Charles Frayssinet

Induction of DNA single-strand breaks by T2 toxin, a trichothecene metabolite of Fusarium: Effect on lymphoid organs and liver

T2 toxin, a trichothecene metabolite produced by Fusarium species, contaminates cereals harvested and stored under damp and cold conditions. These substances are responsible for Alimentary Toxic Aleukia (ATA), a severe human disease, and numerous animal intoxications. The action of T2 toxin on DNA was studied by using Parodis alkaline elution technique coupled with a microfluorimetric determination of DNA. In vivo the effect of the toxin was studied on liver, spleen and thymus, and in vitro on a primary culture of rat hepatocytes and on splenic and thymic lymphocytes stimulated by PHA. Under our experimental conditions, in vivo and in vitro, no damage was observed for the hepatic DNA. By contrast, the DNA of lymphoid organs was severely damaged by the toxin. In vitro, T2 toxin induced severe damage to the DNA molecule with low concentrations (5 ng/ml culture) and for short exposure (2 h). In vivo, a moderate amount of DNA breaks was observed in splenic and thymic lymphocytes 3 h after administration of T2 toxin to mice (3 mg/kg). Reversibility occurred 24 h later under these conditions in vivo, indicating DNA repair. The results agree with the preferential cytotoxicity of T2 toxin for lymphoid cells. The relation between DNA damage, mutagenicity and carcinogenic properties of T2 toxin is discussed.

Protein synthesis inhibition and cardiac lesions associated with deoxynivalenol ingestion in mice.

Deoxynivalenol (DON), an occasional contaminant of foodstuffs, has been implicated in outbreaks of mycotoxicosis. Balb-c mice that had ingested 0.35 mg/kg of DON showed a drastic decrease in food intake and concomitant loss of weight. Severe depletion of the lymphoid organs and liver were also observed. Cardiac lesions, appeared as calcified pericarditis foci in young animals fed a diet contaminated by 10 to 20 ppm of DON for a period of a few weeks. DON inhibited protein synthesis. This inhibition occurred at lower doses for the heart than for the other organs. This preferential effect on cardiac tissue correlated with the cardiotoxicity observed.

Increased endotoxin sensitivity following T-2 toxin treatment is associated with increased absorption of endotoxin

Oral exposure to T-2 Toxin (T-2) in experimental animals results in a syndrome similar to that observed in endotoxemia. Endotoxins are lipopolysaccharide, outer-membrane components of gram-negative bacteria which induce acute, inflammatory responses. In the present study, several aspects of endotoxin pathophysiology were investigated in mice following simultaneous exposure to T-2 and endotoxin, including mortality, hypothermia, tumor necrosis factor-alpha (TNF-alpha) and corticosterone production, and thymic weight. The disposition of endotoxin was also assessed, Acute, simultaneous exposure to T-2 (4 mg/kg, po) and endotoxin (3 micrograms/mouse, ip) resulted in increased mortality, hypothermia, TNF-alpha production, and thymic atrophy compared to treatment with either T-2 of endotoxin alone. Pretreatment of mice with endotoxin, a regime that renders the animals resistant to the effects of endotoxin, reduced many endotoxin effects in animals treated simultaneously with T-2 and endotoxin. Upon further investigation, it was observed that T-2 increased the absorption rate of endotoxin: as the peak height of serum endotoxin increased, the time-to-peak decreased, and the area under the curve was unchanged in animals treated simultaneously with T-2 and endotoxin. It was concluded that increased endotoxin absorption accounted for the increases in mortality, hypothermia, and TNF-alpha associated with T-2 exposure.

Effect of ammoniation on the carcinogenicity of aflatoxin‐contaminated groundnut oil cakes: Long‐term feeding study in the rat

The efficacy of detoxication by ammoniation of aflatoxin-contaminated groundnut oil cakes was determined in long-term (18 months) feeding experiments with rats. The aflatoxin content of the cake was reduced very considerably by the pressurized application of ammonia, dropping from 1000 to 140 ppb at a gas pressure of 2 bar and to 60 ppb at 3 bar. No reversion was noted during the experiment. The percentage of hepatic tumours obtained was very high for the untreated cakes, but fell sharply with medium treatment and was reduced to zero by the treatment at 3 bar. A satisfactory dose-effect relationship was shown between the residual aflatoxin content of the cakes and the observed incidence of tumours. The results show that ammonia treatment is a practical solution to the problem of the carcinogenic potency of contaminated oil cakes.

USE OF CELL CULTURES FOR PREDICTING THE BIOLOGICAL EFFECTS OF MYCOTOXINS

The risk presented by mycotoxins is a toxicological problem. As the data given by physico-chemical analysis may be difficult to translate in terms of toxicity, however, especially when considering multiple contamination, we have developed a system for toxicological analysis of mycotoxins using cell cultures of different origins. The response of several cell types to a number of well defined mycotoxins was obtained in three days. This approach allowed us to: demonstrate and quantify a toxic effect, define some organ specificity related to the preferential action on a particular cell type, and detect an immunosuppressive effect. The results indicate that the system can be used for toxicological screening and that it has a predictive value for the pathological effects of tested products.

Immunosuppressive effects of four trichothecene mycotoxins.

The trichothecenes are a family of mycotoxins contaminating some food and feeds. We have studied the inhibitory effect of four of them including less toxic compounds: T-2 toxin, DAS, DON and Trichodermin, on normal human peripheral blood lymphocytes and murine splenic lymphocytes. The results show that even for the least toxic compound, inhibiting concentrations can be realised in the blood by common alimentary contamination.

Evolution of ornithine decarboxylase activity during the cell cycle of Euglena gracilis Z in synchronous culture Influence of vitamin B-12

Ornithine decarboxylase activity in Euglena gracilis Z was studied during the normal cell cycle and in vitamin B-12 deficiency. The cells were synchronized by means of alternating periods of light and dark. During the normal cell cycle, ornithine decarboxylase activity was very weak in the dark period, while three peaks of activity were recognized in the light period. The first peak, in the G1 phase, occurred when luminous stimulation started; the second preceded the S phase and the third was found in G2. In B-12-deficient cells, ornithine decarboxylase activity was greatly decreased and only the first peak remained. Elimination of the deficiency by addition of vitamin B-12 to the medium induced a very fast and significant increase in ornithine decarboxylase activity.

Use of human lymphoblastoid cells to detect the toxic effect of chloramphenicol and metabolites possibly involved in aplastic anemia in man.

Some Chloramphenicol (CAP) metabolites are suspected to be involved in the etiology of bone marrow aplasia in man. The objective of the present study was to investigate the cytotoxicity as well as the genotoxicity of CAP and six of its metabolites on human bone marrow cells (RiBM cells) and to compare these results with those obtained on human peripheral blood lymphocytes in order to estimate the relative sensitivity of the two types of cells. Three CAP metabolites NO-CAP, DH-CAP and NPAP inhibited 3H thymidine incorporation in RiBM cells at concentrations ranging from 2.10(-5) M to 2.10(-4) M. NO-CAP appeared as the most potent cytotoxic compound. CAP itself and NAPD presented some toxic effect at high concentration (1-2.10(-3) M). CAPG and HAP did not present any cytotoxic effect. By comparison, the response of human lymphocytes to CAP and its metabolites showed a similar pattern but DH-CAP was the most inhibitory compound. Concerning the genotoxic potential, NO-CAP and DH-CAP induced DNA single strand breaks in RiBM cells at concentrations of 1 and 2.10(-4) M with a dose response relationship. CAP and other metabolites were completely devoid of genotoxicity up to 4.10(-3) M. The results clearly showed that RiBM cells were much less susceptible to the genotoxic effect of CAP metabolites than human lymphocytes.

Cytotoxicity and DNA damaging potency of chloramphenicol and six metabolites: a new evaluation in human lymphocytes and Raji cells

Chloramphenicol (CAP) is an antibiotic which has been implicated in the etiology of aplastic anemia in man. This product is also used in veterinary medicine. The medical use of chloramphenicol has been limited to cases where the drug is indispensible but veterinary use may lead to the presence of residues in the meat of treated animals and it is essential to establish acceptable levels of intake of such residues in order to protect human health. CAP is metabolized into at least 6 metabolites: nitroso-CAP (NO-CAP), formed in the liver, 3 excretion products: the glucuronide (CAP-G), the CAP base (NAPD), and an alcoholic derivative, HAP. Dehydro-CAP (DH-CAP) and the dehydro-CAP base (NPAP) are formed by enterobacteria in the large bowel. The objective of the present study was to investigate (1) the cytotoxicity of CAP and its metabolites and (2) their ability to induce DNA damage in human cells. This work was performed with human peripheral blood lymphocytes (PBL) and with a lymphoma cell line (Raji).

Synthesis and biological activity of a fluoroketonucleoside: 7-(3-deoxy-3-fluoro-β-d-glycero-hex-2-enopyranosyl-4-ulose)theophylline

Abstract The title compound 5 was synthesized by direct oxidation of the fluoro precursor and not from an epoxyketo intermediate according to the Paulsen procedure, which has been used until now to obtain bromo- and chloro-ketonucleosides. Antineoplastic activity and immunosuppressive effect of 5 were studied on murin splenic lymphocytes (steady state or stimulated by PHA), and on RAJI and DAUDI cells, and compared with the activity of 3-(3-deoxy-β- d -glycero-hex-2-enopyranosyl-4-ulose)thymine. IC50 values showed that the fluoroderivative 5 has a higher antineoplastic activity and a lower immunosuppressive effect.