Superoxide production by polymorphonuclear leukocytes

Summary Phagocytosis by polymorphonuclear leukocytes triggers a burst of oxidative metabolism resulting in hydrogen peroxide and superoxide production, and these active oxygen species function in the killing of microorganisms. A new cytochemical technique, based on a manganese dependent diaminobenzidine oxidation, has been developed to detect superoxide in these cells. It has been shown that superoxide generation is associated with the plasma membrane in cells activated by particulate (zymosan) and nonparticulate (phorbol myristate acetate) stimuli. This membraned activity is maintained during invagination such that reduced oxygen is generated within the endocytic vacuoles. Reaction product is absent from unstimulated cells; additionally, formation of precipitate is blocked by omission of Mn⁺⁺, low temperature, glutaraldehyde prefixation, and the presence of superoxide dismutase in the incubation medium.In honour of Prof. P. van Duijn     

Ultrastructural detection of the onset of pituitary thyrotroph sensitivity to lowered thyroid hormone concentrations in the fetal sheep

The goitrogen methylthiouracil was administered orally to pregnant ewes of known gestational ages to induce hypothyroidism in both mother and fetus. Developing pituitary thyrotrophic cells were studied using electron microscopy to detect the earliest gestational age at which morphological changes occurred in response to lowered plasma thyroid hormone concentrations. At 50 days of gestation, the pituitaries of fetuses exposed to the goitrogen were indistinguishable from untreated control glands. However, at 58 days and subsequent ages, "thyroidectomy' cells were observed in pituitaries of all hypothyroid fetuses. These findings indicate that fetal sheep pituitary thyrotrophs are sensitive to lowered thyroid hormone concentrations by 58 days of gestation, suggesting that thyroid-thyrotroph interaction exists at this early stage of development.     

The catabolism of plasma glycoproteins in normal and injured rats

The catabolism of (14)C-labelled plasma glycoprotein in rats was studied after injecting homologous plasma protein labelled in the N-acetylglucosamine and sialic acid moieties. In normal animals the catabolism was approximately described by a four-compartment model. The fractional rate of catabolism of the plasma-protein amino sugar was found to be 0.0305hr.(-1), corresponding to the degradation of 2.75mumoles/hr. The (14)C label was eliminated from the animals largely as carbon dioxide with a small proportion appearing in the urine. Freely circulating amino sugars or glycopeptides did not appear in the plasma as a result of the catabolic processes, and there was no evidence that the protein-bound amino sugars were reutilized in biosynthetic processes. A study of the distribution of (14)C label in the carcasses of animals 24hr. after injection provided evidence that the gastrointestinal tract accounted for 25-38% of the total catabolic pool; the lungs, kidneys, spleen and liver also appeared to contribute to catabolism. Studies were conducted with rats that had been treated with turpentine to induce an inflammatory reaction; the results could not be analysed kinetically, since the metabolism of plasma proteins in these animals did not appear to be in a steady state. The injected plasma protein disappeared from the intravascular pool more quickly than in normal animals, but there were no significant differences in the rates of excretion of the (14)C label.     

Nitrogenase activity in extracts of heterocystous and non-heterocystous blue-green algae

Summary Cell-free extracts capable of acetylene reduction and cyanide reduction have been prepared from heterocystous (Anabaena cylindrica) and non-heterocystous (Plectonema boryanum 594) blue-green algae. Extracts from Anabaena were obtained from cultures grown in blulk under aerobic conditions, while the Plectonema cultures were grown in bulk on nitrate-nitrogen, then washed free from nitrate and sparged with A/CO₂ for 40 h after which time maximum nitrogenase activity was detected. The nitrogenases of both algae are similar and resemble in many respects nitrogenases from bacteria and legumes. Activity is located primarily in a 40,000xgx15 min supernatant fraction and the rate of C₂H₂ reduction observed is about 10 per cent of whole cell activity. ATP and a source of reducing power (Na₂S₂O₄) are required for efficient functioning of the enzyme. ATP-dependent hydrogen evolution occurs, the extracts are cold labile and highly sensitive to oxygen and the oxygen inhibition is irreversible.     

In vitro differentiation of rabbit blastocyst cells

Summary Six- and seven-day post-coitus (p.c.) rabbit embryos have been cultured in an attempt to establish a trophectodermal cell line. Results indicate that cells with epithelial characteristics (i.e. positive staining for cytokeratin) will survive in culture until Passage 3. At that time a fibroblastlike cell becomes predominant. In addition, we have found that the presence of the inner cell mass is required for embryo explants often results in the development of cells that spontaneously contract. These cells stain positively for myosin, which indicates that they may be precardiac cells. Maximum diastolic potential was −59±1.2 mV and the threshold potential was −53±2.3 mV. Spontaneously contracting cells did not respond to atropine, acetylcholine, epinephrine, isoproterenol, or propranolol. Action potential seems to be a result of an inward calcium current, because the beating rate is decreased in a dose-related manner with the calcium channel blocker verapamil, whereas the voltage-sensitive sodium channel blocker tetrodotoxin was without effect. This work was supported by grants HD21302, HD07069, DK31091, and HL37320 from the National Institutes of Health, Bethesda, MD, with additional support from a University of Alabama at Birmingham Cardviovascular Research and Training Center Award.