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Biomedical subjects

A Kosaki

Publications and source records attributed to A Kosaki.

30 records · Page 2Linked to original sources
PubMed1991

Growth inhibitory effect of bestatin on choriocarcinoma cell lines in vitro.

Bestatin, one of the biological response modifiers (BRMs), is an inhibitor of aminopeptidase B (AP-B), leucine aminopeptidase (LAP) and aminopeptidase M (AP-M). In this report, we investigated the direct effect of bestatin on the growth of cancer cells in vitro using established four choriocarcinoma cell lines. In vitro chemosensitivity was evaluated by the succinate dehydrogenase inhibition (SDI) test. Bestatin showed the growth-inhibitory effect on all the choriocarcinoma cell lines dose-dependently, especially on NaUCC-4 cells. Both an isomer of bestatin with no inhibitory activity against aminopeptidases, (2R, 3S)-AHPA-(R)-Leu, and another isomer with stronger inhibitory activity against AP-B than bestatin, (2S, 3S)-AHPA-(R)-Leu, did not show growth inhibition on NaUCC-4 cells. So it is suggested that one of the possible mechanisms responsible for the direct action of bestatin on the choriocarcinoma cells may be related to the inhibition of activity of LAP or AP-M rather than that of AP-B. Furthermore, cytotoxicity of actinomycin D on the choriocarcinoma cells was significantly enhanced by combination with bestatin. These results suggest that bestatin has not only an indirect host-mediated anti-tumor activity, but also a direct growth-inhibitory effect on some kinds of cancer cell lines.

Aminopeptidases↗
PubMed1991

Regulation of glucose transporter synthesis in cultured human skin fibroblasts.

The present study was designed to see the effects of glucose on glucose transporter expression and glucose transport activity using cultured human skin fibroblasts. When the cells were incubated with various concentrations of glucose (11.1-44.4 mM), no differences were found in the HepG2 glucose transporter mRNA, protein levels and basal and insulin-stimulated 2-deoxyglucose uptake. Glucose deprivation, however, resulted in approximately 4-fold increases in the mRNA and 3-fold increases in the protein and the basal 2-deoxyglucose uptake. Chronic exposure to insulin increased the glucose transporter protein levels to similar degrees in the cells incubated with 11.1, 22.2 and 44.4 mM glucose accompanied by increases in the glucose transport activity. Effects of insulin on the glucose transporter mRNA and protein levels, however, were not evident in the glucose-deprived cells. It is concluded that glucose transport activity correlates closely with HepG2 glucose transporter expression in cultured human fibroblasts and that glucose (11.1-44.4 mM) does not affect the glucose transporter expression and glucose transport activity.

Blotting, Western↗
PubMed1991

Insulin-stimulated glucose uptake and fasting blood glucose.

Changes in insulin-stimulated glucose metabolism were studied in young and aged subjects, subjects with impaired glucose tolerance, and patients with NIDDM by means of the glucose clamp technique. The diabetic group includes obese and non-obese patients treated without insulin and non-obese patients treated with insulin. The glucose disposal rate (GDR) was decreased in aged subjects (5.8 +/- 0.4 mg/kg/min) compared with young controls (7.4 +/- 0.3 mg/kg/min). In patients with IGT, it was further decreased to 3.6 +/- 0.5 mg/kg/min, which was comparable to the rate in NIDDM without insulin treatment (3.3 +/- 0.4 mg/kg/min). There were no differences in the GDR between obese (3.0 +/- 0.3 mg/kg/min) and non-obese (3.4 +/- 0.6 mg/kg/min) diabetic patients. In insulin-treated diabetic patients, GDR ranged widely, but the mean value was partially normalized (5.2 +/- 0.9 mg/kg/min). In the diabetic group, no correlation was observed between fasting blood glucose and GDR. These results suggest that in the course of developing NIDDM, a decrease in insulin-stimulated glucose uptake precedes a rise in fasting blood glucose. Thus, as previously reported for Caucasian NIDDM patients, resistance to insulin-stimulated glucose uptake may be one of the basic defects in Japanese patients with NIDDM. The degree of glycemia, however, is not directly related to the magnitude of the defect in insulin action.

Adult↗
PubMed1990

Changes in insulin receptor kinase with aging in rat skeletal muscle and liver.

To see if insulin receptor kinase activity alters with aging, the activity of wheat germ agglutinin-purified receptor preparations from liver and skeletal muscle was compared in 2-, 4-, 10-, and 20-mo-old rats. Basal and insulin-stimulated autophosphorylation of liver insulin receptor and its kinase activities toward histone 2b and poly(Glu4Tyr1) did not alter with aging. On the other hand, the muscle insulin receptor showed different results. Insulin-stimulated increases of autophosphorylation and the kinase activity toward histone 2b above basal were comparable in the four groups. However, insulin-stimulated phosphorylation of poly(Glu4Tyr1) was decreased in 20-mo-old rats compared with 10- and 4-mo-old rats. These results indicate that insulin receptor kinase activity could vary under certain conditions, depending on the substrate used to measure the activity. It is concluded that insulin receptor kinase activity does not change markedly during the process of aging, although subtle changes seem to exist.

Adenosine Triphosphate↗
PubMed1990

Binding activity and autophosphorylation of the insulin receptor from patients with myotonic dystrophy.

To clarify a mechanism of insulin resistance associated with myotonic dystrophy, we studied the insulin receptor by using three different types of cells--circulating erythrocytes, cultured skin fibroblasts, and Epstein-Barr virus(EBV)-transformed lymphocytes. In myotonic dystrophy, insulin binding to erythrocytes and fibroblasts was significantly decreased as a result of a reduction of the binding affinity. Insulin binding to EBV-transformed lymphocytes was normal. When the receptors were purified from fibroblasts with wheat germ agglutinin, we could not find a decrease in the binding affinity seen in the intact cells. No difference was observed in the magnitude of basal and insulin-stimulated autophosphorylation of insulin receptors from EBV-transformed lymphocytes between the control and myotonic dystrophy. Southern blot analysis of the insulin receptor gene revealed no restriction fragment length polymorphism associated with myotonic dystrophy. These findings suggest that there is no primary defect of the insulin receptor per se in terms of insulin binding and autophosphorylation in myotonic dystrophy. The reduction of the insulin binding to erythrocytes and fibroblasts may be caused by the plasma membrane abnormality that affects the binding affinity of the receptor.

Adult↗
PubMed1989

Postreceptor defect in insulin action in streptozotocin-induced diabetic rats.

To clarify the mechanism(s) responsible for the insulin resistance in streptozotocin (STZ)-treated diabetic rats, we studied insulin-induced glucose disposal by using the glucose clamp technique and measured insulin receptor and glucose transporter of muscles. The insulin dose-response curve of the metabolic clearance rate (MCR) of glucose revealed a decrease of the maximal response without a rightward shift in STZ rats. Maximal MCR was even lower when clamped at 300 rather than 150 mg/dl of blood glucose levels. Insulin binding to the crude plasma membrane of muscles from STZ rats was increased compared with controls. The number of glucose transporter of the plasma and microsomal membranes were significantly decreased in STZ rats. These in vivo and in vitro studies using skeletal muscles suggest that in STZ-treated diabetic rats 1) a defect or defects exist in the signal transduction mechanism of insulin in postbinding steps, 2) the decreased maximal MCR is related at least partly to the decrease of glucose transporter numbers, and 3) a defect in glucose metabolism (postglucose transport defect) is also present.

Animals↗
PubMed1988

Insulin-resistant diabetes due to a point mutation that prevents insulin proreceptor processing.

A point mutation in the human insulin receptor gene in a patient with type A insulin resistance alters the amino acid sequence within the tetrabasic processing site of the proreceptor molecule from Arg-Lys-Arg-Arg to Arg-Lys-Arg-Ser. Epstein-Barr virus-transformed lymphocytes from this patient synthesize an insulin receptor precursor that is normally glycosylated and inserted into the plasma membrane but is not cleaved to mature alpha and beta subunits. Insulin binding to these cells is severely reduced but can be increased about fivefold by gentle treatment with trypsin, accompanied by the appearance of normal alpha subunits. These results indicate that proteolysis of the proreceptor is necessary for its normal full insulin-binding sensitivity and signal-transducing activity and that a cellular protease that is more stringent in its specificity than trypsin is required to process the receptor precursor.

Adult↗
PubMed1988

Defective processing of insulin-receptor precursor in cultured lymphocytes from a patient with extreme insulin resistance.

We have studied a patient with extreme insulin resistance, acanthosis nigricans, and decreased erythrocyte insulin binding. EBV-transformed lymphocytes from this patient exhibited markedly reduced binding of 125I-insulin. Radioiodination of cell surface receptors followed by immunoprecipitation with anti-receptor antibodies revealed the presence of increased amounts of a 210-kD protein but no detectable alpha or beta subunits. Continuous labeling with 2-[3H]mannose revealed the synthesis of a 190-kD precursor and a 210-kD protein. The 210-kD protein was phosphorylated in an insulin-dependent manner at high insulin concentrations. These results suggest that in this patient the biosynthesis of 190-kD receptor precursor, its terminal glycosylation, and intracellular transport to the cell surface proceed normally, while proteolytic maturation to alpha and beta subunits does not occur. We postulate that this defect either results from mutation(s) within the insulin-receptor gene, which render the precursor resistant to cleavage, or from a defect in the receptor processing enzyme.

Acanthosis Nigricans↗
PubMed1988

Insulin resistance in a boy with congenital generalized lipodystrophy.

We have studied insulin resistance in a 12-year-old Japanese boy who presented with congenital generalized lipodystrophy. Oral glucose tolerance test exhibited a diabetic pattern with normal fasting plasma glucose. Results from euglycemic glucose clamp study showed decreases in both insulin sensitivity and responsiveness. Both the patient's erythrocytes and Epstein-Barr virus transformed lymphocytes showed low-normal insulin binding with a slight reduction in binding affinity in the latter. Insulin binding to the cultured fibroblasts was decreased due to a lowered affinity. In addition, they displayed a rightward shift of the insulin dose-response curve for D-14C-glucose uptake with no decrease in the maximum uptake. Insulin-stimulated autophosphorylation and kinase activity of the wheat germ agglutinin purified receptors from the Epstein-Barr virus-transformed lymphocytes appeared normal. The reason for some discrepancies in insulin binding among the cells remains unknown, and we cannot formulate a conclusion as to whether or not a primary binding defect of insulin receptors exists and contributes to insulin resistance in the patient. The decrease in insulin responsiveness demonstrated in the glucose clamp study may result from a defect at the rate-limiting step in the postbinding process of insulin action, presumably a defect in the glucose transport system in muscle tissues. The defect may be secondary to changes in in vivo circumstances.

Cell Line, Transformed↗
PubMed1988

Regulation of glucose-transporter gene expression by insulin in cultured human fibroblasts.

To clarify the effect of insulin on glucose-transporter (GT) biosynthesis, we determined GT mRNA levels in human cultured skin fibroblasts, using HepG2 GT cDNA as a probe. Insulin specifically increased the GT mRNA level in a time- and dose-dependent manner. Time-course study demonstrated that the mRNA level peaked within 3 h of insulin (1 x 10(-7) M) addition. After remaining elevated for several hours, mRNA decreased and returned to the basal level after 24 h. In the cell strains from seven normal subjects, the mean (+/- SE) GT mRNA level determined after 3 h of treatment with 1 x 10(-7) M insulin was 164.3 +/- 8.5% of the level found in untreated control cells. The insulin dose-response curve of GT mRNA levels showed that the maximum stimulation was elicited at 1 x 10(-7) M, and the half-maximum stimulation occurred at approximately 5 x 10(-10) M. Degradation rates of GT mRNA determined in the presence of actinomycin D were not different between insulin-treated and untreated cells. These results suggest that insulin increases GT gene expression in cultured human fibroblasts.

Cells, Cultured↗