The Effect of Insulin on the Growth and Metabolism of the Human Diploid Cell, Wi-38

1970 ◽  
Vol 7 (2) ◽  
pp. 575-585
Author(s):  
J. B. GRIFFITHS
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Glucose Uptake ◽  
Rna Synthesis ◽  
Cultured Cells ◽  
Cell Sheet ◽  
High Energy ◽  
Contact Inhibition ◽  
Contributory Factor ◽  
Membrane Area

In a confluent culture of WI-38 cells the membrane area available for nutrient uptake is greatly reduced and the possibility exists that this reduction in uptake capacity of the cell is a contributory factor in contact inhibition. Insulin has been reported by many authors to facilitate glucose uptake and also to stimulate protein, DNA and RNA synthesis, glycolysis, pino-cytosis and growth in cultured cells. The effect of insulin on WI-38 cells was determined, therefore, to find out whether it enabled the cell to escape from contact inhibition of growth. The action of insulin was found to be dependent upon medium composition. Growth and protein synthesis were stimulated in Eagle's minimal essential medium, but not when this medium was supplemented with glucose and glutamine. Apparently insulin is only effective when high-energy compounds become limiting. Whilst insulin did not induce any post-confluent division, the protein content of cells was increased by 30%, and this was correlated with an increased rate of protein synthesis. Despite this increased activity in protein metabolism, the utilization of amino acids was less in the presence of insulin indicating that a control mechanism for more economical utilization of amino acids for protein synthesis was activated by insulin. Insulin had no effect on RNA synthesis, and only a slight inhibitory effect on DNA synthesis. Evidence was produced suggesting that insulin blocked cell division and encouraged differentiation. Glucose uptake and incorporation into the cell was stimulated by insulin, and this was especially noticeable after the cell sheet became confluent. The turnover of labelled glucose and derivatives was also enhanced by insulin and this was accompanied by a much higher rate of lactic acid production. It is concluded that insulin does not overcome contact inhibition and permit post-confluent division, but that it does enable the cell to take up and utilize nutrients more efficiently in confluent cultures with a resultant increase in metabolic activity and cell size.

The Effect of Cell Population Density on Nutrient Uptake and Cell Metabolism: a Comparative Study of Human Diploid and Heteroploid Cell Lines

1972 ◽  
Vol 10 (2) ◽  
pp. 515-524
Author(s):  
J. B. GRIFFITHS
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Amino Acid ◽  
Dna Synthesis ◽  
Nutrient Uptake ◽  
Cell Lines ◽  
Acid Concentration ◽  
Contact Inhibition ◽  
Inhibition Of Growth ◽  
Extracellular Amino Acid

The possibility that contact inhibition of growth in cultures of human diploid cells is influenced by the effects of cell crowding on nutrient uptake by the cells was investigated. Two human lung cell lines were compared, the diploid line MRC-5 and the heteroploid line L-132. In pre-confluent cultures the ability of these 2 cell types to accumulate amino acids was very similar. Post-confluent L-132 cells showed very little change from the pre-confluent cultures but the ability of MRC-5 cells in post-confluent cultures was greatly reduced. The intracellular concentrations of various amino acids necessary to achieve the maximum rate of protein synthesis were found. These values were identical for sparse and crowded cultures but due to the reduced uptake ability of crowded MRC-5 cells a far higher external amino acid concentration was required in post-confluent cultures. This meant that although amino acids did not become growth-limiting until over 80% utilized in pre-confluent cultures, in post-confluent cultures they became growth-limiting when only 50% utilized. Although protein synthesis was significantly affected by extracellular amino acid concentration and cell crowding, thus contributing towards the effect of contact inhibition of growth, DNA synthesis was shown to be the major metabolic function in contact inhibition. Increased cell density had a very inhibitory effect on DNA synthesis in MRC-5 cultures, but not in L-132 cultures, and this was unaffected by extracellular amino acid and glucose concentration.

THE EFFECT OF GROWTH HORMONE AND OF CORTISOL ON THE RESPONSE OF ISOLATED RAT DIAPHRAGM TO THE STIMULATING EFFECT OF INSULIN ON GLUCOSE UPTAKE AND ON INCORPORATION OF AMINO ACIDS INTO PROTEIN

1959 ◽  
Vol 18 (4) ◽  
pp. 395-408 ◽  
Author(s):  
K. L. MANCHESTER ◽  
P. J. RANDLE ◽  
F. G. YOUNG
Keyword(s):  
Amino Acids ◽  
Growth Hormone ◽  
Protein Synthesis ◽  
Glucose Uptake ◽  
Carbohydrate Metabolism ◽  
Rat Diaphragm ◽  
Pituitary Growth Hormone ◽  
Isolated Rat ◽  
Stimulation Of

SUMMARY 1. The effect of hypophysectomy, or of adrenalectomy, and injection of pituitary growth hormone (GH) or of cortisol, on the uptake of glucose and the incorporation of glycine into protein by isolated rat diaphragm, and the effect of the addition of insulin in vitro on these processes, has been studied. 2. Both hypophysectomy and adrenalectomy raised the uptake of glucose by isolated diaphragm, while treatment of the intact or of the hypophysectomized rat with GH, or of the intact or of the adrenalectomized rat with cortisol, depressed it. Although hypophysectomy and adrenalectomy did not influence the additional glucose uptake induced by 200 mu./ml. of insulin in vitro, both these operations enhanced the effect of 0·1–1·0 mu./ml. of insulin on glucose uptake by diaphragm in vitro. Treatment of the rat with GH or cortisol diminished the rise in glucose uptake of diaphragm induced by 0·1–1·0 mu./ml. insulin. 3. Hypophysectomy depressed, and administration of GH to the intact or hypophysectomized rat raised, the incorporation of glycine into protein of the isolated diaphragm, but neither of these operations altered the magnitude of the stimulation of incorporation induced by 1·0 mu./ml. insulin. 4. Adrenalectomy raised, and administration of cortisol to the intact or adrenalectomized rat depressed, the incorporation of glycine into protein of the isolated diaphragm; adrenalectomy enhanced, the injection of cortisol diminished, the effect of 1·0 mu./ml. insulin on these processes. 5. The possibility that GH directs insulin towards the stimulation of protein synthesis, in part by restraining the action of insulin on carbohydrate metabolism, is discussed.

Investigation of the mode of nuclear control over protein synthesis in early development of loach and sea urchin

Development ◽  
1972 ◽  
Vol 28 (3) ◽  
pp. 491-509
Author(s):  
Maya R. Krigsgaber ◽  
A. A. Neyfakh
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Ionizing Radiation ◽  
Radiation Damage ◽  
Sea Urchin ◽  
Rna Synthesis ◽  
Developmental Stages ◽  
Subsequent Development ◽  
Long Term Treatment ◽  
Androgenetic Haploid

It is shown that in loach embryos the incorporation of precursors into protein takes place in the blastoderm cells only. The change of the rate of incorporation of labelled amino acids into protein of the blastoderm separated from the yolk at successive developmental stages reflects the changes in the level of protein synthesis in intact embryos of the same developmental stages. Typical periodic changes of the intensity of protein synthesis in early embryo-genesis of the loach are detected: low incorporation of amino acids at blastula stages is followed by an increase of synthesis during gastrulation and by a decrease with the onset of organogenesis. To study the genetic control over protein synthesis at various developmental stages of loach and sea-urchin embryos the effects of ionizing radiation and long-term treatment with actinomycin D have been examined. X-Irradiation doses produce an insignificant direct effect on protein synthesis, while radiation damage of the nuclear apparatus results in a gradual but ever increasing inhibition of protein synthesis. The inhibition of RNA synthesis with actinomycin or ionizing radiation damage of the nuclei produce essentially the same effect on the intensity of protein synthesis. Protein synthesis in androgenetic haploid hybrid embryos (loach ♀ × goldfish ♂) and in loach androgenetic haploid embryos after producing a partial elimination of chromosomes does not differ from ‘enucleated’ loach embryos completely deprived of chromosomes. These data suggest that high-polymeric RNA formed after the elimination of some chromsomes is unable to provide a normal level of protein synthesis. Protein synthesis is not controlled by the nuclei up to the stages of early blastula (sea urchin) and of late blastula (loach), being evidently programmed in oogenesis. To provide a rapid activation of protein synthesis in the course of gastrulation in the loach the function of the nuclei has to be realized during mid-blastula stages. An increase of the rate of the incorporation of amino acids at the stages of mesenchyme blastula in the sea urchin depends on the synthesis of RNA at the early blastula. At the same time protein synthesis during gastrulation in the loach and at mesenchyme blastula in the sea urchin is much less dependent on the simultaneous RNA synthesis. Protein synthesis at these stages seems to be provided by the long-living templates and controlled by non-gene mechanisms of the regulation of translation. Thus early embryonic differentiation in the loach and sea-urchin development is related to the activation of protein synthesis. The latter is provided by the preceding morphogenetic nuclear function, which makes protein synthesis relatively independent of simultaneous synthesis of templates that ensures subsequent development stages.

scholarly journals Genomes contain relics of a triplet code connecting the origins of primordial RNA synthesis to the origins of genetically coded protein synthesis

2021 ◽  
Author(s):  
Geoffrey Henry Siwo
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Genetic Code ◽  
Rna Synthesis ◽  
Peptide Bond ◽  
Amino Acid Sequences ◽  
Peptide Bond Formation ◽  
Rna Sequences ◽  
Synthesis Mechanism ◽  
Triplet Code

Life on earth relies on three types of information polymers- DNA, RNA and proteins. In all organisms and viruses, these molecules are synthesized by the copying of pre-existing templates. A triplet-based code known as the genetic code guides the synthesis of proteins by complex enzymatic machines that decode genetic information in RNA sequences. The origin of the genetic code is one of the most fundamental questions in biology. In this study, computational analysis of about 5,000 species level metagenomes using techniques for the analysis of human language suggests that the genomes of extant organisms contain relics of a distinct triplet code that potentially predates the genetic code. This code defines the relationship between adjacent triplets in DNA/RNA sequences, whereby these triplets predominantly differ by a single base. Furthermore, adjacent triplets encode amino acids that are thought to have emerged around the same period in the earth's early history. The results suggest that the order of triplets in primordial RNA sequences was associated with the availability of specific amino acids, perhaps due to a coupling of a triplet-based primordial RNA synthesis mechanism to a primitive mechanism of peptide bond formation. Together, this coupling could have given rise to early nucleic acid sequences and a system for encoding amino acid sequences in RNA, i.e. the genetic code. Thus, the central role of triplets in biology potentially extends to the primordial world, contributing to both the origins of genomes and the origins of genetically coded protein synthesis.

scholarly journals STUDIES ON THE MECHANISM OF ACTION OF PEDERINE

1968 ◽  
Vol 36 (3) ◽  
pp. 485-496 ◽  
Author(s):  
Agnese Brega ◽  
Arturo Falaschi ◽  
Luigi De Carli ◽  
Mario Pavan
Keyword(s):  
Protein Synthesis ◽  
Dna Synthesis ◽  
Cytotoxic Effect ◽  
Rna Synthesis ◽  
Cultured Cells ◽  
Specific Inhibitor ◽  
Cytological Examination ◽  
Effect Analysis ◽  
Inhibitor Of Protein Synthesis

Pederine, a drug extracted from the coleopter Paederus fuscipes, inhibits the growth of in vitro cultured cell lines at concentrations of the order of 1.5 nanogram/ml. Cytological examination shows a generalized cytotoxic effect. Analysis of macromolecular syntheses by the use of radioactive precursors shows that pederine causes an almost immediate block of protein and DNA synthesis, without affecting RNA synthesis. The effects on the synthesis of the two types of macromolecules remain nearly simultaneous even at the lowest active concentrations of pederine. Studies with cell-free systems show that the drug inhibits protein synthesis, whereas it is ineffective on the DNA-polymerizing activity. It seems, therefore, that the drug acts primarily on the amino acid-polymerizing system, and that the effect on DNA is secondary. This idea is strengthened by the observation that puromycin, a specific inhibitor of protein synthesis, also affects promptly DNA synthesis of in vitro cultured cells. Other authors have shown the same phenomenon with a number of inhibitors of protein synthesis; the properties of pederine support, therefore, the view that continuous protein synthesis is necessary for the maintenance of DNA replication in higher organisms.

scholarly journals Nanoparticle Delivered Human Biliverdin Reductase-Based Peptide Increases Glucose Uptake by Activating IRK/Akt/GSK3 Axis: The Peptide Is Effective in the Cell and Wild-Type and Diabetic Ob/Ob Mice

2016 ◽  
Vol 2016 ◽  
pp. 1-15 ◽  
Author(s):  
Peter E. M. Gibbs ◽  
Tihomir Miralem ◽  
Nicole Lerner-Marmarosh ◽  
Mahin D. Maines
Keyword(s):  
Amino Acids ◽  
Cell Culture ◽  
Glucose Uptake ◽  
Animal Studies ◽  
Cultured Cells ◽  
Kinase Domain ◽  
Terminal Segment ◽  
Wild Type ◽  
Stimulation Of

Insulin’s stimulation of glucose uptake by binding to the IRK extracellular domain is compromised in diabetes. We have recently described an unprecedented approach to stimulating glucose uptake. KYCCSRK (P2) peptide, corresponding to the C-terminal segment of hBVR, was effective in binding to and inducing conformational change in the IRK intracellular kinase domain. Although myristoylated P2, made of L-amino acids, was effective in cell culture, its use for animal studies was unsuitable. We developed a peptidase-resistant formulation of the peptide that was efficient in both mice and cell culture systems. The peptide was constructed of D-amino acids, in reverse order, and blocked at both termini. Delivery of the encapsulated peptide to HepG2 and HSKM cells was confirmed by its prolonged effect on stimulation of glucose uptake (>6 h). The peptide improved glucose clearance in both wild-type and Ob/Ob mice; it lowered blood glucose levels and suppressed glucose-stimulated insulin secretion. IRK activity was stimulated in the liver of treated mice and in cultured cells. The peptide potentiated function of IRK’s downstream effector, Akt-GSK3-(α,β)axis. Thus, P2-based approach can be used for improving glucose uptake by cells. Also, it allows for screening peptidesin vitroand in animal models for treatment of diabetes.

Inhibition of cell division in amoebae: the incorporation of tritiated precursors into Amoeba proteus after the injection of non-homologous cytoplasm

1976 ◽  
Vol 20 (3) ◽  
pp. 525-537
Author(s):  
J.M. Cameron ◽  
S.E. Hawkins
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Rna Synthesis ◽  
Amoeba Proteus ◽  
Supernatant Fraction ◽  
Free Living ◽  
Rna Molecules ◽  
Site Of Action ◽  
Membrane Components ◽  
Inhibitory Molecules

The injection of non-homologous cytoplasm into any strain of large free-living amoebae leads to a 60% inhibition of division amongst recipient cells. When the post-microsomal supernatant fraction of Amoeba discoides was injected into A. proteus, this inhibition of division was as high as 95%. The incorporation of tritiated precursors, either [3H]uridine or 3H-amino acids, into these inhibited amoebae was studied at various times after the injection of the inhibitory material using autoradiography. When cells were grown in [3H]uridine, autoradiographs indicated that RNA synthesis had ceased 2 days after the injection of non-homologous material. However, if [3H]uridine was injected into the inhibited cells, some synthesis of RNA could be detected up to 4 days after the injection of inhibitor. These results suggested that uptake of [3H]uridine was impaired and that one site of action of the inhibitory molecules was RNA synthesis for membrane components. Experiments with a variety of 3H-amino acids suggested that protein synthesis continued for at least 9 days after the injection of non-homologous cytoplasm, and that in these cells some informational RNA molecules were long-lived. There seemed to be accumulation of material containing [3H]lysine in the nuclei of control cells taken at random from cultures, and this was seen in the nuclei of inhibited cells 1 day after injection. However, 2 days after the injection of inhibitor, no accumulation of [3H]lysine-containing material was found in the nuclei.

MICROBIOLOGICAL STUDIES ON THE MECHANISM OF ACTION OF SPARSOMYCIN

1966 ◽  
Vol 12 (4) ◽  
pp. 595-604 ◽  
Author(s):  
Edward R. Bannister ◽  
Dale E. Hunt ◽  
Robert F. Pittillo
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Protein Synthesis ◽  
Mechanism Of Action ◽  
Rna Synthesis ◽  
Primary Site ◽  
Cross Resistance ◽  
E Coli

A primary site of sparsomycin attack in Escherichia coli appears to be inhibition of synthesis of protein, which occurs at concentrations of sparsomycin that do not affect DNA or RNA synthesis. Sparsomycin interferes with the normal excretion of amino acids by E. coli. Some cross-resistance was observed between a culture resistant to sparsomycin and cultures resistant to other inhibitors of protein synthesis.

scholarly journals Physiology of Oil Seeds. V. Germination of NC-13 Virginia-type Peanut Seeds in the Presence of Inhibitors and Ethylene.1,2

1975 ◽  
Vol 2 (2) ◽  
pp. 73-77
Author(s):  
D. L. Ketring
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Total Protein ◽  
Rna Synthesis ◽  
De Novo ◽  
Acid Synthesis ◽  
De Novo Synthesis ◽  
Tracer Studies ◽  
Oil Seeds ◽  
Protein And Rna Synthesis

Abstract Control dormant seeds that imbibed water for 16 hr germinated 100% after 10 μ 1/1 C2H4 was applied for 24 hr. Dormant seeds that imbibed cycloheximide (100 μg/ml), 6-methylpurine (50 μ g/ml) and 6-azauracil (50 μ g/ml) for 16 hr did not germinate at either 24 or 48 hr after 10 μ 1/1 ethylene treatment. Both protein- and nucleic acid-synthesis ihhibitors prevented germination induced by ethylene in these dormant seeds. Imbibition of 20 μ M ABA by dormant seeds prevented germination, but this effect was reversed by ethylene. Tracer studies with 14C-amino acids indicate that ABA does not inhibit total protein synthesis, but it does inhibit emergence in the absence of ethylene. In the presence of ABA plus ethylene, emergence occurred, but no change in total protein synthesis was detected. At 8 weeks after harvest, both germination and incorporation of 2–14C-uracil into RNA were inhibited by ABA and stimulated by ethylene. By 17 weeks after harvest, only the inhibition of germination and its reversal by ethylene were notable. However, at 17 weeks after harvest, ethylene enhanced RNA synthesis when germination and protein synthesis were inhibited by cycloheximide. Development of isocitritase activity in the seeds was inhibited by ABA and the inhibition was reversed by ethylene, indicating that de novo synthesis of protein is inhibited by ABA and activated by ethylene in these seeds. The opposite effects of ABA and ethylene on germination, RNA synthesis and isocitritase activity suggest that germination is controlled at the level of RNA and/or protein synthesis in these seeds. The prevention of germination of dormant seeds in the presence of ethylene by protein- and RNA-synthesis ihhibitors supports this suggestion, but the data do not preclude an action of ABA or ethylene prior to detectable affects on RNA or protein synthesis.

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