The iron chelators desferrioxamine and 1-alkyl-2-methyl-3-hydroxypyrid-4-ones inhibit vascular prostacyclin synthesis in vitro.

The iron chelators desferrioxamine (DFO), 1,2-dimethyl(L1)-, 1-ethyl-2-methyl(L1NEt)- and 1-propyl-2-methyl(L1NPr)-3-hydroxypyrid-4-ones inhibited rat aortic prostacyclin (PGI2) synthesis in vitro (rank order of potency: DFO greater than L1 greater than L1NEt greater than L1NPr) when stimulated with adrenaline, arachidonate and the Ca2+ ionophore A23187. The inhibitory action of the chelators was blocked by Fe3+ and Al3+ and reversed by washing and H2O2, but not by ascorbate. These data suggest that iron chelators inhibit prostanoid synthesis in intact tissue through the removal or binding of Fe3+ linked to cyclo-oxygenase. These iron chelators may be of therapeutic value in the treatment of inflammatory and other diseases via two mechanisms: (1) the inhibition of pro-inflammatory prostanoid synthesis and (2) the inhibition of toxic-free-radical generation by cyclo-oxygenase.     

N-acetylglucosaminyltransferase substrates prepared from glycoproteins by hydrazinolysis of the asparagine-N-acetylglucosamine linkage. Purification and structural determination of oligosaccharides with mannose andN-acetylglucosamine at the non-reducing termini

Sixteen asparagine-linked oligosaccharides ranging in size from (Man)₂(GlcNAc)₂ (Fuc)₁ to (GlcNAc)₆(Man)₃(GlcNAc)₂ were obtained from human ₁-acid glycoprotein and fibrinogen, hen ovomucoid and ovalbumin, and bovine fetuin, fibrin and thyroglobulin by hydrazinolysis, mild acid hydrolysis and glycosidase treatment. The oligosaccharides hadN-acetylglucosamine at the reducing termini and mannose andN-acetylglucosamine residues at the non-reducing termini and were prepared for use asN-acetylglucosaminyltransferase substrates. Purification of the oligosaccharides involved gel filtration and high performance liquid chromatography on reverse phase and amine-bonded silica columns. Structures were determined by 360 MHz and 500 MHz proton nuclear magnetic resonance spectroscopy, fast atom bombardment-mass spectrometry and methylation analysis. Several of these oligosaccharides have not previously been well characterized.Abbreviations bis bisecting GlcNAc - DMSO dimethylsulfoxide - FAB fast atom bombardment - Fuc l-fucose - Gal d-galactose - GLC gas-liquid chromatography - GlcNAc or Gn N-acetyl-d-glucosamine - HPLC high performance liquid chromatography - Man or M d-mannose - MES 2-(N-morpholino)ethanesulfonate - MS mass spectrometry - NMR nuclear magnetic resonance - PIPES piperazine-N,N-bis(2-ethane sulfonic acid) the nomenclature of the oligosaccharides is shown in Table 1.     

Formation of heteroduplex DNA during mammalian intrachromosomal gene conversion.

We have studied intrachromosomal gene conversion in mouse Ltk- cells with a substrate designed to provide genetic evidence for heteroduplex DNA. Our recombination substrate consists of two defective chicken thymidine kinase genes arranged so as to favor the selection of gene conversion products. The gene intended to serve as the recipient in gene conversion differs from the donor sequence by virtue of a palindromic insertion that creates silent restriction site polymorphisms between the two genes. While selection for gene conversion at a XhoI linker insertion within the recipient gene results in coconversion of the nearby palindromic site in more than half of the convertants, 4% of convertant colonies show both parental and nonparental genotypes at the polymorphic site. We consider these mixed colonies to be the result of genotypic sectoring and interpret this sectoring to be a consequence of unrepaired heteroduplex DNA at the polymorphic palindromic site. DNA replication through the heteroduplex recombination intermediate generates genetically distinct daughter cells that comprise a single colony. We believe that the data provide the first compelling genetic evidence for the presence of heteroduplex DNA during chromosomal gene conversion in mammalian cells.     

Adenine photodimerization in deoxyadenylate sequences: elucidation of the mechanism through structural studies of a major d(ApA) photoproduct.

The mechanism of the photodimerization of adjacent adenine bases on the same strand of DNA has been elucidated by determining the structure of one of the two major photoproducts that are formed by UV irradiation of the deoxydinucleoside monophosphate d(ApA). The photoproduct, denoted d(ApA)*, corresponds to a species of adenine photodimer first described by P?rschke (P?rschke, D. (1973) J.Am.Chem.Soc. 95, 8440-8446). From a detailed examination of its chemical and spectroscopic properties, including comparisons with the model compound N-cyano-N1-(1-methylimidazol-5-yl)formamidine, it is deduced that d(ApA)* contains a deoxyadenosine unit covalently linked through its C(8) position to C(4) of an imidazole N(1) deoxyribonucleoside moiety bearing an N-cyanoformamidino substituent at C(5). On treatment with acid, d(ApA)* is degraded with high specificity to 8-(5-amino-imidazol-4-yl)adenine whose identity has been confirmed by independent chemical synthesis. It is concluded that the primary event in adenine photodimerization entails photoaddition of the N(7)-C(8) double bond of the 5'-adenine across the C(6) and C(5) positions of the 3'-adenine. The azetidine species thus generated acts as a common precursor to both types of d(ApA) photoproduct which are formed from it by competing modes of azetidine ring fission.     

Sequence of the OXA2 beta-lactamase: comparison with other penicillin-reactive enzymes

The nucleotide sequence of the unusual plasmid-mediated OXA2 beta-lactamase is presented, and compared with other beta-lactamases. The OXA2 enzyme has similar features at the presumed active site, but no other significant regions of homology with other penicillin-reactive enzymes. The active site homology may therefore represent convergent evolution of otherwise dissimilar genes.     

Effects of nectar concentration on butterfly feeding: measured feeding rates for Thymelicus lineola (Lepidoptera: Hesperiidae) and a general feeding model for adult Lepidoptera

Summary Field observations of the adult European skipper, Thymelicus lineola (Ochs), feeding on concentrated nectars (40–65% sucrose) from a variety of flower species led us to question recent literature stating that butterflies feed primarily, and most effectively, on dilute nectars. Rate of sucrose solution intake, volume consumed and feeding duration were measured for males and females at 25 and 35°C under laboratory conditions. As sucrose concentration increased, the volume of solution ingested per meal first increased and then decreased gradually, while sucrose intake was highest at concentrations 40%. Females fed more than males at all concentrations >10% while temperature had no significant effect on meal size. Feeding duration increased with concentration, was shorter at 35 than at 25°C, and was longer for females than males.The rate of volume intake decreased as concentration incresed, but not nearly as rapidly as predicted by earlier models. Rates did not differ between the sexes but were faster at 35 than 25°C. This increase was contributed to equally by a reduction in viscosity and an increase in power output of the cibarial pump. The form of the relations was similar, with maximum rate of sucrose intake occurring at 40% sucrose.A new mathematical model was developed to describe the rate — concentration relation based on the Hagen-Poiseuille equation for laminar fluid flow through pipes. Our model differs from previous models principally in that the power output of the insect's cibarial pump remains relatively constant while the pressure drop created by the pump to induce suction is highly variable. This change results in a very different feeding rate — sucrose concentration function with the optimal rate of sucrose intake at a concentration of approximately 40%. The model indicates that the same relation should hold for a wide range of proboscis shape and size and type of suction pump, and should therefore be applicable to all other nectar feeders with sucking mouth parts. Independent verifications of the model were carried out by measuring the rate of uptake of sucrose solutions of the adult common armyworm, Pseudaletia unipuncta (Haw.), and of human subjects using a volumetric pipette, both of which gave an excellent fit.Nectar concentrations which correspond to optimal rates of sucrose intake should be highly preferred by insects with high feeding costs, those which are time-limited, or which are very vulnerable while feeding. High transport costs and severe water stress may shift preferences to higher and lower concentrations respectively.     

Cell surface changes in preimplantation mouse embryos during compaction investigated using FITC conjugated lectins after proteolytic enzyme treatment

Summary Fluorescein-conjugated lectins were used to examine the reappearance of glycoproteins on the surface of 8-cell mouse embryos after treatment with proteolytic enzymes. Embryos were decompacted in calcium free medium, treated with various proteases and the process of recompaction monitored. The most effective enzymes in delaying recompaction were subtilopeptidase A and proteinase K at 1 mg/ml; the initiation of recompaction was delayed by about 5 h and 90% recompaction by 14–18 h. Papain and -chymotrypsin were only effective in the absence of calcium. The reappearance of receptors for fluorescein-conjugated Con-A, MPA, RCA-I, FBP, BSL-II and DBA was examined photometrically at 0,8–10 and 17–18 h after proteinase K treatment. There was an increase in binding of MPA, RCA-I, FBP and BSL-II in control embryos during the period of the experiment, between approx. 61 and 80 h post coitum in which embryos passed from the 8-cell stage to the 16–32 cell stage. Con-A binding remained the same and that of DBA decreased. By the time that 50% of enzyme treated embryos had recompacted (8–10 h) binding of Con-A was similar to control embryos. Binding of FBP had almost reached control levels while that of BSL-II, DBA, RCA-I and MPA had reached 60–85% of control levels. When embryos were fully compact (17–18 h) Con-A, FBP and DBA were bound in equal or slightly greater amounts to enzyme treated as to control embryos, and receptors for BSL-II, MPA and RCA-I had recovered almost to control levels. The results clearly show that the recovery of glycoproteins on the surface of 8–16 cell embryos parallels recompaction, providing further evidence for the role of these molecules in compaction.     

Computer-linked HPLC system for feedback control of fermentation substrates

Summary A data acquisition/control microcomputer system was interfaced to a commercial HPLC data transmission module. Control of substrate (ethanol) levels for four 7.5 L fermenters containing 100 g/L wet weight of the yeastCandida norvegensis was accomplished by employing intermittent, automated HPLC monitoring and a BASIC-encoded proportional integral policy for controlling substrate feed rates. Ethanol levels were maintained at 0.25, 0.50, 0.75 and 1.00% w/v.     

Proliferative characteristics of progeny derived from normal human hemopoietic progenitor cells

Cell cycle times of cells derived from human hemopoietic progenitor cells were measured directly by using 5-bromodeoxyuridine (BrdU) incorporation and sister chromatid differential staining. Umbilical cord cells were cultured in the mixed-colony assay for 96 h prior to the addition of BrdU and then harvested at 6-12 h intervals. Individual cell cycle times ranging from 8.5-23.4 h were observed while population mean cell cycle times ranged from 17.5-24 h. These results show that some early progeny of hemopoietic progenitor cells have short cell cycle times and that there is considerable heterogeneity within the proliferating cell population.