Glycogen Synthetase Activity Research Articles

Enzyme histochemical and immunohistochemical characterization of oval and parenchymal cells proliferating in livers of rats fed a choline-deficient/DL-ethionine-supplemented diet

Male outbred Sprague-Dawley rats were fed a choline-deficient diet containing 0.10% DL-ethionine for up to 30 weeks. Liver slices from rats killed 4, 6, 10, 14, 22 and 30 weeks after starting the treatment were histochemically analyzed for the following parameters: basophilia, expression of cytokeratin 19 (which in the liver is bile duct epithelial cell-specific), glycogen content and activities of glycogen synthetase (SYN), glycogen phosphorylase (PHO), glucose-6-phosphatase (G6PASE), glucose-6-phosphate dehydrogenase (G6PDH), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), glycerin-3-phosphate dehydrogenase (G3PDH), 'malic enzyme' (MDH), alkaline phosphatase (ALKPASE) and gamma-glutamyltranspeptidase (GGT). The diet induced necrosis of single parenchymal cells and a massive proliferation of oval cells within 4-6 weeks; thereafter cholangiofibroses, cystic cholangiomas and some cholangiofibromas, but no cholangiocarcinomas, were observed. Oval cells, cholangiofibroses, cystic cholangiomas and cholangiofibromas expressed cytokeratin 19, whereas parenchymal cells, foci of altered hepatocytes and hepatocellular adenomas did not; this observation does not support a precursor-product relationship between oval and parenchymal cells. SYN, PHO, G6PASE, G6PDH, GAPDH, G3PDH, MDH, ALKPASE and GGT activities were detected in oval cells; cholangiofibrotic lesions, cystic cholangiomas and cholangiofibromas stained strongly for GAPDH, G3PDH and MDH. In livers from rats fed the diet for 10 weeks, single hepatocytes storing high amounts of glycogen appeared in the parenchyma. There was no indication of a transition from the oval cell population to hepatocytes storing glycogen in excess. Foci of glycogen-storing cells were scattered all over the lobes after 14 and 22 weeks; they had increased G6PASE, G6PDH, ALKPASE and GGT activities. Mixed cell foci and hepatocellular adenomas developed within 22-30 weeks and exhibited a remarkable decrease of G6PASE activity, a strong increase of G6PDH, GAPDH, G3PDH and MDH activities as well as extremely high ALKPASE and GGT activities. The data support the concept that during hepatocarcinogenesis, a number of sequential changes in the activities of various enzymes involved in carbohydrate metabolism occur and that a correlation between morphology and enzyme pattern in the focal lesions does in fact exist. Furthermore, our results suggest that two different cell lineages are involved in the development of cholangiocellular tumors from oval cells and hepatocellular tumors from hepatocytes.

Insulin-Sparing Effects of Pancreatic Polypeptide in Congenitally Obese Rodents

Chronic treatment with bovine pancreatic polypeptide (bPP) is reported to decrease body weight and reduce fasting glucose and insulin concentrations in congenitally obese mice. The present study examines the effects of acute and chronic bPP treatment on insulin release and glucose clearance in lean and obese rodents. After single injections of 0, 5, 50, or 500 micrograms of bPP/kg of body weight, the insulin response to an intragastric glucose meal (5 g/kg of body weight) was substantially inhibited by the two higher doses of bPP. The change in glucose concentration over time was similar among all animals except those receiving the highest dose of bPP (500 micrograms/kg of body weight); in this group, glucose rose to higher levels and was slower to return to basal levels. Chronic treatment of rats with 200 micrograms of bPP/day/kg of body weight for 5 days did not modify glucose or insulin responses to the glucose meal, but did increase the activity of hepatic glycogen synthetase. In contrast, basal glucose levels were lower in obese mice (ob/ob) treated with bPP and glucose clearance was improved in the treated group after injection of exogenous insulin. Islet hormone concentrations in pancreatic extracts were compared in lean and obese mice treated with and without 200 micrograms of bPP/day/kg of body weight for 5 days. The pancreases of obese mice had higher concentrations of insulin and PP, and treatment with exogenous bPP increased endogenous PP in the pancreases of both phenotypes. Treatment with exogenous bPP also increased the insulin content of obese pancreases, but was without effect in pancreases of lean mice.(ABSTRACT TRUNCATED AT 250 WORDS)

Metabolic effects of bovine growth hormone in the tilapia Oreochromis mossambicus

1. 1. Bovine growth hormone (bGH) was injected into tilapia intramuscularly at a dose of 50 μg/100g/day for a total of five injections. Control fish received saline instead. 2. 2. The serum concentrations of amino acid and glucose were significantly higher and hepatic glycogen concentration and glycogen synthetase activity significantly lower in the bGH-treated fish than those in the control fish. 3. 3. The serum concentrations of protein, lipid and cholesterol, and the hepatic concentrations of protein and lipid, remained unaltered after bGH treatment. 4. 4. The results suggest that bGH exerts anti-insulin effects in tilapia.

Preliminary studies on carbohydrate metabolism changes in domesticated rainbow trout ( Oncorhynchus mykiss) transferred to diluted seawater (12 p.p.t.)

1. 1. Assessment of glycogen and glucose content and glycogen phosphorylase, glycogen synthetase, hexokinase and fructose 1,6-bisphosphatase activity in liver and white muscle of domesticated rainbow trout ( Oncorhynchus mykiss) transferred to diluted seawater (12 p.p.t.). Plasma glucose levels were also determined. 2. 2. A decline in liver glycogen content was observed throughout the 12–120 hr period in diluted seawater (dSW). Liver free glucose also showed a decrease, though only significant at 120 hr in dSW. 3. 3. The activity of liver glycogen phosphorylase a and a + b was higher in dSW animals, but glycogen synthetase was unchanged. 4. 4. No changes in liver hexokinase and fructose 1,6-biphosphatase activity were observed. 5. 5. In white muscle and plasma only white muscle glycogen synthetase activity showed a change, in particular a decline in dSW animals. 6. 6. Although domesticated rainbow trout do not naturally migrate to seawater, these data suggest that a depletion of liver reserves occurred in a way similar to that described for other salmonids.

Endocytosis of insulin receptors is not required for activation or deactivation of the hormone response.

To examine the role of endocytosis in insulin action, hormone responsiveness was studied in transfected Rat 1 fibroblasts stably expressing a noninternalizing insulin receptor. The latter receptor (hIR delta ex16) was engineered by deleting the immediately submembranous 22 amino acids encoded by the 16th exon of the human insulin receptor and has previously been shown not to internalize despite having normal insulin-stimulated tyrosine kinase activity. It is shown in the present study that hIR delta ex16 receptors do mediate insulin action. Insulin dose-response curves both for activation of glycogen synthetase and for mitogenic stimulation demonstrate greater insulin sensitivity in hIR delta ex16 cells compared with untransfected Rat 1 cells. In addition, increases in the absolute levels of glycogen synthetase activity are seen in the hIR delta ex16 cells. Species-specific agonistic antibodies to the insulin receptor also stimulate hIR delta ex16 cells, confirming the activity of the mutant receptor. The non-internalizing receptors are rapidly dephosphorylated after removal of insulin, and the activation of glycogen synthetase decays no more slowly in hIR delta ex16 cells than in cells expressing wild-type receptors. The results demonstrate that receptor endocytosis is not necessary for activation or deactivation of the insulin response.

Hypoglycemic action of the pectin present in the juice of the inflorescence stalk of plantain (Musa sapientum)—Mechanism of action

The pectin isolated from the juice of the inflorescence stalk of plantain (Musa sapientum) has been found to show significant hypoglycemic effect both in normoglycemic and alloxan diabetic rats. After its administration at a dose of 20mg/100g body weight, there was increase in the concentration of hepatic glycogen, increased glycogenesis as evident from the increased activity of glycogen synthetase and in normoglycemic rats increased incorporation of labelled glucose into hepatic glycogen. Glycogenolysis and glyconeogenesis were lower as was evident from the decreased activity of glycogen phosphorylase and gluconeogenic enzymes.

Effects of extracts of high- and low-chromium brewer's yeast on metabolism of glucose by hepatocytes from rats fed on high- or low-Cr diets

Brewer's yeast was grown on a defined medium containing glucose, ammonia, salts and vitamins plus tracer 51Cr without (low-Cr) or with (high-Cr) carrier Cr. The two batches of yeast differed by more than 100-fold in Cr content, containing 80 ng and 10 micrograms Cr/g dry yeast respectively. Extraction and fractionation procedures were designed to isolate Cr complexes with properties similar to those reported for glucose tolerance factor. After weaning, rats were reared on rat cubes (normal diet) or on a diet containing less than 0.1 microgram Cr/kg (low-Cr diet), or on the low-Cr diet supplemented with Cr (1 mg Cr/kg). Hepatocytes from these rats were incubated with [U-14C]glucose and incorporation of 14C into glycogen was measured. Incorporation of glucose-C into glycogen was enhanced by some yeast fractions in the presence of insulin, but had less effect in the absence of insulin. No difference could be detected between the responses to fractions from high- or low-Cr yeast extracts, or between responses by hepatocytes from animals fed on normal or low-Cr diets with or without Cr supplementation. Glycogen synthetase (EC 2.4.1.11) activity (total and percentage in the a form) was similar in hepatocytes isolated from animals on the normal and low-Cr diets. Those yeast fractions which enhanced the response to insulin in the 14C-incorporation experiments also enhanced the percentage of the enzyme in the a form in the presence of insulin, but not in the absence of insulin. The presence in yeast extracts of material which enhances the response to insulin by hepatocytes may help to explain the reported beneficial effects of dietary yeast supplements on glucose tolerance.

Mechanisms of hypoglycemic activity of ganoderan B: a glycan of Ganoderma lucidum fruit bodies.

Ganoderan B increased the plasma insulin level in normal and glucose-loaded mice but elicited no effect on insulin binding to isolated adipocytes. Administration of ganoderan B elicited significant increases of the activities of hepatic glucokinase, phosphofructokinase and glucose-6-phosphate dehydrogenase, decreased the hepatic glucose-6-phosphate and glycogen synthetase activities and did not affect the activities of hexokinase and glycogen phosphorylase. Ganoderan B reduced the glycogen content in the liver but had no influence on total cholesterol and triglyceride levels in the plasma and liver.

Mechanisms of hypoglycemic activity of Aconitan A, a glycan from Aconitum carmichaeli roots

Aconitan A did not affect plasma insulin levels in normal, glucose-loaded and alloxan-induced hyperglycemic mice and gave no influence on insulin binding to isolated adipocytes. Aconitan A exerted no effect on the activities of hepatic hexokinase, glucokinase, glucose-6-phosphatase and glucose-6-phosphate dehydrogenase, whereas it significantly increased hepatic phosphofructokinase activity. Although the activity of hepatic glycogen synthetase showed a tendency to increase, the activity of liver phosphorylase and glycogen content were unchanged by aconitan A. Aconitan A did not change the total cholesterol and triglyceride contents of plasma and liver.

Effect of cyclosporin A on carbohydrate metabolism in the rat.

Liver and kidney carbohydrate metabolism was investigated in rats treated with daily doses of 15 mg/kg body weight cyclosporin A (CyA) for 2 and 8 weeks or of 50 mg/kg body weight CyA for 2 weeks. The higher dosage caused significantly reduced liver glycogen and liver glycogen synthetase activity (of both active I-form and total enzyme activity), whereas the activity of the glycogen-degrading enzyme phosphorylase (active a-form and total activity) remained unchanged. Plasma glucose and glucagon levels, as well as blood ketone bodies of these animals, increased significantly and plasma insulin decreased. In contrast, kidney glycogen and glucose content were higher in rats treated with 50 mg CyA, probably due to enhanced ketone body utilization. Reduced liver glycogen synthetase activity was also found in rats treated with 15 mg CyA. Our data suggest that hypoinsulinemia, induced by CyA, might be a contributing factor to the hyperglycemia, which is mainly due to inhibition of liver glycogen synthesis.

Effect of fasting on glycogen metabolism and activities of glycolytic and gluconeogenic enzymes in the mudskipper Boleophthalmus boddaerti

During starvation, muscle glycogen in Boleophthalmus boddaerti was utilized preferentially over liver glycogen. In the first 10 days of fasting, the ratio of the active‘a’form of glycogen phosphorylase to total phosphorylase present in the liver was small. During this period, the active‘I’form of glycogen synthetase increased in the same tissue. In the muscle, the phosphorylase‘a’activity declined during the first 7 days and increased thereafter while the total glycogen synthetase activity showed a drastic decline during the first 13 days of fasting.The glycogen level in the liver and muscle of mudskippers starved for 21 days increased after refeeding. After 6 and 12 h refeeding, liver glycogen level was 8·5 ± 2·3 and 6·9 ± 4·5 mg·g wet wt 1, respectively, as compared to 5·8 ± l·6mg·g wet wt 1 in unfed fish. Muscle glycogen level after 6 and 12 h refeeding was 0·96±0·76 and 0·82 ± 0·50 mg·g wet wt 1, respectively, as opposed to 0·21 ± 0·12 mg·g wet wt 1 in the 21‐days fasted fish. At the same time, activities of glycogen phosphorylase in the muscle and liver increased while the active‘I’form of glycogen synthetase showed higher activity in the liver. Since glycogen was resynthesized upon refeeding, this eliminated the possibility that glycogen depletion during starvation was due to stress or physical exhaustion after handling by the investigator.Throughout the experimental starvation period, the body weight of the mudskipper decreased, with a maximum of 12% weight loss after 21 days. Liver lipid reserves were utilized at the onset of fasting but were thereafter resynthesized. Muscle proteins were also metabolized as the fish were visibly thinner. However, no apparent change in protein content expressed as per gram wet weight was detected as the tissue hydration state was maintained constant. The increased degradation of liver and muscle reserves was coupled to an increase in the activities of key gluconeogenic enzymes in the liver (G6Pase, FDPase, PEPCK, MDH and PC). The increase in glucose synthesis was possibly necessary to counteract hypoglycemia brought about by starvation in B. boddaerti.

Dietary propylthiouracil and 17β-estradiol benzoate implant effects on liver carbohydrate and lipid metabolism in chicks

1. Forty-eight chicks (21 days old) were implanted (+/- 50 mg 17-beta estradiol; EBI) and fed diets containing +/- 0.1% propylthiouracil (PTU) for 7 days to determine the role of gonadal hormones in the regulation of energy metabolism in the hypothyroid chick. 2. In vitro lipogenesis (IVL) and glucose production (NGP) were measured in liver explants. 3. Liver glycogen (GLY) metabolism was studied in particulate fractions (50,000 x g) of liver. 4. Glycogen synthetase activity (GLYSYN) was assayed in the presence of glucose-6-phosphate (G-6P; 0, 0.15 and 10 mM) to determine activation states. 5. PTU and EBI increased (P less than 0.05) both GLY concentration and GLYSYN activity in chicks but, did not increase G-6P activity ratios or the in vitro activation of GLYSYN. 6. Both PTU and EBI increased (P less than 0.05) IVL and NGP. 7. Estradiol magnifies the effects of PTU in chicks suggesting an interaction between thyroid status and gonadal function.

Mechanisms of hypoglycemic activity of panaxans A and B, glycans of Panax ginseng roots: Effects on the key enzymes of glucose metabolism in the liver of mice

AbstractPanaxan A did not affect the activities of hepatic glucokinase and hexokinase but decreased glucose‐6‐phosphatase activity. Although panaxan B did not influence the activities of these enzymes, it showed a tendency to reduce hepatic glycogen synthetase activity, and panaxans A and B elevated the phosphorylase activity. Panaxan B lowered the glycogen content in the liver. Panaxans A and B significantly increased the activities of glucose‐6‐phosphate dehydrogenase and phosphofructokinase in the liver.

Effect of cyclosporin A on carbohydrate metabolism in the rat

Abstract. Liver and kidney carbohydrate metabolism was investigated in rats treated with daily doses of 15 mg/kg body weight cyclosporin A (CyA) for 2 and 8 weeks or of 50 mg/kg body weight CyA for 2 weeks. The higher dosage caused significantly reduced liver glycogen and liver glycogen synthetase activity (of both active I-form and total enzyme activity), whereas the activity of the glycogen-degrading enzyme phosphorylase (active a-form and total activity) remained unchanged. Plasma glucose and glucagon levels, as well as blood ketone bodies of these animals, increased significantly and plasma insulin decreased. In contrast, kidney glycogen and glucose content were higher in rats treated with 50 mg CyA, probably due to enhanced ketone body utilization. Reduced liver glycogen synthetase activity was also found in rats treated with 15 mg CyA. Our data suggest that hypoinsulinemia, induced by CyA, might be a contributing factor to the hyperglycemia, which is mainly due to inhibition of liver glycogen synthesis.

Abnormalities of carbohydrate metabolism in spontaneously hypertensive rats.

The present study was performed to investigate as to whether peripheral insulin resistance exists in spontaneously hypertensive rats (SHR). After a 12 h fasting period, SHR had significantly higher serum glucose and higher plasma glucagon values in comparison to normotensive control rats (WKY). There was a tendency for higher serum insulin concentrations as well, but this difference did not reach significance. After oral glucose loading or glucose/insulin administration, serum glucose and insulin levels were also higher in SHR compared to WKY rats. Muscle glycogen and glucose concentrations were identical in fasted SHR and WKY rats. With an oral glucose load or glucose/insulin treatment there was a significant increase in muscle glycogen, whereas glucose values declined in skeletal muscle. Both total (a+b-form) phosphorylase activity as well as the active a-form of the enzyme were similar in skeletal muscle of SHR and WKY rats. Glucose/insulin administration or oral glucose loading induced a considerable reduction of both a+b-form and a-form activities. The decrease in muscle phosphorylase activities was almost identical in both groups of animals. There was also a comparable activity of muscle glycogen synthetase activity in all groups of rats. Despite subtile changes of glucose, glucagon and to a lesser degree insulin levels which would be suggestive of insulin resistance, the data obtained from skeletal muscle argue against peripheral insulin resistance in spontaneously hypertensive rats.

Effects of inhibitors of diacylglycerol metabolism on protein kinase C-mediated responses in hepatocytes

In hepatocytes pre-labelled with [3H]glycerol, compound R59022 (6-[2-(4-[(4-fluorophenyl)phenylmethylenel-l-piperidinyl)ethyl-7-methyl-5 H-thiazolo[3,2-alpyrimidin-5-one) and 2-bromooctanoate each increased the amount of radioactivity in diacylglycerols. R59022 mimicked the actions of 12- O-tetradecanoylphorbol 13-acetate in completely abolishing the activation by adrenaline (but not that by vasopressin or glucagon) of glycogen phosphorylase a, and in decreasing the activity of glycogen synthetase. Exogenous dioctanoylglycerol caused a small inhibition of adrenaline-stimulated phosphorylase activity. The concentration of R59022 which gave half-maximal inhibition of adrenaline-stimulated phosphorylase activity was 15 μM. Maximal inhibition was observed within 2 min of addition of R59022. 2-Bromooctanoate activated phosphorylase by a process independent of changes in cyclic AMP and Ca 2+, and decreased glycogen synthetase. It is concluded that in hepatocytes (i) diacylglycerols which accumulate as a result of the inhibition of diacylglycerol kinase by R59022 activate protein kinase C and (ii) 2-bromooctanoate increases diacylglycerols but also has other effects on hepatocyte metabolism.

Glycogen metabolism in human fetal testes

The ontogeny of glycogen synthetase, glycogen Phosphorylase and α-D-glucosidase, enzymes which are associated with glycogen metabolism and glycogen level has been studied in human fetal testes of gestational age ranging from 14–32 weeks. Glycogen synthetase activity reaches the peak value at 17–20 weeks of gestation, thereafter it decreases. α-D-Glucosidase activity increases with the advancement of pregnancy up to 28 weeks of gestation decreasing thereafter very rapidly. Phosphorylase activity remains more or less constant throughout gestation. The maximum increase in glycogen content at early stages of gestation (17–20 weeks) and gradual reduction with the advancement of pregnancy are correlated with histochemical observation by the periodic acid-Schiff technique.

Stimulatory action of progesterone on the synthesis of glycogen in primary cell culture of human endometrium.

AbstractThe effects of progesterone and its antagonist, RU486, on the synthesis of glycogen were investigated using a primary cell culture of human endometrium. Progesterone stimulated the glycogen synthesis in a dose‐dependent manner. The minimal effective dose of progesterone is 10‐9 M with maximal effect occurring at 10‐7 M. Stimulation of glycogen synthesis was apparent as early as after 6 hours of incubation. Progesterone induced the activity of glycogen synthetase and reduced the activity of glycogen phosphorylase. The addition of RU486 with progesterone completely eliminated the action of progesterone in both the synthesis of glycogen and the activity of glycogen synthetase. These results clearly demonstrate the stimulatory action of progesterone in glycogen synthesis in cultured human endometrial cells. Moreover, the antiprogesterone, RU486, is shown to counteract the progesterone action in glycogen synthesis in vitro.

Inhibitory effects of estradiol on glycogen synthesis in primary cell cultures of human endometrium.

The effect of estradiol on glycogen synthesis was examined in a primary cell culture system of the human endometrium. Estradiol inhibited glycogen synthesis in a dose dependent manner with a minimum effective dose of 10(-9)M. Progesterone-induced glycogen synthesis was completely abolished by the simultaneous addition of estradiol. Estradiol did not affect the activity of glycogen phosphorylase in endometrial cells, but the activity of glycogen synthetase was decreased by estradiol irrespective of the presence of progesterone. The inhibitory effect of estradiol on the glycogen synthesis of endometrial cells seemed to be mediated by estradiol receptor, because LY156758, an antiestrogenic drug at receptor site, reversed the inhibitory effect of estradiol.

Glucose metabolism in transdifferentiating and glucose-blocked cultures of chick embryo neuroretinal cells: an inverse relationship between glycogen and δ-crystallin accumulation

Chick embryo neuroretinal (NR) cells transdifferentiate extensively into lens when cultured for several weeks in low-glucose (FH) medium, but fail to do so when high levels of supplementary glucose (FHG) are present. We show here that most aspects of glucose metabolism are promoted in high-glucose cultures, including lactate dehydrogenase (LDH) and glucose-6-phosphate dehydrogenase (G-6-PDH) activities, 2-deoxyglucose uptake, pentose shunt activity and lactate production. Continuous supplementation of high-glucose cultures with low levels of ouabain (FHGO) significantly lowers 2-deoxyglucose uptake, from FHG levels down towards FH levels, especially during the early stages of NR culture. Much later, extensive transdifferentiation into lentoids (with concomitant δ-crystallin accumulation) occurs in these FHGO cultures, which thus resemble FH rather than FHG controls. Another parameter strongly affected by ambient glucose levels is the accumulation of glycogen. Both glycogen itself and glycogen synthetase activity increase steadily in FHG cultures, but decrease slightly under FH conditions. Glycogen accumulation in FHG cultures is largely confined to glial-like cells, particularly those underlying clusters of neurones. Other studies have shown that glial differentiation in vitro is promoted by histotypic interactions with retinal neurones. Thus high glucose may act in concert with neuronal influences to stimulate or stabilize the normal differentiation of retinal glial cells, whose characteristic features in vivo include glycogen synthesis and storage. Furthermore, we show that supplementation of high-glucose cultures with forskolin or dibutyryl cyclic AMP (both of which promote glycogenolysis) results in a slower rate of glycogen accumulation and in enhanced transdifferentiation into lens. In both respects, the forskolin- and dibutyryl cAMP-supplemented FHG cultures are intermediate between FH and FHG controls. Thus the enhancement of normal glial differentiation in NR cultures by high glucose may inhibit or preclude subsequent transdifferentiation into lens.