Roles of surface electrochemistry and macromolecular adsorption in heparin-induced red blood cell aggregation

Red blood cell (RBC) aggregation in heparin-saline solution was quantified by microscopic observation. The adsorption isotherms of heparin onto normal and neuraminidase-treated RBC surfaces were determined by radioactive heparin labeled with 125I-Bolton-Hunter Reagent. RBC aggregation by heparin requires the presence of sialic acids at cell surface and was enhanced by reduction of ionic strength of the suspending medium. Adsorption of heparin onto RBC surface was increased by removal of sialic acids. These findings not only serve to elucidate the basic mechanism of cell-cell interaction mediated by negatively charged macromolecules, but also provide experimental evidence for the possible conformational change of macromolecules at the charged surface.     

Neuronal pathways from low-threshold muscle and cutaneous afferents innervating tail to trunk muscle motoneurons in the cat

We studied neuronal pathways from low-threshold muscle (group I, II) and cutaneous afferents (group A(alpha)beta) innervating the tail to motoneurons innervating trunk muscles (m. iliocostalis lumborum and m. obliquus externus abdominus) in 18 spinalized cats. Stimulation of group I muscle afferents produced excitatory postsynaptic potentials or excitatory postsynaptic potentials followed by inhibitory postsynaptic potentials in all motoneurons innervating the m. iliocostalis lumborum which showed effects (32%), and predominantly inhibitory postsynaptic potentials in motoneurons innervating the m. obliquus externus abdominus (47%). Stimulation of group I+II afferents produced significant increases of the incidence of motoneurons showing postsynaptic potentials (the notoneurons innervating the m. iliocostalis lumborum, 87%; the motoneurons innervating the m. obliquus externus abdominus, 82%). The effects of low threshold cutaneous afferents were bilateral, predominantly producing inhibitory postsynaptic potentials in motoneurons innervating both muscles. These results suggest that neuronal pathways from muscle afferents to back muscle motoneurons mainly increase the stiffness of the trunk to maintain its stability, while those to abdominal muscles help to extend the dorsal column by decreasing their activities. The results also indicate that neuronal pathways from cutaneous afferents to trunk motoneurons functionallY disconnect the tail from the trunk.     

Systemic delivery of an adenovirus expressing EBV-derived vIL-10 in mice infected with Schistosoma mansoni or Leishmania amazonensis: controversial effects on the development of pathological parameters.

Within the context of microorganism/host interactions, those which last over weeks are expected to be sensitive to more or less sustained and targeted immuno-intervention, such as delivery of cytokines known to operate as down-regulators of acute inflammatory processes. IL-10 has received growing attention as a potential tool in immunotherapy, due to its anti-inflammatory and immunosuppressive properties. Therefore, using two experimental models of long-term interactions between parasites and laboratory mice, we monitored some effects of the systemic delivery of an adenovirus (Ad) expressing EBV-derived IL-10 (vIL-10) designated Ad-vIL-10. We first monitored the vIL-10 serum level following intranasal, intraperitoneal, intramuscular and intravenous administration. The i. p. and i.v. delivery of Ad-vIL-10 allowed a high serum level of vIL-10 (= 100 ng/ml), the i.v. route leading to a more sustained expression (up to 3 weeks). As a first model of parasite/mouse interaction, Schistosoma mansoni/C57Bl/6 mouse was selected. Ad-vIL-10 delivery was performed 4 weeks after S. mansoni infection i.e. at the time of egg-laying, and several parameters were monitored: (i) number of adult worms in the mesenteric vein, (ii) number of eggs trapped in the liver and intestine, (iii) liver fibrosis, (iv) serum levels of egg-reactive antibody subclasses, (v) serum content of cytokines, and (vi) cytokine production in the supernatant of antigen-stimulated mesenteric lymph node cells. No apparent effect was observed, either on the different parasitological parameters or on fibrosis development at day 70 of infection. Surprisingly, a marked increase in both Th1 and Th2 type cytokines was observed in the sera of the Ad-vIL-10 injected animals, as well as in the supernatants of their Ag-stimulated mesenteric lymph node cells. Nevertheless, polarization of the humoral response towards a Th2 profile was demonstrated by an increase in the IgE level in the Ad-vIL-10-injected animals. As far as the second model is concerned, namely the Leishmania amazonensis /C57Bl6 mouse interactions, Ad-vIL-10 was delivered intravenously one day before subcutaneous injection of stationary promastigotes and footpad swelling was monitored over 110 days. Under these conditions, vIL-10 exhibited a biphasic effect, decreasing the lesion size at the early stages of infection, but leading to a more pronounced lesion size during the chronic phase. This observation suggests a deactivation of the macrophage host cells under the influence of vIL-10. The results are discussed in the context of immunotherapy and the paradoxical effects observed in immunointervention with vIL-10.     

Plantlet regeneration via somatic embryogenesis in anther callus of Vitis latifolia L.

Anthers of Vitis latifolia L. (wild grape) cultured on Nitsch and Nitsch medium supplemented with 20 μM 2,4-D and 9 μM BAP produced callus after 4–6 weeks. Subculture of callus onto Nitsch and Nitsch medium containing 10 μM NAA produced somatic embryos within 6 weeks. On growth regulator-free Nitsch and Nitsch basal medium somatic embryos converted to plantlets in 6–8 weeks. One gram of callus produced more than 400 somatic embryos with 13.7% being converted to complete plantlets, which were subsequently established in soil. Regenerated plants were found to have mixoploid populations of cells, 2n = 38 and n = 19. Received: 23 May 1998 / Revision received: 21 September 1998 / Accepted: 10 October 1998     

Extracellular secretion of levansucrase from Zymomonas mobilis in Escherichia coli

Secretion of levansucrase from Zymomonas mobilis in Escherichiacoli by glycine supplement was investigated. A significant amount of levansucrase (about 25% of total activity) was found in intact whole-cells. Cell fractionation experiments showed that levansucrase was found both in the periplasmic space and in the cytoplasmic fraction of E. coli. None or only trace amounts of levansucrase was detected in the extracellular culture broth at 24 h of cultivation and it accrued with the increasing concentration of glycine in the culture medium and duration of the culture period. Optimal glycine concentration for the maximum secretion of levansucrase was in the range of 0.8-1%, in which approximately 20-50% of levansucrase was released into the extracellular fraction at 24 h of cultivation, although glycine retarded the bacterial growth.     

Application of fluorescence internal transcribed spacer-PCR (f-ITS) for the cluster analysis of bacteria isolated from air and deteriorated fresco surfaces

The aim of this work was the evaluation of fluorescence ITS-PCR (f-ITS) as a molecular tool to analyze the microbial community involved in the biodeterioration of cultural heritage surfaces. As a case study we analyzed by f-ITS ninety-two bacterial strains isolated from a medieval fresco and the surrounding air environment. The internal transcribed spacer between the 16S and 23S rRNA genes was amplified, and then the fluorescently labeled PCR products were separated by capillary electrophoresis. Bacterial strains were identified by 16S rDNA sequencing. The f-ITS electropherograms showed different profiles coherent with the affiliation of the strains at the genus and species levels. Among the isolates obtained from the fresco surface, those belonging to the genus Bacillus were the most prevailing exhibiting 8 different f-ITS profiles. The airborne bacilli exhibited only 2 of these 8 profiles. Staphylococcus were mostly isolated from air and produced 4 different profiles. Pseudomonas isolates presented 3 different profiles, and one of them was typical of Pseudomonas putida. Members of the other genera produced their distinctive profiles. Our results show that f-ITS is a promising molecular tool for the rapid selection and clustering of strains isolated from different sources.