Biomedical subjects

K Asayama

Publications and source records attributed to K Asayama.

At least 19 recordsLinked to original sources

PubMed1995

Increased Cu/Zn superoxide dismutase-like immunoreactivity in the swollen axons of rats intoxicated chronically with beta,beta'-iminodipropionitrile.

Demonstration of a genetic linkage between the Cu/Zn superoxide dismutase (SOD1) gene and familial amyotrophic lateral sclerosis (ALS) has aroused interest in the role of SOD1 in spinal motoneuronal death. We used chronically beta,beta'-iminodipropionitrile (IDPN)-intoxicated rats as a model of ALS and investigated SOD1 changes in the spinal cord by immunocytochemical and in situ hybridization techniques. Compared with control rats, SOD1-like immunoreactivity (SOD1-IR) increased in swollen axons of the proximal spinal roots, but not in motoneuronal and dorsal root ganglion neuronal cell bodies where SOD1 gene transcription did not increase. The present data indicate that treatment with IDPN induces accumulation of SOD1 in the swollen axons by blocking slow axonal flow, suggesting the possibility that increased SOD1-IR in ALS is induced by axonal flow blockade.

Amyotrophic Lateral Sclerosis↗

PubMed1995

Development of a sandwich enzyme-linked immunosorbent assay for the determination of human heart type fatty acid-binding protein in plasma and urine by using two different monoclonal antibodies specific for human heart fatty acid-binding protein.

We have developed a sandwich enzyme-linked immunosorbent assay (ELISA) for the determination of human heart type fatty acid-binding protein (H-FABP) in human plasma and urine using the combination of two distinct monoclonal antibodies (MAbs) directed against human H-FABP purified from human heart muscle. The total assay time of the ELISA is practically much shorter than that of the competitive enzyme immunoassay (EIA) we previously reported. The immunoreactive mass of human H-FABP was specifically measured using a horseradish peroxidase (HRPO)-labeled anti-human H-FABP MAb as an enzyme-linked MAb, and anti-human H-FABP MAb immobilized on the polystyrene microtiter plate as a solid-phase MAb, and purified human H-FABP as standard materials. The assay range of the ELISA was 0-250 ng/ml of plasma and urine. The ELISA yielded a coefficient of variation of less than 10% in inter- and intra-assays, and the good linearity was obtained in dilution test using clinical samples. Anticoagulants, except sodium fluoride and a high concentration of hemoglobin and bilirubin, did not interfere with the assay of plasma samples. A high concentration of hemoglobin, bilirubin and immunoglobulin, and contamination with seminal plasma did not interfere with the assay of urine samples. The average recovery of purified human H-FABP added to human plasma and urine samples was 98.5% and 97.0%, respectively. Myoglobin and myosin did not crossreact in the ELISA. The minimum detection limit of the ELISA was 1.25 ng/ml. The immunoreactive masses of human H-FABP in plasma and urine samples, obtained from one hundred normal healthy subjects were quantified by the sandwich ELISA. The normal mean (+/- SD) level of human H-FABP mass in plasma was 3.65 +/- 1.81 ng/ml, and that in urine was 3.20 +/- 2.70 ng/ml. In conclusion, this sandwich ELISA is a useful tool for the sensitive and precise determination of human H-FABP in human plasma and urine, and it may be used specifically for clinical investigation and diagnosis of myocardial injury.

Antibodies, Monoclonal↗

PubMed1995

Cerebellar superoxide dismutase expression in Menkes' kinky hair disease: an immunohistochemical investigation.

This comparative immunohistochemical study deals with the expression of the cytosolic Cu/Zn-binding and mitochondrial Mn-dependent superoxide dismutases (SODs) in the cerebella of five patients with Menkes' kinky hair disease (MKHD) and five age-matched controls. Several cell types, including Purkinje cells and reactive astrocytes, of all MKHD patients examined were intensely stained by an antibody to Mn SOD, but not by an anti-Cu/Zn SOD antibody. By contrast, the cells of the five controls reacted very weakly or not at all with the anti-Mn SOD antibody, but were strongly reactive with the antibody to Cu/Zn SOD. These results suggest that the increased Mn SOD immunoreactivity in MKHD reflects enzyme induction as a protective mechanism against the highly toxic superoxide anion generated under the disease conditions.

Adolescent↗

PubMed1995

Cu/Zn superoxide dismutase-like immunoreactivity is present in Lewy bodies from Parkinson disease: a light and electron microscopic immunocytochemical study.

Copper-zinc superoxide dismutase (SOD1)-like immunoreactivity has been demonstrated in Lewy body-like inclusions (LIs) in brain tissues from patients with familial and sporadic amyotrophic lateral sclerosis. Using immunocytochemistry, we studied Lewy bodies (LBs), the original inclusions from which the term LI was derived, in five patients with Parkinson disease (PD). Surprisingly, many LBs were immunostained by an antibody against SOD1. There were two types of staining pattern: a diffuse pattern, and a peripheral pattern with an unstained core. An immunoelectron microscopic study demonstrated that the immunoreactive products were restricted to the fibrillary profiles, sparing the unstructured core. Our results showed that SOD1-like immunoreactivity occurred frequently in LBs and LIs, suggesting that a common cytopathological process is responsible for the formation of LB-type neuronal intracytoplasmic inclusions. Our results also suggest that SOD1 plays a role in the neurodegeneration associated with PD.

Aged↗

PubMed1995

The functional limitations of tetraplegic hands for intermittent clean self-catheterisation.

Since the introduction of intermittent self-catheterisation by Lapides et al, low pressure voiding methods have been adopted actively in our centre. A retrospective review of 47 patients with cervical spinal cord injury (CSCI) who performed independent (self) clean intermittent catheterisation (CIC) on discharge is presented. Their functional abilities were evaluated by both the Frankel and the Zancolli scales. Fourteen out of the 47 patients could manage independent CIC, 17 assisted CIC, nine tapped or voided voluntarily, two were discharged with a diaper, one with a suprapubic catheter, and four with an indwelling catheter. According to Frankel's classification, four patients were in A category, four in B, one in C and five in D among the group of complete CIC. C6A/C5B (right/left hand) by Zancolli's classification was the highest level of ability for all these 14 patients; and one patient (a 30-year-old male, Frankel B classification) could manage independent CIC with the aid of our special device. However, the practical limit of functional hands for independent CIC appeared to be about C6B1.

Adolescent↗

PubMed1995

Relationships between biochemical abnormalities and anthropometric indices of overweight, adiposity and body fat distribution in Japanese elementary school children.

OBJECTIVE: To determine the anthropometric indices linked to the biochemical risk factors for atherosclerosis in Japanese obese elementary school children, ages ranging from 6 to 12 years. DESIGN: Cross-sectional study of obese children based on fasting blood samples. SETTING: Outpatient clinic of University Hospital. SUBJECTS: 65 consecutive patients with simple obesity (38 boys and 27 girls), and age-matched controls, 184 boys and 205 girls. MAIN OUTCOME MEASURES: Percent obesity and body mass index as indices of being overweight; percent body fat and the sum of four skinfold thicknesses as indices of adiposity; waist-to-hip circumference ratio and waist-to-thigh circumference ratio as indices of body fat distribution. The anthropometric indices were standardized by calculating standard deviation scores based on data from control children. RESULTS: In the obese boys, all six anthropometric indices studied correlated closely with serum biochemical indices, and strong correlations were observed among the indices of overweight, adiposity and body fat distribution. In contrast, only the indices of body fat distribution, not those of overweight or of adiposity, were correlated with serum biochemical indices in the obese girls. No relationship was found between the indices of body fat distribution and the other anthropometric indices in the obese girls. Thus, the profile of the obese girls differed from that of their male counterparts. CONCLUSION: The results suggest that body fat distribution is related to certain biochemical complications of childhood obesity, and that androgyny in fat patterns induces metabolic derangements in children.

Adipose Tissue↗

PubMed1994

Purification and immunoelectron microscopic localization of cellular glutathione peroxidase in rat hepatocytes: quantitative analysis by postembedding method.

To measure quantitatively the intracellular distribution of cellular glutathione peroxidase (GPX) in rat hepatocytes, ultrathin sections were stained by a postembedding immunogold technique. GPX had a specific activity of 1670 Units/mg protein, and was purified 2050-fold from rat liver by means of heat denaturation, ammonium sulfate fractionation, and a series of chromatographic procedures including thiol-Sepharose 4B. The purified GPX was shown to be electrophoretically pure, and was a homotetramer of 22 kDa subunits. Monospecific polyclonal antibodies were raised in rabbits by immunization. By immunoblot analysis, both the light mitochondrial the and cytosolic fractions of rat liver homogenate gave a single band with an identical mobility to that of the purified enzyme. Under the light microscope, hepatocytes showed nuclear staining and granular cytoplasmic staining, corresponding to certain intracellular structures. The labeling density (number of gold particles/microns 2) for GPX obtained by immunoelectron microscopy was 11.9 in the nuclei, 19.6 in mitochondria, 3.32 in peroxisomes, 1.95 in lysosomes, and 9.81 in the cytoplasmic matrix. These results suggest that cellular GPX is present in various compartments of rat hepatocytes, and that the GPX occurs in relatively higher amounts in mitochondria.

Animals↗

PubMed1992

Demonstration of Cu-Zn superoxide dismutase in rat liver peroxisomes. Biochemical and immunochemical evidence.

In this study, by using highly purified rat liver peroxisomes, we provide evidence from analytical cell fractionation, Western blot, and immunocytochemical analysis that Cu-Zn superoxide dismutase is present in animal peroxisomes. Treatment with ciprofibrate, a peroxisome proliferator, increased the peroxisomal superoxide dismutase activity by 3-fold with no effect on mitochondrial activity but a marked decrease in cytosolic superoxide dismutase activity, further supporting that besides cytosolic and mitochondrial localization, Cu-Zn superoxide dismutase is present in peroxisomes also. Demonstration of superoxide dismutase in peroxisomes suggests a new role for this organelle in pathophysiological conditions, such as ischemia-reperfusion injury.

Animals↗

PubMed1992

Proliferation of myocardial peroxisomes in experimental rat diabetes: a biochemical and immunocytochemical study.

Myocardial peroxisomes were investigated in normal and diabetic rats. Catalase and acyl-CoA oxidase activities were increased in the diabetic rat heart and immunoblot analysis showed that both enzyme proteins were markedly enhanced in diabetic heart homogenates. After immunoenzyme staining, catalase and acyl-CoA oxidase were localized in fine granules in the myocardium, which were increased in number in diabetic rats. The numerical density of the granules stained for catalase was increased 1.7 times and that for acyl-CoA oxidase 1.8 times, compared with controls. Protein A-gold labeling for catalase and acyl-CoA oxidase was present in myocardial peroxisomes. The labeling density for both enzymes was increased in diabetic rats by 1.6 times for catalase and 1.5 times for acyl-CoA oxidase, compared with controls. The results indicate that myocardial peroxisomes are increased in the diabetic rat and that this proliferation is accompanied by an increase in catalase and acyl-CoA oxidase activities.

Acyl-CoA Oxidase↗

PubMed1992

Effect of dexamethasone on antioxidant enzymes in fetal rat lungs and kidneys.

To determine if an enhancement in the fetal antioxidant enzyme (AOE) system by maternal dexamethasone (DEX) is specific to organ and dose, the lung and kidney of pups, whose mother received DEX (0.2 or 2 mg/kg) twice, were obtained on days 19 and 20 of gestation. Low-dose DEX increased the four AOE in the day-19 lung, but not in day-20 lung. High-dose DEX decreased the copper-zinc superoxide dismutase (SOD) and glutathione peroxidase in the lungs. Thus, the DEX-induced maturation of lung AOE is dependent on dose and timing. DEX enhanced the four AOE in the day-19 kidney at both doses. In the day-20 kidney, DEX enhanced the manganese SOD at the low dose and also catalase at the high dose, suggesting that DEX accelerates the maturation of kidney AOE as well.

Animals↗

PubMed1992

Immunohistochemical localization of superoxide dismutases in fetal and neonatal rat tissues.

We investigated the developmental profile of copper-zinc and manganese superoxide dismutase (CuZnSOD and MnSOD) in tissue sections obtained from fetal (Day 12 to 21 of gestation) and neonatal (Day 0 and 6) rats. Tissues were stained immunohistochemically with specific antisera against the respective rat SODs. There was a general trend towards richness of SODs in the epithelial linings and metabolically active sites, although differential distribution between the two SODs also existed. At Day 12 of gestation, immunoreactivity for both SODs was detected in the cardiomyocytes but not in other tissues. Hepatocytes expressed CuZnSOD at Day 14 and MnSOD at Day 17. By Day 18 CuZnSOD was detected in the epithelial cells of the gastrointestinal tract, respiratory tract, pancreatic islets, kidneys, and adrenals. These tissues exhibited MnSOD staining at Day 19. CuZnSOD occurred in the epithelia of the thyroid, thymus, and salivary glands at Day 19, while MnSOD was seen at Day 21. The increase in intensity of the staining for SODs occurred no later than postnatal Day 0, indicating that most tissues accumulated SODs during late gestation. Breathing atmospheric oxygen during early extrauterine life did not appreciably intensify the SOD staining. These results suggest that perinatal increase in SODs occurs as a general mechanism of preparation for birth.

Animals↗

PubMed1991

Effect of diabetes mellitus induced by streptozotocin on renal superoxide dismutases in the rat. A radioimmunoassay and immunohistochemical study.

Two forms of superoxide dismutase, CuZn-SOD and MnSOD, have been investigated in the kidneys of streptozotocin-induced diabetic rats using both radio-immunoassay and immunoenzyme staining. The rats were killed 2, 8 and 12 weeks after the induction of diabetes mellitus and the kidneys excised. Two weeks after the induction of diabetes, the kidneys were hypertrophied because of the proliferation of renal tubular epithelium. However, the total CuZnSOD content of the kidneys did not increase and, because of the epithelial proliferation, the CuZnSOD concentration in each proximal tubular cell was decreased. Armanni-Ebstein lesions were found in the distal tubules 8 and 12 weeks after the induction of diabetes. The cells in these lesions were intensely stained for CuZnSOD, suggesting an adaptive response to the enhanced oxidative stress. The MnSOD staining in the thick ascending limbs of Henle's loops was enhanced in the diabetic kidneys, while that in the cortical tubules was unaltered. MnSOD was assumed to increase in response to hypermetabolism associated with the proliferation of renal tubules. This was most marked in the cells which were rich in mitochondria, again suggesting an adaptive response to enhanced oxidative stress induced by diabetes mellitus. The glomeruli of both the diabetic and control groups were not stained for SODs, and no significant microscopic change was found even 12 weeks after the induction of diabetes mellitus.

Animals↗

PubMed1991

Peroxisomal oxidases in various tissues of diabetic rats.

The effect of diabetes mellitus induced by streptozotocin on the activities of peroxisomal oxidases and H2O2-metabolizing enzymes, and lipid peroxidation in various rat tissues were investigated. Peroxisomal acyl-CoA oxidase, D-amino acid oxidase and L-alpha-hydroxyacid oxidase were measured by a sensitive spectrophotometric method using dichlorofluorescein/peroxidase as the detector of H2O2. Acyl-CoA oxidase activity was increased most markedly in the heart of diabetic rats, less markedly in the liver, and tended to be increased in the kidneys. The activities of other peroxisomal oxidases were much lower than that of acyl-CoA oxidase in the liver and kidneys, and were undetectable in the heart. Catalase activity was decreased in the liver and kidneys of diabetics, and was increased in the heart. Glutathione peroxidase activity was increased more markedly in the kidneys of the diabetics, and less markedly in the heart than in the liver. Lipid peroxide level was higher in the kidneys of the diabetics than in the controls, unchanged in the heart, and was lower in the liver of the diabetics than in the controls. Thus, peroxisomal beta-oxidation and the H2O2 production coupled with that, were activated in various tissues of diabetic rats, presumably as a part of the overall increase in lipid oxidation. However, they did not appear to contribute to the enhanced oxidative stress induced by diabetes mellitus.

Acyl-CoA Oxidase↗

PubMed1991

Concentration of lipid peroxide in serum lipoproteins of insulin-dependent diabetic children.

Lipids, apolipoproteins and lipid peroxide were measured in the sera of 29 children with insulin-dependent diabetes mellitus. Ten non-diabetic children served as controls. High-density lipoprotein (HDL) was separated by heparin-MnCl2 precipitation. Lipid peroxides in HDL and non-HDL fractions were estimated by fluorimetric measurement of thiobarbituric acid-reactive substances. The patients were normolipidemic, and their HDL-cholesterol was increased. Apo A1 level in the patients was similar to that in the controls, while levels of apo A2 and apo B were decreased in the patients. Concentrations of lipid peroxides in the whole serum and non-HDL were unaltered, while that in the HDL was higher in the patients than in the controls. Hemoglobin AIc in the patients correlated with the triglyceride and urinary excretion rate of N-acetylglucosaminidase (NAG). The NAG correlated with the triglycerides. The triglycerides correlated with the atherogenic index, apo B and total cholesterol. The lipid peroxides in the non HDL correlated with the triglyceride, atherogenic index, and NAG. That in the HDL correlated with the HDL-cholesterol, apo A1 and endogenous creatinine clearance, and inversely with the atherogenic index and apo B. Lipid peroxides in HDL and non-HDL appeared to play different physiological roles from each other, and they have provided evidence suggesting that diabetic microvascular injury is mediated by reactive oxygen species.

Acetylglucosaminidase↗

PubMed1991

Immunohistochemical study on perinatal development of rat superoxide dismutases in lungs and kidneys.

It has been reported that levels of antioxidant enzymes are low in fetal rat lungs and kidneys, and that they increase rapidly during late gestation. Among the antioxidant enzymes, both copper-zinc and manganese superoxide dismutases (CuZnSOD and MnSOD) are assumed to play a key role in protection against oxidative tissue injury. To determine the nature of the rapid perinatal increase in CuZnSOD and MnSOD, immunoenzyme staining was performed in the lungs and kidneys of fetal (d 18 and 20 of gestation) and neonatal (d 22) rats. The CuZnSOD and MnSOD in the homogenates were assayed by RIA, and they were found to be higher in the neonatal organs than in the respective fetal organs. The neonatal bronchiolar epithelium was stained for both CuZnSOD and MnSOD more intensely than the fetal one. The CuZnSOD staining in the neonatal alveolar wall was more intense than that in the fetal one. There was a significant reactivity for MnSOD in the neonatal, but not in the fetal, alveolar walls. In the kidneys, the reactivities for CuZnSOD and MnSOD were confined to the undifferentiated tubules. Although the tubules were increased in numbers in the neonatal kidneys, the intensity of the staining for both CuZnSOD and MnSOD was unchanged. The histochemical study disclosed that CuZnSOD and MnSOD increased in the kidneys in a manner different from that in the lungs. The low concentration of both CuZnSOD and MnSOD in the fetal lung tissues may contribute to the vulnerability to oxygen toxicity. Such changes in the concentrations in specific tissues were not delineated in the kidneys.

Age Factors↗

PubMed1991

Peroxisomal beta-oxidation in liver and muscles of gold-thioglucose-induced obese mice: correlation with body weight.

The effect of obesity on peroxisomal beta oxidation has been studied previously only in the liver of genetically obese animals. We measured activity of peroxisomal acyl CoA oxidase (ACO) in livers, hearts and rectus femoris muscles of gold-thioglucose treated obese mice (n = 17) and control mice (n = 8). Since production of H2O2 by ACO could contribute to oxidative stress, activities of H2O2-metabolizing enzymes, catalase and glutathione peroxidase, were also measured. ACO activity was assayed using dichlorofluorescein and peroxidase as detectors of H2O2. ACO activity was higher in the obese liver and skeletal muscles than in their respective controls, while the heart ACO activity was unaltered. The activities of H2O2-metabolizing enzymes were unchanged or tended to be decreased in the obese tissues. There was a close correlation between the body weight and ACO activity in both liver and rectus femoris muscles. The ACO activity in the liver also correlated with the liver triacylglycerol content. These results suggest that the activation of peroxisomal beta oxidation occurring in both hepatic and extrahepatic obese tissues is closely linked to weight gain (i.e. non-genetic in nature), but that this does not enhance oxidative stress detected as reactive change of the defense system against H2O2.

Acyl-CoA Oxidase↗

PubMed1990

Peroxisomes of the rat cardiac and soleus muscles increase after starvation. A biochemical and immunocytochemical study.

We have investigated the change of catalase activity in the homogenates of rat cardiac and skeletal muscles. After 7 days' starvation, the catalase activity of heart increased about 3-fold and that of soleus muscle enhanced 2-fold higher than that of control rats. Immunoblot analysis of catalase showed a single band in the homogenates of cardiac and soleus muscles and increase of catalase antigen after starvation. Light microscopic immunoenzyme staining showed that after starvation catalase positive granules markedly increased in both the cardiac and soleus muscle. Quantitative analysis of the staining showed that number of the granules per 100 microns 2 of tissue section was about 1.4-fold in the soleus muscle and 1.7-fold in the cardiac muscle after starvation. By electron microscopy of alkaline DAB staining, we confirmed that the granules were peroxisomes, which increased in both number and size. Furthermore, we stained the peroxisomes for catalase by a protein A-gold technique. Labeling density (gold particles/micron 2) of the cardiac and soleus muscles from the starved rat increased approximately 1.4 times as much as that of normal animal. When the numerical density is multiplied by the labeling density, the values are largely consistent with the enhancement of catalase activity. These results show that increase in the catalase activity of the muscle tissue after starvation is caused by increase in number and size of peroxisomes.

Animals↗

PubMed1990

Oxidative muscular injury and its relevance to hyperthyroidism.

In experimental hyperthyroidism, acceleration of lipid peroxidation occurs in heart and slow-oxidative muscles, suggesting the contribution of reactive oxygen species to the muscular injury caused by thyroid hormones. This article reviews various models of oxidative muscular injury and considers the relevance of the accompanying metabolic derangements to thyrotoxic myopathy and cardiomyopathy, which are the major complications of hyperthyroidism. The muscular injury models in which reactive oxygen species are supposed to play a role are ischemia/reperfusion syndrome, exercise-induced myopathy, heart and skeletal muscle diseases related to the nutritional deficiency of selenium and vitamin E and related disorders, and genetic muscular dystrophies. These models provide evidence that mitochondrial function and the glutathione-dependent antioxidant system are important for the maintenance of the structural and functional integrity of muscular tissues. Thyroid hormones have a profound effect on mitochondrial oxidative activity, synthesis and degradation of proteins and vitamin E, the sensitivity of the tissues to catecholamine, the differentiation of muscle fibers, and the levels of antioxidant enzymes. The large volume of circumstantial evidence presented here indicates that hyperthyroid muscular tissues undergo several biochemical changes that predispose them to free radical-mediated injury.

Animals↗