Effect of balanced inhibition of DNA and protein synthesis on proliferative activity of lymphocytes in vitro and in vivo

1979 ◽  
Vol 87 (6) ◽  
pp. 634-637
Author(s):  
V. I. Novikov ◽  
R. V. Petrov ◽  
G. E. Fradkin
Keyword(s):  
Protein Synthesis ◽  
Proliferative Activity ◽  
Dna And Protein Synthesis

L-methionine uptake, incorporation and effects on proliferative activity and protein synthesis in bovine claw tissue explants in vitro

2007 ◽  
Vol 146 (1) ◽  
pp. 103-115 ◽  
Author(s):  
N. L. HEPBURN ◽  
C. H. KNIGHT ◽  
C. J. WILDE ◽  
K. A. K. HENDRY ◽  
H. GALBRAITH
Keyword(s):  
Protein Synthesis ◽  
Culture Medium ◽  
Culture Media ◽  
Tissue Level ◽  
Normal Function ◽  
Regulation Of Proliferation ◽  
Methionine Uptake ◽  
Dna And Protein Synthesis

SUMMARYL-methionine is a sulphur-containing nutritionally essential amino acid. It has a number of important roles in epidermal and dermal tissues of the integument of animals. Failure of normal function of these tissues in the hoof (claw) is a cause of lameness in cattle. Little is known about quantitative relationships between post-absorptive concentrations of nutrients including sulphur-containing amino acids and uptake and utilization by epidermis and dermis of the bovine claw. These parameters were studied at the tissue level by use of an established in vitro claw explant system using tissue from cattle of beef or dairy origin and L-[35S]-labelled methionine as tracer. The results showed that uptake of L-methionine by freshly prepared solear explants in Dulbecco's Modified Eagle Medium/F-12 Nutrient Mix (DMEM/F12) (1:1) medium containing 1·0 mmol L-methionine/litre was concentrative after 5–8 min, essentially linear for up to 10 min and became curvilinear thereafter. Maximum uptake and steady state conditions were obtained at approximately 30 min. Further measurements were made following 21 h incubation in culture medium. Under conditions of varying concentrations of L-methionine and measurement of uptake after 30 min, the presence of a saturable curve, that obeyed Michaelis–Menten kinetics, was demonstrated. Values of 3·61 mmol/litre and 5·84 mmol/kg intracellular water/30 min were obtained for KM and Vmax, respectively. Uptake was not influenced by L-cysteine and L-cystine concentrations in the culture media.Similar culture and incubation conditions were used in subsequent studies of DNA and protein synthesis. These showed that rates of incorporation of L-methionine into protein fractions and stimulation of DNA synthesis measured by methyl-thymidine incorporation were dependent on L-methionine concentrations in the medium. Maximal rates occurred at approximately 50 μmol/litre, which is in the normal physiological range, and at 1% of maximum uptake capacity. Examination of histological sections by autoradiography showed localization of L-[35S]-labelled methionine in basal and suprabasal epidermal cells with limited retention in dermis. Measurement, by a range of histological, immunohistochemical, electrophoretic, western blotting and autoradiographic techniques, provided further evidence of L-methionine-dependent regulation of proliferation, differentiation and synthesis of proteins under physiological concentrations, by epidermal horn-forming cells.A key role for L-methionine is suggested in the production of horn in bovine claw. The extrapolation of these in vitro data provides guidance for strategies to optimize methionine supply to claw tissues in vivo. Such extrapolation suggests the appropriateness of delivery of systemic concentrations of 50 μmol L-methionine/litre to maximize proliferative and protein depositional activity in solear epidermis and dermis in vivo.

The Use of the Epithelial Kidney OK Cell Line for Studying the Nephrotoxicity of Anticancer Platinum Coordination Complexes

1993 ◽  
Vol 21 (1) ◽  
pp. 30-37
Author(s):  
Hervé Toutain ◽  
Françoise Courjault
Keyword(s):  
Protein Synthesis ◽  
Glucose Uptake ◽  
Succinate Dehydrogenase ◽  
Platinum Complexes ◽  
Marker Enzyme ◽  
Ok Cells ◽  
Dna And Protein Synthesis ◽  
Release Method

Nephrotoxicity is one of the most important dose-limiting side-effects of cis-diaminedichloroplatinum (II) (cDDP) in humans. Quiescent OK cells grown in hormonally-defined, serum-free medium in the total absence of antibiotics were used to study the in vitro nephrotoxicity of three platinum complexes which produce different renal toxicity in vivo: cDDP, its stereoisomer trans-diaminedichloroplatinum (II) (tDDP), and cis-diamine-1,1-cyclobutane-dicarboxylateplatinUm (II) (CBDCA). The uptake and cytotoxicity of these compounds at concentrations of 3-l,600μM and their impact on DNA and protein synthesis, glucose uptake, Na+-K+-ATPase and succinate dehydrogenase activities, as well as intracellular total glutathione level, were measured. The results showed that the cytotoxicity ranking of these three compounds, assessed by the LDH release method, was not in agreement with their in vivo nephrotoxic potentials (tDDP > cDDP > CBDCA00 in OK cells versus cDDP > CBDCA > tDDP in vivo). cDDP and tDDP showed similar uptakes at all the concentrations studied, which demonstrated that their cytotoxic potential was not directly related to intracellular levels of platinum. At non-cytotoxic concentrations, both cDDP and CBDCA decreased DNA and protein synthesis and, to a lesser extent, Na+-K+-ATPase activity, whereas no effect on glucose uptake and succinate dehydrogenase activity (a mitochondrial marker enzyme) was observed. Nevertheless, 5–10 times greater concentrations of CBDCA were required to induce effects similar to those induced by cDDP. Our results did not show a rapid and early depletion of intracellular glutathione after exposure to cDDP or CBDCA. tDDP exhibited the characteristics of a non-specific cytotoxic chemical which was unable to markedly inhibit the biochemical and functional parameters studied at non-cytotoxic concentrations. These results underline the key role of the inhibition of synthetic activities in the pathogenesis of cDDP-induced and CBDCA-induced nephrotoxicity in OK cells.

Paraquat Alters Growth, DNA and Protein Synthesis in Lung Epithelial Cell and Fibroblast Cultures

1992 ◽  
Vol 20 (2) ◽  
pp. 251-257
Author(s):  
Frank A. Barile ◽  
Seeta Arjun ◽  
Jean-Jacques Senechal
Keyword(s):  
Protein Synthesis ◽  
Fibroblast Cell ◽  
Exposure Period ◽  
Lung Fibroblasts ◽  
Type I ◽  
Lung Epithelial ◽  
Dna And Protein Synthesis ◽  
Dose Dependent

In vivo exposure to paraquat, a bipyridyl herbicide, is associated with selective necrosis of type I and type II alveolar pneumocytes. We examined the influence of paraquat on cell growth, DNA and protein synthesis in human and rat lung epithelial and fibroblast cell lines, to determine the effects and affinity of the chemical in vitro for cells of various origins. Cells were incubated with [3H]-thymidine or [3H]-proline in the absence or presence of 0.05–10mM paraquat for periods of 6 or 24 hours. Cell cultures grown in the presence of 50–200μM paraquat exhibited significantly lower cell numbers than comparable controls after 3 days. The suppression of DNA synthesis by paraquat was dose-dependent, but did not rely on the length of exposure. Paraquat inhibited protein synthesis and selectively decreased net production of low molecular weight proteins. NADPH levels were the same in both control and treated cultures. [14C]-Paraquat was accumulated during a 4-hour exposure period via an active non-competitive process. The results suggest that lung fibroblasts are as sensitive to the herbicide as epithelial cells. In addition, paraquat appears to interfere with transcription and/or translation by its ability to generate superoxide radicals, rather than by depletion of reducing equivalents.

Effect of microbial isolates on microbial yield estimation in RUSITEC system

1998 ◽  
Vol 22 ◽  
pp. 306-308
Author(s):  
M. D. Carro ◽  
E. L. Miller
Keyword(s):  
Protein Synthesis ◽  
Liquid Phase ◽  
Solid Phase ◽  
Liquid Fraction ◽  
Microbial Protein ◽  
Microbial Protein Synthesis ◽  
Continuous Culture System ◽  
The One

The estimation of rumen microbial protein synthesis is one of the main points in the nitrogen (N)-rationing systems for ruminants, as microbial protein provides proportionately 0.4 to 0.9 of amino acids entering the small intestine in ruminants receiving conventional diets (Russell et al., 1992). Methods of estimating microbial protein synthesis rely on marker techniques in which a particular microbial constituent is related to the microbial N content. Marker : N values have generally been established in mixed bacteria isolated from the liquid fraction of rumen digesta and it has been assumed that the same relationship holds in the total population leaving the rumen (Merry and McAllan, 1983). However, several studies have demonstrated differences in composition between solid-associated (SAB) and fluid-associated bacteria in vivo (Legay-Carmier and Bauchart, 1989) and in vitro (Molina Alcaide et al, 1996), as well in marker : N values (Pérez et al., 1996). This problem could be more pronounced in the in vitro semi-continuous culture system RUSITEC, in which there are three well defined components (a free liquid phase, a liquid phase associated with the solid phase and a solid phase), each one having associated microbial populations.The objective of this experiment was to investigate the effect of using different bacterial isolates (BI) on the estimation of microbial production of four different diets in RUSITEC (Czerkawski and Breckenridge, 1977), using (15NH4)2 SO4 as microbial marker, and to assess what effects any differences would have on the comparison of microbial protein synthesis between diets.This study was conducted in conjunction with an in vitro experiment described by Carro and Miller (1997). Two 14-day incubation trials were carried out with the rumen simulation technique RUSITEC (Czerkawski and Breckenridge, 1977). The general incubation procedure was the one described by Czerkawski and Breckenridge (1977) and more details about the procedures of this experiment are given elsewhere (Carro and Miller, 1997).

scholarly journals Phosphorylation of eukaryotic initiation factor 4E (eIF4E) at Ser209 is not required for protein synthesis in vitro and in vivo

2001 ◽  
Vol 268 (20) ◽  
pp. 5375-5385 ◽  
Author(s):  
Linda McKendrick ◽  
Simon J. Morley ◽  
Virginia M. Pain ◽  
Rosemary Jagus ◽  
Bhavesh Joshi
Keyword(s):  
Protein Synthesis ◽  
Initiation Factor ◽  
Eukaryotic Initiation Factor ◽  
Eukaryotic Initiation Factor 4E

scholarly journals Protein synthesis in chloroplasts. Characteristics and products of protein synthesis in vitro in etioplasts and developing chloroplasts from pea leaves

1975 ◽  
Vol 146 (3) ◽  
pp. 675-685 ◽  
Author(s):  
S G Siddell ◽  
R J Ellis
Keyword(s):  
Energy Source ◽  
Protein Synthesis ◽  
Large Subunit ◽  
Sodium Dodecyl ◽  
Supernatant Fraction ◽  
Pisum Sativum L ◽  
Fraction I ◽  
Plastid Ribosomes

The function of plastid ribosomes in pea (Pisum sativum L.) was investigated by characterizing the products of protein synthesis in vitro in plastids isolated at different stages during the transition from etioplast to chloroplast. Etioplasts and plastids isolated after 24, 48 and 96h of greening in continuous white light, use added ATP to incorporate labelled amino acids into protein. Plastids isolated from greening leaves can also use light as the source of energy for protein synthesis. The labelled polypeptides synthesized in isolated plastids were analysed by electrophoresis in sodium dodecyl sulphate-ureapolyacrylamide gels. Six polypeptides are synthesized in etioplasts with ATP as energy source. Only one of these polypeptides is present in a 150 000g supernatant fraction. This polypeptide has been identified as the large subunit of Fraction I protein (3-phospho-D-glycerate carboxylyase EC 4.1.1.39) by comparing the tryptic ‘map’ of its L-(35S)methionine-labelled peptides with the tryptic ‘map’ of large subunit peptides from Fraction I labelled with L-(35S)methionine in vivo. The same gel pattern of six polypeptides is seen when plastids isolated from greening leaves are incubated with either added ATP or light as the energy source. However, the rates of synthesis of particular polypeptides are different in plastids isolated at different stages of the etioplast to chloroplast transition. The results support the idea that plastid ribosomes synthesize only a small number of proteins, and that the number and molecular weight of these proteins does not alter during the formation of chloroplasts from etioplasts.

Effect of Cycloheximide on In Vitro and In Vivo Protein Synthesis by Certain Tissues of Rats and Pigeons with Special Reference to Muscle

1973 ◽  
Vol 51 (12) ◽  
pp. 933-941 ◽  
Author(s):  
Njanoor Narayanan ◽  
Jacob Eapen
Keyword(s):  
Amino Acids ◽  
Body Weight ◽  
Protein Synthesis ◽  
Amino Acid ◽  
Amino Acid Incorporation ◽  
Contrast Administration ◽  
Acid Incorporation ◽  
Tissue Homogenates

The effect of cycloheximide in vitro and in vivo on the incorporation of labelled amino acids into protein by muscles, liver, kidneys, and brain of rats and pigeons was studied. In vitro incorporation of amino acids into protein by muscle microsomes, myofibrils, and myofibrillar ribosomes was not affected by cycloheximide. In contrast, administration of the antibiotic into intact animals at a concentration of 1 mg/kg body weight resulted in considerable inhibition of amino acid incorporation into protein by muscles, liver, kidneys, and brain. This inhibition was observed in all the subcellular fractions of these tissues during a period of 10–40 min after the administration of the precursor. Tissue homogenates derived from in vivo cycloheximide-treated animals did not show significant alteration in in vitro amino acid incorporation with the exception of brain, which showed a small but significant enhancement.

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