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Biomedical subjects

H Tager

Publications and source records attributed to H Tager.

14 recordsLinked to original sources
PubMed1994

Alterations in immunoreactive proinsulin and insulin clearance induced by weight loss in NIDDM.

Subjects with overt non-insulin-dependent diabetes mellitus (NIDDM) were studied in comparison to obese nondiabetic control subjects and patients with subclinical diabetes. Pancreatic insulin secretion rates were measured by deconvolution of peripheral C-peptide over a 24-h period while subjects consumed an isocaloric mixed diet. Subjects were then placed on caloric restriction for at least 6 weeks, during which time body weight fell by at least 10%. Refeeding with solid mixed meals was then resumed for at least 2 weeks until isocaloric intake was attained, and then the meal profiles were repeated. Before weight loss, insulin, C-peptide, and insulin secretion rates were significantly higher in subjects with subclinical diabetes than in the other two groups. Proinsulin concentrations were significantly greater in the two diabetic groups than in control subjects, but, when expressed as a percentage of the total insulin immunoreactivity, the differences were significant only in the group with overt diabetes. Weight loss because of hypocaloric feeding resulted in a significant increase in the rate of clearance of endogenously secreted insulin but did not affect the clearance of C-peptide. In obese subjects and those with subclinical diabetes, weight loss was associated with a reduction in insulin secretion rates, presumably as a result of improvements in insulin sensitivity. In patients with overt diabetes and hyperglycemia, weight loss improved beta-cell responsiveness to glucose and was associated with an increase in insulin clearance and a reduction in proinsulin immunoreactivity.(ABSTRACT TRUNCATED AT 250 WORDS)

Blood Glucose↗
PubMed1986

The limitations to and valid use of C-peptide as a marker of the secretion of insulin.

The accuracy with which the secretion rate of insulin can be calculated from peripheral concentrations of C-peptide was investigated in conscious mongrel dogs. Biosynthetic human C-peptide and insulin were infused intraportally and their concentrations measured in the femoral artery. During steady-state infusions of C-peptide, the peripheral concentration changed in proportion to the infusion rate and the metabolic clearance rate (5.2 +/- 0.3 ml/kg/min) remained constant over a wide range of plasma concentrations. Application of a two-compartment mathematical model, in which the model parameters were estimated from analysis of C-peptide decay curves after intravenous bolus injections, allowed the intraportal infusion rate of C-peptide to be derived from peripheral C-peptide concentrations, even under non-steady-state conditions. Estimates of the intraportal infusion rate based on this model were 102.4 +/- 2.6% of the actual infusion rate as it was increasing and 102.3 +/- 5.5% of this rate as it was falling. The peripheral C-peptide: insulin molar ratio was influenced by the rate at which equimolar intraportal infusions of C-peptide and insulin were changed. The baseline C-peptide: insulin molar ratio (4.1 +/- 0.9) increased to peak values of 8.2 +/- 0.6, 10.3 +/- 2.0, and 14.9 +/- 1.3 when the infusion rate was increased and then decreased rapidly. Peak values of only 5.7 +/- 1.2 were found if the intraportal infusion rate was changed slowly.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1983

Three mutant insulins in man.

We have previously identified a structurally abnormal insulin in the serum and pancreas of a middle-aged man with diabetes mellitus which arose from a leucine for phenylalanine substitution at position 24 or 25 of the insulin B chain; further analysis of the patient's leukocyte DNA showed that one of the patient's insulin alleles had undergone mutation resulting in loss of an MboII restriction site normally present in the human insulin gene. Two additional and unrelated patients with the same clinical syndrome have now been identified (ref. 4 and unpublished results). All of these patients showed hyperglycaemia typical of diabetes and with marked hyperinsulinaemia typical of insulin resistance, but all three show normal tolerance to exogenously administered insulin. As the opportunity of examining pancreatic tissue from patients suspected of secreting insulin variants is rare, we have developed a method combining HPLC and radioimmunoassay to identify insulin variants isolated from human sera. By this method we have shown that all three patients noted above secrete structurally variant and chemically distinct insulins. In correction of our original assignment, one is identified as [LeuB25]insulin.

Amino Acid Sequence↗
PubMed1983

Identification of a mutant human insulin predicted to contain a serine-for-phenylalanine substitution.

Using information gained from (i) the relative HPLC retention of an abnormal insulin present in the serum of a hyperinsulinemic diabetic patient and (ii) the loss of an Mbo II restriction site in one of the patient's insulin gene alleles, it was recently predicted that the mutant insulin contained a serine-for-phenylalanine substitution at position B24 or B25. We have now prepared human [SerB24]insulin and [SerB25]insulin by solid-phase peptide synthesis and semisynthesis using an improved approach whereby the protecting groups can be removed from the final product in a single step. During reversed-phase HPLC analysis, the two semisynthetic insulins were clearly separated from normal insulin and from each other. Analysis of the patient's immunoaffinity-purified serum insulin by HPLC and radioimmunoassay showed that the insulin eluted at the position of [SerB24]insulin and coeluted with the analog when the two were studied in admixture. Additional studies showed that [SerB24]insulin and [SerB25]insulin have about 16% and 0.5% of the activity of normal insulin, respectively, in stimulating glucose oxidation by isolated rat adipocytes. We conclude that the patient's abnormal insulin (insulin Los Angeles) is human [SerB24]insulin and that abnormal insulins with amino acid replacements at both positions B24 and B25 can be associated with human diabetes.

Amino Acid Sequence↗
PubMed1983

Metabolism of C-peptide in the dog. In vivo demonstration of the absence of hepatic extraction.

The in vivo hepatic metabolism of connecting peptide (C-peptide) in relation to that of insulin has not been adequately characterized. A radioimmunoassay for dog C-peptide was therefore developed and its metabolism studied in conscious mongrel dogs, with sampling catheters chronically implanted in their portal and hepatic veins and femoral artery. The hepatic extraction of endogenous C-peptide under basal conditions was negligible (4.3 +/- 4.5%) and was similar to the hepatic extraction of C-peptide measured during the constant exogenous infusion of C-peptide isolated from dog pancreas. Simultaneously measured hepatic extraction of endogenous and exogenously infused insulin were 43.8 +/- 7.6 and 47.5 +/- 4.4%, respectively. The metabolic clearance rate of infused C-peptide was 11.5 +/- 0.8 ml/kg per min and was constant over the concentration range usually encountered under physiological conditions. In additional experiments, the effect of parenteral glucose administration on the hepatic extraction of C-peptide and insulin was investigated. The hepatic extraction of C-peptide (6.2 +/- 4.0%) was again negligible in comparison with that of insulin (46.7 +/- 3.4%). Parenteral glucose administration did not affect the hepatic extraction of either peptide irrespective of whether it was infused peripherally, intraportally, or together with an intraportal infusion of gastrointestinal inhibitory polypeptide. The fasting C-peptide insulin molar ratio in both the portal vein (1.2 +/- 0.1) and femoral artery (2.1 +/- 0.3) was also unaffected by the glucose stimulus. These results therefore indicate that, since the hepatic extraction of C-peptide is negligible and its clearance kinetics linear, the peripheral C-peptide concentration should accurately reflect the rate of insulin secretion. New approaches to the quantitation of hepatic extraction and secretion of insulin by noninvasive techniques are now feasible.

Animals↗
PubMed1983

Characteristics of binding of a low affinity, noncooperative insulin [( LeuB25]insulin) to IM-9 lymphocytes.

[Leu-B25]insulin is a low affinity insulin analog which does not increase the rate of dissociation of 125I-insulin from insulin receptors (i.e. does not display negative cooperativity). We have studied the characteristics of binding of this analog to IM-9 cultured lymphocytes, in order to determine the contribution of negative cooperativity to the curvilinear nature of Scatchard plots typical of insulin binding data. The affinity of [LeuB25]insulin for receptors was approximately 1% that of insulin, as determined by its ability to inhibit 125I-insulin binding. Monoiodinated preparations of insulin and of [LeuB25]insulin were produced, labeled in the tyrosine at position 14 of the A chain. These 125I-TyrA14-labeled species were used in all studies. Both native insulin and a serum containing antiinsulin receptor antibodies were equally potent at inhibiting binding of 125I-[LeuB25]insulin and 125I-native insulin, suggesting that they bind to the same population of receptors. Native insulin (100 ng/ml) increased the rate of dissociation of both 125I-insulin and 125I-[LeuB25]insulin. However, [LeuB25]insulin (2.5 micrograms/ml) did not increase the rates of dissociation of either 125I-insulin or 125I-[LeuB25]insulin (i.e. it did not display negative cooperativity). Competition curves and Scatchard plots were constructed using 125I-[LeuB25]insulin and unlabeled analog. Half-maximal inhibition of 125I-[LeuB25]insulin binding was seen at a [LeuB25]insulin concentration of approximately 500 ng/ml. More importantly, the Scatchard plot of these binding data was markedly curvilinear, as is typical of insulin binding data. In summary, a non-cooperative insulin analog, [LeuB25]insulin, yielded curvilinear Scatchard plots, indicating that negative cooperativity is not the full explanation of the curvilinear nature of Scatchard plots of insulin binding data. Therefore, an alternative explanation, such as the existence of heterogeneous receptor states, with differing affinities for the hormone, must contribute to the nonlinearity of these plots.

Cell Line↗
PubMed1983

Differential immunoreactivity of plasma glucagon components in man: studies with different glucagon antibodies.

To evaluate the relationship between glucagon antibody antigenic determinants and selective reactivity with plasma void volume (Vo) and lower molecular weight immunoreactive glucagon (IRG) components, we studied plasma IRG levels and molecular profiles in normal subjects and patients with disturbances in plasma glucagon levels using three glucagon antibodies, 30K and P7 raised against the whole peptide and antibody X4 raised against the C-terminal tryptic fragment of glucagon. In normal subjects and pancreatectomized patients, plasma IRG levels were 2- to 3-fold higher with the C-terminus-directed antibody X4 than with either 30K or P7, but in glucagonoma and uremic patients, this discrepancy was smaller. Gel filtration analysis revealed that these antibodies reacted identically with 3500 mol wt IRG and 9000 mol wt IRG in normal, glucagonoma, pancreatectomized, and uremic plasma. Relative immunoreactivity of Vo IRG was approximately 5:2:1 with antibodies X4, 30K, and P7, respectively. In two subjects with unexplained hyperglucagonemia, recovery of IRG was entirely in the Vo, with antiserum X4 reacting one third as well as 30K and P7 not reacting at all. Furthermore, this material did not dilute out in parallel to glucagon standard. These data indicate differential immunoreactivity of the high molecular weight circulating IRG component, with a series of three glucagon antibodies reacting similarly with all other plasma IRG fractions, and suggest that Vo IRG material in plasma is predominantly the result of an immunologically cross-reacting peptide sequence in a plasma protein. The selective immunoreactivity of this component with different antibodies has important implications for the glucagon RIA and may have some bearing on other immunoassays as well.

Antibodies↗
PubMed1980

Identification and localization of glucagon-related peptides in rat brain.

Immunochemical and immunocytochemical techniques have been used to identify and characterize glucagon-related peptides of the rat central nervous system. These peptides show immunoreactivity with antiglucagon sera directed towards the central portion of the hormone, but not with antisera specific for the free COOH terminus of glucagon. Highest concentrations were found in hypothalamus (6.1 +/- 1.6 ng/g wet weight) although lower amounts (approximately 2 ng/g) were found in cortex, thalamus, cerebellum, and brain stem. Gel filtration of brain extracts revealed at least two immunoreactive forms, which have molecular weights of about 12,000 and 8000. Both peptides had radioimmunoassay dilution curves parallel to the curve for glucagon and both had identical counterparts in extracts of rat intestine. Digestion of the brain and intestinal peptides with trypsin plus carboxypeptidase B released the immunoreactive COOH-terminal tryptic fragment of pancreatic glucagon from these larger forms. Immunocytochemical studies using antiglucagon serum and peroxidase-antiperoxidase staining identified glucagon-like material in neuronal cell bodie and processes in the magnocellular portion of the paraventricular nucleus, as well as in scattered cells in the supraoptic nucleus and in fibers in the median eminence. These results suggest that glucagon-containing peptides that have undergone the intestinal type of posttranslational modification are present in neuronal cells of the rat hypothalamus.

Amino Acid Sequence↗
PubMed1980

Semisynthesis and biological activity of porcine [LeuB24]insulin and [LeuB25]insulin.

Two analogs of porcine insulin with substitutions of leucine for phenylalanine in the COOH-terminal region of the insulin B chain have been prepared by a combination of solid-phase synthesis and semisynthesis. Solid-phase synthesis of the substituted octapeptides B23-B30 bearing the trifluoracetyl group on lysine-B29, enzymatic coupling of the octapeptides to bis(tertiary-butyloxycarbonyl)desoctapeptide insulin by trypsin, and deprotection of the corresponding adducts in formic acid and piperidine resulted in two insulin derivatives, one with leucine at position B24 and the other with leucine at position B25. These analogs had only about 10% and 1%, respectively, of the activity of porcine insulin in competing for the binding of [125I]iodoinsulin to both rat adipocytes and human IM-9 lymphocytes. The relative potencies of the analogs in stimulating glucose oxidation by rat adipocytes decreased in the order porcine insulin > [LeuB24]insulin > [LeuB25]insulin. However, at high concentrations both analogs had full agonists activity. Experiments in which the semisynthetic insulins were mixed with the native hormone showed that [LeuB24]insulin, but not [LeuB25]insulin, was an active antagonist of insulin action. These results suggest that the antagonistic activity of a human insulin variant having leucine at position B24 or B25 can be assigned to the molecule with the sequence Gly-Leu-Phe-Tyr (residues B23-B26) in its active site.

Adipose Tissue↗
PubMed1979

A structurally abnormal insulin causing human diabetes.

Insulin isolated from the pancreas of a diabetic patient with fasting hyperinsulinaemia showed decreased activity in binding to cell membrane insulin receptors and in stimulating cellular 2-deoxyglucose transport and glucose oxidation. Chemical studies suggest that the isolated hormone is a mixture of normal insulin and an abnormal variant which contains a leucine for phenylalanine substitution at position 24 or 25 of the insulin B-chain.

Amino Acid Sequence↗
PubMed1977

Factitious hypoglycemia. Diagnosis by measurement of serum C-peptide immunoreactivity and insulin-binding antibodies.

In seven patients with factitious hypoglycemia due to the surreptitious injection of insulin, we made the diagnosis by measurements of plasma insulin and C-peptide immunoreactivity (in seven patients), facilitated by the finding of circulating insulin-binding antibodies (in two patients). The simultaneous demonstration of low plasma glucose, high immunoreactive insulin and suppressed C-peptide immunoreactivity represents a triad of results pathognomonic of exogenous insulin administration. Determination of plasma free C-peptide and free insulin permitted patients with high titers of insulin antibodies, including those with a history of insulin-treated diabetes, to be studied and diagnosed in a way similar to that in subjects who had no circulating insulin antibodies.

Adenoma, Islet Cell↗