Stereoscopic back-scattered electron imaging of silver-stained proteins in nucleoli

By using simultaneously the AgNOR silver staining method, back-scattered electron imaging mode and stereo-tilt in scanning electron microscopy (SEM), it is possible to observe the nucleus through the cell surface, the nucleolus, and the tri-dimensional distribution of the AgNOR-associated acidic proteins. In C3H10T1:2 cells and their 7-12-dimethylbenz--anthracene-treated transformants, the staining demonstrates several intranucleolar silver-staining granules (SSG), surrounded by a weakly staining region. The SSG may represent the fibrillar center (FC) and the weakly staining region, the fibrillar dense component (FD). This component can link several SSG together to form a rope-like structure. In cells with no visible nucleolus and inactive nucleolar organizer regions (NORs) the silver-staining granules are less numerous, close together and the presumed fibrillar dense components are not visible. The SSG are located more peripheraly, and the weakly staining region and the rope-like structure are less prominent in control cell nucleoli than in transformed cells with a comparatively high rate of RNA synthesis.     

Isoproterenol induces a ribosomal modification in the heart and the skeletal muscle of hamsters

The protein synthesis activity of heart, skeletal muscle and liver polysomes from isoprotenerol-treated and control hamsters has been compared in an in vitro non-inititating translation system. Heart and skeletal muscle polysomes from treated hamsters were less active than controls and required a higher magnesium concentration for optimal protein synthesis. These results suggest that there is a conformational modification in heart and skeletal muscle ribosomes from isoprotenerol-treated hamsters. No such change was observed with ribosomes from the liver of isoproterenol-treated hamsters.     

Study model of the mid-range ecotoxicity of chemical substances

Semi-chronic ecotoxic effects are measured by studying mortality and reproduction of daphnids, grown in toxic medium and fed with toxic chlorellae during 4 weeks. Toxic biosorption is controlled analytically. Toxicity of Cd++ at a concentration of an order of magnitude of by liter is revealed after 14 days. Cr6+ appears non toxic at concentrations under 25 micrograms/l.     

The majority of independently transformed BHK cell clones share a single functional lesion which determines anchorage independence and influences tumorigenicity

Summary Expression of the anchorage-independent transformed phenotype in BHK 21/13 cells generally behaves as a recessive trait. When chemically induced and spontaneously arising transformants are fused to the nontransformed parent line, transformation is initially suppressed, reappearing after extended growth of the hybrids. In this paper, complementation for the expression of anchorage independence was sought among a large group of such transformants, all independently derived from BHK 21/13 cells. Tumorigenicity studies on selected hybrids and parental lines indicated that the in vitro trait of anchorage independence is an accurate indicator of in vivo neoplasia for these cells. Seventeen of the 18 clones tested did not complement one or more of three tester strains. This result indicates that anchorage independence arose in these clones as a result of lesions in the same genetic function and suggests that the final step in the progressive changes of carcinogenesis may frequently be restricted to lesions at a single locus. This investigation was supported by National Institutes of Health grant CA27306.     

Operation of the glycolate pathway in isolated bundle sheath strands of maize and Panicum maximum

Operation of the glycolate pathway in isolated bundle sheath (BS) strands of two C₄ species was demonstrated from ¹⁴C incorporation into two intermediates, glycine and serine, under conditions favourable for photorespiratory activity. Isolated BS strands fixing ¹⁴CO₂ under light at physiological rates incorporate respectively 3% (Zea mays L., cv. INRA 258) and 7% (Panicum maximum Jacq.) of total ¹⁴C fixed into glycine + serine, at low bicarbonate levels (less than the Km for CO₂ fixation, 0.8 mM). Higher bicarbonate concentrations depressed the percentage of incorporation into the two amino acids. No labelling was observed in the absence of added glutamate. Oxygen was required for glycine + serine labelling, since ¹⁴C incorporation into glycine was largely depressed by argon flushing, and labelling of the two amino acids was nearly suppressed by the addition of the strong reductant, dithionite, especially in maize. Two inhibitors of the glycolate pathway were tested. With α-hydroxypyridine-methanesulfonic acid, an inhibitor of glycolate oxidase, labelling of glycine and serine remained minimal whereas glycolate was accumulated. Isoniazid, an inhibitor of the transformation of glycine to serine induced a 50% increased labelling of glycine in maize BS, and a large decrease in serine labelling. In Panicum, the increase in [¹⁴C]-glycine was 90%. These results suggest that the pathway glycolate → glycine → serine operates in these plants. However, leakage of metabolites occurs in BS cells, especially in maize and a large part of newly formed glycolate, glycine and serine is exported out of the cells. Operation of ribulose-1,5-bisphosphate oxygenase activity in competition with ribulose-1,5-bisphosphate carboxylase is demonstrated by the lowering of total ¹⁴CO₂ fixation when O₂ is increased at low bicarbonate concentration. An interesting feature observed in maize BS, at low bicarbonate concentration, was an increase in ribulose-1,5-bisphosphate labelling when the O₂ level was decreased. This was accompanied by an increase in CO₂ fixation. This could indicate an increased rate in synthesis of ribulose-1,5-bisphosphate (which accumulated) due to a stimulation of ATP synthesis by cyclic photophosphorylation under anaerobic conditions.     

Inorganic pyrophosphatase activity of the mouse spleen in the immune response and after treatment with bis-phosphonates

The inorganic pyrophosphatase activity was determined in different tissues of mice. The immunization of mice by sheep erythrocytes increased the inorganic pyrophosphatase activity of the spleen. The in vivo administration of bisphosphonates (40 mg per 1 g of mass), which are structural analogs of inorganic pyrophosphate (methylene bisphosphonic acid--MBPA, hydroxyethylidene bisphosphonic acid--HEBPA and aminomethylene bisphosphonic acid--AMBPA), inhibited the inorganic pyrophosphatase activity only by MBPA in the thymus and spleen but not in liver. The addition of MBPA, HEBPA as well as of phosphonoacetic acid, imidobisphosphate, bis(phosphonomethyl)-phosphonic acid, MBPA and phosphoric acid monoanhydride to cytosol from the mouse spleen led to the competitive (relative to the [Mg (PPi)2-] complex) inhibition of the inorganic pyrophosphatase activity. AMBPA didn't possess the analogous effect.     

Clastogen-induced chromosomal breakage as a marker for first trimester prenatal diagnosis of Fanconi anemia

Summary Using cultured trophoblast cells obtained by chorionic villus biopsy, we diagnosed Fanconi anemia (FA) in two pregnancies and excluded it in eight pregnancies at risk for the syndrome. Baseline chromosomal breakage and breakage induced by diepoxybutane (DEB) were analyzed. Increased breakage was used as a marker for the syndrome. Our results were unambiguous and provide a reliable method for prenatal detection of FA in the first trimester of pregnancy.