The actions of vasoactive compounds in the foetus and the effect of perfusion through the placenta on their biological activity

In acute experiments on the in utero foetal lamb, angiotensin II was a more potent pressor agent that either noradrenaline or adrenaline, and the response to angiotensin II was not consistently modified by the combined administration of alpha and beta-adrenergic blocking agents. A significant reduction in the pressor response of the foetus to angiotensin II and noradrenaline occurred with infusion of these compounds to the foetus by the umbilical artery when compared with the response obtained with infusions of the same doses of these drugs by the umbilical vein. Moreover, the concentration of angiotensin II (pg. ml-1) present in the foetal circulation was less following umbilical arterial infusions compared with umbilical vein infusions of the same doses. A similar reduction in the pressor activity of adrenaline and the cardio-stimulant effect of isoprenaline occurred when these drugs were infused by the umbilical artery. It is concluded that the foetus, like the adult animal, is more sensitive to angiotensin II than to catecholamines and that the biological activities of noradrenaline, angiotensin II, adrenaline and isoprenaline are reduced by perfusion through the foetal placenta.     

Solution structure of [d(ATGAGCGAATA)]2. Adjacent G:A mismatches stabilized by cross-strand base-stacking and BII phosphate groups.

The solution structure of a rather unusual B-form duplex [d(ATGAGCGAATA)]2 has been determined using two-dimensional nuclear magnetic resonance (2D-NMR) and distance geometry methods. This sequence forms a stable ten base-pair B-form duplex with 3' overhangs and two pairs of adjacent G:A mismatches paired via a sheared hydrogen-bonding scheme. All non-exchangeable protons, including the stereo-specific H-5'S/H-5'R of the 3G and 7G residues, were assigned by 2D-NMR. The phosphorus spectrum was assigned using heteronuclear correlation with H-3' and H-4' reasonances. The complete assignments reveal several unusual nuclear Overhauser enhancements (NOEs) and unusual chemical shifts for the neighboring G:A mismatch pairs and their adjacent nucleotides. Inter-proton distances were derived from time-dependent NOEs and used to generate initial structures, which were further refined by iterative back-calculation of the two-dimensional nuclear Overhauser enhancement spectra; 22 final structures were calculated from the refined distance bounds. All these final structures exhibit fully wound helical structures with small penalty values against the refined distance bounds and small pair-wise root-mean-square deviation values (typically 0.5 A to 0.9 A). The two helical strands exchange base stacking at both of the two G:A mismatch sites, resulting in base stacking down each side rather than down each strand of the twisted duplex. Very large twist angles (77 degrees) were found at the G:A mismatch steps. All the final structures were found to have BII phosphate conformations at the adjacent G:A mismatch sites, consistent with observed downfield 31P chemical shifts and Monte-Carlo conformational search results. Our results support the hypothesis that 31P chemical shifts are related to backbone torsion angles. These BII phosphate conformations in the adjacent G:A mismatch step suggest that hydrogen bonding of the G:A pair G-NH2 to a nearby phosphate oxygen atom is unlikely. The unusual structure of the duplex may be stabilized by strong interstrand base stacking as well as intrastrand stacking, as indicated by excellent base overlap within the mismatch stacks.     

The use of double staining techniques for investigating bacterial attachment to mucopolysaccharide-coated epithelial cells

Double staining techniques were devised to study Escherichia coli attachment to mucopolysaccharide-coated urinary tract epithelial cells. In addition, vital stains were used to distinguish between viable and nonviable epithelial cells. Alcian blue was used to confirm the presence of glycosaminoglycans and periodic acid Schiff was used to distinguish a further group of polysaccharides, proteoglycans, neutral mucosubstances and glycolipids. The staining methods enabled investigations to be carried out concerning the possible importance of mucopolysaccharides in the attachment of bacteria to mucosal surfaces. Staining techniques were also used to investigate whether the presence of a mucopolysaccharide coat is related to epithelial cell viability. The combinations of vital and mucopolysaccharide stains were found to give reproducible results when cell preparations were evaluated by three individuals. Both in vivo and in vitro certain populations of epithelial cells have been found with a large number of bacteria preferentially attached. The double staining techniques described here may help to reveal the nature of these target cells.     

Erythropoietin-mediated erythrocytosis in rodents after intrarenal injection of nickel subsulfide

Rats and guinea pigs developed pronounced erythrocytosis at one to four months after unilateral intrarenal (ir) injection of nickel subsulfide (Ni3S2). For example, at two months after ir administration of Ni3S2 (5 mg) to rats, blood hematocrit values averaged 70 +/- 3 percent (p less than 0.001 vs. 48 4/- 2 in controls); at two months after ir administration of Ni3S2 (20 mg) to guniea pigs, blood hematocrit values averaged 67 +/- 6 percent (p less than 0.001 vs. 49 +/- 1 percent in controls). Hamsters and gerbils did not develop erythrocytosis after ir injection of Ni3S2 (5 mg/animal). Administration of Ni3S2 to rats by intrasplenic injection did not increase blood hematocrit; splenectomy did not prevent erythrocytosis in rats that received ir injection of Ni3S2. Erythrocytosis in rats was completely blocked by excision of the Ni3S2-injected kidney but was unaffected by excision of the non-injected kidney. Partial inhibition of Ni3S2-induced erythrocytosis in rats occurred after simultaneous ir injection of Mn, Cu, or Al dusts, benzo(a)pyrene, or subcutaneous (sc) infusion of sodium diethyldithiocarbamate. Erythrocytosis induced by ir injection of Ni3S2 was augmented by ir injection of Cr dust or intramuscular (im) administration of iron-dextran. Erythrocytosis occurred in rats after ir implantation of Ni3S2 within semi-permeable cellulose tubules, indicating that phagocytosis of Ni3S2 particles is unnecessary for erythropoietic stimulation. Erythropoietin (Ep) activity in rat serum increased sixfold at two weeks after ir injection of Ni3S2 (p less than 0.001 vs. controls), but Ep activity in pooled extracts of Ni3S2-treated rat kidneys did not increase significantly. This study identifies several factors that influence erythropoietic stimulation by Ni3S2, and furnishes salient information concerning the pathogenesis of Ni3S2-induced erythrocytosis.     

Sequence Diversity of Flagellin (fliC) Alleles in Pathogenic Escherichia coli

To study the molecular evolution of flagellin, the protein subunit specifying flagellar (H) antigens, the fliC genes from 15 pathogenic strains of Escherichia coli were amplified by PCR and sequenced. Comparison of fliC sequences of H6 and H7 strains revealed that alleles have a mosaic structure indicating the occurrence of past horizontal transfer of DNA segments between strains. The close similarity of H7 sequences also indicates the exchange of an entire fliC H7 allele between distant clonal lineages. In addition, the ratio of silent substitutions to amino acid replacements suggests that a short segment in the central region of fliC has been under positive selection in the divergence of H6 and H7 alleles. Phylogenetic analysis demonstrates that the fliC sequences of O157:H7 and O55:H7 serotypes are nearly identical and highly divergent from those of E. coli strains expressing H6 and H2 flagellar antigens. A nonmotile clone of sorbitol-fermenting O157 has rapidly accumulated multiple mutations in fliC, presumably as a result of the silencing of flagellin expression.     

Parasites flicking the NPY gene on the host's switchboard: why NPY?

It was investigated whether up-regulation of the NPY gene by the schistosome Trichobilharzia ocellata in its snail host Lymnaea stagnalis redirects the host's energy flows. We cloned the cDNA encoding Lymnaea NPY (LyNPY), purified and sequenced the peptide, and used synthesized peptide for physiological and morphological studies. Increasing the LyNPY titer in nonparasitized snails (mimicking parasitosis) by 1) implantation of slow-release pellets and 2) injections suppressed reproductive activity and reduced growth in a dose- and time-dependent manner without affecting food intake. When the LyNPY titer was back to normal, reproduction and growth were resumed, coinciding with a transient increase of food intake serving to replenish glycogen stores. Observations on double-immunostained whole mount preparations of brains support these data. A close association was found between LyNPY-positive axons and axons both from ovulation hormone-producing neurons and molluscan insulin-like peptide-producing neurons involved in regulation of growth. As no synaptic(-like) contacts were observed, it is supposed that LyNPY acts nonsynaptically. No morphological interaction was found between LyNPY-positive axons and motoneurons innervating the feeding apparatus. Our data explain why it is an advantageous strategy for endoparasites to up-regulate the highly conserved NPY gene in their host.     

Determination of opiates in urine by capillary electrophoresis

A method for the separation of a mixture of opiates comprising pholcodine, 6-monoacetylmorphine, morphine, heroin, codeine and dihydrocodeine by capillary electrophoresis using a running buffer of 100 mM disodium hydrogenphosphate at pH 6 is described. The characteristics of an analytical method based on this separation for the determination of these drugs following extraction from urine and using levallorphan as internal standard are reported. Detection limits in the region of 10 ng cm⁻³ are achieved when using electrokinetic injection. A comparison is made of the sensitivity and reproducibility of electrokinetic and hydrodynamic injection for these drugs. Data are presented to show the results obtained when the proposed method is applied to urine spiked with all the above opiates and also to urine from a subject following consumption of dihydrocodeine and pholcodine. The concentrations found are compared with those obtained by LC.     

Isolation of Rhodococcus rhodochrous NCIMB13064 derivatives with new biodegradative abilities

Abstract Rhodococcus rhodochrous NCIMB 13064 can dehalogenate and utilise a number of halogenated aliphatic compounds as sole carbon and energy source. Mutants of NCIMB13064 can be easily isolated with an enlarged range of 1-chloroalkane utilising ability. Dehalogenation of 1-chlorononane, 1-chlorodecane and short-chain 1-chloroalkanes (C₃-C₈) is encoded by the same plasmid pRTL1. However, a different genetic element(s) is required for the dehalogenation of 3-chloropropionic acid. Two derivatives (P200 and P400) of R. rhodochrous NCIMB 13064 were isolated which had acquired the ability to utilise naphthalene as sole carbon and energy source. Both strains lost the ability to utilise short-chain 1-chloroalkanes and underwent some rearrangements associated with pRTLl plasmid.