Adenovirus early gene products may control viral mRNA accumulation and translation in vivo

The mechanisms controlling early adenovirus gene expression in vivo have been studied using inhibitors of protein synthesis. When inhibitors were added shortly before or at the onset of infection, viral mRNA from all early regions was transcribed, spliced and accumulated over a 7 hr period. After longer pretreatment, accumulation of several early mRNAs were suppressed. Addition of inhibitors 1 hr after infection enhanced the accumulation of viral mRNA in the cytoplasm. Translation of early mRNA selected on adenovirus DNA in a cell-free system reflected the amount of viral mRNA present. A viral coded product may therefore control accumulation of viral mRNA.A different pattern emerged when inhibitors of protein synthesis were removed at 5 hr postinfection and cells were pulse-labeled in vivo. If inhibitors were introduced at or before infection, early viral proteins were synthesized only after a lag of 1–3 hr. However, if treatment was introduced 1 hr post-infection, reversion of the protein synthesis block was instantaneous. It appears that protein synthesis inhibitors reveal an in vivo translational block for viral mRNA. This block could be overcome by preinfection with a related virus. Furthermore, no block was observed in a virus-transformed human embryonic kidney cell line (293) which expresses early region 1 of the viral genome. Viral gene product(s) encoded in early region 1 may control translation of early adenovirus messenger RNA in vivo.     

Human P301L-Mutant Tau Expression in Mouse Entorhinal-Hippocampal Network Causes Tau Aggregation and Presynaptic Pathology but No Cognitive Deficits

Accumulation of hyperphosphorylated tau in the entorhinal cortex (EC) is one of the earliest pathological hallmarks in patients with Alzheimer’s disease (AD). It can occur before significant Aβ deposition and appears to “spread” into anatomically connected brain regions. To determine whether this early-stage pathology is sufficient to cause disease progression and cognitive decline in experimental models, we overexpressed mutant human tau (hTauP301L) predominantly in layer II/III neurons of the mouse EC. Cognitive functions remained normal in mice at 4, 8, 12 and 16 months of age, despite early and extensive tau accumulation in the EC. Perforant path (PP) axon terminals within the dentate gyrus (DG) contained abnormal conformations of tau even in young EC-hTau mice, and phosphorylated tau increased with age in both the EC and PP. In old mice, ultrastructural alterations in presynaptic terminals were observed at PP-to-granule cell synapses. Phosphorylated tau was more abundant in presynaptic than postsynaptic elements. Human and pathological tau was also detected within hippocampal neurons of this mouse model. Thus, hTauP301L accumulation predominantly in the EC and related presynaptic pathology in hippocampal circuits was not sufficient to cause robust cognitive deficits within the age range analyzed here.     

In situ generation of iminodiacetic acid groups on nanoporous alumina for the reversible immobilization of enzymes and other biomolecules

Nanoporous alumina membranes were silanized with aminopropylsilane and iminodiacetic acid (IDA) groups were generated in situ by reaction with iodoacetate. The membranes were mounted in standard filter holders, connected to a HPLC system and saturated with selected metal ions. Cu(II) allowed the capture of chicken muscle lactate dehydrogenase with such stability, repeatability and reproducibility that Michaelis–Menten kinetics could be studied. The IDA surface was stable for months and could be depleted and regenerated with metal ions multiple times without appreciable loss of capacity. The binding of lactate dehydrogenase influenced the backpressure to the extent that could be expected for a monolayer according to Poiseuilles law.     

Degradation of heparin proteoglycan in cultured mouse mastocytoma cells.

Pulse-labelling of mouse mastocytoma cell cultures, established from ascites fluid, with inorganic [35S]sulphate for 1 h yielded labelled heparin proteoglycan containing polysaccharide chains of Mr 60,000-100,000. After chase incubation for 24 h most of the 35S appeared in intracellular polysaccharide fragments similar in size to commercially available heparin, Mr 5000-25,000, as indicated by gel chromatography. Products isolated from cultures after 6 h of chase incubation consisted of partially degraded free polysaccharide chains and, in addition, residual proteoglycans that were of smaller size than the proteoglycans initially pulse-labelled. The polysaccharide chains released by alkali treatment from the residual chase-incubated proteoglycans were of the same size as the chains derived from proteoglycans after 1 h of pulse labelling. These results suggest that the intracellular degradation of heparin proteoglycan to polysaccharide fragments is initiated by release of intact polysaccharide chains, probably by action of a peptidase, and is pursued through cleavage of these chains by an endoglycosidase. An endoglucuronidase with stringent substrate specificity [Thunberg, Bäckström, Wasteson, Ogren & Lindahl (1982) J. Biol. Chem. 257, 10278-10282] has previously been implicated in the latter step. Cultures of more purified mastocytoma cells (essentially devoid of macrophages) did not metabolize [35S]heparin proteoglycan to polysaccharide fragments, but instead accumulated free intact polysaccharide chains, i.e. the postulated intermediate of the complete degradation pathway. When such purified cells were co-cultured with adherent mouse peritoneal cells, presumably macrophages, formation of polysaccharide fragments was observed. It is tentatively proposed that the expression of endoglucuronidase activity by the mast cells depends on collaboration between these cells and macrophages.     

Isolation and characterization of neurokinin A, neurokinin A(3-10) and neurokinin A(4-10) from a neutral water extract of a metastatic ileal carcinoid tumour

A metastasis to the right liver lobe of an argyrophil/argentaffin midgut carcinoid tumour in a patient with the classical carcinoid syndrome was examined for the presence of tachykinins other than substance P, using a specific antiserum. The extract was initially purified using SepPak cartridges, and subsequently subjected to cation-exchange chromatography on SP Sephadex C-25 which separated the immunoreactive material into two main components (components I and II). Both were further purified by anion-exchange chromatography on DEAE-Sephadex A-25, and by reverse-phase fast protein liquid chromatography. Component II was identified as neurokinin A by its immunochemical and chromatographic properties and amino acid sequence analysis. Component I consisted of two molecular forms which were identified as neurokinin A(3-10) and neurokinin A(4-10) by amino acid sequence analysis. The tumour tissue contained only small amounts of the eledoisin-like peptide that has earlier been demonstrated in mammalian tissues. Although this component behaved like the nonmammalian peptide eledoisin on reverse-phase HPLC and on reverse-phase ion-pair chromatography, eledoisin-specific antiserum E2 indicated that eledoisin-like peptide is not identical to eledoisin. Neurokinin A in carcinoid tumours has an N-terminal heterogeneity; this multiplicity constitutes a further support for the hypothesis that carcinoid tumours produce a number of tachykinins which may be present in different relative amounts in individual patients and may contribute to the individual differences in symptomatology.     

Glycosphingolipid expression in spontaneously aborted fetuses and placenta from blood group p women. Evidence for placenta being the primary target for anti-Tja-antibodies

A 12-week-old fetus and one 17-week-old fetus + placenta were obtained after spontaneous abortions from two women of blood group p. The 17-week-old fetus was dissected into intestine, liver, brain and residual tissue. Nonacid glycosphingolipid fractions were prepared from the tissues. Glycolipid characterization was carried out using thin layer chromatography immunostained with monoclonal antibodies and bacteria and by¹H NMR spectroscopy and mass spectrometry. In the placental fraction substantial amounts of globotetraosylceramide (P-antigen) and globotriaosylceramide (P^k-antigen) were identified. In contrast, the fetuses contained only trace amounts of these structures, as revealed by immunostaining. These results indicate that the primary target for the antibodies of the anti-Tj^a serum is the placenta tissue, resulting in termination of the pregnancy.     

Ethylene effects on cambial activity and cell wall formation in hypocotyls of Picea abies seedlings

Ethylene regulation of cell division in the vascular cambium and cell wall formation was studied in hypocotyls of Norway spruce ( Picea abies [L.] Karst.) seedlings. Cuttings from 6-week-old seedlings were placed in water culture to which compounds affecting the synthesis and action of ethylene were added. After a 3-week treatment period, growth, ethylene production, morphology and cell wall composition of the hypocotyls were determined. Addition of high concentrations of the potent ethylene releasing agent 2-chloroethylphosphonic acid (ethrel), which increased ethylene emission by more than twice compared to control plants, inhibited the expansion of xylem cells while stimulating the incorporation of cell wall material, especially cellulose. Addition of small amounts of ethrel, which slightly stimulated ethylene emission, led to increases in the size of xylem cells, the amount of phloem tissue and the number of intercellular spaces in the cortex, and thus to increased hypocotyl diameter. However, no significant change in cell wall composition was detected. When ethylene production was decreased by adding Co²⁺ to the nutrient solution, differentiation of new xylem was disturbed, but the rate of cell division was not affected. Although the incorporation of cell wall material was inhibited, the proportions of lignin and cellulose in the wall appeared to remain unchanged. Silver ions stimulated the expansion of both xylem and cortex cells, but had no significant effect on cell wall formation. We conclude that ethylene has a role in regulating the incorporation of wall carbohydrates.     

Embryology of Calenduleae (Compositae)

Microsporangial conditions, macrosporogenesis, embryo sac development and in some species endosperm formation have been studied in 5 species of Dimorphotheca Vaill. ex Mnch, 2 species of Castalis Cass., 16 species of Osteospermum L., 6 species of Calendula L., 1 species of Gibbaria Cass., and 1 species of Chrysanthemoides Tourn. ex Fabr.
The microsporangial wall consists of four different layers of cells. Miocrospore formation is simultaneous. The tetrads are mostly arranged in tetrahedrals. A tapetal periplasmodium develops.
The archespore of the macrosporangium is in most cases 1-celled, sometimes 2-or many-celled. The meiosis gives rise to a linear tetrad. Usually the chalazal macro-spore develops. The embryo sac development is monosporic of the Polygonum type. In a few cases in Dimorphotheca venusta T. Norl. a tetrasporic embryo sac development was observed besides the Polygonum type. Castalis tragus (Ait.) T. Norl. may have a bisporic embryo sac development in addition to the Polygonum type. All the Calendula species develop synergid haustoria. Antipode haustoria occur in Osteospermum sinuatum (DC.) T. Norl. and Chrysanthemoides monilifera (L.) T. Norl. Chalazally situated extra embryo sacs below the ordinary one in Dimorphotheca sinuata DC, D. venusta , and Castalis nudicaulis var. graminifolia (L.) T. Norl. are regarded to be aposporic. Six species have more than three antipodes per embryo sac. The antipodes occasionally become 2-or 4-nucleate.     

The Influence of Feeding and Oral Rehydration on the Bioavailability of Oxytetracycline in Calves

The influence of feeding on the bioavailability of oxytetracycline was studied in preruminant calves. Oxytetracycline was given in water as a drench to fasting calves or was mixed in the milk replacer. Compared to water the bioavailability was significantly reduced (53.5%) when oxytetracycline was mixed in the milk re-placer. A further reduction, 83.3 %, occurred when the calves were treated one hour post milk feeding. Also concentrate was found to reduce the bioavailability. Very high serum levels were recorded when the drug was given in an oral rehydration solution, pH 4.9, containing glycine. The values obtained when an alkaline (pH 8.3) solution without glycine was used did not differ from the levels recorded when oxytetracycline was given in water. It was suggested that the use of oxytetra-cyclines in feeds may be questioned because of their well-known complex forming ability.