Biomedical subjects

S Okuno

Publications and source records attributed to S Okuno.

At least 163 records · Page 9Linked to original sources

PubMed1994

Increased insulin responsiveness after CS-045 treatment in diabetes associated with Werner's syndrome.

Werner's syndrome is a rare inheritated disorder characterized by accelerated aging and is often accompanied by diabetes mellitus or impaired glucose tolerance. Previous reports suggest that insulin resistance is involved in the development of diabetes associated with Werner's syndrome. In the present study, CS-045((+/-)-5-[4-(6-Hydroxy-2,5,7,8-tetramethylchroman-2-ylmet hoxy)benzyl] - 2,4-thiazolidinedione, a new oral hypoglycemic agent which reportedly reduces insulin resistance, was administered to 2 Werner's syndrome patients. The patients were hospitalized for the duration of the study. During a pretreatment period lasting 8 weeks the patients received a controlled diet, however, their previous treatment was unchanged. Throughout the 4-week treatment period, each subject's blood glucose level was measured 7 times each day (07:30, 10:00, 11:30, 14:00, 17:30, 20:00, 22:00) for 1 week at 8, 4, and 1 week before treatment and at 2 and 4 weeks after treatment. To assess insulin action, the euglycemic glucose clamp technique was performed in these subjects at insulin infusion rates of 20, 120 and 400 mU/kg/min before and after 4 weeks of treatment. After 4 weeks of treatment with CS-045, the mean blood glucose level at each time point measured in this study was markedly lower compared to the corresponding pretreatment level.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

PubMed1994

New teleocidin-related metabolites, (-)-7-geranylindolactam V and blastmycetin F, from Streptoverticillium blastmyceticum.

Two new teleocidin-related metabolites, (-)-7-geranylindolactam-V [2] and blastmycetin F [3], were isolated from fermentation broths of the actinomycete Streptoverticillium blastmyceticum NA34-17, and their structures were determined by spectroscopic methods. Compound 2 bound strongly to phorbol ester receptors in a mouse epidermal particulate fraction, suggesting that it is a potent in vivo tumor promoter comparable to teleocidins A-1 [4] and B-4 [5].

Animals↗

PubMed1994

cDNA cloning and expression of human calmodulin-dependent protein kinase IV.

Calmodulin-dependent protein kinase IV (CaM-kinase IV) is a Ca(2+)-responsive multifunctional protein kinase which occurs abundantly in the brain and thymus. A human cDNA clone encoding CaM-kinase IV was isolated from a Jurkat cell cDNA library and its nucleotide sequence was determined. The cDNA sequence encoded a protein consisting of 473 amino acids with a molecular weight of 51,925. The nucleotide sequence for the coding region and the deduced amino acid sequence showed 81 and 80% identities with those of the rat enzyme, respectively. Western blot analysis, using a polyclonal antibody raised against the recombinant human CaM-kinase IV, which was expressed in Escherichia coli, revealed two bands corresponding in mobility to molecular weights of 60,000 and 61,000, respectively, in a Jurkat cell extract. The antibody also cross-reacted with both isoforms of CaM-kinase IV from rat cerebellum, the apparent molecular weights being 62,000 and 64,000, respectively.

Amino Acid Sequence↗

PubMed1994

Inactivation of Ca2+/calmodulin-dependent protein kinase II by Ca2+/calmodulin.

Incubation of calmodulin-dependent protein kinase II with Ca2+ and calmodulin resulted in a marked inactivation of the enzyme. Chelation of Ca2+ by EGTA or addition of calmodulin antagonists, W-7 or trifluoperazine, completely blocked this inactivation. The concentration required for the half-maximal inactivation, 127 microM, is three to four orders of magnitude higher than that for the half-maximal activation of the enzyme. The Ca2+/calmodulin-independent activity of the proteolytic fragment of the enzyme, whose calmodulin-binding site involved in the enzyme activation was deleted, was also decreased by incubation with Ca2+ and calmodulin. These results suggest that calmodulin-dependent protein kinase II possesses a second, low-affinity calmodulin-binding site, which is distinct from the calmodulin-binding site involved in the activation of the enzyme, and that the binding of calmodulin to the second binding site causes the inactivation of the enzyme. The inactivation by Ca2+/calmodulin was temperature-dependent. The addition of both 500 microM ADP and 10 mM MgCl2 markedly protected the enzyme against the inactivation, while such a marked protection was not observed after the addition of either of the two alone. The addition of 5 microM autocamtide-2, a synthetic substrate peptide containing the amino acid sequence of the autophosphorylation site (Thr286/Thr287 in alpha/beta, gamma, and delta isoforms) lying within the autoinhibitory domain, also protected the enzyme against the inactivation by Ca2+/calmodulin, while syntide-2, another synthetic substrate peptide corresponding to a phosphorylation site of glycogen synthase, did not protect it even at a concentration as high as 304 microM.

Animals↗

PubMed1994

Purification and characterization of Ca2+/calmodulin-dependent protein kinase IV kinase from rat brain.

Calmodulin-dependent protein kinase IV (CaM-kinase IV) kinase was recently discovered in the rat brain by its activity to activate the inactive recombinant CaM-kinase IV expressed in Escherichia coli [Okuno, S. and Fujisawa, H. (1993) J. Biochem. 114, 167-170]. In the present study, CaM-kinase IV kinase was purified approximately 2,000-fold from rat cerebral cortex by purification procedures including calmodulin affinity chromatography, and its properties were examined. The highly purified CaM-kinase IV kinase gave one major protein band corresponding to a molecular weight of about 66,000 upon SDS-PAGE. The purified CaM-kinase IV kinase phosphorylated and concomitantly activated CaM-kinase IV purified from rat brain as well as the recombinant kinase expressed in Escherichia coli in a Ca2+/calmodulin-dependent manner. The phosphorylation of CaM-kinase IV by CaM-kinase IV kinase occurred on only serine residue(s). Among a number of proteins, including several known to be phosphorylated by the various protein kinases tested, CaM-kinase IV was the best substrate for CaM-kinase IV kinase. Since syntide-2, a synthetic peptide known to be a good peptide substrate for calmodulin-dependent protein kinase II (CaM-kinase II), was a fairly good substrate for CaM-kinase IV kinase, some kinetic properties of CaM-kinase IV kinase were examined using syntide-2 as a substrate. The Km value for the peptide substrate in the presence of Ca2+/calmodulin was almost two orders of magnitude lower than that in its absence, although the Vmax value was almost the same in the presence and absence of Ca2+/calmodulin.

Amino Acid Sequence↗

PubMed1994

Cellular-tissue localization and regulation of the GLUT-1 protein in both the embryo and the visceral yolk sac from normal and experimental diabetic rats during the early postimplantation period.

We investigated the tissue-specific developmental expression and localization of GLUT-1 protein in the rat embryo and visceral yolk sac (VYS) during the organogenic periods of normal rats. The expression of GLUT-1 protein was then compared to that of experimental diabetic rats to test whether the diabetic state would affect the regulation of the glucose transporter during the early postimplantation periods (9.5-14.5 days), as we have previously demonstrated that GLUT-1 protein in embryo and VYS was down-regulated in culture with hyperglycemic medium. In the embryo, GLUT-1 protein was highly expressed during the early stages of organogenesis (between 9.5-12.5 days) and declined thereafter, whereas in the VYS, its strong expression was observed at the later stages (from 12.5-14.5 days). Immunohistochemical localization of the GLUT-1 protein in the embryo during the main periods of neurulation (9.5-11.5 days) showed that GLUT-1 immunoreactivity was principally observed in the neuroepithelial cells of the neural tube and also noted in the primitive heart, primitive gut, otic, and optic vesicles. At 12.5 days, GLUT-1 protein started to be expressed in the microvessels at the cranial portions of the neural tube, although its expression in the neuroepithelial cells still remained at the caudal (tail) portions of the neural tube. In the later stages (13.5-14.5 days) after completion of neural tube formation, GLUT-1 protein immunoreactivity substantially decreased in the neuroepithelial cells and was found mainly in the microvessels of the brain vesicles and spinal cord, whereas it continued to be expressed in the heart and eyes. In the VYS, its immunoreactivity was noticeably confined to the endodermal layer, which started as a simple layer and developed wave-like folds in the later stages. The levels of GLUT-1 protein in embryo and VYS from diabetic rats, determined by Western blot analysis, were not down-regulated compared to those in control rats at the different gestational days. Likewise, comparison of GLUT-1 protein immunoreactivity of various tissues in embryo and VYS, focusing on the neural tube, also revealed no significant differences between the two groups. We demonstrated that GLUT-1 protein is abundantly expressed in embryonic tissues and VYS during the early periods of organogenesis. The lack of down-regulation and the continuous abundant expression of the GLUT-1 protein despite the diabetic state in embryo and VYS during the early postimplantation periods may increase delivery of glucose from the VYS into various differentiating embryonic cells, leading to diabetes-induced congenital malformations.

Animals↗

PubMed1994

Tissue-targeting ability of saccharide-poly(L-lysine) conjugates.

To evaluate the effect of introducing a saccharide moiety to poly(amino acids) on tissue distribution, several glycoconjugates of epsilon-(2-methoxyethoxyacetyl)-poly(L-lysine) of three molecular weights were synthesized using an octylene spacer between the sugar and polymer chain. Methoxyethoxyacetylation of the epsilon-amino group of the lysine unit in poly(L-lysine) was useful for avoiding nonspecific distribution to many tissues as the result of cationic charges. The tissue-targeting ability of each saccharide moiety was considered as the actual amount changed in each tissue caused by saccharide modification. Galactose terminated saccharides such as galactose, lactose and N-acetylgalactosamine accumulated exclusively in the liver, probably by the hepatic receptor. These conjugates could therefore be good carriers for a drug delivery system to the liver. On the other hand, the mannosyl and fucosyl conjugates were preferentially delivered to the reticuloendothelial systems such as those in the liver, spleen and bone marrow. In particular, fucosyl conjugates accumulated more in the bone marrow than in the spleen. Xylosyl conjugates accumulated mostly in the liver and lung. Generally, the accumulated amount in the target tissue increased with increasing molecular weight and an increased number of saccharides on one molecule of polymer.

Animals↗

PubMed1993

Regulation of release of hepatocyte growth factor from human promyelocytic leukemia cells, HL-60, by 1,25-dihydroxyvitamin D3, 12-O-tetradecanoylphorbol 13-acetate, and dibutyryl cyclic adenosine monophosphate.

Hepatocyte growth factor (HGF) secreted from human promyelocytic leukemia cell line, HL-60, is indistinguishable from HGF in human plasma and its release is significantly stimulated by 12-O-tetradecanoylphorbol-13-acetate (TPA), a differentiation-inducer of HL-60 cells into monocytes/macrophages (Nishino T et al: Biochem Biophys Res Commun 181:323, 1991). TPA stimulated HGF release from the cells through an activation of C-kinase, but not through a formation of reactive oxygen species. Furthermore, dibutyryl cAMP (dbcAMP), an activator of A-kinase and granulocyte-inducer, also stimulated HGF release. 1,25-Dihydroxyvitamin D3, another monocyte/macrophage-inducer, abated either TPA- or dbcAMP-stimulated synthesis and release of HGF in a dose-dependent manner probably via its nuclear receptor as reflected by vitamin D analog study. The effects of these three reagents on the steady-state levels of HGF mRNA of 6.0 kb corresponded with their effects on its protein levels. Furthermore, a close correlation between intracellular and extracellular HGF levels strongly suggested that these reagents affected HGF release mainly on its synthesis step. Recombinant human HGF significantly stimulated the proliferation and alkaline phosphatase activity of mouse osteoblastic cell line, MC3T3-E1. In summary, HL-60 cells secrete HGF, whose synthesis is specifically regulated by various reagents independent of their differentiation-inducing effects. Because HGF shows a direct effect on osteoblast-like cells, it might be involved in the interaction of bone marrow cells with bone cells.

Alkaline Phosphatase↗

PubMed1993

Developmental change of facilitative glucose transporter expression in rat embryonal and fetal intestine.

The developmental expression and localization of facilitative glucose transporter protein (GLUT-1, GLUT-2 and GLUT-5) in rat embryonal and fetal intestine were examined using immunohistochemical methods. The intestines were collected from embryo and fetus (term days, 10 to 20 of gestation), newborn and adult rats (16 weeks after birth). GLUT-1 was expressed abundantly in the gut tube on day 10 of gestation. From day 10 to birth, GLUT-1 was expressed continuously but its expression decreased gradually. In the adult, no GLUT-1 was detected. GLUT-2 was not expressed from day 10 to 14, but began to be expressed in the lumen of the gut tube on day 16, before the intestinal villi were evident. On day 18, when the intestinal villi were developing, GLUT-2 was localized to the basolateral membranes of the enterocytes. A similar pattern of GLUT-2 localization was also observed in newborn and adult rat intestines. The expression kinetics of GLUT-5 were almost the same as those of GLUT-2. GLUT-5 began to be expressed in the lumen of the gut tube on day 16. In contrast to GLUT-2, GLUT-5 was localized to the brush border membranes of intestinal villi after day 18. These results indicate that the expression and localization of each member of the glucose transporter family in the embryonal and fetal intestine vary with the gestational phase. GLUT-1 may be required for glucose metabolism early in gestation, and GLUT-2 and GLUT-5 may be required in both late gestational and perinatal stages.

Animals↗

PubMed1993

Effect of substituting fluorine for hydrogen at C-26 and C-27 on the side chain of 1,25-dihydroxyvitamin D3.

Previous reports have demonstrated that introduction of fluorine atoms at C-26 and C-27 of 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) results in the potentiation of various aspects of some biological activities. The higher biological activities of 26,26,26,27,27,27-hexafluoro- 1,25-dihydroxyvitamin D3 (26,27-F6-1,25-(OH)2D3) were accounted for in part by a decrease in metabolic inactivation via the 26- and 27-hydroxylation pathways. In addition to 26,27-F6-1,25-(OH)2D3 not being hydroxylated in the 26 and 27 positions, it did not undergo 24-hydroxylation despite a significant induction by 26,27-F6-1,25-(OH)2D3 of 24-hydroxylase activity in the HL-60 cell system. Another fluorinated vitamin D3 analog, 26,26,26,27,27,27-hexafluoro-1 alpha-hydroxyvitamin D3 (26,27-F6-1 alpha-OH-D3) may not undergo 25-hydroxylation as efficiently as 1 alpha-OH-D3 in vivo because a rise in serum 26,27-F6-1,25-(OH)2D3 levels after injection of 26,27-F6-1 alpha-OH-D3 was delayed significantly with a much smaller amplitude. Furthermore, 26,26,26,27,27,27-hexafluoro-1,23(S),25-trihydroxyvitamin D3 retained full activity in the induction of HL-60 cell differentiation even after 23(S)-hydroxylation, in contrast to 1,23(S),25-(OH)3D3. These data suggested that substitution of fluorines for hydrogens at C-26 and at C-27 positions may result in alteration in chemical reactivity and/or conformation of C-23, C-24 and C-25 positions of the 1,25-(OH)2D3 molecule.

Animals↗

PubMed1993

Expression of GLUT4 glucose transporter mRNA and protein in skeletal muscle and adipose tissue from rats in late pregnancy.

We investigated the influence of pregnancy on the expression of insulin-regulated glucose transporter (GLUT4) mRNA and protein in skeletal muscle and adipose tissue. GLUT4 mRNA expression in quadriceps muscle from control and pregnant rats was similar in the fasted and fed states. When the level of expression was determined as the immunoreactive GLUT4 content per gram of tissue, the relative GLUT4 protein content of red quadriceps from pregnant rats was significantly higher than that for control rats in both the fed and fasted states. GLUT4 protein expression in white quadriceps from pregnant rats was significantly increased in the fed state compared to control rats, but there was no significant difference in the fasted state. In adipose tissue, the relative GLUT4 protein content was significantly lower in pregnant rats than control rats in the fasted state, but there was no significant decrease in the fed state. Although the expression of GLUT4 protein showed these variations in late pregnancy, there was no significant difference of GLUT4 mRNA expression between control and pregnant rats. These findings suggest that GLUT4 kinetics may differ between late pregnancy and the normal state, and these changes may be related to insulin resistance in pregnancy.

Adipose Tissue↗

PubMed1993

Glucose transporter gene expression in rat conceptus during high glucose culture.

We investigated the expression of glucose transporter genes and protein in embryo and yolk sac during organogenesis and the regulation of glucose transporters during culture in hyperglycaemic media. Erythrocyte-type glucose transporter (GLUT 1) and brain-type glucose transporter (GLUT 3) mRNA were expressed in embryo and yolk sac. The expression of GLUT-1 and GLUT-3 mRNA was abundant on day 9-11 and day 9-10 in the embryo, respectively, and day 9-14 and day 10-11 in the yolk sac, respectively. The levels of GLUT-1 protein in the embryo increased in parallel with the expression of GLUT-1 mRNA during the corresponding period. Immunohistochemical staining of GLUT-1 protein was found principally in the neuroepithelial cells surrounding the neural tube in the embryo on day 10 and appeared in the microvessels surrounding the neural tube after day 12. To test whether the expression of glucose transporter genes and protein was suppressed during hyperglycaemia, conceptuses were cultured in high glucose medium. The abundant expression of GLUT-1 protein was not decreased during culture in high glucose media for 24 h (day 9-10) and was only down-regulated by prolonged exposure to this media for 48 h (day 9-11). We have demonstrated the predominant expression of the high affinity glucose transporter (GLUT 1 and GLUT 3) genes and (GLUT 1) protein in embryo during the early period of organogenesis. The persistently abundant expression of glucose transporter during the critical period of neural tube formation (day 9-10) even in the presence of hyperglycaemia may explain one of the mechanism of increased glucose flux into the neuroepithelium, which may lead to neural tube defects.

Animals↗

PubMed1993

Glucose transporter gene expression in rat conceptus during early organogenesis and exposure to insulin-induced hypoglycemic serum.

We investigated the glucose transporter gene and protein expression during early organogenesis in the rat and in rat embryos cultured with hypoglycemic serum. Erythrocyte-type glucose transporter (GLUT-1) mRNA was expressed at a high level in embryos; peak levels were reached at days 10.5-11.5 and decreased as gestational age increased. In contrast, the insulin regulatable glucose transporter (GLUT-4) mRNA was not detected. The levels of GLUT-1 protein determined by Western blot analysis increased in parallel with expression of the glucose transporter (GLUT-1) gene and peak levels were observed on days 10.5 and 11.5, which correspond to the main periods of neural tube formation. Immunohistochemical staining of the embryo on day 10.5 showed that GLUT-1 protein was abundantly located in the tissue of neural tube. When embryos were cultured from day 9.5 to day 10.5 with insulin-induced hypoglycemic serum containing 2-3 mM glucose an increased frequency of anterior neural tube defects was observed in association with a significant reduction of the glycolytic flux. Increased levels of GLUT-1 mRNA and protein were not observed during the culture with hypoglycemic serum compared with the levels in embryos cultured in normal serum. Addition of insulin to normal serum (500 microU/ml) did not affect the GLUT-1 mRNA and protein levels. GLUT-1 mRNA and protein are strongly expressed in the embryo during early organogenesis, especially in the tissues of the neural tube, and the expression of the glucose transporter did not increase in response to prolonged glycopenia. This may account for the vulnerability of embryogenesis to hypoglycemia during these critical developmental periods.

Animals↗

PubMed1993

Possible selective effects of interferon alpha-2b on a malignant clone in a case of polycythemia vera.

A patient with polycythemia vera associated with chromosomal translocation (6;8) (q27;p11) and IgA monoclonal gammopathy was treated with interferon alpha-2b. Interferon alpha-2b induced good hematological control and reduced only IgA levels, but not IgG and IgM levels. In addition, cytogenetic improvement was obtained, the abnormal cells with chromosomal translocation declining from 100% to 50%. Interferon alpha may have selective effects on a malignant clone in polycythemia vera.

Chromosome Banding↗

PubMed1993

Lecithin-cholesterol acyltransferase and lipid transfer protein activities in liver disease.

The activities of lecithin-cholesterol acyltransferase (LCAT) and lipid transfer protein (LTP) were assayed using sensitive radioassay methods in controls (n = 113) and in patients with various liver diseases (n = 72). Plasma LCAT activity decreased with progression of hepatocellular damage. Plasma LTP activity in controls was 216 +/- 68 nmol/mL/h, and there were no significant differences between controls and patients with chronic hepatitis ([CH], 193 +/- 70), compensated liver cirrhosis (LC) with or without hepatocellular carcinoma ([HCC], 197 +/- 48 and 193 +/- 62, respectively), or decompensated liver cirrhosis ([dLC], 182 +/- 65). In acute viral hepatitis, LTP activity decreased significantly; however, the degree of reduction was not as dramatic as that for LCAT. There was no correlation between LCAT and LTP activity both in controls and patients with various liver diseases. LCAT activity was positively correlated with serum albumin (r = .52, P < 0.1) and cholinesterase (r = .37, P < .01) levels, and inversely correlated with serum bilirubin level (r = -.38, P < 0.1); there was no correlation between plasma LTP activity and these parameters of liver function. That plasma LTP activity did not change with hepatocellular damage may indicate that the liver in humans may not be the primary site of LTP production.

Acute Disease↗

PubMed1993

Requirement of brain extract for the activity of brain calmodulin-dependent protein kinase IV expressed in Escherichia coli.

Calmodulin-dependent protein kinase IV (CaM-kinase IV) is a Ca(2+)-responsive multifunctional protein kinase which occurs abundantly in the brain. When cDNA for rat brain CaM-kinase IV was expressed in Escherichia coli, the enzyme was produced in a good yield, but it did not show significant activity. The inactive recombinant CaM-kinase IV was phosphorylated and became highly active on incubation with a rat brain extract in the presence of both Ca2+/calmodulin and ATP/Mg2+. The recombinant CaM-kinase IV-activating activity in brain was one to two orders of magnitude higher than that in the other tissues examined. These observations suggest that CaM-kinase IV may undergo a posttranslational modification, probably Ca2+/calmodulin-dependent phosphorylation by CaM-kinase IV kinase, before exhibiting activity in the central nervous system.

Animals↗

PubMed1993

Effect of uremic serum on 1,25-dihydroxyvitamin D3-induced differentiation of human promyelocytic leukemia cell, HL-60.

The mechanism by which resistance to 1,25-(OH)2D3 occurs in patients with chronic renal failure was studied. 1,25-(OH)2D3 causes the induction of differentiation and of 1,25-(OH)2D3-24-hydroxylase activity in the mitochondria of the human promyelocytic leukemia cell line, HL-60, via a steroid hormone-receptor mechanism. Treatment of these cells with 10(-8) M 1,25-(OH)2D3 for 5 days in a medium containing 10% uremic serum from 4 patients with chronic renal failure resulted in maturation of the cells amounting to 30.3 +/- 18.7 (mean +/- SD) and 32.5 +/- 11.2% maturation by the nitroblue tetrazolium reduction assay and the nonspecific esterase assay, respectively. These values were significantly lower than those obtained with 10% normal serum from 3 normal controls (66.6 +/- 12.8 and 58.3 +/- 10.9%, p < 0.02). The occurrence of resistance to 1,25-(OH)2D3 in uremic serum-treated cells was also confirmed when the effect of 1,25-(OH)2D3 was assessed by the induction of the cell's ability to hydroxylate the C-24 position of 1,25-(OH)2[3H]D3. Treatment of HL-60 cells with a mixture of 5% uremic plus 5% normal serum impaired 1,25-(OH)2D3-induced cell differentiation to the levels as those in 10% uremic serum, strongly suggesting the occurrence of a substance(s) having 1,25-(OH)2D3-inhibitory activity in the uremic serum. A significant reduction in 1,25-(OH)2D3 receptor levels was observed in uremic serum-treated cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Blood Proteins↗

PubMed1993

[Isolated ACTH deficiency associated with Hashimoto disease].

Isolated ACTH deficiency is a rare cause of secondary adrenocortical insufficiency. The diagnosis is made by the demonstration of low cortisol production with low plasma ACTH, absent adrenal responses to stimulation for pituitary or hypothalamus with intact adrenal response to exogenous ACTH, and normal secretory indices of other pituitary hormones. Although the pathogenesis of isolated ACTH deficiency is uncertain in most cases, autoimmune process is intimately involved as suggested by the histological evidence of lymphocytic hypophysitis and frequent observation of circulating antipituitary antibodies. In isolated ACTH deficiency, there have been occasional associations with Hashimoto disease which is characterized by the presence of a autoimmune mechanism. This suggests the possibility of common autoimmune process affecting both the pituitary and the thyroid gland.

Adrenocorticotropic Hormone↗