease with mesangial injury.

Animals↗

Differential adaptation to endurance training between heart and gastrocnemius muscle mitochondria in rats.

The effects of endurance training on mitochondrial gene expression and replication in heart and gastrocnemius muscles were studied in rats. Rats were trained for 12 weeks by treadmill running. Although the heart weight was significantly increased by the training, activity of cytochrome oxidase and concentrations of cytochrome b mRNA and mitochondrial DNA in the heart were not altered by the training. On the other hand, those in gastrocnemius muscle were increased about 1.5 fold by the training. When these values were compared between heart and gastrocnemius muscles, they were 10, 3.3, and 1.7 fold, respectively, higher in the former than in the latter. These results suggest that the mitochondrial gene expression and enzyme activity for the oxidative capacity in heart are fairly high compared to those in skeletal muscle, therefore adaptation to the training in heart is attained by increasing the muscle mass, but not mitochondrial gene expression.

Animals↗

Purification and partial characterization of 3-hydroxyisobutyryl-coenzyme A hydrolase of rat liver.

An unusual feature of valine catabolism is a reaction in which an intermediate of its catabolic pathway, (S)-3-hydroxyisobutyryl-CoA, is hydrolyzed to give the free acid and CoA-SH. The enzyme responsible for this reaction, 3-hydroxyisobutyryl-CoA hydrolase (EC 3.1.2.4), was purified 7200-fold from rat liver in this study. The purified enzyme consists of a single polypeptide with an M(r) of 36,000 in the native and denatured forms. The hydrolase is highly specific for (S)-3-hydroxyisobutyryl-CoA and 3-hydroxypropionyl-CoA (Km, 6 and 25 microM, respectively) with optimal activity around pH 8. The turnover rate of the enzyme for (S)-3-hydroxyisobutyryl-CoA is 270 s-1, which is high relative to other enzymes of the valine pathway. Likewise, activity of the enzyme expressed on a wet weight basis is also very high in the major tissues of the rat. These findings suggest that rapid destruction of (S)-3-hydroxyisobutyryl-CoA produced during valine catabolism is physiologically important. We propose that the need for a mechanism to protect cells against the toxic effects of methacrylyl-CoA, which is maintained in equilibrium with (S)-3-hydroxyisobutyryl-CoA by crotonase, explains why valine catabolism involves this enzyme and why its tissue activity is so high.

Acyl Coenzyme A↗

Enzymatic and genetic adaptation of soleus muscle mitochondria to physical training in rats.

To evaluate the effects of physical training on mitochondrial gene expression and mitochondrial biogenesis in slow-twitch muscle, adult female Sprague-Dawley rats were trained for 3, 6, and 12 wk by running on a motor-driven treadmill (speed of 25 m/min and duration of 90 min/day, 5 days/wk), and the activities of citrate synthase, ubiquinol-cytochrome-c oxidoreductase, cytochrome oxidase, mitochondrial cytochrome b mRNA (by Northern blot analysis), and mitochondrial DNA (by slot-blot and Southern blot analyses) were measured in rat soleus muscle. A DNA probe for detection of mitochondrial mRNA and DNA was prepared from a 1,500-bp fragment of human mitochondrial DNA that included the coding region of the cytochrome b gene. Training for 3, 6, and 12 wk significantly increased the activities of citrate synthase (31, 28, and 47%, respectively), ubiquinol-cytochrome-c oxidoreductase (61, 63, and 77%, respectively), and cytochrome oxidase (25, 26, and 32%, respectively) in muscle. The concentration of cytochrome b mRNA in the muscle was proportionally elevated with the enzyme activities. On the other hand, the mitochondrial DNA concentration in the muscle was not altered by training for 3 or 6 wk but increased significantly after training for 12 wk (35% in the slot-blot analysis and 31% in the Southern blot analysis). These results suggest that an increase in the oxidative capacity of slow-twitch muscle by the relatively short-term training is regulated at the pretranslational step in mitochondrial protein synthesis but that the increase by the long-term training involves mitochondrial replication.

Adaptation, Physiological↗

Branched-chain 2-oxo acid dehydrogenase complex activation by tetanic contractions in rat skeletal muscle.

Branched-chain 2-oxo acid dehydrogenase complex in rat skeletal muscle was activated by muscle contractions elicited by electrical stimulation. This activation was attributed to dephosphorylation of the phosphorylated enzyme complex, and the total enzyme activity was not altered by muscle contractions. The activation of the enzyme complex occurred in the muscle of the electrically stimulated leg, but not in the muscle of the non-stimulated (control) leg, indicating that blood components are not involved in the mechanism of the enzyme activation in the muscle. Adenine nucleotides, branched-chain amino and 2-oxo acids and lactate in the muscle were determined as possible factors modulating the enzyme complex activity through inhibition of branched-chain 2-oxo acid dehydrogenase kinase activity. The profile of enzyme activation induced by muscle contractions was different from the alteration of the adenine nucleotide concentrations but was similar to the alteration of the concentrations of branched-chain amino and 2-oxo acids in the muscle. The lactate concentration in the stimulated muscle was elevated 3-5-fold during the contractions, indicating intracellular acidification. Previous studies have shown that the 2-oxo acid derived from leucine is a potent inhibitor of the kinase. These results suggest that intracellular branched-chain 2-oxo acids increased by muscle contractions accumulate in the mitochondria due to exercise-induced acidification of the muscle cell, resulting in activation of branched-chain 2-oxo acid dehydrogenase complex by inhibition of the kinase.

3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)↗

Plateau pattern of afferent discharge rate from frog muscle spindles.

1. A characteristic plateau pattern was observed in the rate of afferent discharges during ramp-and-hold stretch of spindles isolated from semitendinosus muscles of frogs. The plateau pattern was more frequent in summer frogs (84%) than winter frogs (11%). 2. The plateau pattern consisted of a discharge rate around 120 imp/s at the end of dynamic stretch, followed by second and third steps of plateau rates around 60 and 40 imp/s, respectively. The intervals of impulses in lower steps were approximately n times those of the top step. 3. The plateau pattern was not sensitive to cutting extracapsular myelinated branches or lowering temperature from 23 to 12 degrees C. However, the number of the plateau was reduced in both cases. 4. Application of depolarizing current to the sensory terminal abolished the plateau pattern. In contrast, in spindles that did not show a plateau pattern, hyperpolarizing current induced such a pattern. 5. Calcium channel blockers and protein kinase C inhibitors abolished the plateau pattern. The plateau pattern could be established in quiescent spindles by drugs eliciting Ca2+ entry, raising cytosolic-free Ca2+, and activating protein kinase C. 6. The most striking aspect of the present study is the stability in the discharge rate at each step of the plateau, irrespective of different experimental conditions. This suggests that the spindle sensory terminal possesses a stable intrinsic rhythm generator in excitation, of which maximum frequency is 120 imp/s. The generator seems to be triggered by stretch stimulus and to be regulated by cytoplasmic Ca2+ and protein kinase C.

Afferent Pathways↗

L-gulono-gamma-lactone oxidase is the enzyme responsible for the production of methylguanidine in the rat liver.

A methylguanidine-synthesizing enzyme localized in rat liver microsomes produces methylguanidine via the intermediates creatone A and creatone B from the substrate, creatol, a substance produced from creatinine mainly by reaction with hydroxyl radicals. This enzyme has been identified as L-gulono-gamma-lactone oxidase (EC 1.1.3.8). However, no corresponding activity was found in extracts from human livers.

Amino Acid Sequence↗

Purification of methylguanidine synthase from the rat kidney.

Methylguanidine (MG)-synthesizing enzyme was purified from rat kidney lysosomes and peroxisomes. The enzyme was a flavoprotein with a molecular weight of about 37,000 and oxidized creatol to produce MG. The present results suggest that the reaction mechanism of this enzyme is different from that of L-gulono-gamma-lactone oxidase (EC 1.1.3.8) isolated from rat liver microsomes.

Amino Acid Sequence↗

[Plasma intramembrane structure of overaction of the inferior muscle in humans].

We used the freeze-fracture method to compare the intramembrane structure of the plasma membrane in inferior oblique (IO) muscles from six strabismus patients with normal IO muscles. Inferior oblique myectomy provided us with small pieces of IO muscle from three patients with superior oblique paresis, and from one patient each with V-pattern exotropia, V-pattern esotropia, and V-pattern intermittent exotropia. Samples of normal IO muscle were obtained from two men, aged 33 and 73 years, within 24 hours of their death. The excised muscles were separated at their belly part into orbital and global layers, and small bundles of muscle fiber were fixed and freeze-fractured. The size of intramembrane particles (IMPs) did not differ significantly between normal and strabismus-affected specimens (mean: 9.5 nm). The density of IMP in the 33-year-old man's sample ranged from 800 to 3300/microns2 with a mean of 1823/microns2, in the 73-year-old man's sample ranged from 100 to 2500/microns2 with a mean of 670/microns2, while in the affected tissues it ranged from 100 to 1000/microns2 with a mean 500/microns2. There were no significant differences in caveola size and density between normal and strabismus-affected IO muscles.

Adult↗