An Evaluation of the Metabolic Basis of Aflatoxin B1 Toxicity by Using Buffalo Granulocytes and Agranulocytes In Vitro

2005 ◽  
Vol 33 (4) ◽  
pp. 387-390 ◽  
Author(s):  
Tukaram More ◽  
Golla Ramalinga Reddy ◽  
Sandeep Kumar
Keyword(s):  
Protein Synthesis ◽  
Atpase Activity ◽  
Aflatoxin B1 ◽  
Cell Types ◽  
Leucine Incorporation ◽  
Transferase Activity ◽  
Metabolic Basis ◽  
Aflatoxin B ◽  
In Cells

This study was aimed at monitoring cytotoxic changes in buffalo leukocyte subpopulations exposed to aflatoxin B1 (AFB1), since no such information is available for this species. The effects of AFB1 on glutathione (GSH) S-transferase, Ca2+Mg2+ATPase and protein synthesis in leukocyte subpopulations, namely, mononuclear (MN) cells and polymorphonuclear (PMN) cells isolated from the blood of the domestic buffalo ( Bos bubalis), were studied. The cells were separated by using Ficoll-Paque and incubated in the presence of AFB1. GSH S-transferase activity was found to increase in cells exposed to AFB1, but there was no difference in activity between MN and PMN cells. PMN cell ATPase activity increased after AFB1 treatment, whereas no such effect was observed in the MN cells, which showed higher basal levels of ATPase activity. In the presence of AFB1, all the cells showed significant decreases in 14C-leucine incorporation, but the MN cells showed higher 14C-leucine incorporation than the PMN cells. Nevertheless, both cell types were affected by AFB1 and participated in its detoxification. There was also an appreciable decrease in the release of myeloperoxidase by activated PMN cells in the presence of AFB1.

EFFECT OF ACTINOMYCIN D ON TSH-INDUCED STIMULATION OF THYROIDAL PROTEIN SYNTHESIS IN THE RAT

1974 ◽  
Vol 77 (1) ◽  
pp. 64-70 ◽  
Author(s):  
Gustav Wägar
Keyword(s):  
Protein Synthesis ◽  
Actinomycin D ◽  
The Other ◽  
Leucine Incorporation ◽  
Short Term ◽  
Other Hand ◽  
Thyroid Glands ◽  
Translational Level ◽  
Stimulation Of

ABSTRACT Whether the short-term regulation of thyroidal protein synthesis by TSH occurs at the transcriptional or the translational level was tested by measuring the effect of actinomycin D (act D) on the TSH-induced stimulation of L-14C-leucine incorporation into the thyroidal proteins of rats. TSH was injected 6 h before the rats were killed. The thyroid glands were then removed and incubated in vitro in the presence of L-14C-leucine for 2 h. The pronounced stimulation of leucine incorporation in the TSH-treated animals was depressed as compared with controls but still significant even when the animals had been pre-treated with 100 μg act D 24 and 7 h before sacrifice. On the other hand, act D strongly decreased incorporation of 3H-uridine into RNA. Short-term regulation of thyroidal protein synthesis by TSH appears to be partly but not wholly dependent on neosynthesis of RNA. Hence regulation may partly occur at the translation level of protein synthesis.

Secondary biosynthesis of aflatoxin B1 in Aspergillus parasiticus

1970 ◽  
Vol 16 (10) ◽  
pp. 959-963 ◽  
Author(s):  
R. W. Detroy ◽  
C. W. Hesseltine
Keyword(s):  
Protein Synthesis ◽  
Aflatoxin B1 ◽  
Aspergillus Parasiticus ◽  
Synthetase Activity ◽  
Macromolecular Synthesis ◽  
High Concentration ◽  
Methyl Group ◽  
Aflatoxin B ◽  
Concentration Levels

The effect of two inhibitors on the formation of aflatoxin B1 synthetase activity in strain NRRL 2999 Aspergillus parasiticus has been studied. Aflatoxin B1 synthesizing activity was measured in vivo by incorporation of the 14C-methionine methyl group into aflatoxin B1. Cycloheximide at a concentration of 150 μg/ml blocks protein synthesis completely. If addition of cycloheximide is made before B1 synthetase appears, no activity accumulates; if added during accumulation, activity is frozen at the level reached at the time of addition. The cycloheximide effect is reversible since morphogenesis, total protein synthesis, and aflatoxin B1 synthetase activity all resume after removal of the inhibitor.DL-p-Fluorophenylalanine partially inhibits aflatoxin B1 synthesis in vivo; however, its effect upon macromolecular synthesis is incomplete even at high concentration levels. Once formed, the aflatoxin synthetase appears to maintain B1 synthesis when further protein synthesis is blocked; i.e., it is not rapidly degraded.

Use of an in Vitro Protein Synthesizing System to Test the Mode of Action of Chloracetamides

Weed Science ◽  
1980 ◽  
Vol 28 (3) ◽  
pp. 334-340 ◽  
Author(s):  
Luanne M. Deal ◽  
J. T. Reeves ◽  
B. A. Larkins ◽  
F. D. Hess
Keyword(s):  
Protein Synthesis ◽  
Sand Culture ◽  
Leucine Incorporation ◽  
Insoluble Protein ◽  
In Vitro System ◽  
A Cell ◽  
Protein Incorporation ◽  
Vitro Protein

The effects of chloracetamides on protein synthesis were studied both in vivo and in vitro. Four chloracetamide herbicides, alachlor [2-chloro-2′,6′-diethyl-N-(methoxymethyl)acetanilide], metolachlor [2-chloro-N-(2-ethyl-6-methylphenyl)-N-(2-methoxy-1-methylethyl)acetamide], CDAA (N–N-diallyl-2-chloroacetamide), and propachlor (2-chloro-N-isopropylacetanilide) were tested for inhibition of [3H]-leucine incorporation into protein. Incorporation of3H-leucine into trichloroacetic acid (TCA)-insoluble protein was inhibited in oat (Avena sativaL. ‘Victory’) seedlings grown in sand culture and treated 12 h at 1 × 10−4M with these chloracetamides. The herbicides were also tested in a cell-free protein synthesizing system containing polyribosomes purified from oat root cytoplasm. These herbicides had no effect on the rates of polypeptide elongation nor on the synthesis of specific polypeptides when herbicides (1 × 10−4M) were added directly to the system. Polypeptide formation was inhibited 89% when 1 × 10−4M cycloheximide was added during translation. Cytoplasmic polyribosomes were isolated from oat roots treated 12 h with 1 × 10−4M herbicide. Translation rates and products were not altered when these polyribosomes were added to the in vitro system. Protein synthesis is inhibited when tested in an in vivo system; however, the inhibition does not occur during the translation of mRNA into protein.

Activation of Protein Synthesis in Aging Potato Tuber Slices

1971 ◽  
Vol 26 (10) ◽  
pp. 1064-1067 ◽  
Author(s):  
Günter Kahl
Keyword(s):  
Protein Synthesis ◽  
Potato Tuber ◽  
Actinomycin D ◽  
Leucine Incorporation ◽  
Amino Acid Incorporation ◽  
Acid Incorporation ◽  
Fresh Potato ◽  
Polyuridylic Acid ◽  
Storage Organs

Whereas ribosome preparations of freshly sliced potato disks do not show appreciable activity in an in-vitro amino acid incorporation system, aging of the tissue leads to a greatly enhanced incorporation activity which reaches its maximum 24 hours after slicing. If ribosomes from freshly excised disks are provided with polyuridylic acid, their activity in the incorporation of phenylalanine is increased about 8 fold.Moreover, an RNA-fraction can be dissociated by EDTA from ribosomes of aged potato tuber slices, which sediments at 15 —18S, has a base composition different from that of 16S — rRNA, 5S-and 4S —RNA, and is not present on ribosomes of fresh slices. Its appearance is inhibited by actinomycin D and therefore most probably dependent on transcription. This compound, purified from sucrose gradients, enhances in vitro leucine incorporation into peptide material by ribosomes of fresh potato slices.The possibility is discussed that this fraction-among other factors-is responsible for the enhanced protein synthesis after slicing plant storage organs, and is indicative of a general derepression phenomenon in these tissues.

scholarly journals Intestinal Mucus Alters the Ability of Probiotic Bacteria To Bind Aflatoxin B1 In Vitro

2004 ◽  
Vol 70 (10) ◽  
pp. 6306-6308 ◽  
Author(s):  
S. Gratz ◽  
H. Mykkänen ◽  
A. C. Ouwehand ◽  
R. Juvonen ◽  
S. Salminen ◽  
...  
Keyword(s):  
Aflatoxin B1 ◽  
Probiotic Bacteria ◽  
Dependent Manner ◽  
Surface Binding ◽  
Intestinal Mucus ◽  
Aflatoxin B ◽  
Strain Dependent

ABSTRACT Several probiotics are known to bind aflatoxin B1 (AFB1) to their surfaces and to adhere to intestinal mucus. In this study, preincubation of two probiotic preparations with either AFB1 or mucus reduced the subsequent surface binding of mucus and AFB1, respectively, in a strain-dependent manner.

scholarly journals Phosphorylation and internalization of gp130 occur after IL-6 activation of Jak2 kinase in hepatocytes.

1994 ◽  
Vol 5 (7) ◽  
pp. 819-828 ◽  
Author(s):  
Y Wang ◽  
G M Fuller
Keyword(s):  
Protein Synthesis ◽  
Cell Surface ◽  
De Novo ◽  
Cell Types ◽  
Surface Expression ◽  
Cell Surface Expression ◽  
Protein Synthesis Inhibitors ◽  
Different Cell Types ◽  
Kinetics Of

Recent evidence has shown that members of the Jak kinase family are activated after IL-6 binds to its receptor complex, leading to a tyrosine phosphorylation of gp130, the IL-6 signal-transducing subunit. The different members of the IL-6 cytokine subfamily induce distinct patterns of Jak-Tyk phosphorylation in different cell types. Using monospecific antibodies to gp130, Jak2 kinase, and phosphotyrosine, we investigated the kinetics of IL-6 stimulation of members of this pathway in primary hepatocytes. Our findings show that Jak 2 is maximally activated within 2 min of exposure to IL-6, followed by gp130 phosphorylation that reaches its peak in another 2 min then declines to basal level by 60 min. In vitro phosphorylation experiments show that activated Jak 2 is able to phosphorylate both native gp130 and a fusion peptide containing its cytoplasmic domain, demonstrating gp130 is a direct substrate of Jak 2 kinase. Experiments designed to explore the cell surface expression of gp130 show that > or = 2 h are required to get a second round of phosphorylation after the addition of more cytokines. This finding suggests that activated gp130 is internalized from the cell surface after IL-6 stimulation. Additional experiments using protein synthesis inhibitors reveal that new protein synthesis is required to get a second cycle of gp130 phosphorylation indicating gp130 must be synthesized de novo and inserted into the membrane. These findings provide strong evidence that down regulation of the IL-6 signal in hepatocytes involves the internalization and cytosol degradation of gp130.

Properties of skeletal muscle mitochondria isolated from subsarcolemmal and intermyofibrillar regions

1993 ◽  
Vol 264 (2) ◽  
pp. C383-C389 ◽  
Author(s):  
A. M. Cogswell ◽  
R. J. Stevens ◽  
D. A. Hood
Keyword(s):  
Skeletal Muscle ◽  
Protein Synthesis ◽  
Mitochondrial Protein ◽  
Mitochondrial Fraction ◽  
Biochemical Properties ◽  
Leucine Incorporation ◽  
Mitochondrial Protein Synthesis ◽  
Skeletal Muscle Mitochondria ◽  
Mitochondrial Fractions

Two mitochondrial fractions, termed intermyofibrillar (IMF) and subsarcolemmal (SS), were isolated from skeletal muscle, and their biochemical properties were related to differences in respiration and mitochondrial protein synthesis. State III respiration was 2.3- to 2.8-fold greater in IMF than in SS mitochondria. Site 1 inhibition of respiration with rotenone reduced this difference to 1.4-fold. When sites 1 and 2 were inhibited with antimycin, the 1.4-fold differences remained. The activities of cytochrome-c oxidase (CYTOX) and succinate dehydrogenase (SDH) could account for some of these differences, since CYTOX was 20% greater (P < 0.05) in IMF mitochondria, and SDH was 40% greater (P < 0.05) in SS mitochondria. Cytochromes a, b, c, and c1 contents were similar in the two fractions. Cardiolipin (CL) content was higher (P < 0.05) in SS mitochondria, indicating a less dense mitochondrial fraction with respect to CL. In vitro [3H]leucine incorporation was 1.8-fold higher (P < 0.05) in IMF than in SS mitochondria. Thus compositional differences between IMF and SS fractions exist, perhaps representing mitochondria at different stages of biogenesis. The biochemical and functional differences could not solely be due to differences in mitochondrial protein synthesis but could also be due to nuclear-directed protein synthesis specific to each mitochondrial fraction.

Uptake of [3H]Uridine into Precursor Pools and RNA in Osteogenic Cells

1967 ◽  
Vol 2 (1) ◽  
pp. 39-56
Author(s):  
MAUREEN OWEN
Keyword(s):  
Protein Synthesis ◽  
Mammalian Cells ◽  
Rna Synthesis ◽  
Degradation Products ◽  
Cell Types ◽  
Water Soluble ◽  
Radioactive Label ◽  
Cytoplasmic Rna ◽  
Nuclear Rna

Young rabbits were given a single intraperitoneal injection of [3H]uridine. Using the technique of water-soluble autoradiography a study was made of the uptake of the radioactive label into soluble precursors and RNA in cells on an actively growing bone surface. Labelling of the soluble intracellular pools was immediate, but incorporation of label from these pools into RNA was not completed until 24 h after injection. At this time all the label in the sections was in RNA but this represented only 30% of the total label initially in the soluble pools. This means that 70% of the label is lost from the cell in the first 24 h either as degradation products of RNA synthesis or by other as yet unknown mechanisms. The pattern of labelling of the RNA was similar to that previously found for other mammalian cells in vivo or in vitro. There was a rapid uptake of label into nuclear RNA which reached a maximum by 2 h after injection and a slower uptake into cytoplasmic RNA which reached a maximum by 24 h after injection. There was a slow loss of label from the cells after 24 h indicating a half-life of about 8 days for this relatively stable RNA. A comparison was made of RNA synthesis in the proliferating preosteoblasts and the highly differentiated non-dividing osteoblasts. Labelling of the nuclear RNA for the two cell types was identical. The rate of labelling of the cytoplasmic RNA was similar for the two cell types but the maximum level of labelling in the cytoplasm of the osteoblasts was 2 to 3 times that in the preosteoblasts. This could be correlated with the more active protein synthesis by the osteoblasts. There was a slow loss of labelled RNA by the osteoblasts and preosteoblasts and a rapid loss by the osteocytes after the cells had been incorporated within the bone. It was suggested that this loss paralleled the decline in the rate of protein synthesis by the cells as their environment changed.

THYROXINE STIMULATION OF AMINO ACID INCORPORATION INTO MITOCHONDRIAL PROTEIN: DIFFERENCES BETWEEN IN VIVO AND IN VITRO EFFECTS

1973 ◽  
Vol 72 (4) ◽  
pp. 684-696 ◽  
Author(s):  
Amirav Gordon ◽  
Martin I. Surks ◽  
Jack H. Oppenheimer
Keyword(s):  
Protein Synthesis ◽  
Amino Acid ◽  
Mitochondrial Protein ◽  
Leucine Incorporation ◽  
Mitochondrial Protein Synthesis ◽  
Amino Acid Incorporation ◽  
Acid Incorporation ◽  
Stimulation Of

ABSTRACT The in vivo and in vitro stimulation of rat hepatic mitochondrial protein synthesis by thyroxine (T4) was compared. In confirmation of Buchanan & Tapley (1966). T4 added to isolated mitochondria rapidly stimulated [14C] leucine incorporation into mitochondrial protein. The in vitro stimulation was reversed after T4 was removed by incubating the mitochondria with bovine serum albumin (BSA). The decrease in T4 stimulation of protein synthesis appeared proportional to the T4 removed by BSA. Thus, it appears probable that exchangeable T4 controls the in vitro system. In contrast, the increase in mitochondrial protein synthesis which was observed 3 to 4 days after pretreatment of hypothyroid rats with labelled and non-radioactive T4 was not reversed by BSA treatment. Moreover, mitochondrial radioactivity could not be extracted with albumin. The in vivo phenomenon does not, therefore, appear to be related to exchangeable hormone in the mitochondria. Furthermore, the estimated quantity of T4 associated with mitochondria after in vivo stimulation was at least two orders of magnitude less than that required to produce comparable stimulation of mitochondrial protein synthesis in vitro. These findings strongly suggest that in vitro and in vivo stimulation of amino acid incorporation by T4 may be mediated by different biochemical mechanisms.

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