Quantitative study of natural antioxidant systems for cellular nitrofurantoin toxicity

The toxicity of nitrofurantoin was studied on human WI-38 fibroblasts: this chemical was lethal when added at concentrations higher than 5·10⁻⁵ M in the culture medium. The protection afforded by anitoxidants was then tested: α-tocopherol gave at 10⁻⁴ M a light protection in contrast to ascorbic acid which even became toxic at high concentrations. We also tested catalase, superoxide dismutase and glutathione peroxidase introduced intracellularly by the microinjection technique. On a molecular basis, glutathione peroxidase was 23-times more efficient than catalase and 3000-times more than superoxide dismutase. The results also showed that a similar range of enzyme concentrations was found for the protection against high oxygen pressure. This suggests that, in the case of both oxygen and nitrofurantoin toxicity, the peroxide derivatives are the most toxic intermediates of the free radical attacks.     

Intergenerational effect of an adverse intrauterine environment on perturbation of glucose metabolism.

Human epidemiological and animal studies have revealed the late consequences of malnutrition during gestation and early life on the health of the offspring. These studies have highlighted the inverse relationship between birth weight and the incidence of insulin resistance and type 2 diabetes later in life. The aim of this paper is to review the different means of achieving foetal malnutrition and its consequences even for a next generation, in animal models and to identify key area for further research. We address the impact of two models of maternal malnutrition (protein restriction and caloric restriction) as well as the impact of maternal diabetes, the three maternal conditions leading to perturbed foetal nutritional environment. Particular emphasis is given to the endocrine pancreas and the insulin sensitive tissues. More specifically, alterations of the foetal nutritional environment perturb the development of the endocrine pancreas and target the ss cell mass at birth. Some adaptations later in life may take place but stress situations such as pregnancy and ageing precipitate the animals to glucose intolerance and insulin resistance. Even the next generation features alterations in the development of the endocrine pancreas. Some mechanisms by which the foetal ss cell mass is altered are approached in this review and specific attention is paid to the amino acid profile. The preventive role of taurine is discussed.     

Comparative study of the enzymatic defense systems against oxygen-derived free radicals: the key role of glutathione peroxidase

Human WI-38 diploid fibroblasts have been cultivated under high toxic O2 pressure, and their survival curves are reported. Superoxide dismutase, catalase, or glutathione peroxidase provided some protection when injected in the cells exposed to O2. This protective effect, recorded after 3 or 4 days of incubation, was the most pronounced when cells were injected just before oxygen exposure. Quantitative injection assays have been performed for the three enzymes. Surprisingly, glutathione peroxidase was found to be much more effective than both catalase and superoxide dismutase, the latter being particularly inefficient.     

Inhibition of platelet-activating factor biosynthesis via the acetyltransferase by arachidonic and oleic acids in ionophore A23187-stimulated bovine neutrophils

Platelet-activating factor (PAF) is a phospholipid mediator of inflammation and allergy that is synthesized by several inflammatory cells including neutrophils. Addition of exogenous arachidonic acid to ionophore A23187-stimulated bovine neutrophils led to the inhibition of PAF biosynthesis assayed by incorporation of [3H]acetate into PAF and by bioassay; under the same conditions, leukotriene B4 (LTB4) formation was not decreased. The activities of the PAF metabolism enzymes indicated that the PAF synthesis inhibition by arachidonic acid is mediated via the acetyltransferase inhibition which is the last enzyme of the PAF formation. Another unsaturated fatty acid, oleic acid, exhibited the same inhibitory effect on [3H]acetate-PAF formation; however, the saturated stearic acid did not lead to any inhibition. These findings suggest that liberation of unsaturated fatty acids from membrane phospholipids, as a consequence of phospholipase A2 activation, would modulate PAF formation via inhibition of the acetyltransferase. In addition, the utilization of arachidonic acid oleic acids in activated neutrophils furnishes an easy means of blocking PAF synthesis in order to understand the role of this mediator in cellular processes.     

Comparative study of conjugation between horseradish peroxidase and D-cytochrome b5. Incorporation into subcellular membranes

Horseradish peroxidase was conjugated to D-cytochrome b5 by three different two-step methods. The yield of conjugates based on the peroxidase enzymatic activity recovered after gel filtration was very low in the glutaraldehyde method, but higher in the N-succinimidyl 3-(2-pyridyldithio)propionate (SPDP) and periodate methods. The molecular size of the conjugates was analyzed by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. Monomeric conjugates were mostly formed via the glutaraldehyde and SPDP methods in the presence of appropriate molar ratios of proteins. Most of the conjugates formed via the periodate method were polymers. The conjugate preparations of the three methods could be incorporated into microsomal membranes. Conjugate polymers, however, appeared less able to be incorporated then monomers. There was a nonpreferential incorporation of free or conjugated D-cytochrome b5 contained in the conjugate preparation of the glutaraldehyde method. In conclusion, this study gives preference to the glutaraldehyde method for the preparation of conjugates that will subsequently be used as an in vivo marker of the D-cytochrome b5 incorporation into membranes.     

The stroma-vascular fraction of rat inguinal and epididymal adipose tissue and the adipoconversion of fat cell precursors in primary culture

The stroma-vascular fraction (SVF) of inguinal and epididymal fat pads of 4 week-old rats was studied by electron microscopy. Among the various cell types, endothelial cells and preadipocytes were found in both SVF, while mesothelial cells were only detected in the epididymal SVF. The resulting heterogeneity of primary culture and the adipoconversion of the fat cell precursors were studied in a serum-supplemented medium enriched with insulin (14.5 nM) and exogenous triglycerides. Despite the heterogeneity of the inoculum, the primary cultures were rather homogeneous, fat cell precursors being the main cell type. Distinctive contaminant fibroblast-like cells were observed in both cultures, whereas epithelial-like cells, which correspond most probably to mesothelial cells, were only found in epididymal cultures. Differentiation of fat cell precursors was assessed by the appearance of lipoprotein lipase (LPL) and glycerol-3-phosphate dehydrogenase (GPDH). LPL activity was found in the same level in cells of both deposits while GPDH activity was elevated in inguinal vs epididymal derived stroma-vascular cells. The different adipose conversion pattern of both cultures was confirmed by morphological quantification: the maturation of epididymal fat cell precursors was faster but less extensive. These differences could be related mainly to regional localization rather than to different maturation of the two fat deposits.     

Mutations affecting the mitochondrial genes encoding the cytochrome oxidase subunit I and apocytochrome b of Chlamydomonas reinhardtii

Mitochondrial mutants of the green alga Chlamydomonas reinhardtii that are inactivated in the cytochrome pathway of respiration have previously been isolated. Despite the fact that the alternative oxidase pathway is still active the mutants have lost the capacity to grow heterotrophically (dark + acetate) and display reduced growth under mixotrophic conditions (light + acetate). In crosses between wild-type and mutant cells, the meiotic progeny only inherit the character transmitted by the mt ^– parent, which indicates that the mutations are located in the 15.8 kb linear mitochondrial genome. Two new mutants (dum-18 and dum-19) have now been isolated and characterized genetically, biochemically and at the molecular level. In addition, two previously isolated mutants (dum-11 and dum-15) were characterized in more detail. dum-11 contains two types of deleted mitochondrial DNA molecules: 15.1 kb monomers lacking the subterminal part of the genome, downstream of codon 147 of the apocytochrome b (COB) gene, and dimers resulting from head-to-head fusion of asymmetrically deleted monomers (15.1 and 9.5 kb DNA molecules, respectively). As in the wild type, the three other mutants contain only 15.8 kb mitochondrial DNA molecules. dum-15 is mutated at codon 140 of the COB gene, a serine (TCT) being changed into a tyrosine (TAC). dum-18 and dum-19 both inactivate cytochrome c oxidase, as a result of frameshift mutations (addition or deletion of 1 bp) at codons 145 and 152, respectively, of the COX1 gene encoding subunit I of cytochrome c oxidase. In a total of ten respiratory deficient mitochondrial mutants characterized thus far, only mutations located in COB or COXI have been isolated. The possibility that the inactivation of the other mitochondrial genes is lethal for the cells is discussed.     

The binding of cytochrome b5 to plasma membranes of rat liver. Its implication for membrane specificity and biogenesis

The in vitro incorporation of cytochrome $\text{b}{}_5$ into purified plasma membranes was investigated by biochemical and immunological methods. Plasma membrane preparations incorporated three times less cytochrome $\text{b}{}_5$ than did microsomal preparations; 60% of this cytochrome $\text{b}{}_5$ could not be reduced by the NADH-cytochrome $\text{b}{}_5$ reductase and was considered as being bound to the plasma membrane. The morphological observations made after the immunochemical labeling of cytochrome $\text{b}{}_5$ clearly showed a good but asymmetrical distribution of the ferritin labeling: only the inner face of the plasma membrane incorporated cytochrome $\text{b}{}_5$. These results are discussed with respect to theories which concern the subcellular membrane relationships in the cell.     

Gonads ofCarassius auratus (teleosts) in organ culture: A new technique and the effects of testosterone

Summary The use of a semi-natural medium (Eagle’s MEM supplemented with chicken embryo extract) appears to be satisfactory for ovarian and testicular fragment cultures ofCarassius auratus. The male germ cells show a normal degeneration which might be attributed to a lack of pituitary hormones: gonadotropic and, possibly, somatotropic hormone. The ovarian fragments maintain a normal organization during 21 days of culture. Only the occytes in late vitellogenesis massively atresiate at the beginning of culture. The addition of fetal bovine serum to this medium produces surprising effects on the testicular explants, particularly on the spermatogonial mitoses. This might be attributed to the presence of STH and androgen in the serum. The modifications of ovarian explants, on the other hand, are identical to those observed with Eagle’s MEM supplemented with chicken embryo extract. M 199 has been tested as a synthetic medium; the preliminary results are encouraging. Testosterone, when added to the semi-natural medium, permits complete spermatogenesis in the testicular fragments during the 21 days of culture, but does not produce any effect on the ovarian explants.     

Variability in host plant resistance of Sorghum to Striga Hermonthica infestation in West Africa

Fourteen elite sorghum lines were evaluated for their resistance to Striga hermonthica at three locations in Nigeria and Mali. Results showed that many of the lines especially MALISOR 84-1, SAMSORG 41, 97-SB-F5DT-64 (Keninkédié) and the check SRN 39 remained resistant to Striga in all locations with low emerged Striga counts, while SAMSORG 14 had the highest Striga infestation in all locations. Considerable variation in reaction to Striga infestation was observed on Séguètana, 97-SB-F5DT-63 (Wasa), 97-SB-F5DT-65, CMDT 38, CMDT 39 and CMDT 45 which were susceptible to Striga at Samaru, Nigeria but were resistant to Striga at both locations in Mali. Based on low Striga resistance and high grain yield, lines MALISOR 84-1, SAMSORG 41, 97-SB-F5DT-64, 97-SB-F5DT-65, CMDT 39 and SAMSORT 14 have been nominated for wider evaluation across more West African countries.