The in vitro effects of histamine and metiamide on neutrophil motility and their relationship to intracellular cyclic nucleotide levels.

Histamine at concentrations of 1 x 10(-5) M to 5 x 10(-5) M consistently increased neutrophil movement as measured in Boyden chambers. This effect was entirely caused by stimulation of chemokinesis (stimulated random migration) and true chemotaxis was inhibited by these concentrations. This inhibition of chemotaxis could be abolished by pretreatment with metiamide, an H-2 receptor antagonist, and levamisole, but not by diphenylhydramine, an H-1 receptor antagonist. Metiamide at similar concentrations produced a mild stimulation of chemokinesis but has no effect on true chemotaxis. The histamine effects on neutrophil motility were associated with increased levels of intracellular cAMP wehreas cAMP levels were unaffected. Agents known to elevate intracellular cAMP levels produced effects on neutrophil motility similar to those of histamine. It is suggested that histamine exerts a 2-fold effect on neutrophil motility mediated via an H-2 receptor site and associated with elevated levels of cAMP.     

Cyclins A and B associate with chromatin and the polar regions of spindles, respectively, and do not undergo complete degradation at anaphase in syncytial Drosophila embryos

Maternally contributed cyclin A and B proteins are initially distributed uniformly throughout the syncytial Drosophila embryo. As dividing nuclei migrate to the cortex of the embryo, the A and B cyclins become concentrated in surface layers extending to depths of approximately 30-40 microns and 5-10 microns, respectively. The initiation of nuclear envelope breakdown, spindle formation, and the initial congression of the centromeric regions of the chromosomes onto the metaphase plate all take place within the surface layer occupied by cyclin B on the apical side of the blastoderm nuclei. Cyclin B is seen mainly, but not exclusively, in the vicinity of microtubules throughout the mitotic cycle. It is most conspicuous around the centrosomes. Cyclin A is present at its highest concentrations throughout the cytoplasm during the interphase periods of the blastoderm cycles, although weak punctate staining can also be detected in the nucleus. It associates with the condensing chromosomes during prophase, segregates into daughter nuclei in association with chromosomes during anaphase, to redistribute into the cytoplasm after telophase. In contrast to the cycles following cellularization, neither cyclin is completely degraded upon the metaphase-anaphase transition.     

Differences between the endogenous and exogenous DNA sequences of Rous-associated virus-O.

DNA sequences related to the endogenous retrovirus of chickens, Rous-associated virus-O (RAV-O), have been examined using site-specific DNA endonuclease analysis of cellular DNA derived from line 15 and line 100 chickens. Individual embryos from both inbred lines were used as a source of embryonic fibroblasts from which cellular DNA was isolated. Analysis of DNA containing either endogenous RAV-O sequences alone or both endogenous and exogenous RAV-O sequences produced identical patterns of RAV-O-specific DNA fragments after digestion with the endonucleases Eco RI, Hind III, BgI II, Bam HI or Xho I. Similar analysis with endonucleases Hinc II or Hha I, however, produced several RAV-O-specific DNA fragments which were derived from cellular DNA containing both endogenous and exogenous RAV-O sequences but not from cellular DNA containing only endogenous sequences. Although some differences exist between the DNA fragments specific for the endogenous viral sequences of line 15 and line 100 cellular DNA, the DNA fragments specific for the exogenous viral sequences were identical between the two inbred lines. Cleavage of an unintegrated linear RAV-O DNA molecule with Hinc II or Hha I produced DNA fragments identical to those specific for the exogenously acquired RAV-O provirus. This suggests that these characteristic fragments contain no cellular DNA. The potential DNA junction fragments containing both viral and cellular DNA, identified after analysis of DNA that contains both endogenous and exogenous viral sequences, were identical to those observed after analysis of DNA containing only endogenous viral sequences. These results support the following conclusions. First, exogenous proviral sequences are integrated into chicken cell DNA following an interaction between viral and cellular DNA that is specific with respect to the virus and nonspecific with respect to the cell. Second, both the free linear RAV-O DNA intermediate and the newly integrated exogenous provirus contain specific endonuclease sites that are not found in endogenous RAV-O DNA sequences. These results suggest that the formation of the exogenous DNA provirus involves specific alteration of the endogenous viral DNA sequences before reinsertion of the sequences as the exogenous RAV-O DNA provirus. It is possible that newly integrated exogenous RAV-O sequences are characterized by specific differences in the pattern of base methylation and a limited sequence arrangement.     

Ecological studies of spores of aquatic hyphomycetes in the cringle brook,lincs

The aquatic Hyphomycete spora of the Cringle Brook, Lincs, was examined by foam sampling and by the use of cellophane impaction traps between August, 1968 and January, 1970.The species most frequently found (Tetracladium marchalianum, Alatospora acuminata and Flagellospora curvula) were generally in agreement with those found by other workers in temperate areas. Impaction trap samples generally contained fewer species than foam samples but filiform spore types such as Flagellospora were more frequently found on traps than in foam suggesting that impaction is more selective towards the filiform spore type than is foam. Many species increased in frequency in autumn accompanying and following leaf fall, and the winter spora was dominated by Alatospora acuminata, Clavariopsis aquatica, Clavatospora stellata, Flagellospora curvula and Lemonniera aquatica. During the summer the spora was dominated by Tetracladium marchalianum.The role of foam and impaction in the balance of aquatic spore populations is discussed in relation to techniques available for their study.     

Luminescence-based nonextractive technique for in situ detection of Escherichia coli in soil.

Measurement of light output by luminometry was used to estimate quantitatively the cell concentrations of luminescent strains of Escherichia coli in liquid culture and inoculated into soil. Strains were constructed in which luciferase production was autoinducible or constitutive. In the former, light output per cell varied considerably during growth but was constant in constitutive strains. In liquid culture, the lower detection limit was in the order of 10(2) cells ml-1. Sensitivity was reduced by approximately 1 order of magnitude for cells inoculated into soil, when 2 x 10(2) to 6 x 10(3) cells g of soil-1 could be detected. Light output measurements were obtained within 5 min of sampling, and luminometry therefore potentially offers a rapid and sensitive detection technique for genetically engineered microorganisms.     

Trap‐shyness subsidence is a threshold function of mark–recapture interval in brown mudfish Neochanna apoda populations

The influence of capture interval on trap shyness, and temperature, rainfall and drought on capture probability (p) in 827 brown mudfish Neochanna apoda was quantified using mark–recapture models. In particular, it was hypothesized that the loss of trapping memory in marked N. apoda would lead to a capture‐interval threshold required to minimize trap shyness. Neochanna apoda trap shyness approximated a threshold response to capture interval, declining rapidly with increasing capture intervals up to 16·5 days, after which p remained constant. Tests for detecting trap‐dependent capture probability in Cormack–Jolly–Seber models failed to detect trap shyness in N. apoda capture histories with capture intervals averaging 16 days. This confirmed the applicability of the 16 day capture‐interval threshold for mark–recapture studies. Instead, N. apoda p was positively influenced by water temperature and rainfall during capture. These results imply that a threshold capture interval is required to minimize the trade‐off between the competing assumptions of population closure and p homogeneity between capture occasions in closed mark–recapture models. Moreover, environmental factors that influence behaviour could potentially confound abundance indices, and consequently abundance trends should be interpreted with caution in the face of long‐term climate change, such as with global warming.     

Pharmacological rescue of the mutant cystic fibrosis transmembrane conductance regulator (CFTR) detected by use of a novel fluorescence platform

Numerous human diseases arise because of defects in protein folding, leading to their degradation in the endoplasmic reticulum. Among them is cystic fibrosis (CF), caused by mutations in the gene encoding the CF transmembrane conductance regulator (CFTR ), an epithelial anion channel. The most common mutation, F508del, disrupts CFTR folding, which blocks its trafficking to the plasma membrane. We developed a fluorescence detection platform using fluorogen-activating proteins (FAPs) to directly detect FAP-CFTR trafficking to the cell surface using a cell-impermeant probe. By using this approach, we determined the efficacy of new corrector compounds, both alone and in combination, to rescue F508del-CFTR to the plasma membrane. Combinations of correctors produced additive or synergistic effects, improving the density of mutant CFTR at the cell surface up to ninefold over a single-compound treatment. The results correlated closely with assays of stimulated anion transport performed in polarized human bronchial epithelia that endogenously express F508del-CFTR. These findings indicate that the FAP-tagged constructs faithfully report mutant CFTR correction activity and that this approach should be useful as a screening assay in diseases that impair protein trafficking to the cell surface.     

A novel method for purification of prephenic acid.

Prephenic acid accumulated in culture filtrates of Neuro-spora crassa has been purified in 66% yield utilizing adsorption chromatography on Sephadex G-10 in the major purification steps.