Studies on the nature of the seromucoid and haptoglobin responses to experimental inflammation

1968 ◽  
Vol 46 (5) ◽  
pp. 477-481 ◽  
Author(s):  
M. Maung ◽  
D. G. Baker ◽  
R. K. Murray
Keyword(s):  
Protein Synthesis ◽  
Actinomycin D ◽  
De Novo ◽  
De Novo Synthesis ◽  
Experimental Inflammation ◽  
Haptoglobin Level

The effects of the administration of actinomycin D, ethionine, and puromycin on the elevations of the total seromucoid fraction and of one of its components (haptoglobin) occurring during experimental inflammation have been studied. All three inhibitors of protein synthesis abolished the elevation of haptoglobin level. Ethionine and puromycin also completely suppressed the elevation of total seromucoid level, whereas actinomycin D only partially suppressed it. The seromucoid and haptoglobin levels in control animals injected with only the inhibitors of protein synthesis were not in general significantly different from those of the animals injected with turpentine and these agents. The results are consistent with the concept that the elevation of various plasma glycoproteins occurring during inflammation is principally due to de novo synthesis of these proteins rather than release of preformed proteins from tissue pools.

Recovery of normal protein synthesis in heat-shocked chicken myotubes by liposome-mediated transfer of mRNAs

1985 ◽  
Vol 63 (3) ◽  
pp. 231-235 ◽  
Author(s):  
Jnanankur Bag
Keyword(s):  
Protein Synthesis ◽  
Heat Shock ◽  
Rna Synthesis ◽  
Actinomycin D ◽  
De Novo ◽  
Critical Role ◽  
Recovery Process ◽  
De Novo Synthesis ◽  
Normal Pattern ◽  
Large Unilamellar Vesicles

Exposure of chicken myotube culture to 45 °C induced the synthesis of three heat-shock polypeptides of 25 000, 65 000, and 81 000 daltons. Recovery to the normal pattern of protein synthesis was judged by the decrease in the synthesis of heat-shock polypeptides. This recovery to normal protein synthesis required de novo synthesis of mRNAs for normal cellular proteins. Inhibition of RNA synthesis by actinomycin D during recovery at 37 °C blocked the recovery process and resulted in the continued synthesis of heat-shock polypeptides. Large unilamellar vesicles were used to examine the effect of delivery of mRNAs isolated from both normal and heat-shocked myotubes on the recovery of these cells from heat-shock treatment. The results presented here show that liposome-mediated delivery of normal mRNAs to heat-shocked cells relieved the block of recovery by actinomycin. On the other hand, when mRNAs from heat-shocked cells were used during recovery, the synthesis of heat-shock polypeptides was stimulated. These observations suggest that the relative abundance of mRNAs in the cytoplasm plays a critical role in regulating protein synthesis in chicken myotube cultures.

scholarly journals Evidence for the De Novo Synthesis of Erythropoietin in Hypoxic Rats

Blood ◽  
1972 ◽  
Vol 40 (5) ◽  
pp. 662-670 ◽  
Author(s):  
J. C. Schooley ◽  
L. J. Mahlmann
Keyword(s):  
Rna Synthesis ◽  
Actinomycin D ◽  
De Novo ◽  
Protein S ◽  
Male Rats ◽  
Hypoxic Exposure ◽  
De Novo Synthesis ◽  
Serum Erythropoietin

Abstract Significant increases in the serum erythropoietin of male rats occur after the end of a brief hypoxic exposure. These increases in the hormone are almost completely abolished when the kidneys are removed after the hypoxic exposure. Injection of puromycin or cycloheximide after the hypoxic exposure significantly decreases the subsequent increases in serum erythropoietin titers, whereas injections of actinomycin D at this time have no significant effect on erythropoietin levels. Injections of actinomycin D before the hypoxic exposure prevent the increase in serum erythropoietin that normally occurs. These findings suggest that a brief period of hypoxia initiates a DNA-dependent RNA synthesis that regulates the de novo ribosomal synthesis of protein(s) involved in the biogenesis of erythropoietin and that the kidney is essential for these reactions to occur.

Glucocorticoids inhibit IL-1β-induced GM-CSF expression at multiple levels: roles for the ERK pathway and repression by MKP-1

2010 ◽  
Vol 427 (1) ◽  
pp. 113-124 ◽  
Author(s):  
Robert Newton ◽  
Elizabeth M. King ◽  
Wei Gong ◽  
Christopher F. Rider ◽  
Karl J. Staples ◽  
...  
Keyword(s):  
Gene Expression ◽  
Protein Synthesis ◽  
Transcriptional Repression ◽  
Actinomycin D ◽  
De Novo ◽  
A549 Cells ◽  
Mitogen Activated Protein Kinase ◽  
Erk Pathway ◽  
Gm Csf ◽  
Erk Phosphorylation

In the present study, IL (interleukin)-1β increased GM-CSF (granulocyte/macrophage colony-stimulating factor) expression from pulmonary A549 cells and primary HBE (human bronchial epithelial) cells. These responses were repressed by the glucocorticoid dexamethasone, allowing the use of A549 cells as a relevant model. IL-1β induced GM-CSF release into the culture medium by 6 h and in cell lysates (cytosolic) at 2 h. These effects were profoundly inhibited by dexamethasone, yet IL-1β-induced GM-CSF mRNA and unspliced nRNA (nuclear RNA; a surrogate of transcription rate) were modestly inhibited by dexamethasone at times up to 2 h. Although this indicates an effect on protein synthesis, actinomycin D chase experiments also indicated post-transcriptional repression by dexamethasone. Dexamethasone-dependent mRNA repression increased with time and was prevented by translational blockade. In addition, dexamethasone and the dissociated steroid RU24858 repressed GM-CSF release in an actinomycin D-sensitive manner, thereby implicating glucocorticoid-induced gene expression. At 2 h, IL-1β-induced expression of GM-CSF protein, but not mRNA, was sensitive to the MEK [MAPK (mitogen-activated protein kinase)/ERK (extracellular-signal-regulated kinase) kinase] inhibitors PD098059 and U0126. Although this indicates a role for the MEK/ERK pathway in GM-CSF translation, PD098059 subsequently destabilized GM-CSF mRNA. Dexamethasone and RU24858 both reduced IL-1β-induced ERK phosphorylation and increased MKP-1 (MAPK phosphatase-1) expression. Inhibition of ERK phosphorylation was reproduced by MKP-1 overexpression and prevented by MKP-1-targeting siRNA (small interfering RNA). Since MKP-1 prevented GM-CSF expression by transcriptional, post-transcriptional and translational processes, we propose that glucocorticoids induce MKP-1 expression to reduce both MEK/ERK activation and GM-CSF protein synthesis. Thus de novo gene expression, particularly of MKP-1, is involved in the repressive effects of glucocorticoids.

scholarly journals EFFECTS OF CYTOSINE ARABINOSIDE ON DIFFERENTIAL GENE EXPRESSION IN EMBRYONIC NEURAL RETINA

1974 ◽  
Vol 61 (3) ◽  
pp. 688-700 ◽  
Author(s):  
R. E. Jones ◽  
A. A. Moscona
Keyword(s):  
Cytosine Arabinoside ◽  
Rna Synthesis ◽  
Actinomycin D ◽  
De Novo ◽  
Neural Retina ◽  
Regulatory Genes ◽  
De Novo Synthesis ◽  
Macromolecular Synthesis ◽  
Differential Gene ◽  
Inhibition Of Dna Synthesis

The analogue of cytidine, cytosine arabinoside (Ara-C), elicited a significant increase in the level of glutamine synthetase (GS) in embryonic chick neural retina in the absence of the steroid inducer of the enzyme. The increase was due to de novo synthesis of GS and was mediated by RNA which accumulated in the presence of the effective concentration of Ara-C. Accumulation of GS did not result from the inhibition of DNA synthesis for which Ara-C is best known. This new effect of Ara-C involves differential suppression of macromolecular synthesis in this system: the concentration of Ara-C which caused maximum GS accumulation suppressed overall protein and RNA syntheses 65–75% without inhibiting the transcription and translation of templates essential for GS synthesis. Withdrawal of Ara-C resulted in restoration of RNA synthesis and cessation of GS accumulation, even though preformed templates for the enzyme were present; however, if all RNA synthesis was arrested with actinomycin D at the time of Ara-C withdrawal, GS continued to accumulate. The results are consistent with the hypothesis that Ara-C differentially affects the activity of structural and regulatory genes involved in the regulation of GS levels in the retina: Ara-C allows transcription of the enzyme-specific templates, but reversibly inhibits the expression of regulatory genes which limit the accumulation of GS.

Kallikrein release by vascular tissue

1993 ◽  
Vol 265 (4) ◽  
pp. H1209-H1214 ◽  
Author(s):  
H. Nolly ◽  
O. A. Carretero ◽  
A. G. Scicli
Keyword(s):  
Protein Synthesis ◽  
Vascular Tissue ◽  
De Novo ◽  
Vena Cava ◽  
Lactic Dehydrogenase ◽  
De Novo Synthesis ◽  
Kinin System ◽  
Vascular Rings ◽  
Small Vessels ◽  
Kallikrein Kinin System

Vascular tissue contains kallikrein and kallikrein mRNA, suggesting a vascular kallikrein-kinin system. We questioned whether 1) kallikrein concentration varies among large and small vessels; 2) kallikrein is released by vascular tissue; and 3) blocking protein synthesis inhibits release, suggesting de novo synthesis. Using rat vascular rings and isolated-perfused hindquarters, we examined kallikrein in the bath and perfusate. Active kallikrein was higher in tail arteries than the aorta (P < 0.001); tail veins had six times more kininogenase than the vena cava (P < 0.001). Total kallikrein showed a similar pattern, being highest in tail vessels. Arterial rings released active and total kallikrein. After 1, 2, and 3 h incubation, cumulative release was as follows: active, 90 +/- 13, 201 +/- 25, and 311 +/- 41 pg.h-1 x mg tissue-1; total, 170 +/- 14, 366 +/- 24, and 537 +/- 40 pg.h-1 x mg tissue-1, indicating constant release up to > or = 3 h. In contrast, lactic dehydrogenase fell from 6.7 +/- 2.5 to 2.5 +/- 0.4 U.h-1 x mg tissue-1. Total kallikrein in the rings was 302 +/- 51 pg bradykinin/mg wt tissue before 3 h and 298 +/- 68 afterward. Kallikrein released by the hindquarters after 3 h was as follows: active, 6.2 +/- 2.8 ng bradykinin.min-1 x kg.body wt-1; total, 85.2 +/- 17 ng bradykinin.min-1 x kg body wt-1. Puromycin pretreatment (10 mg ip) reduced total perfusate kallikrein from 105 +/- 19 to 8.5 +/- 3.6 (P < 0.005).(ABSTRACT TRUNCATED AT 250 WORDS)

Transmembrane transport and intracellular kinetics of amino acids in human skeletal muscle

1995 ◽  
Vol 268 (1) ◽  
pp. E75-E84 ◽  
Author(s):  
G. Biolo ◽  
R. Y. Fleming ◽  
S. P. Maggi ◽  
R. R. Wolfe
Keyword(s):  
Skeletal Muscle ◽  
Amino Acids ◽  
Protein Synthesis ◽  
Amino Acid ◽  
Human Skeletal Muscle ◽  
De Novo ◽  
Transmembrane Transport ◽  
Acid Transport ◽  
De Novo Synthesis ◽  
Intracellular Amino Acid

We have used stable isotopic tracers of amino acids to measure in vivo transmembrane transport of phenylalanine, leucine, lysine, alanine, and glutamine as well as the rates of intracellular amino acid appearance from proteolysis, de novo synthesis, and disappearance to protein synthesis in human skeletal muscle. Calculations were based on data obtained by the arteriovenous catheterization of the femoral vessels and muscle biopsy. We found that the fractional contribution of transport from the bloodstream to the total intracellular amino acid appearance depends on the individual amino acid, varying between 0.63 +/- 0.02 for phenylalanine and 0.22 +/- 0.02 for alanine. Rates of alanine and glutamine de novo synthesis were approximately eight and five times their rate of appearance from protein breakdown, respectively. The model-derived rate of protein synthesis was highly correlated with the same value calculated by means of the tracer incorporation technique. Furthermore, amino acid transport rates were in the range expected from literature values. Consequently, we conclude that our new model provides a valid means of quantifying the important aspects of protein synthesis, breakdown, and amino acid transport in human subjects.

scholarly journals Cell-shape-dependent modulation of p52(PAI-1) gene expression involves a secondary response pathway

1995 ◽  
Vol 306 (2) ◽  
pp. 497-504 ◽  
Author(s):  
P J Higgins ◽  
L Staiano-Coico ◽  
M P Ryan
Keyword(s):  
Protein Synthesis ◽  
Transcriptional Activation ◽  
Actinomycin D ◽  
De Novo ◽  
Rat Kidney ◽  
Secondary Response ◽  
Protein Levels ◽  
Run Off ◽  
Dependent Pathway ◽  
Pai 1

Expression of the rat p52(PAI-1) gene is positively regulated by agents that influence cellular microfilament organization and/or cell-to-substrate adhesion [e.g. cytochalasin D (CD) and sodium n-butyrate (NaB)] [Higgins, Chaudhari and Ryan (1991) Biochem. J. 273, 651-658; Higgins, Ryan and Providence (1994) J. Cell. Physiol. 159, 187-195]. As shape-responsive genes may be subject to inducer-specific controls, the biochemical mechanisms underlying the shape-dependent pathway of p52(PAI-1) gene regulation were examined in v-ras-transformed rat kidney (KNRK) cells. NaB and/or CD effectively stimulated p52(PAI-1) run-off transcription and augmented de novo p52(PAI-1) mRNA and protein synthesis in KNRK cells; induction at both the mRNA and protein levels was inhibited by actinomycin D. Pretreatment with cycloheximide (CX) markedly attenuated NaB- and/or CD-stimulated p52(PAI-1) expression. CX alone, however, induced low levels of p52(PAI-1) mRNA; increased p52(PAI-1) protein synthesis was evident after release of KNRK cells from CX blockade. Such CX-mediated induction was also sensitive to actinomycin D. Full stimulation of p52(PAI-1) expression in KNRK cells in response to the shape modulators NaB and/or CD involves transcriptional activation of the p52(PAI-1) gene, requires de novo RNA synthesis and occurs through a secondary-response (i.e. protein-synthesis-dependent) pathway.

scholarly journals OBSERVATIONS ON THE ACID PHOSPHATASES OF EUGLENA GRACILIS

1965 ◽  
Vol 24 (2) ◽  
pp. 223-234 ◽  
Author(s):  
Jacob J. Blum
Keyword(s):  
Cell Division ◽  
Acid Phosphatase ◽  
Phosphatase Activity ◽  
Acid Phosphatase Activity ◽  
Actinomycin D ◽  
De Novo ◽  
Fluoride Concentration ◽  
Acid Phosphatases ◽  
De Novo Synthesis ◽  
Increased Activity

When a bleached strain of Euglena is maintained in a medium containing very low con centrations of phosphate, the acid phosphatase activity increases. The increase in acid phosphatase activity is prevented by Actinomycin D and by p-fluorophenylalanine (PFA), indicating that the increased activity is due to de novo synthesis of acid phosphatase. When phosphate is replenished, the acid phosphatase activity decreases to the level characteristic of uninduced cells before there is any appreciable cell division. When cell division resumes in the presence of PFA, the level of acid phosphatase activity remains approximately constant. This indicates that there are two different phosphatases: a constitutive enzyme, whose synthesis is insensitive to the presence of PFA, and an induced enzyme, whose synthesis is sensitive to PFA. These enzymes are not equally sensitive to changes in pH and in fluoride concentration, thus permitting them to be assayed individually in whole toluene-treated cells. Induced cells also acquire the ability to remove phosphate from the medium very rapidly.

Angiotensin II AT1 receptor internalization, translocation and de novo synthesis modulate cytosolic and nuclear calcium in human vascular smooth muscle cells

2003 ◽  
Vol 81 (3) ◽  
pp. 274-287 ◽  
Author(s):  
G Bkaily ◽  
S Sleiman ◽  
J Stephan ◽  
C Asselin ◽  
S Choufani ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Smooth Muscle ◽  
Angiotensin Ii ◽  
Smooth Muscle Cells ◽  
Vascular Smooth Muscle Cells ◽  
Nuclear Translocation ◽  
De Novo ◽  
Ang Ii ◽  
De Novo Synthesis ◽  
Nuclear Compartments

The present study was designed to verify if human (h) Angiotensin II (Ang II) type-1 receptor (hAT1R) undergoes internalization, nuclear translocation, and de novo synthesis in primary culture of human aortic vascular smooth muscle cells (hVSMCs) and if overexpression of this receptor modulates sustained free cytosolic ([Ca]c) and nuclear ([Ca]n) calcium. 3-dimensional (3-D) confocal microscopy was used to monitor free intracellular Ca2+ and hAT1R-green fluorescence protein (GFP) fusion protein in cultured hVSMCs. Immunofluorescence studies showed the presence of hAT1R and the absence of hAT2R in normal hVSMCs. Using 3-D imaging technique, hAT1 receptors were localized at the sarcolemma and in the cytosolic and nuclear compartments. In native as well as in normal hAT1R or hAT1R –GFP overexpressing hVSMCs, Ang II (10–9 and 10–4 M) induced internalization and nuclear translocation of this type of receptor. The internalization of hAT1Rs is mediated via clathrin-coated pits and vesicles pathway. This phenomenon of trancellular trafficking of receptors was associated with an increase of hAT1R. The Ang II induced increase of hAT1R density was prevented by the protein synthesis inhibitor cycloheximide. Overexpression of hAT1R and hAT1R–GFP decreased both basal cytosolic and nuclear Ca2+. In normal hVSMCs and low hAT1R–GFP overexpressing hVSMCs, Ang II (10–15 to 10–4 M) induced a dose-dependent sustained increase of [Ca]c and [Ca]n with an EC50 near 5 × 10–11 and 5 × 10–9 M, respectively. Our results suggest that hAT1Rs are the predominant type of Ang II receptors in aortic hVSMCs and are present in the sarcolemma, the cytosolic and the nuclear compartments. Ang II rapidly induces hAT1R internalization, nuclear translocation, as well as nuclear de novo synthesis of this receptor. The hAT1R overexpression in hVSMCs modulates sustained [Ca]c and [Ca]n.Key words: angiotensin, calcium, protein synthesis, nucleus, AT1 receptor, nuclear de novo synthesis.

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