Insulin and Amino-acid Regulation of Polysomes in Perfused,Diabetic Rat Liver

IN normal mammalian liver, protein synthesis and concomitant polyribosome assembly are controlled by amino-acids^1–4. A similar control may be exerted by insulin in liver⁵ and muscle⁶. Both the effect of amino-acids on liver and that of insulin on muscle can be demonstrated with isolated, perfused tissue^4,7, but the interrelationship between these agents has not been clarified either in liver or in muscle. We have used perfused livers from diabetic rats to establish whether amino-acids and insulin promote polysomal assembly independently or in concert.     

Amino acid--induced insulin release from the perfused rat pancreas. The influence of phenylalanine and tyrosine

The effects of L or D phenylalanine and L tyrosine on insulin release from the perfused rat pancreas were investigated. It was found that in the presence of D-glucose, all three amino-acids stimulate insulin secretion. After L-Phe had been removed from perfusate in the presence or absence of L-Tyr, the secondary rise of insulin release (an "off response") was noticed. This phenomenon did not follow to either D-Phe or L-Tyr.     

Transport processes through the blood-brain barrier

The existence of the blood-brain barrier is due to tight junctions between endothelial cells preventing the passage of liquid and solute material at the capillary level. Substances can thus pass across the blood-brain barrier if they are lipophilic or if they have transport systems in the membranes of endothelial cells. The luminal membrane brings metabolites needed for the brain function, the abluminal one plays an important part in removing substances from brain, this can happen against a concentration gradient and thus needs energy. Ions are transported differently by the 2 membranes. Sodium and chloride have carriers and potassium is transported very actively by the sodium-potassium ATPase of the abluminal membrane. Blood-brain glucose influx is very important and happens by carrier transport at the 2 membranes. Efflux seems to use the same transport system as the influx. Transport of ketone bodies seems to happen only from blood to brain, the carriers being reversibly used for brain-blood transport of pyruvic and lactic acid. Amino-acid transport is very different on the luminal and abluminal membranes. On the luminal membrane there are 2 transport systems, one for basic amino acids, the other one, the L system, for neutral amino-acids. All neutral amino-acids are transported through the abluminal membrane by the L, A and ASC systems. There exists a system of transport for basic amino-acids, and a very active one for acid amino-acids. Some systems for the transport of hormones, vitamins and for some peptides exist also at the blood-brain barrier which thus plays a very important role in the regulation of brain metabolism.     

Oligopeptides and plant morphogenesis: A working hypothesis

The two plant hormones—the cytokinins and the auxins—which can induce or enhance morphogenetic events as bud-initiation and root-initiation, contain strong structural relations to amino-acids. Since oligopeptides gathered more and more importance as molecular signals in animal physiology within the last few years, oligopeptides may have comparable functions in plant physiology and especially in plant morphogenesis. The working hypothesis describes bud formation as regulated by an oligopeptidic bud activator with auxins as bud inhibitor, and root formation as regulated by an oligopeptidic root activator and cytokinins as root inhibitor. Some possible proofs of this hypothesis are discussed.     

Effect of hydrocortisone and insulin on (14C)-glycine and (3H)-methionine incorporation into spleen proteins in alloxan-diabetic rats

The intensity of labelled amino acid incorporation into spleen total proteins of albino rat with experimental diabetes was studied as influenced by hydrocortisone, insulin and both hormones simultaneously. Hydrocortisone inhibits and insulin stimulates the [14C] glycine and [3H] methionine incorporation into total proteins of alloxan-diabetic rat spleen. Under simultaneous administration of both hormones the inhibitory hydrocortisone effect is allayed by insulin. It is suggested that an increased level of glucocorticoid hormones in blood is one of the reasons of protein metabolism disturbance under diabetes mellitus.     

Effects on induction of tyrosine aminotransferase in fetal mouse liver in vitro of prednisolone,insulin and thyroxine

The hormonal requirements for formation of tyrosine aminotransferase (EC 2.6.1.5) in fetal mouse liver were investigated in organ culture using chemically defined medium. The hormones tested were insulin, thyroxine and prednisolone. Prednisolone alone resulted in a two-fold increase in tyrosine amino-transferase activity in explanted liver in hormone-free medium on day 6, and its effect was dose dependent, but neither insulin nor thyroxine alone induced the enzyme. Addition of prednisolone plus thyroxine and prednisolone plus insulin increased the enzyme activity 1.4- and 1.3-fold, respectively, over that of explants with prednisolone alone. These three hormones together had the greatest effect, causing induction of 1.5-fold more activity than that with prednisolone plus insulin or plus thyroxine. The three hormones were not all needed continuously during the culture period: prednisolone and insulin were required during the early part of cultivation and thyroxine during the later part. The effects of these hormones were blocked by actinomycin D or puromycin, suggesting that these hormones increase de novo synthesis of tyrosine aminotransferase. Phase-contrast microscopy showed that prednisolone stimulated liver epithelial cell outgrowth, probably acting with insulin.     

Effect of ovarian suppression with gonadotropin-releasing hormone agonist on glucose disposal and insulin secretion

Several lines of evidence suggest that ovarian hormones influence glucose homeostasis, although their exact role in humans has not been clearly defined. In the present study, we sought to test the hypothesis that ovarian hormones regulate glucose homeostasis by examining the effect of pharmacologically induced ovarian hormone deficiency on glucose disposal and insulin secretion. Young, healthy women with regular menstrual patterns were studied during the follicular and luteal phases of their cycle at baseline and after 2 mo of treatment with gonadotropin-releasing hormone agonist (GnRHa; n = 7) or placebo (n = 6). Using hyperglycemic clamps, in combination with stable isotope-labeled (i.e., (13)C and (2)H) glucose tracers, we measured glucose disposal and insulin secretion. Additionally, we assessed body composition and regional fat distribution using radiologic imaging techniques as well as glucoregulatory hormones. Ovarian hormone suppression with GnRHa did not alter body composition, abdominal fat distribution, or thigh tissue composition. There was no effect of ovarian suppression on total, oxidative, or nonoxidative glucose disposal expressed relative to plasma insulin level. Similarly, no effect of ovarian hormone deficiency was observed on first- or second-phase insulin secretion or insulin clearance. Finally, ovarian hormone deficiency was associated with an increase in circulating adiponectin levels but no change in leptin concentration. Our findings suggest that a brief period of ovarian hormone deficiency in young, healthy, eugonadal women does not alter glucose disposal index or insulin secretion, supporting the conclusion that ovarian hormones play a minimal role in regulating glucose homeostasis. Our data do, however, support a role for ovarian hormones in the regulation of plasma adiponectin levels.     

Protein anabolic effects of insulin and IGF-I in the ovine fetus

We determined the effect of insulin and/or recombinant human (rh)IGF-I infusion on ovine fetal phenylalanine kinetics, protein synthesis, and phenylalanine accretion. The chronically catheterized fetal lamb model was used at 130 days gestation. All studies were performed while fetal glucose and amino acid concentrations were held constant. Experimental infusates were 1). saline, 2). rhIGF-I plus a replacement dose of insulin (40 nmol), 3). insulin (890 mIU/h), and 4). IGF-I plus insulin (40 nmol IGF-I/h and 890 mIU insulin/h). Both hormones increased glucose and amino acid utilization, with insulin having a greater effect. The major effect on phenylalanine kinetics was a pronounced fall in phenylalanine hydroxylation, again with insulin having the greatest effect. Whole body protein breakdown was not significantly altered by either hormone; whole body protein synthesis was significantly increased during the combined infusion. Protein accretion was increased by both hormones, with the greatest increase during combined infusion. The fractional synthetic rate (FSR) of circulating albumin was increased by IGF-I but not by insulin. Both hormones significantly increased skeletal muscle FSR without a synergistic effect. The anabolic effects of insulin and IGF-I were more pronounced in these studies than in previous studies where amino acid concentrations were not maintained. The present data also suggest that insulin and IGF-I promote fetal growth through distinct, organ-specific mechanisms.     

Effect of hyperthyroidism and hypothyroidism on rat red blood cell insulin receptors and catecholamines: relationship with cellular ageing

Insulin receptor activity and its relationship with catecholamines in rat young, middle aged and old red blood cells were investigated in experimental hypothyroidism and hyperthyroidism. In control animals, a loss of insulin receptor activity was found with cellular ageing and increased levels of norepinephrine, epinephrine and glycosylated hemoglobin. There was down regulation of insulin receptors together with alterations in membrane bound catecholamines in thyroid hormones imbalances. These results suggest that loss of insulin receptor in cellular ageing is probably part of a more generalised alteration and rat serves as an excellent model in defining the role of thyroid hormones in carbohydrate tolerance.     

Precocious differentiation of chick embryo pancreas in vitro: Roles of prednisolone,insulin and L-thyroxine

The hormonal requirements for functional differentiation of chick embryo pancreas were investigated by using organ cultures in chemically defined medium. The hormones tested were prednisolone, insulin and thyroxine, and the parameters examined were α-amylase (EC 3.2.1.1) and chymotrypsinogen (EC 3.4.4.5) activities, and the ultrastructure of the tissues. Addition of prednisolone alone to explants from 14-day-old chicken embryo pancreas for 3 days increased the activities of amylase and chymotrypsinogen in the tissues by 3.4- and 6.6-fold, respectively, those of tissues before cultivation. Neither thyroxine or insulin alone, nor both hormones together affected pancreatic exocrine differentiation. Thyroxine enhanced the effect of prednisolone on both enzymes, but insulin did not. When the explants were cultured in the medium containing all three hormones, maximum enzyme activities were observed; amylase or chymotrypsinogen activity being 7- or 18-fold, respectively, that of tissues before cultivation. But these three hormones were not simultaneously necessary. Morphological differentiation was also observed in explants cultivated in medium containing these three hormones. These results suggest that glucocorticoids are essential for normal differentiation of chick pancreas during the late fetal period, possibly with insulin and thyroxine, and also support the idea that pancreatic enzymes are controlled separately.     

Cell surface changes associated with EGF and insulin-induced cell adhesion

The ability of insulin and epidermal growth factor (EGF) to restore cell surface function in cells damaged by serum deprivation has been examined. Both insulin (10^?7 M) and EGF (10^?8 M), when added for 2 h, resulted in reattachment of cells to the culture dish; with insulin, attachment was associated with increased amino acid uptake. Both hormones caused an increase in sialic acid and in free sulfhydryl groups associated with the cell membrane. The mobility of concanavalin A (Con A) on the upper cell surface was increased by EGF, and to a lesser extent by insulin. The effects of insulin and EGF on adhesion appear to be regulated by specific receptors for these hormones.     

Precocious differentiation of chick embryo pancreas in vitro. Roles of prednisolone, insulin and L-thyroxine.

The hormonal requirement for functional differentiation of chick embryo pancreas were investigated by using organ cultures in chemically defined medium. The hormones tested were prednisolone, insulin and thyroxine, and the parameters examined were alpha-amylase (EC 3.2.1.1) and chymotrypsinogen (EC 3.4.4.5) activities, and the ultrastructure of the tissues. Addition of prednisolone alone to explants from 14-day-old chicken embryo pancreas for 3 days increased the activities of amylase and chymotrypsinogen in the tissues by 3.4- and 6.6-fold, respectively, those of tissues before cultivation. Neither thyroxine or insulin alone, nor both hormones together affected pancreatic exocrine differentiation. Thyroxine enhanced the effect of prednisolone on both enzymes, but insulin did not. When the explants were cultured in the medium containing all three hormones, maximum enzyme activities were observed; amylase or chymotrypsinogen activity being 7- or 18-fold, respectively, that of tissues before cultivation. But these three hormones were not simultaneously necessary. Morphological differentiation was also observed in explants cultuvated in medium containing these three hormones. These results suggest that glucocorticoids are essential for normal differentiation of chick pancreas during the late fetal period, possibly with insulin and thyroxine, and also support the idea that pancreatic enzymes are controlled separately.     

A test for hormonal responsiveness in a mammary epithelial cell line, NMuMg

The NMuMG cell line derived from normal mouse mammary epithelial cells was tested for responsiveness to hormones. The hormones studied included insulin, glucocorticoids (cortisol and dexamethasone), and prolactin. In addition to membrane bound insulin receptors and prolactin receptors, the cells had 2 X 10(4) cytoplasmic glucocorticoid receptors per cell. Morphological changes were observed in response to hormones. Clusters of cells appeared with greatly increased diameter, and the number of cells per plate was reduced. The rate of DNA synthesis, corrected by cell number, indicates that cell division, and hence cell turnover, was increased by the combination of all three hormones. Insulin greatly enhanced protein synthesis, but glucocorticoid and prolactin did not further increase the rate. The combination of three hormones produced a change in the synthesis of histones, consistent with the increase in cell turnover. There were substantial responses of enzyme activities to hormonal treatment of the cells. Insulin by itself induced a doubling of the activity of glyceraldehyde phosphate dehydrogenase and perhaps a modest increase in NADH-cytochrome c reductase. Lactose synthetase activity showed a three- to fourfold induction of both A and B subunits of the enzyme when the cells were treated with insulin, glucocorticoid, and prolactin, and the effect of the latter two hormones was shown to be additional to that of insulin.     

Hormonal regulation of some steps of thyroglobulin synthesis and secretion in bicameral cell culture

Porcine thyroid cells were cultured for 15 days on porous bottom chambers with or without different mixtures of hormones added to serum-free basal medium. Assays with 10% serum were also performed for comparison with previously published results. The effects of the hormones, particularly insulin, TSH and hydrocortisone, were studied on total RNA content, thyroglobulin mRNA level, the amount of thyroglobulin secreted into the apical medium and on glycosylation. Insulin and TSH similarly increased the total RNA content, and their effects were additive. Thyroglobulin mRNA content was increased twofold by insulin and threefold by TSH. When they were added simultaneously, the maximal level of thyroglobulin mRNA was reached, showing that TSH and insulin effects on thyroglobulin gene expression were additive. Hydrocortisone alone did not modify total RNA or thyroglobulin mRNA content but the hormone amplified total RNA when insulin and TSH were present together. The basal level of thyroglobulin secreted into the apical medium was increased threefold by insulin and fourfold by TSH. The effects of these two hormones added together appeared to be additive. Hydrocortisone had no effect alone or even when combined with insulin or TSH. However, when the three hormones were added together, the hormonal response was amplified. TSH effect and insulin effect on the incorporation of ³H-mannose into thyroglobulin as well as on the anionic residue content of the molecule were additive. © 1994 Wiley-Liss, Inc.     

Use of a monoclonal antibody to distinguish between precursor and mature forms of human lysosomal alpha-glucosidase

The levels of functional mRNA encoding glucose-6-phosphate dehydrogenase (G6PDH; EC 1.1.1.49) were examined in hepatocytes from fasted and fasted/carbohydrate-refed rats and in hepatocytes inoculated into primary culture. Functional G6PDH mRNA was assessed in a cell-free protein synthesis system in vitro. We observed that hepatocytes from fasted/carbohydrate-refed rats had a 12-fold higher level of mRNA than did hepatocytes from fasted rats. The possibility that the adrenal glucocorticoids and insulin were responsible for the increase in G6PDH mRNA in refed rats was examined by studying the effect of insulin and the synthetic glucocorticoid, dexamethasone, on the level of functional G6PDH mRNA in primary cultures of rat hepatocytes maintained in a chemically defined medium. Hepatocytes from fasted rats were inoculated into primary culture and maintained for 48 h either in the absence of hormones or in the presence of insulin alone, dexamethasone alone or both hormones together. We observed that dexamethasone alone caused a fourfold increase in G6PDH mRNA while insulin caused about a twofold increase. Both hormones together elicited an increase that was additive. A comparison of functional G6PDH mRNA levels with the effect of the hormones on G6PDH activity and relative rate of enzyme synthesis suggests that the glucocorticoid elevates the level of G6PDH mRNA within the cell without causing a concommitant increase in the rate of synthesis of the enzyme or the level of G6PDH activity. The results obtained with the primary cultures of hepatocytes indicate that insulin and the glucocorticoids are probably involved with the regulation of hepatic G6PDH mRNA. However, involvement of other hormones, such as thyroid hormone, seems likely since the induced levels of G6PDH mRNA in hepatocytes in culture was one-third of that observed in refed rats.     

Incretin hormone receptors are required for normal beta cell development and function in female mice

The incretin hormones, glucose dependent insulinotropic polypeptide (GIP) and glucagon-like peptide 1 (GLP-1), potentiate insulin secretion and are responsible for the majority of insulin secretion that occurs after a meal. They may also, however, have a fundamental role in pancreatic beta cell development and function, independently of their role in potentiating insulin secretion after a meal. This has led to observations that a loss of GIP or GLP-1 action affects normal beta cell function, however each one of the incretin hormones may compensate when the action of the other is lost and therefore the overall impact of the incretin hormones on beta cell function is not known. We therefore utilized a mouse line deficient in both the GLP-1 and GIP receptor genes, the double incretin receptor knockout (DIRKO), to determine the consequences of a lifelong, complete lack of incretin hormone action on beta cell function, in vivo, in intact animals. We found that DIRKO mice displayed impaired glucose tolerance and insulin secretion in response to both oral glucose and mixed meal tolerance tests compared to wild-type mice. Assessment of beta cell function using the hyperglycemic clamp technique revealed an 80% decrease in first phase insulin response in DIRKO mice, but a normal second phase insulin secretion. A similar decline was seen when wild-type mice were given acute intravenous injection of glucose together with the GLP-1 receptor antagonist Ex9-39. Ex vivo assessments of the pancreas revealed significantly fewer islets in the pancreata of DIRKO mice despite no differences in total pancreatic mass. Insulin secretion from isolated islets of DIRKO mice was impaired to a similar extent to that seen during the hyperglycemic clamp. Insulin secretion in wild-type islets was impaired by acute treatment with Ex9-39 to a similar extent as the in vivo intravenous glucose tolerance tests. In conclusion, a loss of the action of both incretin hormones results in direct impairment of beta cell function both in vivo and in vitro in a process that appears to be independent of the intestinally secreted incretin hormones. We therefore conclude that the incretin hormones together significantly impact both beta-cell function and beta-cell development.     

Regulation of insulin sensitivity by adipose tissue-derived hormones and inflammatory cytokines

PURPOSE OF REVIEW: The aim of this review is to assess the role of adipose tissue-derived hormones and inflammatory cytokines in the pathogenesis of obesity-linked type II diabetes, with a special focus on articles published between December 2002 and December 2003. RECENT FINDINGS: Insulin resistance is widely recognized as a fundamental defect seen in obesity and type II diabetes. Although the molecular mechanisms triggering the development of insulin resistance remain elusive, recent studies have suggested that adipose tissue and adipose tissue-derived hormones and inflammatory cytokines play essential roles in the overall insulin sensitivity in vivo. Dysfunctions of adipose tissue can lead to systemic insulin resistance. SUMMARY: Understanding the regulation of the metabolic and secretory functions of adipose tissue, as well as its subsequent impact on overall insulin sensitivity, is becoming increasingly important given the therapeutic potential of targeting the root causes of insulin resistance in the treatment of type 2 diabetes and its associated complications, such as cardiovascular and cerebrovascular diseases.     

Regulation of adenylyl cyclase signaling system by insulin, biogenic amines and glucagon at their separate and combined action in muscle membranes of mollusc Anodonta cygnea

In the smooth muscles of mollusc Anodonta cygnea the regulatory action of hormones on adenylyl cyclase signaling system (ACSS) are realized through the receptors of serpentine type (biogenic amines, isoproterenol, glucagon) and receptor tyrosine kinase (insulin) type. Intracellular mechanisms of their interaction are interconnected. Application of hormones, their antagonists and pertussis toxin in combination with insulin and biogenic amines or glucagon on adenylyl cyclase (AC) activity allows revealing the possible sites of cross-linking in the mechanisms of their action. Combined influence of insulin and serotonin or glucagon leads to decreased stimulation of adenylyl cyclase (AC) by these hormones, whereas combined application of insulin and isoproterenol suppresses AC-stimulating effect of insulin, but AC-inhibiting effect of isoproterenol is maintained in the presence and absence of non-hydrolysable analog of GTP—guanylyl imido diphosphate (GIDP). The specific blockage of AC-stimulating effect of serotonin by cyproheptadine—antagonist of serotonin receptors, did not change AC stimulation by insulin. Beta-adrenoblockers (propranolol and alprenolol) prevent inhibition of AC activity by isoproterenol, but did not change AC stimulation by insulin. Pertussis toxin blocked AC-inhibiting effect of isoproterenol and weakened AC-stimulating action of insulin. Thus, in the muscles of Anodonta cygnea negative interaction between ACS have been revealed, which are realized under combined influence of insulin and serotonin or glucagon, most probably, at the level of receptor of serpentine type (serotonin, glucagon), whereas under action of insulin and isoproterenol at the level of G_i protein and AC interaction.     

Explanation for Unusual Potency of Salmon Calcitonin

THE calcitonins are polypeptide hormones of thirty-two amino-acids which lower serum calcium in mammals by inhibiting bone resorption¹. During evaluation of these hormones as a means of treating skeletal disorders in man, particularly Paget's disease of bone^{2, 3}, the surprising observation was made that calcitonin from the salmon (SCT) is 20–200 times more potent than porcine calcitonin (PCT) and at least ten times more potent than human calcitonin^{3, 4}. SCT is far more potent than any mammalian calcitonin yet tested in a wide variety of animal species^{5, 6}. This unusual potency of salmon calcitonin could reflect either a greater hormone affinity for receptor sites or a greater resistance to metabolic destruction. We now report evidence which supports the latter possibility, infused SCT disappears from the circulation of the dog much more slowly than does PCT.