A ground-based model to study the effects of weightlessness on lymphocytes

The mitogenic response of human lymphocytes was found to be markedly reduced in weightlessness conditions as compared to normal gravity. One possible explanation is that due to the non-existent sedimentation in space the lymphocytes could not adhere and spread on a substratum. Thus, we investigated the effect of substratum adhesiveness on lymphocyte responsiveness by reducing and blocking cell adhesion with poly-HEMA in a simple on-ground system. Lymphocyte adhesiveness was assessed by measuring the proportion of non-adhesive, slightly, and strongly adhesive 51Cr-radiolabelled cells on uncoated and poly-HEMA coated plastic. The amount of cell spreading on surfaces with varying adhesiveness was determined by measuring the area of cells. Cells grown on medium and thick poly-HEMA films were rounded in shape. By contrast, on tissue culture plastic, they showed clear signs of spreading. The mitogenic response of lymphocytes grown on thick poly-HEMA films was reduced by up to 68% of the control (tissue culture plastic). Interferon-gamma production was virtually nil when the cells were grown on the least adhesive substratum. These results show that activated lymphocytes need to anchor and spread prior to achieving an optimal proliferation response. We conclude that decreased lymphocyte adhesion could contribute to the depressed in vitro lymphocyte responsiveness found in the microgravity conditions of space flight.     

Fluidity of detergent micelles plays an important role in muscarinic receptor solubilization

In order to find a suitable reagent for extracting the muscarinic receptor from rat brain membranes 14 different detergents were tested. Only the plant glycoside digitonin efficiently solubilized the receptor protein in its native form. At the same time microviscosity of detergent micelles was determined by measuring the fluorescence polarization of a hydrophobic fluorescent probe diphenylhexatriene incorporated into the micelles. In the case of digitonin the polarization value was close to the corresponding value obtained for rat brain membrane fragments, while for the other detergents studied it remained considerably lower. The results obtained indicate that the fluidity of detergent micelles may play an important role in retaining the active conformation of the solubilized muscarinic receptor.     

DNA encapsidation by viruslike particles assembled in insect cells from the major capsid protein VP1 of B-lymphotropic papovavirus.

Capsids of polyomaviruses--small, nonenveloped DNA viruses--consist of the major structural protein VP1 and the minor structural proteins VP2 and VP3. The contributions of the individual capsid proteins to functions of the viral particle, such as DNA encapsidation, cell receptor attachment, entry, and uncoating, are still not clear. Here we show that viruslike particles assembled in nuclei of insect cells from VP1 of the monkey B-lymphotropic papovavirus (LPV) are sufficient to unspecifically encapsidate DNA. LPV VP1 expressed in large amounts in insect cells by a baculovirus vector assembled spontaneously in the nuclei to form viruslike particles. After metrizamide equilibrium density gradient purification and nuclease digestion, a fraction of these particles was shown to contain VP1-associated linear, double-stranded DNA with a predominant size of 4.5 kb. The fraction of DNA-containing VP1 particles increased with time and dose of baculovirus vector infection. The DNA-containing particles, further purified by sucrose gradient centrifugation, appeared as "full" particles in negative-staining electron microscopy. As shown by DNA hybridization, the encapsidated DNA consisted of insect cell and baculoviral sequences with no apparent strong homology to LPV sequences. Three non-LPV VP1-derived host proteins with apparent molecular masses of approximately 14, 15, and 16 kDa copurified with the DNA-containing particles and may represent insect cell histones encapsidated together with the DNA. A similar species of host DNA was also found in purified LPV wild-type virions. These data suggest that LPV VP1 alone can be sufficient to encapsidate linear DNA in a sequence-independent manner.     

Experimental revegetation of the regulated lake Ontojärvi in northern Finland

Water level regulation causes large-scale ecological changes in the littoral areas of lakes in northern Finland. If the summertime water level is raised, intensive erosion processes begin, causing a sudden decline in shore vegetation. The need for shore protection is obvious in areas of high recreational value. At lake Ontojärvi, planting experiments with littoral helophytes and bushes were carried out during the years 1990–92. All the experiments were carried out in the eroded sandy areas, which were partly protected by mechanical barriers. Several plant species were planted on the shore which had been treated with different peat mixtures, etc. The frequencies of the different species were followed monthly. After the first summer, the average survival rates were about 45 % due to the drying of seedlings. A rapid decrease in the survival rates took place during the high water level period in 1991 at which point only 20% of the planted individuals were alive. The best results were obtained for the helophyte Carex rostrata Stokes, of which 30% had survived erosion. Tall willows (Salix phylicifolia L.) were also erosion-resistant with a survival rate of 80%.     

The Saccharomyces Cerevisiae Ku Autoantigen Homologue Affects Radiosensitivity Only in the Absence of Homologous Recombination

In mammalian cells, all subunits of the DNA-dependent protein kinase (DNA-PK) have been implicated in the repair of DNA double-strand breaks and in V(D)J recombination. In the yeast Saccharomyces cerevisiae, we have examined the phenotype conferred by a deletion of HDF1, the putative homologue of the 70-kD subunit of the DNA-end binding Ku complex of DNA-PK. The yeast gene does not play a role in radiation-induced cell cycle checkpoint arrest in G(1) and G(2) or in hydroxyurea-induced checkpoint arrest in S. In cells competent for homologous recombination, we could not detect any sensitivity to ionizing radiation or to methyl methanesulfonate (MMS) conferred by a hdf1 deletion and indeed, the repair of DNA double-strand breaks was not impaired. However, if homologous recombination was disabled (rad52 mutant background), inactivation of HDF1 results in additional sensitization toward ionizing radiation and MMS. These results give further support to the notion that, in contrast to higher eukaryotic cells, homologous recombination is the favored pathway of double-strand break repair in yeast whereas other competing mechanisms such as the suggested pathway of DNA-PK-dependent direct break rejoining are only of minor importance.     

Zooplankton of Lake Peipsi-Pihkva in 1909–1987

164 taxa were identified in the net zooplankton of the pelagial of L. Peipsi-Pihkva in 1909–1987, including 3 species of protozoans, 74 species of rotifers, 58 species of cladocerans, 28 species of copepods and 1 mollusc. One rotifer species, Ploesoma peipsiense Mäemets et Kutikova, has been described as new for science here. The zooplankton of L. Peipsi-Pihkva is remarkably rich in species including rarities in Estonia: Limnosida frontosa, Drepanothrix dentata, Bythotrephes longimanus, B. cederstroemi etc. Due to its large surface area, L. Peipsi-Pihkva provides a large scale of biotopes of a diverse trophic state and humic content, which support species with different ecological requirements. Most of the aquatory of the lake has lately been mesotrophic, favouring the coexistence of indicators of oligo- and mesotrophic state and species preferring a higher trophic state. The occurrece of 10 species of the genus Bosmina including B. berolinensis, B. gibbera, B. lilljeborgi, B. thersites and B. crassicornis, sparse in Estonian lakes, is the most noteworthy feature of the zooplankton of L. Peipsi-Pihkva. The coexistence of B. coregoni and B. berolinensis, B. gibbera, B. lilljeborgi etc. which were earlier regarded as subspecies of B. c. coregoni proves that they are different species producing usually no hybrids. The species composition was subjected to certain changes during the years under consideration. Larvae of Dreissena were first found in zooplankton in 1962. The oligo-mesotrophic indicator Holopedium gibberum occurred in the lake in 1909–1964, but was lacking in later samples.     

Variable effects of air-drying on leaching losses from tree leaf litter

Leaching of soluble substances may be an important first step in leaf litter decomposition in small streams, but recent research has suggested that large leaching losses (up to 30% of initial mass in 48 h) may be an artifact created by using air-dried leaves in decomposition experiments. In laboratory experiments, we compared 3 d leaching losses from freshly fallen and air-dried senescent leaves of 27 tree species from different regions across Canada. Air-dried leaves from all species leached measurable amounts of original mass (3.6–32.8% dry mass), but leaching losses from fresh leaves (0–35%) were detectable in all but two species. Air-drying increased leaching losses in many species, but in others it reduced leaching losses or had no measurable effect. Results for leaves of the same species collected in different regions or in different years were generally similar, but species within the same genus often behaved very differently. Neither moisture content (fresh or air-dried), leaf thickness, nor cuticle thickness proved of any value as predictors of leaching losses or the effect of air-drying. The propensity of autumn-fallen leaves to leach, whether fresh or air-dried, appears to be a property of the individual tree species.     

A cladistic analysis ofAnisopappus (Asteraceae: Inuleae)

A cladistic analysis of the genusAnisopappus (Asteraceae: Inuleae) has been undertaken. A hypothesis of species interrelationships in the genus is presented for the first time. The analysis also includedArctotis (Arctoteae), used as outgroup, and five additional genera from theInuleae: Geigeria, Calostephane, Asteriscus, Buphthalmum, Pulicaria, andInula. It is concluded thatAnisopappus is a monophyletic group situated at the base of the tribe, diagnosed by, e.g., their obtuse stylar sweeping-hairs. The species with acute sweeping-hairs were found to be derived within the genus. Problems concerning species delimitation, biogeography and character evolution in the genus are briefly discussed.     

The nervous system of Tricladida. III. Neuroanatomy ofDendrocoelum lacteum andPolycelis tenuis (Plathelminthes,Paludicola): an immunocytochemical study

The organization of the nervous system ofDendrocoelum lacteum (Tricladida, Paludicola, Dendrocelidae) andPolycelis tenuis (Tricladida, Paludicola, Planariidae) was studied by immunocytochemical double staining, using neuropeptide RFamide and serotonin (5-HT) antisera on cryosections. The study confirmed the status of the main nerve cords (MCs) as the most important and stable of the longitudinal cords and supported the hypothesis of a common phylogenetic origin of the MCs in flatworms. The ganglion-like structures along the MCs at the beginning of transverse commissures and laterla branches showed a close contact with ventral fibres of the submuscular nerve plexus indicating an origin from crossing points of insunken ring commissures. The distributional pattern and morphology of the RFamide-IR cell bodies inD. lacteum corresponded to that of neurosecretory cells. Most RFamide-IR cells were unipolar and rounded while 5-HT-IR cells were uni- bi- and multipolar. The neutropile consisted of a dense RFamide-IR and a loose 5-HT-IR network. RFamide dominated in all parts of the genital plexus.     

Microbial oxidation of ebastine

The microbial oxidation of ebastine to carebastine was investigated. Among the 15 micro-organisms examined, only theCunninghamella strains showed the desired biotransformation.Cunninghamella blakesleeana oxidised the substrate within 7 days, via the intermediates alcohol and aldehyde, mainly to carebastine, the corresponding carboxylic acid. Optimisation of the culture conditions increased the yield from initially 10% up to a reproducible 40%. For the synthesis of carebastine a substrate concentration of 200 mg/l, a starting pH of 5.0 and the addition of 1% poly(vinyl alcohol) is favourable. The results achieved in experiments with shaking flasks are transferable to the fermentation scale and yielded 270 mg carebastine in a 3-l fermentation of 600 mg ebastine. The progress of the reaction was detected by TLC and HPLC, the products were identified by mass spectrometry and NMR.