Biomedical subjects
S Akazawa
Publications and source records attributed to S Akazawa.
The heterogeneity of whole islet cell cytoplasmic antibodies evidenced by absorption test with glutamic acid decarboxylase.
A recent report identified two islet cell cytoplasmic antibody subclasses using an immunohistochemical method. The islet cell cytoplasmic antibody subclass which reacts with only Beta-cells was termed 'Beta-cell islet cell cytoplasmic antibodies' and another islet cell cytoplasmic antibody subclass which reacts with Beta and non-Beta cells was called 'whole islet cell cytoplasmic antibodies'. The whole islet cell cytoplasmic antibody reactivity with pancreatic islets has been shown not to be abolished by pre-incubation with rat brain homogenate. In this study, we examined the inhibitory effect of purified glutamic acid decarboxylase to islet cell cytoplasmic antibody reactivity among whole islet cell cytoplasmic antibodies and assessed the heterogeneity of islet cell cytoplasmic antibodies. Auto-antibodies to 64,000 Mr islet cell protein (64 K antibodies) were also determined by conventional method. Sera from 17 Type 1 (insulin-dependent) diabetic patients containing whole islet cell cytoplasmic antibodies with more than 20 Juvenile Diabetes Foundation units were used. In 11 (78.6%) of 14 sera positive for 64 K antibodies, the reactivity of islet cell cytoplasmic antibodies was markedly blocked by pre-incubation with purified glutamic acid decarboxylase. In contrast, none of the 64 K antibody-negative sera were blocked. All of the patients showed similar clinical characteristics regardless of the inhibitory effect of glutamic acid decarboxylase on islet cell cytoplasmic antibodies, except for islet cell cytoplasmic antibody titer and glutamic acid decarboxylase antibody titer. The mean log 2 islet cell cytoplasmic antibody titer was 2.4 +/- 0.6 (mean +/- SD) JDF unit in the 'markedly blocked' group and 1.6 +/- 0.3 (mean +/- SD) in the 'never blocked' group. The islet cell cytoplasmic antibody titer was significantly higher (P < 0.05) in the former, and the mean glutamic acid decarboxylase antibody titer was 624 +/- 127.0 (mean +/- SE) units in the 'markedly blocked' group and 127 +/- 55.5 (mean +/- SE) in the 'never blocked' group. The glutamic acid decarboxylase antibody titer was also significantly higher (P < 0.05) in the former. We demonstrated here that some whole islet cell cytoplasmic antibodies are absorbed by purified glutamic acid decarboxylase, suggesting heterogeneity of islet cell cytoplasmic antibodies among the 64 K glutamic acid decarboxylase antibody positive group.
Evaluation of islet-specific autoantibodies in Japanese patients with insulin-dependent diabetes mellitus: a comparison between autoantibodies to glutamic acid decarboxylase, autoantibodies to 64 kDa islet cell protein and islet cell antibodies.
Autoantibodies to glutamic acid decarboxylase (GAD), autoantibodies to 64 kDa islet cell protein and islet cell antibodies (ICA) were measured in 79 Japanese patients with insulin-dependent diabetes mellitus (IDDM). The overall prevalences of GAD antibodies, 64K antibodies, and ICA in these patients were 69.6% (55/79), 48.1% (38/79), and 46.8% (37/79), respectively. However, in a subset of these patients with recent onset IDDM (< 1 year) the prevalences of GAD antibodies, 64K antibodies, and ICA were 78.8% (26/33), 66.7% (22/33), and 78.8% (26/33), respectively. Furthermore, the prevalences of GAD antibodies, 64K antibodies, and ICA were significantly decreased in patients with long standing diabetes at 60.9% (28/46), 34.8% (16/46), and 23.9% (11/46), respectively. However, when these patients were divided into two groups by the presence or absence of organ-specific autoimmune disease (OSAD), the mean levels of GAD antibodies and ICA in the patients who gave a positive result were significantly higher in patients with OSAD (397 units and 98 JDF units, respectively) than in patients without OSAD (74 units and 39 JDF units, respectively). These results demonstrate that it is important to evaluate the prevalences and levels of islet-specific autoantibodies when considering disease duration and co-existence of autoimmune disease in patients with IDDM.
Glucose metabolism in separated embryos and investing membranes during organogenesis in the rat.
Glucose metabolism provides an essential energy source in the mammalian embryo. We used the rat embryo culture system to investigate the activity of several critical pathways for glucose metabolism during early organogenesis, before (day 10 of gestation) and during (day 11) the establishment of the chorioallantoic circulation and closure of the neural tube. We studied glucose metabolism in the intact conceptus, the separated embryo, and its investing membranes, including the visceral yolk sac, allantois, and amnion. Short-term incubations were performed for 4 hours in culture media containing U-14C-, 14C-1-, or 14C-6-labeled D-glucose on day 10 and day 11 of gestation, and the rates of glucose utilization by glycolysis and oxidative metabolism, including the pentose phosphate pathway (PPP), were measured. Glycolytic metabolism, estimated by the accumulation of lactate, was high on day 10 in the intact conceptus, embryo, and membranes (92 +/- 6, 63 +/- 5, and 99 +/- 8 nmol/micrograms protein/4 h, respectively) and decreased by two thirds by day 11 in each tissue. Of note, the rate of glycolysis was greater in membranes than in the embryo on both days (P < .01). On the other hand, oxidative metabolism, reflected in the production of 14CO2, was relatively low on day 10 (0.78 +/- 0.04, 0.73 +/- 0.05, and 0.6 +/- 0.06 nmol/micrograms protein/4 h for intact conceptus, embryo, and membranes, respectively) and increased significantly in each tissue by day 11 (P < .01 for each).
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)
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.
Autoantibodies to glutamic acid decarboxylase in patients with IDDM and autoimmune thyroid disease.
Autoantibodies to glutamic acid decarboxylase (GAD), previously reported to be the 64,000-M(r) (64K) islet cell protein, were measured by a radioimmunoassay using purified pig brain GAD in 29 insulin-dependent diabetes mellitus (IDDM) patients with autoimmune thyroid disease (AITD) and in 29 sex- and disease duration-matched IDDM patients without AITD. Islet cell antibodies (ICAs) and 64K antibodies were also determined. In IDDM patients with short-duration diabetes (< 1 year), the prevalence and levels of GAD antibodies were 100% (8 of 8) and 609 +/- 166 U (means +/- SE), respectively, in IDDM patients with AITD and 81.8% (9 of 11) and 90 +/- 51 U, respectively, in patients without AITD. In patients with long-standing IDDM (3-22 years), the prevalence and levels of GAD antibodies were 76.2% (16 of 21) and 193 +/- 66 U, respectively, in patients with AITD and 50.0% (9 of 18) and 36 +/- 14 U, respectively, in patients without AITD. For up to 6 years after the onset of IDDM, the levels of GAD antibodies in IDDM patients with AITD were significantly higher than in IDDM patients without AITD. A close and significant correlation was found between GAD antibodies and ICA or 64K antibodies in IDDM patients with AITD. Our results demonstrate that high levels of GAD antibodies were present in IDDM patients with AITD. The observed differences in GAD immunoreactivity between IDDM patients with and without AITD might help evaluate the role of GAD antibodies in IDDM.
[Effects of repeated sevoflurane anesthesia on hepatic and renal function in a pediatric patient].
A 10-yr-old boy with an injured lower extremity received sevoflurane anesthesia 5 times within 40 days. Laboratory tests for hepatic and renal function i.e., serum transaminase (glutamic oxaloacetic transaminase, glutamic pyruvic transaminase, gamma-glutamyl transpeptidase), serum cholinesterase, plasma protein, serum cholinesterase, serum bilirubine, serum lactic dehydrogenase, serum prothrombin time, blood urea nitrogen, serum creatinine, beta 2-microglobulin, N-acetyl-D-glucosamidase and 24 hr-creatinine clearance remained within normal ranges throughout his perioperative period. Repeated sevoflurane anesthesia did not exert any adverse effect on hepatic and renal function in this patient.
[Usefulness of continuous intra-arterial blood gas monitoring in a patient undergoing tracheal transection and reconstruction].
Usefulness of a new continuous intra-arterial blood gas monitoring system (PB3300, Puritan-Bennett, Carlsbad, California) was evaluated in a patient with tracheal cancer who was undergoing tracheal transection and reconstruction. The PB3300 detected continuous changes in pH, PaCO2, and PaO2 during the surgical intervention and provided reliable information to take appropriate therapeutic measures. PB3300 was superior to pulse oximetry because the latter can not detect changes in PaO2 more than 100 mmHg accompanying pulmonary dysfunction caused by surgical procedures. We conclude that the PB3300 is a useful monitor when used during anesthetic management of the patient who needs frequent arterial blood gas analyses.
[A method of tracheal intubation tried in a patient with multicentric reticulohistiocytosis].
A 48-year-old woman had suffered from multiple destructive arthritis, skin nodules and skin ulcer of bilateral legs caused by multicentric reticulohistiocytosis for 16 years. Skin transplantation and total hip joint replacement were scheduled. Tracheal intubation was anticipated to be difficult because of destructive arthritis of the temporomandibular joint. We tried a new method to intubate the trachea through a laryngeal mask. After topical application of 8% lidocaine to the oropharynx, a size 4 laryngeal mask was inserted and its cuff was inflated by 20 ml of air. Anesthesia was induced with 4% sevoflurane in 50% nitrous oxide via laryngeal mask. A fiberscope through a tracheal tube was inserted through the laryngeal mask into the trachea. A tracheal tube with the internal diameter of 6.5 mm, was successfully intubated. This method gave little discomfort to the patient.
A phase II study of combination therapy with 5'-deoxy-5-fluorouridine and cisplatin in the treatment of advanced gastric cancer with primary foci.
BACKGROUND: 5'-Deoxy-5-fluorouridine (5'-DFUR, doxifluridine) is a recently developed prodrug of oral 5-fluorouracil (5-FU), which is used clinically in Japan for the treatment of gastric, colorectal, and breast cancer. 5-FU has been reported to act synergistically with cisplatin (CDDP) in experimental and clinical studies. The authors conducted a multicenter Phase II study of combination therapy with 5'-DFUR and CDDP to evaluate the therapeutic usefulness of this regimen in the treatment of unresectable and advanced gastric cancers with primary foci. None of the patients had previously undergone chemotherapy. Their ages ranged from 27 to 75 years and performance status was grade 0 to 3. METHODS: 5'-DFUR (1400 mg/m2/d) was administered orally on days 1 through 4 and 15 through 18, and CDDP (80 mg/m2/d) was injected intravenously on day 5. This treatment cycle was repeated every 4 weeks. An independent panel of specialists evaluated the clinical response. RESULTS: Fifty-one patients were studied. Clinical evaluation of response was possible in 43 patients who met the protocol requirements. The overall response rate was 50.0% (14 of 28, 95% confidence limits, 30.7%-69.4%) for patients with measurable lesions. The median duration of response was 5.2 months (156 days). The overall median survival time was 8.9 months (268 days) for evaluated patients. Therapeutic toxicity of World Health Organization (WHO) grade > or = 3 was manifested as anorexia and nausea or vomiting in 20.9% and 18.6% of the patients, respectively. However myelotoxicity and nephrotoxicity of WHO grades 3 and 4 occurred in less than 10% of the patient group. No drug-related mortality occurred. CONCLUSIONS: Combined therapy with 5'-DFUR and CDDP is a safe and effective treatment regimen for advanced gastric cancers with primary foci which stresses the patient's quality of life, especially when used in an outpatient setting.
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.
Cloning and expression of large isoform of glutamic acid decarboxylase from human pancreatic islet.
Glutamic acid decarboxylase (GAD) catalyzes formation of gamma-aminobutyric acid from glutamic acid and is a major autoantigen in insulin-dependent diabetes mellitus. Its two isoforms, GAD65 and GAD67, are encoded by two separate genes. We prepared human islet cDNA library and screened it with cDNA probes of rat brain GAD67. We cloned the cDNA for GAD67, the large isoform of glutamic acid decarboxylase, and determined its nucleotide sequence. Sequencing of the resulting clone identified a 1,785 residue open-reading frame encoded a 594 amino acid polypeptide that showed a 99.4% similarity with GAD67 from human brain. The bacterially expressed human islet GAD67 protein was enzymatically active and immunoreactive. The isolation of cDNA for this additional islet GAD isoforms will be important in studying the etiology and pathogenesis of IDDM.
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.
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.
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.
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.
[Rapid induction of anesthesia with inhalation of sevoflurane].
Induction of anesthesia with inhalation of sevoflurane was evaluated in 45 patients. Inhalation anesthesia was induced by a single vital capacity breath followed by spontaneous breathing. The compositions of anesthetic gases were as follows: 5% sevoflurane in oxygen, 5% sevoflurane in 67% nitrous oxide and oxygen, and 7% sevoflurane in 67% nitrous oxide and oxygen. The time (mean +/- SEM) necessary for the loss of consciousness was 85 +/- 6, 56 +/- 4, and 42 +/- 3 seconds, and breathing frequency was 8 +/- 1, 5 +/- 0.6, 1.4 +/- 0.2, in each composition of anesthetic gasses, respectively. Changes in blood pressure and heart rate were relatively small. The technique was found to be acceptable to all of the patients studied. We conclude that the technique of single breath induction with sevoflurane is a safe and acceptable alternative to intravenous induction in cooperative adult patients.