Phosphoinositide hydrolysis in mitogen-stimulated human peripheral-blood T lymphocytes.

Both phytohaemagglutinin and antibodies to the CD3 molecule induced proliferation and phosphoinositide hydrolysis in human peripheral-blood T lymphocytes, but the magnitude of the inositol phosphate response was small and the rate of accumulation slow [significant increases in Ins(1,4,5)P3 were observed only after 10 min]. Hence this response differs from the well-characterized Ins(1,4,5)P3 responses of many other systems. This slow response, its abrogation in Ca2+-depleted medium, the slow and maintained increase in Ca2+ as measured by Quin-2, and the ability of the Ca2+ ionophore A23187 to stimulate Ins(1,4,5)P3 accumulation all suggest that the increase in Ins(1,4,5)P3 occurs, at least in part, as a result of receptor-mediated Ca2+ influx in mitogen-stimulated T lymphocytes.     

Electric shock-mediated transfection of cells. Characterization and optimization of electrical parameters.

The effect of various parameters on the electric shock-mediated permeabilization and transfection of CHO cells has been investigated. Up to 70% of the cells can be maintained transiently permeable to erythrosin B for periods of at least 1 h at 20 degrees C. Electrical conditions optimal for transient permeabilization were also optimal for efficient DNA transfection by pSV2neo. However, the DNA must be present during exposure to the electric field for efficient transformation. The same requirement existed for voltage-induced DNA toxicity. The results suggest that DNA moves into the cells by electrophoresis, not by simple diffusion. Based on these observations a simple, rapid procedure for optimizing the conditions for electric shock-mediated DNA transfer into cells has been developed.     

Failure to detect platelet-activating factor using the splenectomized mouse bioassay

The ability of purified preparations of platelet-activating factor (PAF), from three different suppliers, to induce thrombocytopaenia in mice after splenectomy and to activate mouse platelets in vitro was examined. Although the PAF preparations were potent activators of horse and cow platelets in vitro, injections of up to 1 microgram PAF failed to elicit thrombocytopaenia responses in either CD1 or Swiss Webster random-bred mice. However, when thrombin was injected into Swiss Webster mice, a dose-dependent decrease in the concentration of platelets was observed. Furthermore, isolated platelets from these strains and from 3 inbred lines (C3H/He, BALB/c, C57BL/6) of mice, were not aggregated by PAF in vitro but were sensitive to adenosine diphosphate and thrombin. No change in circulating platelet concentrations was observed over the initial 7 days of gestation in intact Swiss Webster and C57BL/6 or splenectomized C57BL/6 mice, suggesting either an absence of PAF production during early pregnancy in these strains or insensitivity of their platelets to PAF. These results suggest that many mouse strains are unsuitable for the bioassay of PAF.     

Triploidy induction in Nile tilapia,Oreochromis niloticus L. using pressure,heat and cold shocks

Summary The results of a study aimed at the identification of treatment optima for triploidy induction in recently fertilised Oreochromis niloticus L. eggs by altering the intensity, duration and timing of application of pressure, heat and cold shocks are reported. Preliminary, but not directly comparable, trials suggested the following treatments to be close to the individual agent optima. Pressure: 8,000 psi 2-min duration applied 9 min after fertilisation (a.f.); heat: 41 °C, 3.5-min duration applied 5 min a.f., cold: 9°C, 30-min duration applied 7 min a.f. In a directly comparable trial in which the eggs of eight different females were separately exposed to the optimum shocks listed above, individual triploid yields were more variable following cold shocks and mean triploid yields were, therefore, higher following pressure and heat shock. These and other results obtained are presented and the light they shed on the timing of the second meiotic division in this species is discussed.     

Human serum does contain a high molecular weight hepatocyte growth factor: studies pre- and post-hepatic resection

Levels of a high molecular weight hepatotrophin were measured in human serum taken from patients before and 24 hours after undergoing major hepatic resection. In in-vitro rat hepatocyte cultures a 'hepatotrophin' enriched fraction of human serum induced the incorporation of tritiated thymidine into DNA in both pre and post-operative patients. Levels after hepatic resection were 2-3 fold higher than those achieved at the same protein concentration before operation in the same patient. The hepatotrophic factor had an apparent molecular weight of approximately 150,000 daltons, and was an anionic protein.     

Evidence for a pool of non-recycling transferrin receptors in peripheral sheep reticulocytes

Sheep reticulocytes from phlebotomized animals have a total transferrin binding potential that may exceed by an order of magnitude the surface binding capacity. Steady state uptake of transferrin at 37 degrees C is generally less than 50% of the total transferrin binding capacity. During long-term incubation of the reticulocytes, all transferrin binding ability is lost, the ability to internalize being lost most rapidly. The loss in ability to bind transferrin during long-term incubation is independent of the number of surface transferrin binding sites, since removal of surface receptors with pronase does not affect the rate of loss of the internal pool of receptors during long-term incubation. Moreover, after removing surface receptors with pronase, only a fraction of the original number of receptors is restored to the surface, despite the presence of a large pool of internal receptors. These data suggest that only a fraction of the internal pool of receptors is capable of recycling to the cell surface in sheep reticulocytes.     

Life-cycles of some invertebrate taxa in a small pond together with changes in their numbers over a period of three years

1. Monthly quantitative samples of the invertebrate fauna (except Protozoa) in a small pond were taken over a period of three years. During one year, insect emergence traps were in operation. Water temperatures were recorded during the investigation.
2. The most abundant organisms in the pond were Phaenocora typhlops, Limnodrilus hoffmeisteri and Chaoborus crystallinus. Certain species of Micro-Crustacea and Chironomidae were also abundant but these groups have been dealt with elsewhere (see p. 66). Dendrocoelum lacteum, Polycelis nigra, Helobdella stagnalis, Lumbriculus aariegatus, Tubifex tubifex, Planorbis complanatus, and Asellus meridianus also occurred in considerable though lower numbers; other species occurred in low numbers.
3. The life-cycles and changes in numbers of the more numerous species are considered. The life-histories of D. lacteum, P. nigra, H. stagnalis, P. complanatus, A. meridianus are in agreement with published information. P. typhlops is seasonal in occurrence, being active from May to Sept. inclusive. Times of emergence of adults of various insect species agree with information available in the literature.
4. The life-cycle of L. hoffmeisteri in the pond is as follows: young worms hatch in spring/summer and form the bulk of the population from April to July/Aug; they mature from Aug. onwards and breeding starts in earnest from Feb./March. The life-cycle of T. tubifex is as follows: breeding starts in Feb., recruitment of young takes place from April till June, and these start to mature in Nov./Dec. It is not certain if some animals which breed in the spring/early summer survive to breed the following year.
5. The life-cycle of C. crystallinus appears to be as follows: first instars present from May to Oct., second instars from May to Dec., third instars from June till following Jan., fourth instars all the year round, pupae from May till Aug., and eggs from May to Sept. Adult emergence takes place from late April till mid-Sept.
6. A six-week drought in Oct/beginning Nov. in the second year of the investigation caused considerable mortality in most species, but most survived with only a few exceptions.

     

Recovery of Colony-Forming Ability and Genetic Marker Activity by UV-Damaged Hemophilus influenzae

The rate of recovery of UV-irradiated Hemophilus influenzae from acriflavine-sensitized loss of colony-forming ability was studied at various acriflavine concentrations, UV doses, and temperatures. This rate (as calculated from an equation based upon certain assumptions) was on the order of 0.07 per minute per cell at 37°C. This did not vary greatly with UV dose or acriflavine concentration, but did with temperature, giving a ΔH‡ of about 16 kcal/mole. In another set of experiments, cells bearing two genetic markers (resistance to 2000 μg/ml streptomycin and to 2.5 μg/ml novobiocin) were irradiated and then incubated without acriflavine. DNA extracts made from samples taken after various periods of incubation time were assayed on antibiotic-sensitive cells using acriflavine to inhibit repair during and following transformation. It was found that both in vivo irradiated markers were reactivated in the donor to approximately the same extent (with a rate constant of 0.04 per minute). This result was in contrast to the results obtained when extracted DNA bearing the same markers was irradiated in vitro and used to transform cells. In this latter case the streptomycin marker was much more sensitive than the novobiocin marker. This difference is interpreted as being due to the mechanics of the transformation system.     

Phosphorylation of choline and ethanolamine in Ehrlich ascites-carcinoma cells

1. Ehrlich ascites-cell extracts convert choline and ethanolamine approximately equally well into their respective phosphoryl derivatives. 2. Choline is a potent inhibitor of ethanolamine phosphorylation, but ethanolamine has little effect on choline phosphorylation. 3. 2,3-Dimercaptopropanol, cysteine and Ca(2+) inhibit ethanolamine phosphorylation, but have no detectable effect on choline phosphorylation. 4. Choline-phosphorylating activity in Ehrlich ascites-cell extracts is more stable during storage than ethanolamine-phosphorylating activity. 5. Choline phosphorylation is stimulated in the presence of benzoylcholine, succinylcholine, butyrylcholine and propionylcholine, whereas ethanolamine phosphorylation is inhibited. This relationship is reciprocal: the compounds causing the greatest stimulation of choline phosphorylation bring about the greatest inhibition of ethanolamine phosphorylation.