No abstract

The present study was conducted to determine the extent of insulin deficiency and glucagon excess in the hyperglycemia of type 2 diabetes in children. The incidence of type 2 diabetes mellitus in children and adolescents has increased substantially over the past several years. Because insulin and glucagon action both regulate blood glucose concentration, we studied their responses to mixed meals in children with type 2 diabetes. Subjects were 24 patients with type 2 diabetes compared with 24 controls, aged 9--20 yr (predominantly African-Americans), matched for body mass index and sexual maturation. All of those with diabetes were negative for antibodies to glutamic acid decarboxylase. Plasma glucose, glucagon, and serum C-peptide concentrations were measured at 0, 30, 60, 90, and 120 min after a mixed liquid meal (Sustacal) ingestion (7 mL/kg body weight; maximum, 360 mL). The area under the curve (AUC) was calculated by trapezoidal estimation. The incremental C-peptide (Delta CP) in response to the mixed meal was calculated (peak -- fasting C-peptide). The plasma glucose AUC was significantly greater in patients than in controls (mean +/- SEM, 1231 +/- 138 vs. 591 +/- 13 mmol/L x min; P < 0.001). The Delta CP was significantly lower in those with diabetes than in controls (1168 +/- 162 vs. 1814 +/- 222 pmol/L; P < 0.02). Glucagon responses did not differ between the two groups. Hyperglycemia is known to inhibit glucagon secretion. Therefore, our patients with substantial hyperglycemia would be expected to have decreased glucagon responses compared with controls and are thus relatively hyperglucagonemic. Patients were divided into poorly and well controlled subgroups (glycosylated hemoglobin A(1c), > or =7.2% and <7.2%, respectively). There were no significant differences in the Delta CP and glucagon responses between these two subgroups. We next analyzed the data in terms of duration of diabetes (long term, > or =1 yr; short term, <1 yr). The CP was significantly lower in long- vs. short-term patients (768 +/- 232 vs. 1407 +/- 199 pmol/L; P < 0.05). The plasma glucagon AUC was significantly higher in the long- vs. short-term patients (9029 +/- 976 vs. 6074 +/- 291 ng/L x min; P < 0.001). Hemoglobin A(1c) did not differ between long- vs. short-term patients. Our results indicate that relative hypoinsulinemia and hyperglucagonemia represent the pancreatic beta- and alpha-cell dysfunctions in children with type 2 diabetes. The severity of both beta- and alpha-cell dysfunctions appears to be determined by the duration of diabetes.

Two N-linked sites of glycosylation in the insulin receptor were examined for their contribution to insulin binding, tyrosine kinase activity, and receptor biosynthesis. Asn397 and Asn418 were replaced by Gln using site-directed mutagenesis either as single mutations, i.e., Q-397 and Q-418, or as a double mutation in which both sites were removed (Q-D). The mutations were transiently expressed in COS cells and the findings compared with cells that transiently expressed the wild-type human insulin receptor. Q-397 and Q-418 mutant insulin receptors had insulin-binding characteristics similar to the wild-type human insulin receptor, whereas no insulin-binding activity could be detected above the control level in cells transfected with Q-D. Flow cytometry with antibodies against the human insulin receptor indicated the presence of Q-397, Q-418, and wild-type human insulin receptors in the surface of COS cells and failed to demonstrate a Q-D receptor. Insulin-induced autophosphorylation was similar in Q-397, Q-418, and wild-type human insulin receptors as was their ability to phosphorylate an artificial substrate, poly Glu-Tyr (4:1). Our inability to detect Q-D receptors was not caused by a lack of Q-D mRNA. COS cells transfected with Q-D cDNA generated as much Q-D mRNA as the amount of wild-type human insulin receptor mRNA present in cells transfected with wild-type receptor cDNA. Finally, pulse-chase experiments with [35S]Met were able to detect 190,000-M(r) proreceptors and the alpha-subunits for Q-397, Q-418, and wild-type human insulin receptors.(ABSTRACT TRUNCATED AT 250 WORDS)

The beta-adrenergic blocking agent propranolol was shown in previous studies to increase orthotopic bone formation in rats. To understand the cellular mechanisms underlying this observation, propranolol was tested for its effects on osteoblastic cells, which possess adenylate cyclase-coupled beta-adrenergic receptors. The ability of propranolol to modulate parathyroid hormone (PTH) and isoproterenol effects on adenylate cyclase activity and on alkaline phosphatase expression was studied in the osteoblast-like rat osteosarcoma cell line ROS 17/2.8. At concentrations between 0.1 and 10 microM, DL-propranolol specifically inhibited adenylate cyclase stimulation by the beta-adrenergic agonist isoproterenol, but did not alter either basal or PTH-stimulated activity. At these concentrations, propranolol also blunted the inhibition of alkaline phosphatase activity by isoproterenol but not PTH. Propranolol alone had minimal effects on ROS alkaline phosphatase activity at low concentrations (0.1-1 microM), but became inhibitory at high concentrations (10-100 microM). Thus, the direct effects of physiologically relevant propranolol concentrations on osteoblastic cells can be attributed principally to beta-adrenergic blockade. These findings further suggest that propranolol may enhance bone formation by preserving osteoblastic activity in the face of inhibition by beta-adrenergic agonists.