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Biomedical subjects

A H Rubenstein

Publications and source records attributed to A H Rubenstein.

At least 109 records · Page 6Linked to original sources
PubMed1981

Expression of major histocompatibility antigens on pancreatic islet cells.

Insulin-dependent diabetes mellitus is often accompanied by manifestations of autoimmunity and is frequently associated with certain HLA haplotypes, predominantly DR3 and DR4. Because the major histocompatibility antigens are important determinants of the immune response in various tissues, we have investigated their expression on the pancreatic islet cells. Human, mouse, or rat islets of Langerhans, as well as lymphocytes or other differentiated cells, were biosynthetically labeled with radioactive amino acids, lysed in detergent, and immunoprecipitated with several antisera specific for major histocompatibility antigenic groups. The immunoprecipitates were analyzed by NaDodSo4/polyacrylamide gel electrophoresis under reducing conditions followed by autoradiography. The major histocompatibility antigens corresponding to the H-2 K,D molecules in mice, the H1-A in rats, and the HLA-A, -B, and -C in humans were precipitated from both islet and lymphocyte lysates and were accompanied by beta 2-microglobulin. Binding of H-2 antibodies to islet cells was also confirmed by a radioligand assay using 125I-labeled protein A and by indirect immunofluorescence. Analyses in the fluorescence-activated cell sorter revealed that greater than 95% of the cells in the beta-cell-rich fraction were fluorescent, providing further evidence that the pancreatic beta cells express the major histocompatibility antigens. Monoclonal antibodies or mouse alloantisera against HLA-DR or Ia antigens did not react with labeled pancreatic islet cell proteins.

Animals↗
PubMed1981

The protein-sparing action of protein feeding: absence of relationship to insulin secretion.

The influence of a 2-day fast was compared with that of a modified fast (1.5 g beef protein hydrolysate per kg body wt) on nitrogen metabolism and insulin secretion in three normal young men. During the fast urinary urea excretion indicated a negative nitrogen balance of 10 to 12 g daily in each subjects. With protein feeding nitrogen equilibrium was approached in each instance. Insulin secretion as indicated by the urinary excretion of C-peptide dropped profoundly with fasting to levels which averaged just 13 to 22% of the control base-lines and remained at about these levels during the protein feeding. The fact that this sensitive indicator of insulin secretion disclosed no significant difference during fasting and protein feeding provides strong evidence that the protein-sparing influence of protein feeding cannot be ascribed to the decline in insulin secretion.

C-Peptide↗
PubMed1981

Hepatic metabolism of glucagon in the dog: contribution of the liver to overall metabolic disposal of glucagon.

The hepatic extraction (HE) of glucagon (G) and insulin (I) was measured in 27 dogs, using peripheral infusion of the hormones following elimination of endogenous secretion by pancreatectomy (Px) or somatostatin (S) infusion. HE(G) was 22.5 +/- 1.7%, and HE(I) was 45.1 +/- 3%. HE(G) in seven Px dogs was 27.9 +/- 4.2%, not significantly different from the value of 20.6 +/- 1.6% in 20 S-infused dogs, with corresponding values for HE(I) being 44.9 +/- 6 and 46.0 +/- 3.6%, respectively, suggesting that S does not affect HE of either hormone. HE of endogenous G (22.1 +/- 2.8%) was similar to that of exogenously infused G (19.1 +/- 1.9). HE(G) was nonsaturable in the physiologic and pathophysiologic range of plasma G levels, but there was evidence of saturability in the pharmacologic range. Comparison of simultaneously measured parameters of I and G metabolism indicated independence of the metabolic processes of these two islet hormones, despite distinct similarities in their overall patterns of metabolic disposal. Metabolic clearance rates (MCR) for G and I were 12.6 +/- 0.8 and 19.5 +/- 1.0 ml . kg-1 . min-1, while simultaneously measured hepatic HE rates were 4.2 +/- 0.3 and 8.1 +/- 0.6 ml . kg-1 . min-1, respectively. MCR(G) was independent of arterial G levels. Half-life of infused G and I was 5.5 +/- 0.5 and 4.1 +/- 0.3 min, respectively. The liver accounted for 34.7 +/- 2.4% of the MCR(G) and 42.0 +/- 2.9% of MCR(I). The liver is thus an important site for G removal. However, HE(G) varies widely in different animals, and it is therefore not possible to predict portal vein G concentrations or G secretion rates from G levels in peripheral vessels.

Animals↗
PubMed1981

Insulin binding, internalization, and receptor regulation in cultured human fibroblasts.

Insulin binding to receptors was studied using monolayers of cultured normal human fibroblasts. Binding was rapid and inversely related to the incubation temperature; prolonged periods of steady-state binding were achieved at all temperatures studied and the amount of degradation of extracellular insulin was minimal. Competition curves demonstrated half-maximal inhibition of 125I-insulin binding at an unlabeled insulin concentration of 125I-insulin binding at an unlabeled insulin concentration of 7 ng/ml. Scatchard plots of the binding data were curvilinear and revealed that fibroblasts contained about 7,000 receptor sites per cell. Bound 125I-insulin dissociated from fibroblasts with a t 1/2 of 10 min at 30 degrees C and 35 min at 16 degrees C. After 60 min dissociation at 30 degrees C, 45% of the dissociated radioactivity consisted of 125I-insulin degradation products, whereas only 8% of the dissociated material was in the form of degraded products after 60 min of dissociation at 16 degrees C. This indicates that fibroblasts possess a temperature-sensitive receptor-mediated process for insulin degradation. Preincubation of the monolayers with insulin led to a hormone-induced loss of insulin receptors. Thus, incubating cells with 25 ng/ml insulin for 6 h at 37 degrees C caused a 50% reduction in subsequently measured 125I-insulin binding. This hormone-induced receptor loss was sensitive to physiologic insulin levels, with approximately 5 ng/ml causing a half-maximal receptor loss. When monolayers were treated with the lysosomotropic agent chloroquine and subsequently incubated with 5 X 10(-11) M 125I-insulin, a 130% increase in cell-associated radioactivity was observed after 120 min at 30 degrees C. In summary, 1) cultured normal human fibroblasts possess insulin receptors that exhibit kinetic properties and specificity identical to that of other insulin target cells; 2) incubation of fibroblasts with physiologic concentrations of insulin causes a marked loss of cell-surface insulin receptors; and 3) receptor-bound 125I-insulin is internalized through an energy-dependent endocytotic pathway and subsequently degraded by a chloroquine-sensitive reaction.

Cells, Cultured↗
PubMed1981

Block in insulin release from column-perifused pancreatic beta-cells induced by islet cell surface antibodies and complement.

Dispersed rat pancreatic islet cells were mixed into a short column of Bio-Gel P-2 polyacrylamide beads and perifused with an antiserum containing islet cell surface antibodies. The release of radioactive chromium from prelabeled cells, as a measure of cell membrane permeability, was not affected by cell surface antibodies alone, but increased dramatically in the presence of complement. While there was an eightfold increase in glucose-stimulated insulin release from beta-cells exposed to control serum and complement, insulin release was completely blocked from beta-cells exposed to islet-cell-specific antibodies and complement. These findings suggest that islet cell surface antibodies can mediate complement-dependent cytotoxicity.

Animals↗
PubMed1981

Hepatic extraction of plasma immunoreactive glucagon components. Predilection for 3500-dalton glucagon metabolism by the liver.

This study examines quantitatively the extraction of plasma immunoreactive glucagon (IRG) components by the liver. It was shown that the liver has a predilection for removal of the 3500-dalton biologically active IRG component with virtually no extraction of the other IRG fractions. Hepatic extraction of whole plasma IRG was 25.2 +/- 2.5%. Analysis of the hepatic extraction of the four plasma immunoreactive components, separated by gel filtration, revealed variable but quantitatively insignificant extraction of all components other than the 3500-dalton fraction, which was 33.4 +/- 3.2% (P less than 0.001). Hepatic extraction of whole plasma IRG was significantly less than that of the 3500-dalton component during periods of basal glucagon secretion when IRG fractions other than the 3500-dalton fraction contribute substantially to the whole plasma IRG level. However, during periods of stimulation of glucagon secretion by arginine or arginine plus cholecystokinin-pancreozymin, when the 3500-dalton component accounts for virtually all of the whole plasma IRG level, hepatic extraction of whole plasma IRG was similar to that of the 3500-dalton fraction.

Animals↗
PubMed1980

Formation of biologically active peptides.

Many small biologicaly active peptides are derived from larger precursor forms which fulfil a variety of roles in the synthesis, segregation and intracellular migration of secretory products. Limited proteolysis may occur at several stages during this process, giving rise to products that are either degraded (e.g. the prepeptides) or discharged coordinately from their cells of origin during exocytosis (e.g. insulin and C-peptide). Molecular defects have recently been found to occur at cleavage sites in proinsulin as well as in other proproteins, and these point mutations may, in some instances, be responsible for familial metabolic disorders. The nature and cell specificity of the proteolytic enzymes involved in the conversion of the various precursor forms remains unresolved. Recent studies in our laboratory have led to the identification of precursors of glucagon and somatostatin in rat islets of Langerhans. Analysis of tryptic maps of these precursors has shown that a trypsin-like enzyme would be sufficient to cleave the C-terminally located somatostatin sequence from its precursor (relative molecular mass 12,500), but that both trypsin-like and carboxypeptidase B-like enzymes would be necessary to cleave the internal glucagon sequence from its prohormone (relative molecular mass 18,000). Molecular cloning techniques have provided valuable new approaches to analysing the structures of a variety of precursor forms, including those for insulin, gastrin, growth hormone, adrenocorticotropic hormone and the endorphins, and in the future will undoubtedly shed more light on the structures of their chromosomal genes, the mechanisms regulating their expression, and their evolutionary origins.

Amino Acid Sequence↗
PubMed1980

Diabetes due to secretion of an abnormal insulin.

A 51-year-old, nonobese man with diabetes mellitus had marked hyperinsulinemia (70 to 120 muU per milliliter; 502 to 860 pmol per liter) and fasting hyperglycemia (140 to 170 mg per 100 ml; 7.8 to 9.4 mmol per liter). Plasma proinsulin, glucagon, growth hormone, and cortisol levels were normal; insulin antibodies and insulin-receptor antibodies were not detected. The patient showed relatively normal insulin sensitivity, and insulin receptors on circulating monocytes were within the normal range. Insulin from the patient's serum bound to IM-9 lymphocytes and rat adipocytes approximately 40 per cent as well as insulin standards. Its biologic activity on rat adipocytes averaged 15 per cent of that expected from its immunologic concentration. The impaired biologic activity of this patient's circulating insulin was probably due to a structural abnormality. Subsequent studies of the patient's insulin (fortuitously obtained from his pancreas during a laparotomy for a pancreatic cyst) have confirmed this conclusion. (N Engl J Med 302:129-135, 1980).

Blood Glucose↗
PubMed1980

Prospective evaluation of some candidate tumor markers in the diagnosis of pancreatic cancer.

As part of a prospective diagnostic protocol, patients suspected of having pancreatic cancer had systemic and portal venous blood samples assayed, in coded batches, for peptide hormones and enzymes thought to be of potential value as tumor markers. An average of 111 patients were tested for each candidate marker. Results were analyzed by dividing patients into three groups according to the definitive diagnoses. These were pancreatic cancer (32% of patients), other cancers (27%), and benign diseases (41%). Although elevated mean levels of fasting plasma glucose and serum alkaline phosphatase were found in the pancreatic cancer group, there were no significant differences in the mean levels of any of the candidate markers studied in the three groups. The diagnostic values of normal and elevated levels of each candidate marker studied have been calculated. None has proven to be as useful as the serum level of pancreatic oncofetal antigen, fasting plasma glucose, or serum alkaline phosphatase in the diagnosis or exclusion of pancreatic cancer.

Alkaline Phosphatase↗
PubMed1980

Diagnosis of pancreatic islet hyperplasia causing hypoglycemia in a patient with portacaval anastomosis.

A patient with biopsy-proved biliary cirrhosis and previous gastrojejunostomy and portacaval anastomosis experienced episodes of severe hypoglycemia. She was found to have hyperinsulinemia and hyperglucagonemia. An oral glucose tolerance test showed postgastrectomy hypoglycemia. Results of the intravenous tolbutamide test were diagnostic for insulinoma, but results of the intravenous glucagon test and prolonged fast (96 hours) were not. Failure, on two occasions, to suppress C-peptide normally during insulin-induced hypoglycemia led to a diagnosis of pancreatogenous hyperinsulinemia. The pancreas showed a 10-fold increase in islet volume, with intensely positive staining with anti-insulin and anti-glucagon antiserums in addition to anti-somatostatin and anti-pancreatic polypeptide antiserums. Incidental findings at pancreatic exploration were a mesothelioma, which did not stain with anti-insulin antiserum, and, at autopsy one year later, a hepatoma.

Blood Chemical Analysis↗
PubMed1980

Uridine triphosphate and RNA synthesis during diabetes-induced renal growth.

UTP, CTP, and RNA synthesis were studied in the renal cortex of diabetic and control rats in vivo. The incorporation of UTP into RNA (nmol/h DNA) was used as estimate of RNA synthesis rate. Two to three days after streptozotocin injection, UTP and CTP ppol size and orotate incorporation into UTP and RNA were greater in diabetic animals than in controls. In addition, RNA content and RNA synthesis rate were increased. These changes were corrected by insulin infusion. In diabetic animals, additional increases in UTP pool, RNA content, and RNA synthesis rate followed contralateral nephrectomy. This increase in RNA content was greater than in uninephrectomized controls. The changes in the diabetic renal cortex were not accompanied by increased plasma concentrations of growth hormone. The increase in RNA content in the diabetic renal cortex is probably due to increased RNA synthesis. Increased synthesis of pyrimidines and expansion of the UTP pool may make this substrate more readily available for the synthesis of UDP sugars and may facilitate the synthesis of basement membrane in diabetes.

Animals↗
PubMed1980

Effect of bovine pancreatic polypeptide on isolated rat liver cells.

Pancreatic polypeptide (PP) is a newly isolated protein found in the islets of Langerhans of many species. No clear metabolic function of PP has yet been determined. We have investigated the possible metabolic effect of bovine PP (bPP) on rat liver cells. bPP had no effect on glucose or urea production when liver cells were incubated with a variety of substrates, epinephrine, or glucagon. In addition, there was no appreciable effect of bPP on ketone body production or cAMP levels. Though species differences between the polypeptide and the cells may account for our results, it would seem that further efforts in the search for metabolic actions of PP should be directed toward possible effects on the periphery, i.e. fat and muscle tissue.

Amino Acids↗
PubMed1980

Characterization and measurement of human apolipoprotein A-II by radioimmunoassay.

The development of a radioimmunoassay for apolipoprotein A-II (apo A-II) is described. Initial studies revealed a lack of immunological identity between purified apo A-II used as the standard and serum or HDL. Extensive testing of different buffers, standards, antisera, tracers, utilization of a detergent, and heating of sera failed to resolve the problem. Gel filtration of iodinated and non-iodinated apo A-II on Sephadex G-100 columns showed that apo A-II, in dilute solution, elutes in a higher molecular zone than expected with a broad, assymetrical profile. The use of a subfraction of the tracer in the assay resulted in parallelism in the serum and standard dilution curves. The apo A-II assay was sensitive, specific, and reproducible. Apo A-II added to sera was fully recovered and delipidation did not affect the immunoreactivity of either serum or HDL. Apo A-II contributed approximately 20% to the protein mass of HDL. Comparison of these results with those obtained by radial immunodiffusion, and with previously reported data, indicates that the reactivity of apo A-II in its native and delipidated forms may be markedly influenced by different immunologic methodologies and their specific reagents. Caution should thus be shown at present in assigning absolute concentrations to apo A-II in serum or HDL.

Adult↗