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Biomedical subjects

K Utsumi

Publications and source records attributed to K Utsumi.

At least 73 records · Page 4Linked to original sources
PubMed1998

Differentiation elicits negative regulation of human beta-galactoside alpha2,6-sialyltransferase at the mRNA level in the HL-60 cell line.

We studied the regulation of the beta-galactoside alpha2,6-sialyltransferase (hST6Gal I) gene during HL-60 cell differentiation induced with dimethyl sulfoxide (DMSO), all transretinoic acid (ATRA), and phorbol myristate acetate (PMA). During HL-60 cell line differentiation, cell surface levels of alpha2,6-sialic acids expression decreased, as measured by flow cytometric analysis using Sambucus nigra agglutinin (SNA). Activities of hST6Gal I and levels of hST6Gal I mRNA dramatically decreased after 1 day of stimulation. Using reverse transcription polymerase chain reaction (PT-PCR), we found the major hST6Gal I mRNA isoform in HL-60 cells contains 5'-untranslated exons Y and Z. These results suggest that the expression of cell surface alpha2,6-sialic acids is controlled at the mRNA level, which is regulated by a promoter located 5'-upstream of exon Y.

Base Sequence↗
PubMed1998

Oxidative damage in the senescence-accelerated mouse.

The senescence-accelerated mouse (SAM) exhibited a shortened life span (about 18 months) and early manifestation of various signs of senescence, including changes in physical activity, skin, and spinal curvature. The mechanism of senescence acceleration in SAM is thought to be related to free radical damage. Oxidative phosphorylation was estimated in liver mitochondria from SAMPS and the senescence-resistant subtrain, SAMR1. The respiratory control ratio decreased during aging, and the ATP/O, an index of ATP synthesis, was depressed at 18 months of age in SAMPS. DNP-dependent uncoupled respiration in liver mitochondria was markedly decreased, and active uptake of calcium was markedly dysfunctional with aging. These findings suggest that the functional disorders in mitochondria may be closely related to the shorter life span of SAMPS. White-footed (WF) mice can live at least to 5.5 years, when some animals are still capable of reproducing and their external body condition remains healthy. The mitochondrial functions were examined in the same way as in the SAM experiments. However, no particular finding responsible for their longevity was observed in WF mice at 3 and 12 months old. More comprehensive examinations on more aged WF mice are needed for explanation of their greater longevity.

Aging↗
PubMed1998

Activation of caspase-3-like protease by digitonin-treated lysosomes.

Apoptosis, a naturally occurring programmed cell death or cell 'suicide', has been paid much attention as one of the critical mechanisms for morphogenesis and tissue remodeling. Activation of cysteine aspartases (caspases) is one of the critical steps leading to apoptosis. Although a mitochondria-mediated pathway has been postulated to be one of the activation mechanism of caspase-3, another subcellular compartment might be involved in the activation of the enzyme. The present study shows that the supernatant fraction of digitonin-treated lysosomes strongly activates Ac-DEVD-CHO inhibitable caspase-3-like protease. Activation of caspase-3-like protease by digitonin-treated lysosomal fractions was specifically suppressed by leupeptin and E-64, inhibitors of cysteine protease. These results indicate that leakage of lysosomal cysteine protease(s) into the cytosolic compartment might be involved in the activation of caspase-3-like protease.

Animals↗
PubMed1998

Protective protein/cathepsin A loss in cultured cells derived from an early-infantile form of galactosialidosis patients homozygous for the A1184-G transition (Y395C mutation).

Galactosialidosis is a human autosomal recessive lysosomal storage disease caused by a genetic defect of protective protein/cathepsin A (PPCA). The patients in a Japanese family with the severe early-infantile form of galactosialidosis were revealed to be homozygous for the A1184-G transition in the PPCA gene in both alleles, which leads to the Y395C substitution. The acid carboxypeptidase (cathepsin A) and lysosomal neuraminidase activities were markedly decreased in cultured fibroblasts and chorionic villus cells derived from the patients, although the decrease in beta-galactosidase activity was less. Immunoblot and immunocytochemical analyses showed that neither the precursor nor the mature form of the PPCA gene product was present in the cultured cells. The Y395C mutation was revealed to cause the loss of the translated product, that determines the severity of the clinical phenotype.

Amino Acid Substitution↗
PubMed1998

Immunohistochemical characterization of transgenic mice highly expressing human lysosomal alpha-galactosidase.

Human lysosomal alpha-galactosidase predominantly hydrolyzes ceramide trihexoside. A transgenic mouse line, C57BL/6CrSIc-TgN(GLA) 1951 Rin, highly expressing human alpha-galactosidase, has been established and investigated biochemically and immunohistochemically in order to clarify the distribution of the expressed enzyme proteins and to evaluate it as a donor model of organ transplantation therapy for Fabry disease caused by a genetic defect of alpha-galactosidase. In these transgenic mice, about five copies of the transgene were integrated, and alpha-galactosidase activity was expressed in liver, kidney, heart, spleen, small intestine, submaxillary gland, skeletal muscle, cerebrum, cerebellum, bone marrow cells and serum. The enzyme activity was about 22 to 11,080-fold higher than that in non-transgenic mice. In liver, heart and kidney tissues, which are important organs for transplantation studies, sufficient amounts of alpha-galactosidase mRNAs were transcribed, and the expressed enzymes, with molecular weights of 54-60 kDa, are abundant in the liver (enzyme activity: 53,965 nmol h-1 mg-1 protein) and heart (39,906 nmol h-1 mg-1 protein), followed by in the kidney tissue (9177 nmol h-1 mg-1 protein), respectively. An immunohistochemical microscopic study clearly demonstrated the distribution of the expressed enzyme proteins in kidney and liver tissues. Highly expressed alpha-galactosidase was detected in glomerular cells, tubular cells and hepatocytes. These transgenic mice will be useful as a donor model for experimental organ transplantation, and also it will enable recurrent biopsies and long-term observation. The organ transplantation data on mice will provide us with important information.

Animals↗
PubMed1998

Adult Sandhoff's disease: R505Q and I207V substitutions in the HEXB gene of the first Japanese case.

We describe a 31-year-old Japanese man with adult Sandhoff s disease presenting as spinocerebellar degeneration. There was a marked cerebellar atrophy on MRI, and proliferation of abundant PAS-positive foamy macrophages in the rectal mucosa. The activities of total beta-Hex, beta-Hex A, and beta-Hex B in leucocytes of the patient were 14%, 15%, and 6% of control values, respectively. However, oligosacchariduria or ultrastructural storage materials in liver tissue were nil. Direct sequencing of cDNA and genomic DNA, and restriction digestion revealed two different homozygous base substitutions in the HEXB gene: the G1514-->A substitution (R505Q) and the A619-->G substitution (1207V). The parents were consanguineous. His healthy mother, an enzymatic heterozygous carrier, was homozygous for 1207V, but heterozygous for R505Q mutation. Thus, the patient is probably homozygous for both base substitutions and a R505Q mutation may be linked to the phenotype of adult Sandhoff's disease.

Adult↗
PubMed1998

Mitochondrial dysfunction in the senescence accelerated mouse (SAM).

Oxidative damage to DNA, proteins, and lipids in mitochondria caused by free radicals may be one factor in aging. Oxidative phosphorylation was estimated in liver mitochondria from senescence accelerated mice (SAMP8) and a senescence resistant substrain (SAMR1). The respiratory control ratio decreased in liver mitochondria of SAMP8 during aging, and it was estimated that at 18 months of age this respiratory control value suggested that it might be insufficient to provide ATP synthesis necessary for normal cell metabolism. In addition, the ADP/O, an index of efficiency of ATP synthesis, was depressed at 18 months of age. Dinitrophenol-dependent uncoupled respiration in liver mitochondria of SAMP8 mice was markedly decreased with aging, suggesting a dysfunctional energy transfer mechanism in mitochondria of aged SAMP8 mice. Active uptake of calcium in liver mitochondria was markedly dysfunctional in SAMP8 mice with aging, and uncoupling of respiration was induced more easily in aged mitochondria. Milder effects on these functional parameters were observed in SAMR1 mice. A similar dysfunction was also observed in heart mitochondria of SAMP8 mice at 12 months of age. The amount of Bcl-x in liver mitochondria was slightly decreased in SAMP8. We suggest that these changes in mitochondrial function may be related to the shorter life span of the senescence accelerated mouse.

Aging↗
PubMed1998

Role of glutathione in nitric oxide-dependent regulation of energy metabolism in rat hepatoma cells.

Previous studies in this laboratory revealed that nitric oxide (NO) reversibly inhibits the respiration of isolated mitochondria and ascites hepatoma (AH-130) cells by an oxygen concentration-dependent mechanism. The inhibitory effect of NO on the respiration of AH-130 cells was enhanced by treating with digitonin that selectively permeabilized plasma membranes and released cytosolic low-molecular-weight compounds. Reduced glutathione (GSH) is the most abundant cytosolic thiol that easily reacts with NO. To elucidate the mechanism by which digitonin enhanced the inhibitory action of NO, the effect of GSH and related thiols was studied with AH-130 cells and their mitochondria. The inhibitory effect of NO on the respiration of digitonin-treated cells was suppressed by either GSH, L-cysteine, or N-acetylcysteine, but not by oxidized glutathione. The inhibitory effect of NO on the respiration of their mitochondria was also decreased by GSH. In contrast, the inhibitory effect of NO was markedly enhanced with AH-130 cells obtained from animals that were pretreated with L-buthionine sulfoximine (BSO), a specific inhibitor for GSH synthesis. Kinetic analysis revealed that NO dose-dependently decreased GSH levels in AH-130 cells with concomitant generation of S-nitrosothiols. Although S-nitrosoglutathione (GSNO), a slow releaser of NO, also inhibited the respiration of tumor cell mitochondria, its effect was significantly lower than that of NO. These results suggest that cellular GSH might play pivotal roles in the regulation of energy metabolism in hepatoma cells by modulating free forms of NO.

Acetylcysteine↗
PubMed1998

Macrophage-derived nitric oxide induces apoptosis of rat hepatoma cells in vivo.

Although nitric oxide (NO) has been postulated to play important roles in host defense mechanisms against tumor cells, a direct evidence supporting this hypothesis is lacking. To obtain molecular insights into the antitumor action of NO, its metabolism and effect on ascites hepatoma (AH-130) cells were investigated in tumor-bearing rats. Kinetic analysis revealed that substantial amounts of nitrite and nitrate, metabolites of NO, appeared in plasma and ascites of AH-130-inoculated rats. Western blot analysis revealed that a large number of macrophages that expressed inducible type of NO synthase (iNOS) appeared in cancerous ascites, particularly during 1 to 2 weeks after inoculation of AH-130 cells. When NO generation by peritoneal macrophages increased, a significant fraction of AH-130 in ascites fluid underwent apoptosis as judged from the fragmentation of their nuclear DNA. Kinetic analysis revealed that NO strongly inhibited mitochondrial electron transport and changed calcium status in AH-130 cells, particularly under low oxygen tensions such as in cancerous ascites. Intraperitoneal injection of NO donor strongly enhanced DNA fragmentation of AH-130 cells. Antimycin A, a specific inhibitor for mitochondrial electron transport, also induced DNA fragmentation of AH-130 cells by a mechanism that was inhibited by adding ascorbate and tetramethyl-p-phenylene diamine (TMPD) as electron donors. These results indicate that NO derived from peritoneal macrophages inhibits mitochondrial electron transport and disturbs calcium homeostasis in ascites hepatoma AH-130 cells, thereby inducing their apoptosis in vivo.

Animals↗
PubMed1998

[Safety and indication of open heart surgery without blood transfusion for congenital cardiac defects in infants].

Between January 1994 and June 1997, 16 cases of ventricular septal defect (VSD) and endocardial cushion defect (ECD) with pulmonary hypertension (PH), each weighing from 5 to 9 kg, underwent definitive surgery at Matsudo Municipal Hospital. We classified them into 2 groups; Group N: 8 cases without blood transfusion, Group H: 8 cases with blood transfusion. Cardiopulmonary bypass (CPB) system was a closed circuit and priming volume was 370 to 500 ml. There was no difference between the 2 groups in operative age, body weight, preoperative state, operation time, CPB time, aortic cross clamp time, bleeding and postoperative state. In Group N, CPB blood was returned to the patient as soon as possible after CPB was weaned, and postoperative hemodynamics were stable in both groups. In Group N, hematocrit (Ht) values were consistently lower than in Group H, from initiation of CPB to leaving the hospital. To accomplish safe CPB, we measured systemic venous oxygen saturation (SvO2). In 6 cases of Group N, SvO2 during rewarming was 48.1 +/- 16.0% and Ht value was 13.2 +/- 1.5%. It is thought that the safe CPB could be conducted in Group N. In addition, in Group N, respiratory index showed better values than in Group H during the postoperative period. It is thought that CPB without blood transfusion may be favorable to prevent lung injury after CPB. Retrospectively, it is thought that, to accomplish safe CPB without blood transfusion, preoperative Ht values of over 30% are desirable in patients weighing 6 kg and those of over 35% are desirable in patients weighing 5 kg.

Blood Transfusion↗
PubMed1998

[Open heart surgery without blood transfusion for cyanotic congenital cardiac defects].

Between November 1994 and January 1997, 42 cases of cyanotic congenital cardiac defects underwent definitive surgery at Matsudo Municipal Hospital. We evaluated 30 cases, each weighing from 7 to 20 kg. The procedures were performed at the age of 9 months to 6 years (mean age-2.4 years). The body weights were 7.7 to 20 kg (mean weight-11.4 kg). The preoperative diagnoses were Tetralogy of Fallot (TOF) in 19 cases, Fontan candidates in 6 and the others in 5. We classified them into 3 groups; Group A--15 cases were completed with non-blood transfusion, Group B--8 cases used only plasma protein fraction and Group C--7 cases used blood transfusion. Cardiopulmonary bypass (CPB) system is a semi-closed circuit and priming volume is 400 to 600 ml. There is no difference among the 3 groups in operative age, body weight, operation time, CPB time, aortic cross clamp time, bleeding and postoperative state. The same results were obtained in minimum base excess and urine output during CPB and the changes of hematocrit and total protein. In Groups A and B, CPB blood was returned to the patient as soon as possible after CPB was weaned, but in Group C, blood transfusion was performed without the return of CPB blood. In all groups, hemodynamics were stable. Retrospectively, it is thought that blood transfusion was not necessary in Group C and the use of the plasma protein fraction was not needed in Group B. In conclusion, the open heart surgery can be performed safely without blood transfusion for cyanotic congenital cardiac defects.

Blood Transfusion↗
PubMed1998

Effects of cumulus cell density during in vitro maturation of the developmental competence of bovine oocytes.

To determine the role of cumulus cells in oocyte maturation, we carried out an investigation on the effects of addition of cumulus cells to the maturation medium on the developmental competence of corona-enclosed oocytes and oocytes denuded from their somatic cells. The addition of cumulus cell (1.6 x 10(6) cells/mL) improved the development of bovine corona-enclosed oocytes, however, addition of a similar number of cumulus cells as cumulus-oocyte-complexes (COCs, cumulus cell density: 4.2 x 10(6) cells/mL) had no effect on the development of oocytes denuded from their somatic cells. To determine if corona-enclosed oocytes can obtain developmental competence without the addition of extra cumulus cells, the effects of cell density during in vitro maturation on the developmental competence were studied. A density of 1.6 to 3.2 x 10(6) cumulus cells/mL was the most effective for in vitro maturation of oocytes with intact gap junctions. The effects of the medium conditioned by COCs on the developmental competence of oocytes was also examined. It was demonstrated that COC-conditioned medium improved the development of bovine oocytes to the blastocyst stage. These data suggest that the developmental competence of bovine oocytes surrounded with corona cells is supported in a cell density-dependent manner in the maturation medium. In addition, the data indicate that cumulus cells benefit bovine oocyte development either by secreting soluble factors which induce developmental competence or by removing an embryo development-suppressive component from the medium.

Animals↗
PubMed1998

Alpha-fetoprotein producing pulmonary blastoma in a patient with systemic sclerosis: pathogenetic analysis.

We describe a rare case of pulmonary blastoma complicated with systemic sclerosis. The serum alpha-fetoprotein level was elevated at presentation and the biopsied tumour stained positively against anti-alpha-fetoprotein antibody. The alpha-fetoprotein produced autonomously by tumour cells was of yolk-sac origin. Although the pathogenesis of pulmonary blastoma has not been clarified, we suggest that this pulmonary blastoma is a type of yolk-sac tumour.

Female↗
PubMed1998

Biochemical properties of human oral polymorphonuclear leukocytes.

Polymorphonuclear leukocytes (PMN) isolated from the oral cavity of healthy human volunteers, spontaneously generated superoxide, nitric oxide (NO) and other reactive oxygen species (ROS) which exhibited strong luminol chemiluminescence (LCL). To understand the physiological roles of oral PMN (OPMN), biochemical properties of the cells were analyzed. Biochemical analysis revealed that OPMN were already primed under physiological conditions. Western blot analysis revealed that they strongly expressed the inducible type of NO synthase (NOS II) and exhibited the activity to catalyze tyrosine phosphorylation of various proteins including a 115 kDa protein (cbl product). OPMN also generated H2O2 and .OH by some superoxide dismutase (SOD)-sensitive mechanism and released myeloperoxidase (MPO). Kinetic analysis using specific inhibitors revealed that OCl- generated by OPMN was predominantly responsible for the enhanced LCL. During the incubation under standard culture conditions, OPMN underwent apoptosis which proceeded more rapidly than that of the circulating PMN (CPMN). Immunochemical analysis revealed that expression of apoptosis-related gene products, such as Bcl-2, Bcl-xL and Bax, was below detectable levels with both cell types. However, caspase-3 but not caspase-1 was markedly activated in OPMN. These results indicate that the primed OPMN spontaneously generate ROS and play an important role in the defense mechanism in the oral cavity and that the generated ROS activate caspase-3 thereby inducing apoptosis of the cells.

Apoptosis↗
PubMed1998

[Management for the postoperative mediastinitis in infancy].

Postoperative mediastinitis is a rare but life-threatening complication after cardiac surgery. We successfully managed three infants with postoperative mediastinitis. When the postoperative mediastinitis was suspected, intravenous infusion of antibiotics (Vancomycin) and local irrigation were performed. The reoperation for closure was planned when the value of c-reactive protein decreased to 1.0-2.0. An application of a pectoral musculocutaneous flap was effective when the sternum was destroyed by infection.

Anti-Bacterial Agents↗
PubMed1997

Generation and characterization of transgenic mice expressing a human mutant alpha-galactosidase with an R301Q substitution causing a variant form of Fabry disease.

Transgenic mice expressing a human mutant alpha-galactosidase with an R301Q substitution, which was found in a patient with a variant form of Fabry disease, were established. The mice transcribed a sufficient amount of alpha-galactosidase mRNA, but the steady-state levels of the enzyme protein were decreased in liver, kidney and heart, only residual activity being detected in these tissues. The mice will be useful for the clarification of the defective regulation of the structurally altered enzyme protein expressed by the mutant gene at the organ or individual level as well as for the evaluation of drugs that stabilize and/or activate the mutant alpha-galactosidase.

Animals↗
PubMed1997

Evaluation of acrosomal status of bovine spermatozoa using concanavalin a lectin.

We report here that fluorescein isothiocyanate-conjugated concanavalin A (FITC-ConA) specifically labels the acrosomal region of acrosome-reacted bovine spermatozoa. This labeling is found to be useful in evaluating the acrosome status of bovine spermatozoa. When fresh bovine spermatozoa that had been fixed with 4% formaldehyde, smeared on glass slides and then air-dried were stained by FITC-ConA, weak fluorescence was observed on the acrosomal region, although almost all the spermatozoa appeared to be acrosome-intact. However, when fresh sperm suspensions were incubated with FITC-ConA and then mounted on glass slides, no fluorescence was observed on the acrosomal region. Therefore, in the ensuing experiments, both the fixation and the FITC-ConA staining of spermatozoa were done in suspension. When ethanol-treated spermatozoa, whose outer membrane may be permeabilized, were stained with FITC-ConA, the fluorescence was extensively observed on the inner acrosomal region. This fluorescence was inhibited in the presence of 0.2 M D-mannose, a competitive sugar, suggesting that FITC-ConA binds specifically to glycocomponents on the inner acrosomal membrane. We next tried to stain fresh or frozen-thawed spermatozoa from 3 different bulls that had been treated with the calcium ionophore A23187, which is known to induce acrosome reaction of bovine spermatozoa, with FITC-ConA. A significant correlation between the percentage of ConA-labeled spermatozoa and that of rose bengal stained negative ones at various time points during A23187 incubation was achieved. Furthermore, suitability of dual staining to distinguish between physiological acrosome reaction (acrosome-lost and live) and degenerative acrosomal loss (acrosome-lost and dead) using FITC-ConA and Hoechst bis-benzimide 33258 (H258) supravital stain was also confirmed. From these results, it was concluded that the FITC-ConA labeling procedure is a feasible and reliable method for the assessment of physiological acrosome reaction of bovine spermatozoa.

Journal Article↗