Glycogen Content Of Cells Research Articles

Gluconeogenesis, ketogenesis, and ureogenesis in isolated hepatocytes from diabetic rats: effects of insulin

The intermediary metabolism in hepatocytes isolated from diabetic rats has been studied. The incubation medium is a Krebs-Henseleit buffer containing albumin-bound oleate (1 mmol/l). The high rates of gluconeogenesis, ureogenesis and ketogenesis were consistent with the diabetic state of the rats. Insulin (8 X 10(-7) mol/l) decreased glucose and urea productions from alanine (10 mmol/l) by 30%; beta-hydroxybutyrate production, oleate utilization and malate efflux (calculated) from mitochondria were also decreased. No effect of insulin was found with lactate (10 mmol/l) as gluconeogenic substrate. The glycogen content of cells was constant during the time of incubation (4h). These data suggest that the oxidation-reduction state of mitochondria and the cytoplasmic oxaloacetate concentration could be important factors in the action of insulin.

Glycogen in Methanolobus and Methanococcus

Glycogen was isolated from Methanolobus tindarius and 4 species of Methanococcus. It was identified by the iodine reaction, determination of glucose, maltose and isomaltose in acid hydrolysates and partial acid hydrolysates, respectively, determination of erythritol and glycerol after periodate oxidation and treatment with amyloglucosidase. Glycogen particles in the cells are distributed throughout the cytoplasm. Ml. tindarius can use elemental nitrogen and cysteine as nitrogen source. The glycogen and polyphosphate content of cells of Ml. tindarius increased under nitrogen limitation. During incubation without external energy source, glycogen decreased and methane formation was correlated to the glycogen content of the cells.

Glycogen inMethanolobusandMethanococcus

Abstract Glycogen was isolated from Methanolobus tindarius and 4 species of Methanococcus . It was identified by the iodine reaction, determination of glucose, maltose and isomaltose in acid hydrolysates and partial acid hydrolysates, respectively, determination of erythritol and glycerol after periodate oxidation and treatment with amyloglucosidase. Glycogen particles in the cells are distributed throughout the cytoplasm. Ml. tindarius can use elemental nitrogen and cysteine as nitrogen source. The glycogen and polyphosphate content of cells of Ml. tindarius increased under nitrogen limitation. During incubation without external energy source, glycogen decreased and methane formation was correlated to the glycogen content of the cells.

Somatomedin-C stimulates glycogen synthesis in fetal rat hepatocytes.

The effects of somatomedin-C/insulin-like growth factor I (Sm-C) on glycogen metabolism in cultured hepatocytes from 20-day-old rat fetuses have been examined and compared with the effects of insulin. Sm-C (25-375 ng/ml; 3.25-50 nM) stimulated dose-dependent increases in [14C]glucose incorporation into glycogen (14.4-72.9%; P less than 0.001) and total cell glycogen content (10.6-34.3%; P less than 0.01). Maximal stimulation of glycogen synthesis by Sm-C occurred at 2-4 h of incubation. Insulin (10 nM to 10 microM) also stimulated [14C]glucose incorporation but its potency was only 1/20th that of Sm-C. The time course of stimulation of glucose incorporation by insulin was identical to that of Sm-C, the dose-response curves of the two hormones were parallel, and the maximal effects of insulin were not enhanced by simultaneous exposure of cells to Sm-C. These findings suggest that Sm-C and insulin stimulate glycogenesis in fetal liver through similar or identical mechanisms. Since the potency of Sm-C was 20 times greater than that of insulin, the glycogenic action of insulin in fetal liver may be mediated through binding to a hepatic receptor which also binds Sm-C. In addition to having mitogenic effects on fetal tissues, Sm-C may have direct anabolic effects on fetal carbohydrate metabolism.

452 SOMATOMEDIN-C STIMULATES GLYCOGEN SYNTHESIS IN FETAL RAT HEPATOCYTES

The role of somatomedin-C/IGF-I (Sm-C) in the regulation of fetal metabolism is poorly understood. Since Sm-C has insulin-like effects on tissues of postnatal animals, we have compared the effects of highly purified Sm-C and insulin on glycogen metabolism in cultured hepatocytes from 20 day old fetal rats. Sm-C (50–750 ng/ml, 6.5–100 nM) stimulated dosedependent increases in 14C-glucose incorporation into glycogen (14.4–72.9%, p<0.001) and total cell glycogen content (10.6–34.3%, p<0.01). Maximal stimulation of glycogen synthesis by Sm-C occurred at 2–4 hrs of incubation. Insulin (10 nM–10 uM) also stimulated 14C-glucose incorporation but its potency was only one tenth that of Sm-C. The time course of stimulation of glucose incorporation by insulin was identical to that of Sm-C, the dose-response curves of the two hormones were parallel, and the maximal effects of insulin were not enhanced by simultaneous exposure of cells to Sm-C. These findings suggest that Sm-C and insulin stimulate glycogenesis in fetal liver through similar or identical mechanisms. Since the potency of Sm-C was greater than 10 times that of insulin, the glycogenic action of insulin in fetal liver may be mediated through binding to a hepatic receptor which also binds Sm-C. In addition to its mitogenic effects on fetal tissues, Sm-C may have direct anabolic effects on fetal carbohydrate metabolism. Supported by NIH HD07447 and HD06301.

Insulin-induced receptor regulation in cultured Zajdela rat hepatoma cells and relationship to the stimulation of glycogen synthesis.

Insulin receptors were measured in cultured Zajdela rat hepatoma cells (ZHC cells), a stable cell line which presents differentiated hepatic functions. The number of sites was 50,000/cell at 2 C, and the dissociation constant for high affinity binding was 1.6 x 10(-10) M. Down-regulation of receptors occurred rapidly when cells were treated with insulin; this process was related to ambient insulin concentrations and led to a decrease in the number of insulin receptors from 50,000 to 30,000/cell. Cycloheximide prevented part of this regulation. When down-regulated cells were incubated in standard medium devoid of insulin, the number of receptor sites gradually increased and attained control values within 7 h; cyclohexamide inhibited this process. Insulin markedly enhanced glycogen synthesis in ZHC cells, with an ED50 of 1.0 x 10(-9) M, leading to an increase in the total cell glycogen content. In addition, the predicted righthand shift of the dose-response curve was observed for insulin-treated cells. These findings provide evidence of insulin-induced receptor regulation in cultured ZHC cells which is related to the biological effect of the hormone on glycogen synthesis.

AN ORGAN CULTURE MODEL FOR THE STUDY OF FETAL LUNG MATURATION

We have developed an organ culture model for the study of biochemical and morphological maturation in late gestation fetal rat lung. Lungs from 18 or 19 day fetal rats are chopped into cubes with 0.8 mm sides using a McIlwain tissue chopper. The explants are placed on a millipore filter supported on a stainless steel grid and cultured in F12 medium in 95%O2, 5%CO2 at 37'C. Use of an air and CO2 environment resulted in areas of central necrosis. Morphologic maturation continues in culture with the progressive development of larger and more numerous alveoli, flatter alveolar lining cells and less stromal tissue. There is a progressive decrease in type II cell glycogen content and an increase in the number of lamellar bodies. Vascular elements are less well preserved. Biochemical findings after varying periods in culture are tabulated below. Values are expressed as a percentage of those obtained with fresh lung tissue from 18 or 19 day fetuses. This data indicates that fetal lung remains viable in short term organ culture, a useful model for the study of hormonal influences on lung development. Supported by HL19752.

Studies on the effect of collagenase and hyaluronidase on glycogen content of isolated rat liver parenchymal cells.

SummaryRat liver parenchymal cells were isolated with collagenase and hyaluronidase from fed and fasted rats. Addition of hyaluronidase significantly decreased intracellular glycogen content of cells in fed rats, although there was no effect on cell yields. The incorporation of 14C isoleucine, 14C-valine, and 14C-phenylalanine into proteins was significantly higher in younger as compared to older rats. The incorporation of these various 14C amino acids was lower in cells from fasted rats when compared to those from fed rats.

Studies on the differential response to glucagon concentration on gluconeogenesis in isolated hepatocytes containing high and low glycogen

Rat liver parenchymal cells were isolated with (a) collagenase alone and (b) with both collagenase and hyaluronidase. Addition of hyaluronidase significantly decreased intracellular glycogen content of cells from fed rats. Effects of various concentrations of glucagon on gluconeogenesis were also studied in isolated hepatocytes from fed and fasted rats. Glucagon at the concentration of 10 −12M to 10 −10M stimulated gluconeogenesis in fed rats. Higher concentrations (10 −8M) had no further stimulating effect. In fasted rats, glucagon at the concentrationsof 10 −12M had no effect whereas at 10 −10M to 10 −8M concentrations, it stimulated gluconeogenesis by 2 fold. These studies suggest that glucagon functions in gluconeogenesis both in the fed and fasted state.

Phosphorylase and glycogen synthetase during myoblast differentiation

Summary The variations of the ratios of a to b form of phosphorylase, and I to D form, of glycogen synthetase have been studied during differentiation of myoblasts of the established line L 6 D, under standard medium conditions, and after a shif to a medium of low glucose concentration. The specific activities of phosphorylase and glycogen synthetase increase several folds during the differentiation process, without appreciable changes in the ratio of the phosphorylated to the non phosphorylated forms of these enzymes, phosphorylase being mainly in the a form and glycogen synthetase in the D form. The glycogen content of the cells, which is already appreciable in non differentiated myoblasts, increases 2 to 3 fold during differentiation. A shift of differentiating myoblasts to a medium of low glucose concentration induces a rapid decrease in the cell's glycogen content and a reversal of the ratio of the phosphorylated to the non phosphorylated form of the two enzymes. These results are in agreement with the hypothesis that the cell's glycogen content exerts a control on the phosphorylation of these two enzymes « in vivo .

Interaction between glycogen and glycogen phosphorylase of Tetrahymena

The glycogen phosphorylase of Tetrahymena pyriformis complexes with glycogen as judged by its elution pattern from columns of Sepharose 6B. Complex formation does not occur with starch, amylose, or amylopectin, and neither do these polyglucans serve as primers for the enzyme. To study the association between the phosphorylase and glycogen particles in situ, Tetrahymena were grown under differing physiological conditions, phosphorylase was isolated and chromatographed on a Sepharose 6B column. Phosphorylase activity isolated from cells grown in the absence of glucose was only partially associated with glycogen, while in cells exposed to glucose for 30 min or more all the phosphorylase activity was associated with glycogen. The effects of culture age and anaerobiosis on the relative amounts of free and glycogen-bound enzyme in the cells were also studied. It was concluded from the in vivo experiments that there was no simple relation between the fraction of enzyme bound to glycogen and between cell glycogen content.

Changes in the ultrastructure of Nematospiroides dubius (nematoda) intestinal cells during development from fourth stage to adult.

The general fine structure of intestinal cells and changes which occur in ultrastructure during development from fourth-stage to adult N. dubius are reported. In fourth-stage worms pigment granules are prominent in intestinal cells. In adults the number of pigment granules appears to be reduced and phagolysosomes containing membranous profiles and pigment material increase in number. Another reorganization of cell structure involves mitochondria which are randomly distributed in the cytoplasm of cells in fourth-stage worms, concentrated in the apical cytoplasm in worms in the molting process, and confined to the base of cells in adult worms. Other changes involved structure of the nucleus, rough endoplasmic reticulum and glycogen content of cells.

On the regulation of glycogen metabolism in Tetrahymena

The glycogen phosphorylase and glycogen synthetase activities of Tetrahymena pyriformis were assayed after growth of cells under various conditions. Cells exposed to low concentrations of theophylline or to low oxygen tension during growth in proteose-peptone medium had increased synthetase and phosphorylase activities. Supplementing the medium with glucose increased the synthetase activity. Growth in the presence of both glucose and theophylline markedly increased the synthetase activity and decreased the phosphorylase activity. Neither reserpine nor dichloroisoproterenol affected the synthetase activity, but either or both drugs increased the phosphorylase activity. Triiodothyronine did not affect the phosphorylase activity but inhibited the synthetase activity. It was also found that Tetrahymena had 3′,5′-cyclic AMP phosphodiesterase activity and that caffeine and theophylline inhibited this activity. The effects of glucose supplementation, low oxygen tension, and of the drugs mentioned above were correlated with their effects on cell glycogen content, and the implications of these findings with respect to the control of glycogen metabolism are discussed.

Corticosteroid-like substances detected histochemically in the ovaries of women suffering from the Stein-Leventhal syndrome

1 N T H E L A S T few years great interest has been accorded to the histochemical investigation of the ovaries of normal, mature women and those of women suffering from the Stein-Leventhal syndrome. Interesting histochemical findings were disclosed about the former in regard to cell glycogen content, ascorbic acid, acid and alkaline phosphatase, neutral lipids, cholesterol and acid, and neutral mucopolysaccharides2, 3 These results were interpreted as the result of altered cell activity, of a deviation in the normal pathway of synthesis, or the degradation of some substance, probably of steroid origin. In order to determine the more exact chemical characteristics of these substances further histochemical investigations were undertaken. In 1949 Ashbel and Selingmanlp 5 proposed a new histochemical technique for the detection of all 17-ketosteroids with an active carbonyl ( > C =O) group at the C,, C17, and C,, positions. The reaction is not specific, but positive with all substances having a ketonic grouping as in the adrenal steroids, the male and female sex hormones, and their metabolic products. If the ovaries of women suffering from the