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  • Articles  (744)
  • gene expression  (307)
  • Triticum aestivum  (229)
  • fish  (211)
  • Springer  (744)
  • 1995-1999  (744)
  • 1
    Electronic Resource
    Electronic Resource
    Springer
    Entomologia experimentalis et applicata 91 (1999), S. 403-412 
    ISSN: 1570-7458
    Keywords: aphid ; Diuraphis noxia ; Triticum aestivum ; endosymbiont ; insect-plant interactions ; phloem ; amino acids
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The feeding behavior of Diuraphis noxia Mordvilko (Homoptera: Aphididae) on susceptible hosts causes both ultrastructural and tissue level damage which may affect phloem composition. Genetic evidence suggests that endosymbiotic bacteria in most aphids overproduce limiting amino acids to benefit hosts but that D. noxia depends less on endosymbionts for these nutrients, possibly due to an enriched diet. To determine whether D. noxia feeding damage results in higher concentrations of essential amino acids, stylet exudates were analyzed from wheat (Triticum aestivum) damaged to different degrees. Comparison of samples from undamaged and damaged susceptible wheat revealed changes in amino acid composition and an increase in levels of essential amino acids, indicating a nutritionally enhanced ingesta. The changes in stylet exudates paralleled changes in leaf exudates, indicating that the effects are systemic. Feeding damage is not observed on a resistant wheat host, var. Halt, and leaf exudates from infested Halt did not show changes in amino acid composition. Mean relative growth of nymphs was significantly lower on Halt than on susceptible Arapahoe, indicating that Halt is a less suitable host. Both varieties show similar amino acid levels in non-infested samples, suggesting that D. noxia infestation does not enhance the phloem environment in Halt. This study provides evidence that aphid feeding can generate a nutritionally enhanced phloem diet.
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  • 2
    ISSN: 1570-7458
    Keywords: sieve element ; salivation ; aphid ; plant resistance ; wheat ; Sitobion fragariae ; Triticum aestivum
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Extended sieve element salivation (E1 waveform in the electrical penetration graph) is a characteristic activity during early sieve element punctures, particularly in resistant plants. In order to explore a chemically-mediated mechanism of resistance associated with sieve element salivation, we compared the pattern of feeding behaviour of the aphid, Sitobion fragariae (Walker), on two cultivars of the wheat Triticum aestivum L., with different concentrations of hydroxamic acids (Hx). During 24 h of electronic monitoring, aphids dedicated over 50% of the total time to phloem ingestion from the sieve elements. Total time allocated to E1 in the experiment, time to first E1 within the experiment, time allocated to E1 before a sustained phloem ingestion (E2) and the contribution of sieve element salivation to the phloem phase (E1/[E1+E2]) were significantly higher in the high-Hx cultivar. The increased salivation in plants with higher contents of Hx suggests the existence, at least in this system, of a chemically-mediated sieve element constraint.
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  • 3
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    Cellular and molecular life sciences 52 (1996), S. 942-949 
    ISSN: 1420-9071
    Keywords: Apoptosis ; transglutaminase ; signalling ; gene expression ; promoter elements ; retinoic acids
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Clarification of the molecular details of forms of natural cell death, including apoptosis, has become one of the most challenging issues of contemporary biomedical sciences. One of the effector elements of various cell death pathways is the covalent cross-linking of cellular proteins by transglutaminases. This review will discuss the accumulating data related to the induction and regulation of these enzymes, particularly of tissue type transglutaminase, in the molecular program of cell death. A wide range of signalling pathways can lead to the parallel induction of apoptosis and transglutaminase, providing a handle for better understanding the exact molecular interactions responsible for the mechanism of regulated cell death.
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  • 4
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    Cellular and molecular life sciences 51 (1995), S. 1116-1123 
    ISSN: 1420-9071
    Keywords: Antisense RNA ; gene expression ; insertional mutagenesis ; physical mapping ; reporter genes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Over the past ten years, powerful molecular genetic techniques have been developed to analyze gene function inDictyostelium. DNA-mediated transformation using a variety of selections and vectors has allowed the introduction of wild-type or modified genes that are under various forms of transcriptional control. Homologous recombination is efficient and can be used to modify the genome in precise ways. In addition, it is now possible to clone genes based on their mutant phenotype alone, either by insertional mutagenesis, or by screening antisense expression cDNA libraries. Finally, a nearly complete physical map of the genome is available and so genes are easily mapped by physical techniques. We discuss many of these advances within the context of major research problems presently under study.
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  • 5
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    Cellular and molecular life sciences 51 (1995), S. 606-611 
    ISSN: 1420-9071
    Keywords: Metallothionein ; isometallothioneins ; gene expression ; rabbit kidney cell-line ; cadmium adaptation ; zinc adaptation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract We explored the molecular genetics underlying the massive induction of isoMTs by Zn2+ or Cd2+ in metal tolerant rabbit kidney (RK-13) sub-line cells, using band shift assays and Southern blotting analysis. In sub-line cells accommodated to intermediate metal concentrations (100 μM Zn2+; 1–20 μM Cd2+) evidence suggested that the increase in the capacity for isoMT synthesis is brought about by an increased binding activity of the nuclear transcription factors MTF-1 and Sp1. Using quantitative band shift analysis with a mouse MRE-d oligonucleotide probe, the binding of both transcription factors was found to be enhanced two to three times over the binding activity measured in the unexposed parental RK-13 cells. Their increase in binding activity is probably the cause of the overexpression of MT genes and the development of metal tolerance in these cells. In cells tolerant to the highest concentrations of metal the analysis of Southern blot signals revealed MT gene amplification to be the most probable cause of the increased MT production. Thus, in cells of sub-lines growing in the presence of 350 μM Zn2+, two of the isoMT genes were coordinately triplicated and in cells tolerant to 150 μM Cd2+ one isoMT gene was amplified two-fold.
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  • 6
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    Cellular and molecular life sciences 51 (1995), S. 768-774 
    ISSN: 1420-9071
    Keywords: Cortisol ; stress ; heat ; Antarctic ; fish
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Radioimmunoassay was used to determine levels of the stress-inducible glucocorticoid, cortisol, circulating in the plasma of the extremely stenothermal Antarctic fishPagothenia borchgrevinki at rest and after heat stress. Fish sampled immediately after capture (−1.9°C) had low cortisol levels (10.4±1.4 ng ml−1, mean±SEM) as did fish which were laboratory rested for 3 days. Sudden exposure to 5°C (48h) resulted in a peak cortisol value after 3 h (69.9±6.8 ng ml−1) whereas exposure to 8°C (6h) resulted in a peak value after 1 h (73.5±8.0 ng ml−1). At both temperatures levels remained significantly elevated (p〈0.05) for the entire period of exposure. Increased temperature also resulted in a significant change in haemoglobin, haematocrit and mean cell haemoglobin concentration (MCHC) (p〈0.05). Plasma lactate was significantly elevated only after exposure to 8°C (p〈0.05). Plasma cortisol levels fromP. borchgrevinki are reported here for the first time and show this cryopelagic Antarctic species to have an unusual hormonal stress profile.
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  • 7
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    BioMetals 11 (1998), S. 345-358 
    ISSN: 1572-8773
    Keywords: calcium ; CREB ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Through the evolution of multicellular organisms, calcium has emerged as the preferred ion for intracel-lular signalling. It now occupies a pivotal role in many cell types and nowhere is it more important than in neurons, where it mediates both the relaying and long-term storage of information. The latter is a process that enables learning and memory to be formed and requires the activation of gene expression by calcium signals. Evidence from a number of diverse organisms shows that transcription mediated by the transcrip-tion factor CREB is critical for learning and memory. Here we review the features of CREB activation by calcium signals in mammalian cells. In contrast to other transcription factors, its regulation is dependent on an elevation of nuclear calcium concentration, potentially placing this spatially distinct pool of calcium as an important mediator of information storage.© Kluwer Academic Publishers
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  • 8
    ISSN: 1432-1890
    Keywords: Key words Glomus mosseae ; Hydroponics ; Nitrate uptake ; Root respiration ; Triticum aestivum
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  Oxygen and CO2 fluxes were measured in hydroponically grown mycorrhizal and non-mycorrhizal Triticum aestivum L. cv. Hano roots. The NO3 – uptake of the plants was used to estimate the amount of root respiration attributable to ion uptake. Plants were grown at 4 mM N and 10 μM P, where a total and viable mycorrhizal root colonisation of 48% and 18%, respectively, by Glomus mosseae (Nicol. and Gerd.) Gerd. and Trappe (BEG 107) was observed. The O2 consumption and NO3 – uptake rates were similar and the CO2 release was higher in mycorrhizal than in non-mycorrhizal wheat. This resulted in a significantly higher respiratory quotient (RQ, mol CO2 mol–1 O2) in mycorrhizal (1.27±0.13) than in non-mycorrhizal (0.79±0.05) wheat. As the biomass and N and P concentrations in mycorrhizal and non-mycorrhizal wheat were the same, the higher RQ resulted from the mycorrhizal colonisation and not differences in nutrition per se.
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  • 9
    ISSN: 1432-0789
    Keywords: 15N-labelled fertilizer ; Added N interaction Fertilizer N uptake ; Soil N uptake ; Wheat ; Triticum aestivum
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Geosciences , Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract The effects of 15N-labelled ammonium nitrate, urea and ammonium sulphate on yield and uptake of labelled and unlabelled N by wheat (Triticum aestivum L. cv. Mexi-Pak-65) were studied in a field experiment. The dry matter and N yields were significantly increased with fertilizer N application compared to those from unfertilized soil. The wheat crop used 64.0–74.8%, 61.5–64.7% and 61.7–63.4% of the N from ammonium nitrate, urea and ammonium sulphate, respectively. The fertilizer N uptake showed that ammonium nitrate was a more available source of N for wheat than urea and ammonium sulphate. The effective use of fertilizer N (ratio of fertilizer N in grain to fertilizer N in whole plant) was statistically similar for the three N fertilizers. The application of fertilizer N increased the uptake of unlabelled soil N by wheat, a result attributed to a positive added N interaction, which varied with the method of application of fertilizer N. Ammonium nitrate, urea and ammonium sulphate gave 59.3%, 42.8% and 26.3% more added N interaction, respectively, when applied by the broadcast/worked-in method than with band placement. A highly significant correlation between soil N and grain yield, dry matter and added N interaction showed that soil N was more important than fertilizer N in wheat production. A values were not significantly correlated with added N interaction (r=0.719). The observed added N interaction may have been the result of pool substitution, whereby added labelled fertilizer N stood proxy for unlabelled soil N.
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  • 10
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    Biology and fertility of soils 23 (1996), S. 327-331 
    ISSN: 1432-0789
    Keywords: CO2 emission ; Field method ; Soil respiration ; Triticum aestivum ; Soil moisture ; Carbon reservoirs ; Greenhouse effect ; Grey forest soil ; Mineralization
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Geosciences , Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract The dynamics of the rate of CO2 evolution from soil in fallow and croplant under spring wheat (Triticum aestivum L.) was studied in a crop rotation in grey forest soil of the Baikal forest-steppe during the growing season and in different years. It was shown that the regional characteristics of soils and hydrothermal conditions in different years affect the rate of CO2 evolution in agroecosystems. The seasonal dynamics of CO2 is characterized by insignificant changes in the autumn to spring period and enhanced emission in hot and dry summers. CO2 evolution is assumed to increase due to enhanced mineralization and partial diffusion from the carbonate horizon at the depth of the seasonal frost. During the growing season the dynamics of CO2 evolution depends on the soil moisture regime. There was a strong correlation between the rate of CO2 emission and soil moisture in the particularly dry year of 1993 (η=0.86) and a moderate correlation in the other years (η=0.38–0.54). The effect of the previous crop and fertilizer application on the rate of CO2 emission was insignificant. In a continuous fallow the total carbon release into the atmosphere varied throughout the years studied from 558 to 1880 kg ha-1. Humus losses varied from 0.9% to 3.1%.
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  • 11
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    Biology and fertility of soils 23 (1996), S. 327-331 
    ISSN: 1432-0789
    Keywords: Key words CO2 emission ; Field method ; Soil respiration ; Triticum aestivum ; Soil moisture ; Carbon reservoirs ; Greenhouse effect ; Grey forest soil ; Mineralization
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Geosciences , Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract The dynamics of the rate of CO2 evolution from soil in fallow and cropland under spring wheat (Triticum aestivum L.) was studied in a crop rotation in grey forest soil of the Baikal forest-steppe during the growing season and in different years. It was shown that the regional characteristics of soils and hydrothermal conditions in different years affect the rate of CO2 evolution in agroecosystems. The seasonal dynamics of CO2 is characterized by insignificant changes in the autumn to spring period and enhanced emission in hot and dry summers. CO2 evolution is assumed to increase due to enhanced mineralization and partial diffusion from the carbonate horizon at the depth of the seasonal frost. During the growing season the dynamics of CO2 evolution depends on the soil moisture regime. There was a strong correlation between the rate of CO2 emission and soil moisture in the particularly dry year of 1993 (η=0.86) and a moderate correlation in the other years (η=0.38–0.54). The effect of the previous crop and fertilizer application on the rate of CO2 emission was insignificant. In a continuous fallow the total carbon release into the atmosphere varied throughout the years studied from 558 to 1880 kg ha–1. Humus losses varied from 0.9% to 3.1%.
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  • 12
    ISSN: 1432-0789
    Keywords: Key words Phosphate-solubilizing microorganisms ; Mussoorie rock phosphate ; Vesicular-arbuscular mycorrhizae ; Triticum aestivum ; Nutrient-deficient soils
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Geosciences , Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract  The effect of inoculating wheat (Triticum aestivum L.) with the PO4 3–-solubilizing microorganisms (PSM) Bacillus circulans and Cladosporium herbarum and the vesicular-arbuscular mycorrhizal (VAM) fungus Glomus sp. 88 with or without Mussoorie rock phosphate (MRP) amendment in a nutrient-deficient natural sandy soil was studied. In the sandy soil of low fertility root colonization by VAM fungi was low. Inoculation with Glomus sp. 88 improved root colonization. At maturity, grain and straw yields as well as N and P uptake improved significantly following inoculation with PSM or the VAM fungus. These increases were higher on combined inoculation of PSM and the VAM fungus with MRP amendment. In general, a larger population of PSM was maintained in the rhizosphere of wheat in treatments with VAM fungal inoculation and MRP amendment. The results suggest that combined inoculation with PSM and a VAM fungus along with MRP amendment can improve crop yields in nutrient-deficient soils.
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  • 13
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    Biology and fertility of soils 23 (1996), S. 273-281 
    ISSN: 1432-0789
    Keywords: Wheat ; Triticum aestivum ; Rhizosphere ; Soil microflora ; Gram-negative bacteria ; API 20 NE ; Flavobacterium spp ; Cytophaga
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Geosciences , Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract We identified 161 Gram-negative bacterial strains isolated from the root surface of wheat grown under different soil conditions. The strains were divided into seven groups based on major morphological and physiological properties. Taxonomic allocation of the groups was verified by guanine+cytosine contents of DNA. Except for one group, which may be assumed to include bacteria belonging to the genera Flavobacterium and Cytophaga, the various groups were taxonomically united. The distribution of the groups changed with soil improvement. Pseudomonads predominated in unimproved soil, but Flavobacterium and Cytophaga spp. were predominant in the most improved soil. As all the strains were non-fermentative by Hugh and Leifson's test, API 20NE identification was applied. However, many strains were misidentified by this system, especially in the Flavobacterium and Cytophaga spp. group. For ecological studies, the strains were classified to species level by the API 20 NE system and by the results of a combination of guanine+cytosine (mol%) and isoprenoid quinone data. The pattern of distribution of the bacteria on the root surface of wheat varied at species level within one genus depending on soil conditions.
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  • 14
    ISSN: 1432-0975
    Keywords: Key words Dispersal ; coral reefs ; fish ; swimming ; Larvae ; Juveniles
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Geosciences
    Notes: Abstract  Characteristics of the pelagic stages of reef fishes have generally been investigated at the family level, which may mask important differences among species. Here the variation in sustained swimming ability of the late pelagic stages is examined among species, within two families (Pomacentridae and Chaetodontidae). The pomacentrids displayed a 7.5-fold difference in sustained swimming ability across 24 species, while the chaetodontids displayed a 2-fold difference across 10 species. The variation within the Pomacentridae was not related to pelagic larval duration, post-settlement habitat or taxonomy. There was, however, a significant correlation between sustained swimming ability and total length (TL) of individuals (r=0.435, P〈0.0001). Differences in the mean distance swum by pomacentrid species, however, was most strongly related to differences in mean wet weight (r=0.814, P〈0.0001). When the mean distance swum by species was scaled with respect to mean TL there was still a strong correlation with mean wet weight (r=0.644, P〈0.005). Among chaetodontid individuals TL and sustained swimming ability were not correlated (r=−0.004, P=0.978). Furthermore, sustained swimming ability was not significantly related to the trans-oceanic distribution of species in either family. The variation in sustained swimming ability, however, may contribute to explanations of the observed levels of gene flow within populations.
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  • 15
    ISSN: 1436-5073
    Keywords: mercury ; fish ; collaborative study ; atomic absorption spectrometry
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract One of the major analytical problem in the analysis of fish tissues for total mercury is the dissolution of the sample. This paper compares two different methods of wet digestion (microwave and closed pressurized vessel), followed by FIAAS as final determination and a Zeeman-solid-sample-AAS determination method without previous mineralization. Six fish samples were selected for this study. The top, the central and the bottom portions of the fish were analyzed separately. The mercury contents in the bottom portions determined by Zeeman-solid-sample-AAS were significantly higher than those obtained by the other two systems. A good correlation was observed among the results of the different techniques.
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  • 16
    ISSN: 1436-5073
    Keywords: microwave digestion ; fish ; mercury ; CVAAS
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract Food samples digestion, in view to quantify total mercury, is the first step to perform before measuring mercury by cold vapour atomic absorption spectrometry (CVAAS). We have compared two microwave digestion systems, one working at atmospheric pressure (open system) and the other one under pressure (closed system). Results obtained for fish muscle samples by the two methods are in good agreement. However, fat is not digested in the open system, whereas in the closed system no more fat remains in the final solution. During the quantification step, applying the CVAAS technique, we have noticed that the choice and the concentration of the reductant solution is very important to obtain good results. Elaborated methods have been validated using certified reference materials.
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  • 17
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    Plant molecular biology reporter 16 (1998), S. 323-339 
    ISSN: 1572-9818
    Keywords: Aux/IAA genes ; gene expression ; gene families ; RT-PCR ; tomato
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We have developed an improved method for determination of gene expression levels with RT-PCR. The procedure is rapid and does not require extensive optimization or densitometric analysis. Since the detection of individual transcripts is PCR-based, small amounts of tissue samples are sufficient for the analysis of expression patterns in large gene families. Using this method, we were able to rapidly screen nine members of the Aux/IAA family of auxin- responsive genes and identify those genes which vary in message abundance in a tissue- and light-specific manner. While not offering the accuracy of conventional semi-quantitative or competitive RT-PCR, our method allows quick screening of large numbers of genes in a wide range of RNA samples with just a thermal cycler and standard gel analysis equipment.
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  • 18
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    Plant molecular biology reporter 17 (1999), S. 371-383 
    ISSN: 1572-9818
    Keywords: epidermal peel ; extraction ; gene expression ; stomata ; tree tobacco
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Stomatal guard cells are critical for maintenance of plant homeostasis and represent an interesting cell type for studies of leaf cell differentiation and patterning. Here we describe techniques for the isolation of guard cell RNA and protein from blended epidermal peels of Nicotiana glauca. The RNA isolation procedure is a modification of the hot borate method, which is particularly well-suited for recalcitrant tissues. Protein was extracted by disrupting guard cell-enriched epidermis with a French® press. This system offers the following advantages: relatively high yield, low or no contamination by other cell types, fresh tissue as a source of RNA and protein rather than protoplasts, and a plant species that is readily transformable. These techniques will allow for cloning and analysis of genes expressed in guard cells, application of traditional biochemical techniques to guard cell proteins, as well as characterization of genetic manipulation of guard cell function in transgenic plants.
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  • 19
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    Nutrient cycling in agroecosystems 53 (1999), S. 157-175 
    ISSN: 1573-0867
    Keywords: Brassica napus ; Cicer arietinum ; current P ; Lens culinaris ; Lupinus albus ; Lupinus angustifolius ; P concentration response ; P content response ; Pisum sativum ; previous P ; sigmoid response ; single superphosphate ; Triticum aestivum ; Vicia faba ; yield response
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Phosphorus (P) is a major deficiency of soils of south-western Australia (WA). The fertilizer P requirements are not known for grain legumes being evaluated for neutral to alkaline, fine textured soils in WA. To rectify this, glasshouse and field experiments were undertaken to compare the responses of several grain legume species, wheat and canola to applications of single superphosphate and the results are reported in this paper. The glasshouse experiments measured responses of dried tops, harvested at 26 to 42 days after sowing, to P that was freshly-applied (current P) and previously-applied (previous P). Responses in the glasshouse were measured using yield, P concentration and P content (P concentration multiplied by yield) of oven dried tops of the following: wheat (Triticum aestivum), canola (Brassica napus), faba bean (Vicia faba), chickpea (Cicer arietinum), lentil (Lens culinaris), field pea (Pisum sativum), albus lupin (Lupinus albus) and narrow leaf lupin (Lupinus angustifolius). Field experiments in 1994 and 1995 compared seed (grain) yield responses of faba bean, chickpea, lentil, albus lupin and wheat to applications of current P. The P was banded (drilled) with the seed while sowing at 5 cm depth. Canola and wheat produced very large yield responses to increasing applications of current P. Responses were much smaller for albus lupin, faba bean and chickpea. Responses for lentil, narrow leaf lupin and field pea, fell in between responses of the small and large seeded species. Similar trends for responses were obtained as measured using yield, P concentration, or P content. For soils treated with previous P, similar trends were observed as for current P, but differences in yield responses between species were much less marked and the response curves tended to become more sigmoid. In the field experiments, grain yield responses to current P of albus lupin and chickpea were less than that for wheat. Relative to wheat, faba bean was the most responsive grain legume to applications of current P, with lentil producing similar responses to wheat in one experiment at a newly cleared, P deficient site.
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  • 20
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    Nutrient cycling in agroecosystems 44 (1995), S. 205-215 
    ISSN: 1573-0867
    Keywords: Incubation of phosphorus in soil ; relative effectiveness ; superphosphate ; ×Triticosecale ; Triticum aestivum
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Single superphosphate was incubated for six months at 25°C in soil which had been subject to one of three moisture treatments. These were: dried in a glasshouse, dried at a constant temperature of 25°C, or moist soil. Phosphorus (P) effectiveness was then compared with effectiveness of P from freshly-applied superphosphate using yields of wheat (Triticum aestivum) and triticale (×Triticosecale) tops in pot experiments. Incubation in soil which had been dried at 25°C did not decrease the effectiveness of the P. Incubation in moist soil decreased it to about 20% of the effectiveness of freshly-applied P in one case and to about 50% in the other case. Incubation in soil which had been dried in a glasshouse also decreased its effectiveness. The decrease varied with conditions, but in two cases the P was 70% as effective as freshly-applied P, and in one case only 45% as effective. Presumably sufficient moisture was present in the soil dried in the glasshouse to enable water-soluble P present in the fertilizer to react with the soil.
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  • 21
    ISSN: 1572-977X
    Keywords: culverts ; culvert trap ; mosquito impoundment ; seine ; fish
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Fish population dynamics in a 24.3 ha mangrove-dominated mosquito impoundment in east-central Florida were examined by seining and culvert traps before and after installation of culverts that established estuarine connection for the first time in 39 years. In a 27-day period following the culvert opening, fish species increased from 9 to 21, while total number of fish in the impoundment decreased. Movement of fishes through culverts in both directions commenced immediately following culvert opening. Recruitment of transient species into the impoundment appeared to key on a single wind-driven high tide event. Such short-term events may be important cues for fish movement into and out of impounded salt marshes.
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  • 22
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    Cellular and molecular life sciences 52 (1996), S. 888-891 
    ISSN: 1420-9071
    Keywords: Ageing ; rat ; brain ; gene expression ; differential display
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract We have used the polymerase chain reaction (PCR)-based technique of differential display to analyse changes in gene expression during ageing of the rat brain. In this approach we have compared three young adult (6 months) with three old adult (20 months) animals. RNA preparations from the homogenised brains were subjected to reverse transcriptase (RT)-PCR using 36 different combinations of primer pairs. Any PCR product which was consistently found to be more prominent in the three young brains compared to the three old brains, and vice versa, was scored as potentially representing a gene which was differentially expressed during the ageing of this tissue. Out of a possible 2000+PCR products we identified 44 that might represent genes that exhibit differential expression during ageing of the rat brain. An initial screen of these fragments, by Southern-blotting the PCR products and hybridising them with cDNA probes derived from either young or old brain RNA preparations, indicated that 40% of them represented genes that were differentially expressed. This approach is likely to prove invaluable for identifying cohorts of genes that show differential expression during the ageing process.
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  • 23
    ISSN: 1420-9055
    Keywords: Phoxinus phoxinus ; fish ; Diplostomum phoxini ; Macrolecithus papilliger ; parasitism
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The influence of parasites on the breeding biology of the European minnow (Phoxinus phoxinus) was studied by examining 268 adult and 59 young-of-the-year minnows from an artificial Swiss Alpine lake for parasites during the breeding seasons in 1990 and 1991. Two trematode parasites;Diplostomum phoxini, in their brains, andMacrolecithus papilliger, in their guts, were found. The prevalence ofD. phoxini was 100% in both years. The mean abundance was 231 in 1990 and 448 in 1991; the difference was significant. Prevalence (and abundance) ofM. papilliger was 37.5% (1.04±0.39) in 1990 and 59.5% (2.68±0.55) in 1991; also significantly higher in 1991 than in 1990. No differences in prevalence or abundance between males and females were found. Numbers of both parasites were significantly correlated with fish length. There was seasonal variation in the abundance ofD. phoxini metacercariae, with a peak in June/July in both years. The prevalence ofM. papiller decreased after July in both years. Distribution of the two trematodes was overdispersed as indicated by the variance to mean (abundance) ratio. The observed seasonal variation inD. phoxini abundance also suggests that sublethal parasites could influence the population dynamics of their hosts.
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  • 24
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    Aquatic sciences 57 (1995), S. 106-118 
    ISSN: 1420-9055
    Keywords: Benthos ; fish ; vertical distribution ; predation risk ; selective fishing ; Lake Constance
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract To compare the vertical distributions of benthos and fish species of Upper Lake Constance (Bodensee), 552 benthos samples were taken with an Auerbach grab at four transects (10–250 m). An additional 54 samples (1–10 m) were collected elsewhere. Complementary data was taken from the literature. In winter, most fish feed on benthos, mainly chironomids, but in summer less than 5% do so. Formerly the bottom gill-netting aimed mainly at catchingCoregonus pidschian, but it now aims atPerca fluviatilis. Contrary to expectation, in winter the greatest abundance ofP. fluviatilis is much deeper (55 m) than that of its preferred food, i.e. chironomids (〈20 m). Comparably,C. pidschian also stayed deeper than its main food source (chironomids and mollusks).Lota lota, Salvelinus profundus andS. alpinus — unlike the unwanted cyprinids — also stay or stayed deeper than the attractive chironomids. It is argued that the fish reduce their “predation risk”, i.e. the risk of being gillnetted in the upper 50 m, by “selecting” the deeper less dangerous, but also less food-abundant habitat. Balancing predation risk against food abundance may have led to “survival of the deepest”.
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    Sexual plant reproduction 8 (1995), S. 266-272 
    ISSN: 1432-2145
    Keywords: Triticum aestivum ; Alloplasmic lines ; Apomixis ; Parthenogenesis ; Protein patterns ; 2-D gel electrophoresis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract An in vivo model system to study the initiation of embryo development is presented. From the so-called Salmon system of wheat (alloplasmic lines with a 1BL-1RS chromosome translocation), three completely isogenic and homozygous lines were produced by selection for uniformity in about 20 selfing/backcross generations as well as between sublines of doubled haploids. The line (aestivum)-Salmon is male fertile and sexual. The lines (caudata)-Salmon and (kotschyi)-Salmon are male sterile and have a parthenogenetic capacity of about 90%. The expression of nuclear-cytoplasmic male sterility is different for the two parthenogenetic lines. The initiation of autonomous embryo development at defined developmental stages of the ovaries and the maximum degree of parthenogenesis are identical in both parthenogenetic lines as proved by the auxin test and progeny analyses. The protein patterns from ovary extracts of the three isogenic lines were identical for more than 200 spots of 2-D polyacrylamide gels, confirming their homogeneity. However, one protein (P 115.1) was found 3 days before and during anthesis only in ovaries of the parthenogenetic lines. It seems to be involved in the initiation of parthenogenesis.
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  • 26
    ISSN: 1432-2145
    Keywords: Key words Male sterility ; Starch ; Triticum aestivum ; Water stress ; Anther ; Pollen
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  Water deficit during meiosis in microspore mother cells of wheat (Triticum aestivum L.) induces male sterility, which reduces grain yield. In plants stressed during meiosis and then re-watered, division of microspore mother cells seems to proceed normally, but subsequent pollen development is arrested. Stress-affected anthers generally lack starch. We employed light microscopy in conjunction with histochemistry to compare the developmental anatomy of water-stress-affected and normal anthers. The earliest effects of stress, detectable between meiosis and young microspore stages, were the degeneration of meiocytes, loss of orientation of the reproductive cells, and abnormal vacuolization of tapetal cells. Other effects observed during subsequent developmental stages were deposition of starch in the connective tissue where it is normally not present, hypertrophy of the middle layer or endothecial cells, and deposition of sporopollenin-like substances in the anther loculus. The resulting pollen grains lacked both starch and intine. These results suggest that abnormal degeneration of the tapetum in water-stressed anthers coupled with a loss of orientation of the reproductive cells could be part of early events leading to abortion of microspores.
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  • 27
    ISSN: 1432-203X
    Keywords: glyphosate ; gene expression ; gene amplification ; cell culture ; resistance
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The stability and expression of amplified 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) genes was examined in glyphosate resistant tobacco cells grown in glyphosate-free medium, and in plantlets regenerated from resistant cells. Amplified DNA was maintained in resistant cells grown in the absence of glyphosate for three years. Amplified EPSPS genes were retained in regenerated plantlets at levels comparable to those observed in the resistant cells, and EPSPS mRNA was overexpressed (compared to unselected plantlets). However, glyphosate resistance in cell lines grown in glyphosate-free medium declined 7-fold, and in regenerated plantlets approximately 20-fold, compared to resistant cells maintained under glyphosate selection. In plantlets, reduced resistance correlated with lower levels of EPSPS mRNA. Plantlets regenerated from resistant cells exhibited morphological variation, and had an approximate doubling of their nuclear genome size.
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    Plant cell reports 16 (1997), S. 663-667 
    ISSN: 1432-203X
    Keywords: Key words Embryogenesis ; Ovule culture ; Triticum aestivum ; Zygote
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Ovules of the wheat breeding line Veery #5 were excised and transferred to culture within 24 h after pollination. When ovules were cultured on Phytagel-solidified medium, and the pericarp removed exclusively at the micropylar tip and the abaxial side, zygotes from up to 79.2% of the ovules underwent embryogenesis with the same developmental pattern as found in planta. Embryos from more than 50% of the cultured ovules germinated when transferred to regeneration medium. More than 100 plantlets were randomly chosen for transfer to soil, all of which developed to phenotypically normal and fertile plants. With this system, the entire process of zygotic embryogenesis can be studied using living material. Furthermore, the method could be used as an embryo rescue technique for plant breeding purposes.
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  • 29
    ISSN: 1432-203X
    Keywords: Key words Abscisic acid ; Anther culture ; Light ; Metallothionein ; Pollen embryogenesis ; Triticum aestivum
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A cloned cDNA to the wheat (Triticum aestivum) early cysteine-labeled metallothionein has many characteristics of a molecular marker for pollen embryogenesis in this plant. This transcript was not detected in uninucleate microspores at the time of culture or in pollen at any stage during normal ontogeny; its mRNA did begin to increase in embryogenic microspores within 6 h of culture, peaked at around 24 h, declined, then leveled off through the 21-day-old embryoid stage. Additionally, the accumulation of the embryoid-abundant EcMt gene transcript showed a direct and positive correlation with an increase of ABA in embryogenic microspores and developing pollen embryoids. Irradiating cultures with high intensity white light or with far-red, or blue light, suppressed EcMt transcript accumulation and the ability of microspores to form embryoids; however, light did not affect ABA concentrations during the early stages of culture. These results suggest that although a promoter of pollen embryogenesis in bread wheat, ABA alone cannot maintain the sporophytic differentiation of microspores subjected to inhibitory regimes of light in vitro. Whether or not light acts directly or indirectly in suppressing EcMt gene expression and pollen embryogenesis remains unknown.
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    Journal of thermal analysis and calorimetry 57 (1999), S. 283-291 
    ISSN: 1572-8943
    Keywords: DSC ; fish ; general proteolytic activity ; pyloric caeca ; ripening ; salted herring product
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract The thermoanalytical behaviour of pyloric caeca during salting and ripening was investigated using a Perkin Elmer DSC 7. Not only the thermal stability of the muscle proteins was influenced by salting but also that of pyloric caeca. It was recognised that the salting itself leads to a remarkable increase of the transition temperature compared with raw herring. An influence of the salt:fish ratio could be observed. The higher the salt content the higher the increase of the denaturation temperature. During ripening the transition temperature remained on a high level or showed only a slight decrease during the investigation period. The dependency from the salt content remained evident. The increase of the transition temperature was accompanied by a decrease of the transition enthalpy. The increase of thermal stability is connected with a decrease of the general proteolytic activity in pyloric caeca. Possibly, the enzymes are diffusing from the pyloric caeca into the muscle and cause there an increase of enzymatic activity observable in North Sea herring accompanied by a decrease of activity in pyloric caeca itself. Simultaneous the thermal stability of pyloric caeca is lowered. The reason for the differences in ripening could be seen in some enzyme-inhibiting factors unknown until now.
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    Water, air & soil pollution 85 (1995), S. 303-308 
    ISSN: 1573-2932
    Keywords: Acidification ; fish ; forestry ; water-chemistry ; management ; calcium ; aluminium ; TOC
    Source: Springer Online Journal Archives 1860-2000
    Topics: Energy, Environment Protection, Nuclear Power Engineering
    Notes: Abstract The soils and waters in the Galloway hills of south-west Scotland have undergone considerable acidification during the present century. The aims of this study were to examine the extent to which afforestation can influence water chemistry, and the occurrence of fish stocks within an individual catchment in this area. Studies were carried out at 45 sites in the Bladnoch catchment during 1994. River water samples were collected at all sites and determinations made of pH, total organic carbon, calcium and monomeric aluminium concentrations. In order to quantify the size and abundance of fish populations, electrofishing was carried out at 21 of the sites. Estimates of afforestation were made by a combination of field surveys and the use of 1∶:10 000 scale ordnance survey maps. The presence of coniferous trees had a substantial affect on water chemistry. In areas where the whole catchment was afforested, the water pH was 0.7 units lower than in unafforested areas (p〈0.001). Afforestation was also correlated with low concentrations of calcium, and increases in the concentrations of monomeric aluminium and total organic carbon. There was a clear relationship between fish populations and water chemistry. Numbers of trout were positively correlated with pH (r 2=0.69; p〈0.001), and with concentrations of calcium (r 2=0.55; p〈0.001). The study has shown that salmonid fish populations are sensitive to acidity and the associated changes in river water chemistry; these problems were exacerbated by afforestation.
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  • 32
    ISSN: 1573-2932
    Keywords: Na+ flux ; Cl− regulation ; chloride cells ; anal papillae ; acid-sensitivity ; crustaceans ; insect larvae ; fish
    Source: Springer Online Journal Archives 1860-2000
    Topics: Energy, Environment Protection, Nuclear Power Engineering
    Notes: Abstract Fishes, Zooplankton, insect larvae, and benthic invertebrates differ in their sensitivity to acidic waters. Some species are able to survive and complete their life-cycle below pH 3.5 while others are eliminated once the pH drops below 5.5. Generally, acid-sensitive fauna are highly water permeable and have difficulty regulating osmotically essential ions, such as sodium and chloride, at low pH. Increased permeability during certain stages of a life-cycle (post-molt crayfish, for example) are often associated with increased acid-sensitivity. Special adaptations, including enlarged anal papillae in Chironomids and the number and morphology of chloride cells in such diverse organisms as crustaceans, insect larvae, and fishes, may enhance acid-tolerance. To test the hypothesis that Na+ regulation can be used to predict relative acid-sensitivity of aquatic fauna we will need a mechanism for standardizing Na+ regulatory capability.
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    Water, air & soil pollution 85 (1995), S. 889-894 
    ISSN: 1573-2932
    Keywords: acidification ; acid neutralizing capacity ; calcite ; fish ; liming ; mitigation ; ecosystems
    Source: Springer Online Journal Archives 1860-2000
    Topics: Energy, Environment Protection, Nuclear Power Engineering
    Notes: Abstract The Experimental Watershed Liming Study (EWLS) evaluated the application of CaCO3, to a forested watershed to mitigate the acidification of surface water. During October 1989, 6.9 Mg CaCC3/ha was applied by helicopter to two subcatchments of about 50% (102.5 ha) of the Woods Lake watershed area. The EWLS team investigated the response to treatment of soils (chemistry and microbial processes), vegetation, wetland, stream and lake waters, and phytoplankton and fish, and applied the Integrated Lake Watershed Acidification (ILWAS) model in predicting a watershed treatment duration of up to 50 years. Observations showed a gradual change in pH, acid neutralizing capacity (ANC) and Ca2+ in the water column; direct lake additions of CaCO3 (three different times) were characterized by abrupt changes following base addition and subsequent rapid reacidification. Moreover, the watershed treatment eliminated the snowmelt acidification of the near-shore region of the lake observed during direct lake treatments. Positive ANC water in the tributary and near-shore area improved conditions for fish reproduction and for a viable fish population. Budgets for 12-month periods before and after the watershed treatment showed that the lake shifted from a source of ANC to a sink due to retention of elevated inputs of Ca2+ from the watershed CaCO3 application.
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  • 34
    ISSN: 1573-4919
    Keywords: vasoactive intestinal peptide ; ulcerative colitis ; gene expression
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    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The availability of colon provides a ready source of human neurons. Among the products of nerve cell bodies, vasoactive intestinal peptide is a neuropeptide that serves as a marker of non-adrenergic, non-cholinergic inhibitory nerves in colon. These nerves have been proposed to be involved in regulation of immune function, secretion, and smooth muscle function. In previous work, we identified decreased tissue levels of vasoactive intestinal peptide in a disorder of chronic colonic mucosal inflammation, ulcerative colitis. We hypothesized that diminished gene expression of vasoactive intestinal peptide could result in decreased tissue levels of this neuropeptide. Sigmoid colon was obtained at surgery from controls (n=6) and patients with ulcerative colitis (n=6). Vasoactive intestinal peptide mRNA was quantified by Northern blot hybridization and tissue levels of vasoactive intestinal peptide were determined by radioimmunoassay. Tissue vasoactive intestinal peptide was decreased only in the mucosalsubmucosal layer of ulcerative colitis (p=.02). There was a single 1.7 kbase vasoactive intestinal peptide transcript identified in both control colon and ulcerative colitis. Normalized vasoactive intestinal peptide mRNA levels were increased by 260% in ulcerative colitis compared to controls (p〈.01). These observations suggest that decreased vasoactive intestinal peptide gene expression or abnormal post-transcriptional processing are not primary defects in this disorder of chronic inflammation. The findings support the alternative hypothesis that axonal degeneration in ulcerative colitis could result in increased expression of neuronal vasoactive intestinal peptide mRNA.
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  • 35
    ISSN: 1573-4919
    Keywords: regucalcin ; calcium-binding protein ; insulin ; calcium ; gene expression ; rat liver
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    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The effect of refeeding on the expression of Ca2+-binding protein regucalcin mRNA in the liver of fasted rats was investigated. When rats were fasted overnight, the hepatic regucalcin mRNA level was reduced about 70% of that in feeding rats. Refeeding produced a remarkable elevation of hepatic regucalcin mRNA level (about 150–170% of fasted rats). Liver regucalcin concentration was appreciably increased by refeeding, although it was not altered by fasting. The oral administration of glucose (2 g/kg body weight) to fasted rats caused a significant increase in hepatic regucalcin mRNA level. Moreover, hepatic regucalcin mRNA level was clearly elevated by a single subcutaneous administration of insulin (10 and 100 U/kg) to fasted rats. The hormonal effect was not further enhanced by the simultaneous administration of calcium chloride (250 mg Ca/kg) to fasted rats, although calcium administration stimulated regucalcin mRNA expression in the liver. The present study suggests that the expression of hepatic regucalcin mRNA stimulated by refeeding is significantly involved in the action of insulin and/or calcium as stimulating factors.
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    Molecular and cellular biochemistry 143 (1995), S. 67-71 
    ISSN: 1573-4919
    Keywords: regucalcin ; calcium-binding protein ; gene expression ; gene distribution
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The existence and expression of gene encoding the Ca2+-binding protein regucalcin in various species and tissues were investigated with Southern and Northern hybridization analyses using regucalcin cDNA (0.9 kb of open reading frame). Genomic Southern hybridization analysis demonstrated that regucalcin gene was widely conserved among higher animals including human, monkey, rat, mouse, dog, bovine, rabbit and chicken. The gene was not found in yeast. The Northern blot analysis of poly (A)+RNAs extracted from the liver of various species showed that regucalcin mRNA was predominantly expressed in rat and mouse, although the expression was also seen in human, bovine and chicken. Furthermore, the enzyme-linked immunoadsorbent assay (ELISA) with rabbit-anti-regucalcin IgG indicated that hepatic regucalcin concentration was most pronounced in rat as compared with that of guinea pig, mouse and chicken. These observations show that the gene expression of regucalcin and its protein synthesis is unique in the liver of rats, suggesting the existence of a specific mechanism in demonstrating regucalcin synthesis from gene.
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    Molecular and cellular biochemistry 143 (1995), S. 137-141 
    ISSN: 1573-4919
    Keywords: regucalcin ; calcium-binding protein ; estrogen ; gene expression ; rat liver
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The effect of nuclear receptor-related hormones on the expression of hepatic calcium-binding protein regucalcin mRNA in rats was investigated. The change of regucalcin mRNA levels was analyzed by Northern blotting using liver regucalcin cDNA (0.9 kb of open-reading frame). A single subcutaneons administration of 17β-estradiol (0.5, 1.0 and 2.0 mg/kg body weight) in rats induced a remarkable increase of regucalcin mRNA in liver; the level was about 200% of control at 24 h after the administration of 2.0 mg/kg. The increase showed about 350% even at 6 h after the administration. Meanwhile, hepatic regucalcin mRNA level was not appreciably altered by a single subcutaneous administration of thyroxine (T4) (20, 40 and 80 mg/kg) or hydrocortisone (10 and 30 mg/kg) in rats. The present study demonstrates that the expression of hepatic regucalcin mRNA is stimulated by estrogen action in the liver nuclei of rats.
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    Molecular and cellular biochemistry 144 (1995), S. 105-108 
    ISSN: 1573-4919
    Keywords: fatty acid synthase ; gene expression ; and thyroid hormone
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The effect of triiodothyronine (T3) on regulation of fatty acid synthase in chicken liver was investigated. In hypothyroid animals, enzyme activity was about one half of that in euthyroid animals. T3 treatment increased the enzyme activity in hypothyroid animals. There is little difference in both the mRNA concentration and the transcription rate between euthyroid and hypothyroid animals. T3 treatment markedly decreased both the mRNA concentration and the transcription rate in euthyroid and hypothyroid animals. These results suggested that T3 maintained the normal level of enzyme expression primarily by stimulating the post-transcriptional step, while the transcription of the gene was inhibited by hyperthyroidism.
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    Molecular and cellular biochemistry 155 (1996), S. 85-90 
    ISSN: 1573-4919
    Keywords: regucalcin ; calcium-binding protein ; gene expression ; rat hepatoma ; Morris hepatoma cells
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Whether the gene expression of hepatic Ca2+-binding protein regucalcin is altered in hepatomas was investigated. The change in regucalcin mRNA levels was analyzed by Northern blotting using liver regucalcin complementary DNA (0.9 kb). Rat hepatoma was induced by continuous feeding of basal diet containing 0.06% 3′-methyl-4-dimethylaminoazobenzene (3′-Me-DAB). After 35 weeks feeding, rats were sacrificed, and the non-tumorous and tumorous tissues of the livers were removed. In individual rats, the regucalcin mRNA levels in the tumorous tissues were generally decreased in comparison with that of the non-tumorous tissues of the chemical-fed rats, although the chemical administration might decrease the mRNA expression in normal rat liver, suggesting that the chemical administration causes a suppresive effect on the mRNA expression. When the genomic DNA extracted from the liver tumorous tissues was digested with restriction enzymes (EcoRI, BamHI and HindIII) and analyzed by Southern blotting, no rear-ranged band was found in the regucalcin gene from the hepatoma. Interestingly, in the transplantable Morris hepatoma cells, the regucalcin mRNA was markedly expressed, while the albumin mRNA was expressed only slightly. The present study demonstrates that regucalcin mRNA is clearly expressed in the transformed cells (Morris hepatoma cells).
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    Molecular and cellular biochemistry 155 (1996), S. 105-111 
    ISSN: 1573-4919
    Keywords: regucalcin ; calcium-binding protein ; aldosterone ; estrogen ; dexamethasone ; gene expression ; rat kidney cortex
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The effect of various steroid hormones on the expression of calcium-binding protein regucalcin mRNA in the kidney cortex of rats was investigated. The change of regucalcin mRNA levels was analyzed by Northern blotting using rat liver regucalcin complementary DNA (0.9 kb of open-reading frame). Regucalcin mRNA was expressed in the kidney cortex but not the medulla. Rats received a single subcutaneous administration of steroid; the animals were sacrificed 60 min after the treatment of aldosterone (2.5, 5.0 and 10 μg/100 g body weight) or 6 h after the treatment of estrogen (17β-estradiol; 0.05, 0.1 and 0.2 mg/100 g), hydrocortisone (0.5, 1.0 and 3.0 mg/100 g) and dexamethasone (50, 100 and 150 μg/100 g). Regucalcin mRNA levels in the kidney cortex were clearly diminished by the administration of aldosterone or estrogen, while hydrocortisone administration had no effect. The administration of dexamethasone (100 μg/100 g) caused a remarkable increase of regucalcin mRNA levels in the kidney cortex. The dexamethasone-induced increase in regucalcin mRNA levels was completely blocked by the simultaneous administration of cycloheximide (150 μg/100 g), although the drug administration had no effect on the mRNA levels in control rats. Meanwhile, the dexamethasone administration did not cause an appreciable alteration of calcium content in the kidney cortex. The present study demonstrates that, of the various steroid hormones used, dexamethasone uniquely has a stimulatory effect on regucalcin mRNA expression in the kidney cortex of rats. The steroid effect may be mediated through a newly synthesized protein.
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    Molecular and cellular biochemistry 160-161 (1996), S. 307-313 
    ISSN: 1573-4919
    Keywords: myocardium ; hypertension ; gene expression ; estrogens ; cardiac hypertrophy ; signal transduction ; genetic program
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Gender specific differences in cardiovascular disease are largely mediated by sex hormones. The use of estrogens significantly reduces the overall incidence of heart disease in postmenopausal women. Beneficial effects of estrogens on plasma lipoprotein levels are clearly established. However, these do not explain the magnitude of risk reduction seen in clinical studies. Thus, additional and currently unknown functions of estrogens must be operative. Elucidation of the exact estrogen action in the heart will have important implications in the treatment of cardiovascular disease. It will probably enhance the therapeutic repertoire in treating heart disease, the most common cause of death in industrialized countries. We will review the current understanding of the function of estrogens in the heart and discuss potential strategies on how to apply these data to clinical practice.
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    Molecular and cellular biochemistry 162 (1996), S. 139-144 
    ISSN: 1573-4919
    Keywords: regucalcin ; calcium-binding protein ; gene expression ; saline ingestion ; hypertensive rats ; kidney cortex
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The effect of adrenalectomy (ADX) or saline ingestion, which is a hypertensive factor, on the expression of calcium-binding protein regucalcin mRNA in the kidney cortex of rats was investigated. The change of regucalcin mRNA levels was analyzed by Northern blotting using rat liver regucalcin complementary DNA (0.9 kb of open-reading frame). Regucalcin mRNA was expressed in the kidney cortex but not the medulla. Rats were adrenalectomized, and 48 h later they were sacrificed. ADX caused a reduction of regucalcin mRNA levels in the kidney cortex, suggesting that adrenal glands participate in the regulation of the mRNA expression. This reduction was not restored by the subcutaneous administration of dexamethasone with an effective dose (1 mg/kg body weight), which can stimulate kidney regucalcin mRNA expression. Regucalcin mRNA levels in the kidney cortex of rats were markedly suppressed by the ingestion of saline for 7 days. The ADX-induced decrease of renal cortex regucalcin mRNA levels was not appreciably restored by saline ingestion. Moreover, regucalcin mRNA levels in the kidney cortex of spontaneous hypertensive rats (SHR) were clearly decreased as compared with that of control (Wistar-Kyoto) rats. Meanwhile, calcium content in the kidney cortex was not significantly decreased by ADX or saline ingestion. The present study suggests that the expression of regucalcin mRNA in the kidney cortex of rats is suppressed by saline administration.
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  • 43
    ISSN: 1573-4919
    Keywords: calreticulin ; gene expression ; steroid receptor
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Calreticulin is a ubiquitously expressed Ca2+ binding protein of the endoplasmic reticulum which inhibits DNA binding and transcriptional activation by steroid hormone receptors. In this study the effects of calreticulin on tyrosine aminotransferase (TAT) gene expression in cultured McA–RH7777 hepatocytes was investigated. McA–RH7777 cells were stably transfected with calreticulin expression vector to generate cells overexpressing the protein. The transcriptional activity of the TAT gene, which is glucocorticoid–sensitive and cAMP–dependent, was investigated in the mock transfected McA–RH7777 and in cells overexpressing calreticulin (designated McA–11 and McA–17). In the presence of dexamethasone or the cAMP analog (CTP–cAMP) expression of the TAT gene was induced in mock transfected McA–RH7777 cells by approximately 4.5 and 5 fold, respectively. In McA–11 and McA–17 cells, overexpressing calreticulin, glucocorticoi ever, the CTP–cAMP–dependent expression of the TAT gene was not affected. The ability of calreticulin to inhibit glucocorticoid–sensitive TAT gene expression but not the cAMP–dependent expression of the gene suggests that the protein affects specifically the action of transcription pathways involving steroid receptors or transcription factors containing KxFF(K/R)R–like motifs. Calreticulin may play an important role in the regulation of glucocorticoid–sensitive pathway of expression of the hepatocytes specific genes during development.
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    Molecular and cellular biochemistry 148 (1995), S. 45-57 
    ISSN: 1573-4919
    Keywords: manganese ; superoxide dismutase ; gene expression ; hyperoxide lung injury ; nuclear factor kappa B
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract TNFα and IL-1 each can activate NF-κB and induce gene expression of manganese superoxide dismutase (MnSOD), a mitochondrial matrix enzyme which can provide critical protection against hyperoxic lung injury. The regulation of MnSOD gene expression is not well understood. Since redox status can modulate NF-κB and potential κB site(s) exist in the MnSOD promoter, the effect of thiols (including NAC, DTT and 2-ME) on TNFα and IL-1 induced activation of NF-κB and MnSOD gene expression was investigated. Activation of NF-kB and increased MnSOD expression were potentiated by thiol reducing agents. In contrast, thiol oxidizing or alkylating agents inhibited both NF-κB activation and elevated MnSOD expression in response to TNFα or IL-1. Since protease inhibitors TPCK and TLCK can inhibit NF-κB activation, we also investigated the effect of these compounds on MnSOD expression and NF-κB activation. TPCK and TLCK each inhibited MnSOD gene expression and NF-κB activation. Since the MnSOD promoter also contains anAP-1 binding site, the effect of thiols and thiol modifying agents on AP-1 activation was investigated. Thiols had no consistent effect onAP-1 activation. Likewise, some of the thiol modifying compounds inhibited AP-1 activation by TNFα or IL-1, whereas others did not. Since diverse agents had similar effects on activation of NF-κB and MnSOD gene expression, we have demonstrated that activation of NF-κB and MnSOD gene expression are closely associated and that reduced sulfhydryl groups are required for cytokine mediation of both processes.
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  • 45
    ISSN: 1573-4919
    Keywords: DNA adducts ; liver ; fish ; 32P-postlabelling ; polycyclic aromatic hydrocarbons ; genotoxic biomarker
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The levels of DNA adducts in the hepatic tissue of the white sucker fish speciesCatostomus commersoni were determined by32P-postlabelling. The fish were caught at four sites: two sites near the city of Windsor (Québec, Canada) on the St. François River, a downstream tributary of the St. Lawrence River, and two sites in the St. Lawrence River itself, near the city of Montréal (Québec, Canada). The latter sites are known to be contaminated by many pollutants including polycyclic aromatic hydrocarbons. Total adduct levels in all fish ranged from 25.1–178.0 adducts per 109 nucleotides. White sucker from the selected sites of the St. Lawrence River had a significantly higher mean level of DNA adducts than those of the St. François River (129.4 vs 56.8, respectively). These results suggest that the effluents of many heavy industries (e.g. from a Soderberg aluminium plant) flowing in the St. Lawrence River are more likely to produce genotoxic damage to fish than those released in one of its tributary, and mainly associated to the activities of a small town and a nearby pulp and paper mill.
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  • 46
    ISSN: 1573-4919
    Keywords: regucalcin ; calcium-binding protein ; gene expression ; calmodulin ; spontaneous hypertensive rats ; rat kidney cortex
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The expression of calcium-binding protein regucalcin mRNA in the kidney cortex of rats ingested with saline was investigated. The alteration in regucalcin mRNA levels was analyzed by Northern blotting using liver regucalcin complementary DNA (0.9 kb of open reading frame). Rats were freely given saline as drinking water for 7 days. Regucalcin mRNA levels in the kidney cortex were suppressed by saline ingestion. When calcium chloride (10 mg Ca/100 g body weight) was intraperitoneally administered to rats ingested with saline for 7 days, the effect of calcium administration to increase regucalcin mRNA levels was weakened by saline ingestion. Such effect was also seen by the administration of 2.5 and 5 mg Ca/100 g. Regucalcin mRNA levels in the kidney cortex of spontaneous hypertensive rats (SHR) were not appreciably increased by the administration of calcium (10 mg/100 g). Meanwhile, calcium content in the kidney cortex was significantly elevated by the administration of calcium (10 mg/100 g) to normal rats. This increase was weakened in saline-ingested rats. Moreover, Ca2+/calmodulin-dependent protein kinase activity in the cytosol of kidney cortex was significantly decreased by saline ingestion. These results suggest the possibility that saline ingestion-induced suppression of regucalcin mRNA expression in the kidney cortex is partly involved in the attenuation of Ca2+ signalling.
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  • 47
    ISSN: 1573-4919
    Keywords: microbodies ; diabetes mellitus ; steroid hormone receptor ; β-oxidation ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract To determine whether the increased fatty acid β-oxidation in the peroxisomes of diabetic rat liver is mediated by a common peroxisome proliferation mechanism, we measured the activation of long-chain (LC) and very long chain (VLC) fatty acids catalyzed by palmitoyl CoA ligase (PAL) and lignoceryl CoA ligase and oxidation of LC (palmitic acid) and VLC (lignoceric acid) fatty acids by isotopic methods. Immunoblot analysis of acyl-CoA oxidase (ACO), and Northern blot analysis of peroxisome proliferator-activated receptor (PPAR-α), ACO, and PAL were also performed. The PAL activity increased in peroxisomes and mitochondria from the liver of diabetic rats by 2.6-fold and 2.1-fold, respectively. The lignoceroyl-CoA ligase activity increased by 2.6-fold in diabetic peroxisomes. Palmitic acid oxidation increased in the diabetic peroxisomes and mitochondria by 2.5-fold and 2.7-fold, respectively, while lignoceric acid oxidation increased by 2.0-fold in the peroxisomes. Immunoreactive ACO protein increased by 2-fold in the diabetic group. The mRNA levels for PPAR-α, ACO and PAL increased 2.9-, 2.8- and 1.6-fold, respectively, in the diabetic group. These results suggest that the increased supply of fatty acids to liver in diabetic state stimulates the expression of PPAR-α and its target genes responsible for the metabolism of fatty acids.
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  • 48
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    Molecular and cellular biochemistry 155 (1996), S. 153-162 
    ISSN: 1573-4919
    Keywords: apolipoprotein B and E ; lipid ; gene expression ; rat ; mouse
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The aim of the present investigation was to study the regulation of apolipoprotein E by two dietary nutrients, saturated fat and cholesterol, known to raise plasma cholesterol levels. ApoE is a protein component of several classes of lipoproteins including VLDL and HDL, and dietary lipids may regulate VLDL and apoE-containing HDL particles through their effects on apoE gene. Male rats and mice were fed the following 4 diets: control diet (C); high cholesterol diet with 0.5% cholesterol (HC); high fat diet with 20% hydrogenated coconut oil (HF); and high fat plus high cholesterol diet with 0.5% cholesterol and 20% fat (HF/C). Plasma cholesterol levels remained unchanged on HC diet, but in mice VLDL-cholesterol increased by 31%. HF diet increased VLDL and LDL by 15–17% in rats, and 21% in mice. A combination of fat and cholesterol diet showed pronounced effects on plasma lipoprotein concentrations, raising apoB-containing particles by 21% and 44% in mice and rats, respectively. Plasma apoE levels increased significantly on all diets. The mechanism of regulation of increased plasma apoB and apoE levels was examined. Quantification of hepatic apoB mRNA showed a lack of correlation between plasma apoB and hepatic apoB mRNA levels, suggesting that posttranscriptional regulation increased plasma apoB-containing lipoproteins in animals fed saturated fat diets. Hepatic apoE mRNA levels increased significantly in animals fed cholesterol-rich diets. However, despite increased plasma apoE levels on diet containing only saturated fat, hepatic apoE mRNA did not change. Synthesis of apoE on the liver polysomes increased selectively on cholesterol-rich diets. These results suggest that cholesterol-rich diets altered apoE, in part, by transcriptional mechanism, and saturated fat-rich diets increased plasma apoE levels by posttranscriptional mechanism, possibly decreased receptor-mediated uptake of apoE-containing particles. The regulation of LDL receptor was also studied since plasma apoB and E levels may be altered by LDL receptor-mediated uptake by the hepatocytes. As expected, high cholesterol diet decreased LDL receptor mRNA by 30–40%. However, the LDL receptor protein on liver membranes did not change on any of the test diets in both animal species. Hepatic cholesterol content increased several fold selectively on high cholesterol diets. These findings suggest that: 1) both transcriptional and posttranscriptional mechanisms are important in regulating plasma apoB and E containing lipoproteins; 2) dietary cholesterol regulates apoE gene by a transcriptional mechanism anddietary saturated fat by posttranscriptional mechanism; and 3) changes in the hepatic apoE and LDL receptor mRNA are associated with the changes in intracellular cholesterol concentrations.
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  • 49
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    Molecular and cellular biochemistry 146 (1995), S. 71-77 
    ISSN: 1573-4919
    Keywords: regucalcin ; calcium-binding protein ; gene expression ; rat kidney cortex
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The expression of calcium-binding protein regucalcin mRNA in the kidney cortex of rats was investigated. The change of regucalcin mRNA levels was analyzed by Northern blotting using liver regucalcin complementary DNA (0.9 kb of open-reading frame). Regucalcin mRNA was expressed in the kidney cortex, and this expression was clearly increased by a single intraperitoneal administration of calcium chloride solution (5–15 mg Ca/100 g body weight) in rats; this increase was remarkable at 60–120 min after the administration. Thyroparathyroidectomy (TPTX) caused a slight decrease of regucalcin mRNA levels in the kidney cortex. However, the administration of calcium (10 mg/100 g) in TPTX rats produced a clear increase of regucalcin mRNA levels in the kidney cortex. The subcutaneous administration of calcitonin (10–100 MRC mU/100 g) or parathyroid hormone [1–34] (1–10 U/100 g) in TPTX rats which received calcium (10 mg/100 g) administration did not cause an appreciable alteration of regucalcin mRNA levels in the kidney cortex, suggesting that the mRNA expression is not stimulated by calcium-regulating hormones. The administration of trifluoperazine (TFP; 5 mg/100 g), an inhibitor of Ca2+/calmodulin action, completely blocked the expression of regucalcin mRNA stimulated by calcium administration. Now, calcium content in the kidney cortex was significantly elevated by a single intraperitpneal administration of calcium (10 mg/100 g) in rats. The present study clearly demonstrates that the expression of regucalcin mRNA in the kidney cortex is stimulated by calcium administration in rats. This expression may be mediated through Ca2+/calmodulin action in the kidney cortex.
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  • 50
    ISSN: 1573-4919
    Keywords: fatty acid transport protein ; gene expression ; subtractive hybridization ; oxidative stress ; ischemia/reperfusion ; ischemic preconditioning
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract In this study, ischemia and oxidative stress-inducible gene expression in heart was examined by subtractive hybridization technique. Total RNA was isolated from ventricular muscle fragments of normal and oxidative stress-induced hearts. Poly (A)+ RNA was purified followed by the construction of a plasmid cDNA library. This was followed by the subtractive screening of oxidative stress-induced cDNA library. The positive colonies were amplified and the plasmid isolated. An aliquot was subjected to restriction cutting with Bam H1 and EcoRl; the fragments corresponding to cDNA insert were separated by electrophoresis, radiolabeled by random-primed DNA synthesis, and used as probes in standard Northern blotting experiments. An aliquot containing the plasmid from the confirmed positives was then subjected to bidirectional partial DNA sequencing (using M13 and T7/T3α primers) by the chain-extension/chain termination method. These sequences were subjected to a computerized search for homologies against all sequences in the updated worldwide Gen Bank and EMBL sequence databases followed by restriction mapping and reading frame identification. Out of 24 putative positive colonies screened, one clone was matched with 〉 97% homology with FAT gene that has been implicated in binding or transport of long chain fatty acids. cDNA probe synthesized from this clone identified two major transcripts of 4.8 and 2.9 kb. Additional experiments were then performed where isolated perfused rat hearts were subjected to the following treatments: (1) 5 min ischemia; (2) 10 min ischemia; (3) 20 min ischemia; (4) 5 min ischemia followed by 10 min reperfusion (ischemic preconditioning); and (5) 5 min ischemia followed by 10 min reperfusion, repeated four times (4 × preconditioning). RNAs were extracted from these hearts and hybridized with the FAT cDNA probe. The results indicated that FAT gene was induced by oxidative stress, ischemic preconditioning, but not by ischemia. (Mol Cell Biochem 160/161: 241–247, 1996)
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  • 51
    ISSN: 1573-4919
    Keywords: regucalcin ; calcium-binding protein ; gene expression ; diabetic state ; ethanol ; liver injury
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The alteration in calcium-binding protein regucalcin in the liver and serum of rats with streptozotocin (STZ)-diabetic state or ethanol ingestion was investigated. STZ (6.0 mg/100 g body weight) was subcutaneously administered in rats, and 1 or 3 weeks later they were sacrificed by bleeding. Liver regucalcin mRNA levels were not clearly altered by the diabetic state, as evidenced by Northern blotting using regucalcin cDNA (0.9 kb of open reading frame). Based on enzyme-linked immunoadsorbent assay (ELISA) with rabbit-anti-regucalcin IgG, hepatic regucalcin concentration was decreased about 50% of control levels by STZ treatment. However, serum regucalcin concentration was not significantly altered by STZ treatment. Meanwhile, when rats ingested ethanol (10 and 30%) in the drinking water for 2 weeks, liver regucalcin mRNA levels were clearly increased, although hepatic regucalcin concentration was significantly decreased. Serum regucalcin concentration was not appreciably altered. Serum transaminases (GOT and GPT) activities were significantly increased at 1 or 3 weeks after STZ administration in rats, while their activities were not altered by ethanol ingestion. The present study demonstrates that hepatic regucalcin concentration is decreased independent of mRNA expression in the STZ-diabetes and during ethanol ingestion in rats.
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  • 52
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    Molecular and cellular biochemistry 163-164 (1996), S. 231-237 
    ISSN: 1573-4919
    Keywords: extracellular matrix ; angiotensin II ; fibrillar collagen ; cardiac fibrosis ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Recent studies suggest that angiotensin II (angiotensin) may be involved in the regulation of metabolism of the cardiac extracellular matrix (ECM). Two major components of ECM are collagen types I and III which play an important role in maintaining the structure and function of the heart. Although the cellular metabolism of collagen is very complex (especially at the posttranslational level), we chose to address events that occur relatively early in the synthesis of cardiac collagen molecules. To gain an understanding of the role of angiotensin in the regulation of cardiac collagen gene expression, we studied the effect of three different doses of angiotensin (12, 24, and 48 μg/kg/h) on adult heart and cultured neonatal cardiac fibroblasts. The steady-state mRNA abundance of collagen types I and III was monitored using Northern blot analysis in both left and right ventricular samples at day 3 of angiotensin infusion and in cultured cardiac fibroblasts stimulated with angiotensin. In all mRNA abundance studies, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) signal was used to normalize the data for possible differences in loading and/or transfer of total RNA. Both collagen types I/GAPDH and III/GAPDH mRNA signal ratios were increased significantly in left ventricle in all dose regimens used for angiotensin infusion. Only the collagen type I/GAPDH mRNA signal ratio was increased in right ventricle with angiotensin infusion. Angiotensin (10−7-10−5 M) had no effect on the steady-state mRNA abundance of collagen genes in cultured neonatal cardiac fibroblasts after 24 h treatment in serum-free conditions. Our results confirm that infusion of angiotensin may upregulate steady-state collagen gene mRNA abundance in the heart. Angiotensin had no observable effect on collagen mRNA abundance in neonatal fibroblast culture. An explanation for the current results may be that angiotensin causes the release of undefined factors from cardiac myocytes, and that these secondary factors may be involved in either the activation of collagen gene transcription or in alteration of stability of collagen mRNA transcripts via a paracrine mechanism. Although our results indicate hemodynamic loading may potentiate the action of angiotensin, this scenario is unlikely as collagen type I gene expression was increased in the normotensive right ventricle.
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  • 53
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    Molecular and cellular biochemistry 167 (1997), S. 169-177 
    ISSN: 1573-4919
    Keywords: tamoxifen ; interferon ; gene expression ; breast cancer
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The molecular basis for the enhanced growth inhibition of MCF-7 human breast cancer xenografts by a combination of human interferon-β (IFN-β) and tamoxifen was investigated. Treatment of MCF-7, MDA-MB-231, and BT-20 cells with the combination of IFN-β and tamoxifen resulted in enhanced antiproliferative effects in vitro. Treatment with the combination of IFN-β and tamoxifen enhanced the expression of several IFN-β-inducible genes in human breast carcinoma cell lines relative to levels induced by IFN-β alone. Tamoxifen alone did not induce transcription of IFN-stimulated genes (ISGs). Augmentation of ISG expression by the combination of IFN-β and tamoxifen was noted in breast tumor cell lines irrespective of their functional estrogen receptor (ER) status or their dependence on estradiol for growth, suggesting that upregulation of ISGs was independent of ER status. Enhancement of IFN-stimulated gene expression by tamoxifen occurred at the transcripti onal level. Expression of transfected reporter genes under the control of IFN-α/β regulated promoters was also enhanced in IFN-β and tamoxifen-treated cells. Similarly, transcriptional induction of chimeric reporter plasmids driven by an IFN-γ inducible promoter (GAS; IFN-γ activated site) was also enhanced by the combination of IFN-γ and tamoxifen. In tamoxifen treated cells, IFN-β and IFN-γ readily activated transcription factors ISGF-3 and GAF, respectively. Therefore, augmentation of ISG expression by tamoxifen is an early event in the antitumoral activity of this drug combination. (Mol Cell Biochem 167: 169-177, 1997)
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  • 54
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    Molecular and cellular biochemistry 172 (1997), S. 47-57 
    ISSN: 1573-4919
    Keywords: smooth muscle ; gene transfer ; DNA ; RNA ; ribozyme ; liposome ; lipoxygenase ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Chemically synthesized hammerhead-type ribozymes targeted against the porcine leukocyte-type 12-lipoxygenase (LO) have been developed and studied. One chimeric ribozyme consists of DNA in the non-enzymatic portions, and RNA in the enzymatic core as well as two phosphorothioate internucleotide linkages at 3′ terminus. The second ribozyme consists of ribonucleotide sequences generated by in vitro transcription. In this chapter we describe methodologies to first analyze the ribozyme catalytic activity in vitro by studying cleavage of target RNA in vitro. The subsequent sections will describe how to target the catalytic ribozyme and deliver it to porcine vascular smooth muscle cells (PVSMC) by a liposome-mediated method. Finally ways to evaluate its activity to inhibit expression of the 12-LO mRNA will be presented. These results demonstrate the feasibility of using ribozymes as novel candidates for therapeutic agents to block specific gene expression in vascular cells.
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  • 55
    ISSN: 1573-4919
    Keywords: heart ; DNA ; library ; gene expression
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    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The availability of high quality cDNA libraries is often crucial to the successful identification and characterization of genes. The concepts and potential pitfalls of constructing cDNA libraries are presented. Various applications requiring high quality cDNA libraries are outlined, including large-scale single pass sequencing of cDNA clones to generate expressed sequence tags (ESTs) and differential screening of cDNA libraries. The usefulness of combining such approaches for the discovery of novel disease-related and cardiovascular-based ESTs (CVBest) is discussed.
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  • 56
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    Molecular and cellular biochemistry 173 (1997), S. 59-69 
    ISSN: 1573-4919
    Keywords: hydrogen peroxide ; oxidative stress ; gene expression ; lens epithelial cells ; N-acetylcysteine ; pyrrolidine dithiocarbamate
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The involvement of H2O2 in cataract development has been established inboth human patients and animal models. At the molecular level H2O2 has beenobserved to cause damage to DNA, protein and lipid. To explore the oxidativestress response of the lens system at the gene expression level, we haveexamined the effects of H2O2 on the mRNA change of the proto-oncogenes,c-jun, c-fos and c-myc in a rabbit lens cell line, N/N1003A. H2O2 treatmentof the rabbit lens epithelial cells for 60 min induces quick up-regulationof both c-jun and c-fos mRNAs. The maximal induction is 38 fold for c-jun at150 µM and 72 fold for c-fos at 250 µM H2O2. Treatment ofN/N1003A cells with 50-250 µM H2O2 for 60 min leads to a 2-5 foldincrease of the c-myc mRNA level. H2O2 also induces an up-regulation intransactivity of the activating protein-1 (AP-1) as shown with a reportergene driven by a prolactin gene promoter with 4 copies of AP-1 binding sitesinserted in the upstream of the promoter. Maximal induction occurs with 150µM H2O2. In the same system, the antioxidants, N-acetyl-cysteine (NAC)and pyrrolidine dithiocarbamate (PDTC) at concentrations shown toup-regulate the mRNAs of both c-jun and c-fos, also enhance thetransactivity of AP-1. NAC and PDTC have different effects in modulating theinduction of AP-1 activity by H2O2 and TPA. These results reveal thatoxidative stress regulates expression of various regulatory genes in lenssystems, which likely affects cell proliferation, differentiation andviability and thus affect normal lens functions.
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  • 57
    ISSN: 1573-4919
    Keywords: regucalcin ; calcium-binding protein ; cDNA cloning ; gene expression ; mouse liver
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The molecular cloning of the cDNA coding for a Ca2+-binding proteinregucalcin and its mRNA expression in mouse liver were investigated. ThecDNA clone encoding a regucalcin was isolated from a mouse liver cDNAlibrary and sequenced. Analysis of the sequence of the cloned cDNA showedthat the cDNA encoded the complete amino acid sequence of the mouseregucalcin molecule; the cDNA had an open reading frame of 897 bp. Mouseregucalcin was composed of 299 amino acid residues, and its molecular weightwas estimated to be 33,406 Da. The amino acid sequence of mouse regucalcinhad 94% homology, as compared with that of rat regucalcin. Northernblot analysis with the mouse liver cDNA probe revealed that mouse regucalcinmRNA was mainly present in the liver but only slightly in the kidney with asize of 1.8 kb. Hepatic regucalcin mRNA level of male mouse was higher thanthat of female mouse. A single intraperitoneal administration of calciumchloride (5, 15, and 30 mg Ca2+/100 g body weight) to mice induced aremarkable increase in regucalcin mRNA in the liver; the increase inregucalcin mRNA levels at 30 min after calcium administration wasdose-dependent. The present results demonstrate that regucalcin mRNA in miceis uniquely expressed in the liver, and that its expression is stimulated bycalcium administration.
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  • 58
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    Molecular and cellular biochemistry 176 (1997), S. 273-279 
    ISSN: 1573-4919
    Keywords: cardiac hypertrophy ; myosin heavy chain ; gene expression ; adrenergic system
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Growth of the heart in hypertrophy is accompanied by changes in the phenotypic expression of cardiac genes. To explore the molecular basis of cardiac hypertrophy, we have analyzed the regulation of myosin heavy chain gene (MHC) expression. In one set of experiments, pressure overload on the rat heart was produced by constriction of the abdominal aorta. Changes in the α and β-MHC mRNA were then studied in overloaded hearts and following load removal. Pressure overload resulted in down-regulation of the α-MHC with corresponding up-regulation of the steady state level of β-MHC mRNA. Load removal (debanding) resulted in regression of cardiac hypertrophy and a rapid return of α-MHC mRNA to normal values. In contrast, the recovery in β-MHC mRNA was much slower to the extent that it remained substantially elevated compared to respective sham controls even after 7 weeks of post-debanding. These results suggest that putative load-related signals independently regulate two genes. Several lines of evidence indicate that adrenergic nervous system plays an important role in the induction and maintenance of cardiac hypertrophy and in the redistribution of myosin isoforms. We have analyzed the effect of cAMP inducing agents on the regulation of a-MHC gene in primary cultures of the fetal (18 day) rat cardiac myocyte. Inclusion of 8 Br-cAMP in the culture media increased the expression of α-MHC promoter/reporter construct comprising of 2.9 kb upstream sequence of the α-MHC gene. Several deletion mutations in the α- MHC gene promoter defined the cAMP responsive boundaries to be a 32 bp region comprising of -71 to -40 bp sequences. Deletion of this region resulted in loss of cAMP response as well as in basal expression of α-MHC promoter/reporter construct. These data suggest a role of β-adrenergic pathway in the modulation of α-MHC gene expression.
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  • 59
    ISSN: 1573-4919
    Keywords: regucalcin ; Ca2+-binding protein ; protein kinase C ; Ca2+signaling ; gene expression ; H4-II-E hepatoma cells
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The expression of hepatic Ca2+-binding protein regucalcin in the cloned rat hepatoma cells (H4-II-E) was investigated. The change in regucalcin mRNA levels was analyzed by Northern blotting using rat liver regucalcin complementary DNA (0.9 kb of open reading frame). Regucalcin mRNA was expressed in H4-II-E hepatoma cells. This expression was clearly stimulated in the presence of serum (10% fetal bovine serum). Bay K 8644 (2. 5 × 10-6 M), a Ca2+ channel agonist, significantly stimulated regucalcin mRNA expression in the absence or presence of 10% serum. Dibutyryl cyclic AMP (10-3 M) did not have a stimulatory effect on the regucalcin mRNA expression. The presence of phorbol 12-myristate 13-acetate (PMA; 10-6 M) or estrogen (10-8 M) caused a significant increase in regucalcin mRNA levels in the hepatoma cells cultured in serum-free medium, while insulin (5 × 10-9 M) or dexamethasone (10-6 M) had no effect. Bay K 8644-stimulated regucalcin mRNA expression in the hepatoma cells was completely blocked in the presence of trifluoperazine (10-5 M), an antagonist of calmodulin, or staurosporine (10-7 M), an inhibitor of protein kinase C. The stimulatory effect of PMA was clearly inhibited in the presence of stauroporine. The present study demonstrates that regucalcin mRNA is expressed in the transformed H4-II-E hepatoma cells, and that the expression is stimulated through Ca2+-dependent signaling factors.
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  • 60
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    Molecular and cellular biochemistry 199 (1999), S. 189-200 
    ISSN: 1573-4919
    Keywords: lung ; cancer ; urokinase ; receptor ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The urokinase-type plasminogen activator (uPA) interacts with its receptor (uPAR) to promote proteolysis as well as cell proliferation and migration. These functions contribute to the pathogenesis of neoplastic growth and invasiveness. Expression of uPAR in tumor extracts also inversely correlates with prognosis in many forms of cancer. In this study, we sought to determine if differences in uPAR expression were distinguishable between cultured human lung carcinoma and malignant mesothelioma subtypes. We also sought to determine if, as in malignant mesothelioma cells, uPAR expression is regulated at the posttranscriptional level in cultured malignant lung carcinoma cells. Using 125I-uPA binding and ligand blotting techniques, uPAR was expressed by phenotypically diverse lung carcinoma cell lines, including the H460, H157 and H1395 non-small cell lines and the H146 small cell lung carcinoma line. Increased uPAR expression was also detected in spindle-shaped (M33K) and epithelioid (M9K and MS-1) malignant mesothelioma cells. Selected mediators, including TGF-β, TNF-α, LPS and PMA, uniformly enhanced uPAR expression in each of the tumor cell lines. Steady state uPAR mRNA expression was determined by RNase protection assay and correlated directly with the changes in cell surface uPAR expression. By gel mobility shift and UV-cross linking assays, a uPAR mRNA binding protein (uPAR mRNABp) implicated in the posttranscriptional control of message stability, was identified in each of the cell lines. Expression of uPAR and its message in cultured lung carcinoma and malignant mesothelioma cells is similarly influenced by effectors present in the tumor microenvironment. Regulation of the uPAR message occurs at the posttranscriptional level in cultured small and non-small cell lung carcinoma cells as well as spindle-shaped and fibrous malignant mesothelioma cell lines. Posttranscriptional regulation of uPAR in all these cells involves the interaction of the uPAR mRNABp with uPAR mRNA, which promotes uPAR mRNA destabilization.
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  • 61
    ISSN: 1573-4927
    Keywords: glycophorins ; gorilla ; evolution ; gene family ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Homologues of MN blood group antigens, encoded by members of the glycophorin A (GPA) gene family, are expressed in man, anthropoid apes, and some species of Old World monkeys. Previous studies had shown that a three-gene framework, most closely related to that in man, is present in the chimpanzee. Here we report the genomic structure, transcript map, and protein expression of the GYPA locus in gorillas. Compared to the corresponding human and chimpanzee homologues, gorilla GPA, GPB, and GPB/E genes each showed a high degree of sequence identity, with the same exon-intron organization. However, the expression of exons III, IV, or V encoding the extracellular or membrane domains of homologous glycophorins varied among the three species. Gorilla GPA and GPB/E genes were unique in that the former occurred in two allelic forms with or without the expression of exon III, whereas the latter contained one (ψ exon III) instead of two silenced exons (ψ exons III and IV). Differences from human but not chimpanzee GPA also included the presence of a hybrid M/N epitope and the absence of the sequon for N-glycosylation. Owing to the retention of a functional exon III, gorilla GPB was more similar to chimpanzee GPB than human GPB. A transspecies allele was identified in the gorilla that gave rise to the Henshaw (He)-like antigen similar to that found in man. These results provide further insight into the model for evolution of the GPA gene family, indicating that the mechanisms underlying inter- and intraspecific polymorphism of glycophorins could predate the divergence of gorillas as the consequence of gene duplication and diversification.
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    Molecular and cellular biochemistry 172 (1997), S. 37-46 
    ISSN: 1573-4919
    Keywords: gene transfer ; gene expression ; adenovirus ; blood vessel
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Adenovirus-mediated gene transfer is a promising method for studies of vascular biology and potentially for gene therapy. Intravascular approaches for gene transfer to blood vessels in vivo generally require interruption of blood flow and have several limitations. We have used two alternative approaches for gene transfer to blood vessels in vivo using perivascular application of vectors. First, replication-deficient adenovirus expressing nuclear-targeted bacterial b-galactosidase was injected into cerebrospinal fluid via the cisterna magna of rats. Leptomeningeal cells over the major arteries were efficiently transfected, and adventitial cells of large vessels and smooth muscle cells of small vessels were occasionally stained. When viral suspension was injected with the rat in a lateral position, the reporter gene was expressed extensively on the ipsilateral surface of the brain. Thus, adenovirus injected into cerebrospinal fluid provides gene transfer in vivo to cerebral blood vessels and, with greater efficiency, to perivascular tissue. Furthermore, positioning of the head may ‘target’ specific regions of the brain. Second, vascular gene delivery was accomplished by perivascular injection of virus in peripheral vessels. Injection of the adenoviral vector within the periarterial sheath of monkeys resulted in gene transfer to the vessel wall that was substantial in magnitude although limited to cells in the adventitia. Approximately20% of adventitial cells expressed the transgene, with no gene transfer to cells in the intima or media. These approaches may provide alternative approaches for gene transfer to blood vessels, and may be useful for studies of vascular biology and perhaps vascular gene therapy.
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    Molecular and cellular biochemistry 172 (1997), S. 111-120 
    ISSN: 1573-4919
    Keywords: differential display ; cardiac development ; gene expression
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    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract An estimated 15,000 different mRNA species are expressed in a typical mammalian cell. The differential expression of mRNAs in both a temporal and cell-specific manner determines the fate of the cell and creates the organism. Analysis of this differential gene expression has become a central aim of many laboratories attempting to understand the mechanisms underlying various biological processes. Currently, we are using a technique called differential display to analyze the differential expression of genes in cardiomyocytes. Differential display is a rapid and powerful technique that was introduced by Liang and Pardee in 1992. Since that time, it has been successfully applied by several groups, and it is quickly becoming a standard method for studying differential gene expression. Here, we present a detailed article discussing the differential display methodology and how we have utilized it to identify potential genes involved in cardiomyocyte proliferation. Furthermore, we have provided a list of materials and supplied examples of data obtained, in an effort to allow the reader to perform the technique with success in their own laboratory.
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  • 64
    ISSN: 1573-4919
    Keywords: plasminogen activators ; plasminogen activator inhibitors ; gene expression ; left ventricular hypertrophy ; pressure overload
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    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract In the early stages of left ventricular hypertrophy (LVH) acute adaptive changes occur in the coronary vasculature as it remodels. Plasminogen activators (PAs) and inhibitors (PAIs) have the potential effects of proteolytic degradation that is relevant to tissue remodeling and angiogenesis. Our study focused on the possible roles of PAI-1, PAI-2, uPA and tPA in myocyte hypertrophy and angiogenesis in the early and late stages of pressure overload induced left ventricular hypertrophy (LVH). We divided seventeen adult swine, weighing 24.2 ± 6.5 kg, into four groups: control, sham-operated, early LVH and late heart failure LVH group. At surgery we placed a fixed constrictor on the ascending aorta immediately above the aortic valve. This increased LV systolic pressure from 133 ± 15 to 193 ± 24 mm Hg after the surgery. We subdivided the early group into groups of 3 animals each that we euthanized at 8, 24 and 72 h after operation and obtained heart samples for analysis. In the late heart failure group individual animals were euthanized at 55, 59, 62 and 72 days after the detection of congestive heart failure. We also obtained tissue samples from the control and sham-operated swine. Sections for histologic analysis were fixed in 10% buffered formalin. We isolated RNA, size fractionated it using 1% formaldehyde-agarose gel electrophoresis and then did Northern blots. The mRNAs from both PAI-1 and PAI-2 showed a remarkable increase at 8 and 24 h after acute aortic constriction and returned to control by 72 h. Regional differences showed that most of the increases were in the endocardium. Three animals in the late heart failure LVH group were determined to be in congestive heart failure at about 2 months after the onset of aortic constriction. In these animals PAI-1 and PAI-2 were increased in both the left and right ventricles but remained low in an animal of the same elevation in aortic pressure seen by the LV who did not have congestive failure. These data suggest that PA and PAI gene expressions change before morphologic changes occur in the early stages of developing LVH. Also at the time of onset of congestive heart failure this increased expression reappears. PAs and PA inhibitors mRNA levels vary in the different regions of the heart reflecting changing wall stresses. Thus, the PAs and PA inhibitors may play an important role in angiogenesis that occurs during the early stages of LVH. The increased expression in the late stage of LVH may reflect further changes in wall stresses since these animals also showed overt clinical signs of heart failure.
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  • 65
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    Molecular and cellular biochemistry 178 (1998), S. 157-162 
    ISSN: 1573-4919
    Keywords: protein tyrosine phosphatases ; gene expression ; degenerate deoxyoligonucleotides ; RT-PCR ; Swiss 3T3 fibroblasts
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The aim of this study was to identify protein tyrosine phosphatases (PTPs) expressed in Swiss 3T3 fibroblasts and to examine their expression levels as well as to characterize quantitative aspects of RT-PCR based on degenerate deoxyoligonucleotides. By using an RT-PCR assay based on degenerate deoxyoligonucleotide primers, expression of mRNAs for two cytoplasmic- and six transmembrane-type PTPs in Swiss 3T3 cells was detected. The sequences of two of them are new. Among nine analyzed PTPs expressed to widely varied extends, only three have mRNA levels high enough to be seen on Northern blots with 10 µg of total RNA per lane. The frequencies with which the examined PTPs are represented among the PCR amplification products, correlate stronger with the primer fidelity, defined as the number of mismatches between the primer- and the cDNA target-sequences, rather than with the PTP expression levels. In conclusion, an RT-PCR assay based on degenerate primers can be successfully used to sample the expressed PTPs and to identify new members of this gene family. However, reliable quantification of their mRNA levels can only be achieved using the classical approaches, like Northern, RNase protection assay or non-degenerate quantitative RT-PCR.
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  • 66
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    Molecular and cellular biochemistry 162 (1996), S. 51-58 
    ISSN: 1573-4919
    Keywords: metabolism ; glucose transporter ; adipocytes ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract We tested the hypothesis that the constitutive glucose transporter (GLUT 1) in 3T3-L 1 adipocytes belongs to the family of glucose-regulated proteins which are transcriptionally regulated by glucose deprivation. Using cDNA probes for both GRP78 (BiP) and GLUT1, we show that the level of GRP78 mRNA increased by 15-fold within 24 h of glucose deprivation with little change in GLUT1 mRNA. The elevated GRP78 mRNA in turn led to a time-dependent increase in GRP78 protein. While glucose deprivation did not alter the expression of the normal glycoform of GLUT 1, a lower molecular weight glycoform accumulated with extended deprivation. Mannose and fructose, but not galactose, prevented the induction of GRP78 and accumulation of the abnormal GLUT1. Because GRP78 acts as a chaperone in other cell systems, we also sought evidence to support this activity in 3T3-L1 adipocytes. Using the technique of co-immunoprecipitation, we demonstrate that GRP78 bound several proteins unique to the glucose-deprived state. No deprivation-specific proteins could be detected in association with GLUT 1. These data lead us to conclude that GLUTl does not display characteristics of the glucose-regulated proteins, at least in 3T3-L1 adipocytes, a widely used model for differentiation, hormone action, and nutrient control. However, the mechanisms for activating traditional members of this family appear intact.
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  • 67
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    Molecular and cellular biochemistry 186 (1998), S. 43-51 
    ISSN: 1573-4919
    Keywords: myocardial ischemia ; gene expression ; growth factors ; phospholamban ; calsequestrin heat shock proteins ; preconditioning ; stunning
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Brief periods of coronary occlusion render the affected myocardium more tolarant to the otherwise devastating effects of long coronary occlusion. Besides this phenomena, called ischemic preconditioning, short periods of ischemia cause a regional dysfunction, namely myocardial stunning. The molecular mechanisms of both syndromes are not very well understood. We therefore investigated the expression of genes which may be involved in cardioprotection or repair processes.Using our porcine model of ischemia and reperfusion we were able to show an induction of genes coding for transcription factors (proto-oncogenes), for proteins involved in repair processes (heat shock genes), for proteins implicated in the calcium homeostasis (calcium-handling genes) and for growth factors. We could show that the increased mRNA levels are due to an enhanced transcriptional activity and not to a prolonged half-life of the transcripts. The angiogenic growth factor vascular endothelial growth factor (VEGF) represents an exception. It exhibits - in addition to a HIF-motif (Hypoxia Inducible Factor) in its promoter/enhancer - a protein binding region in its 3′ UTR which when occupied renders the mRNA more stable. However to what extent the expression of the distinct genes contributes to the cardioprotective effect of ischemic preconditioning or myocardial stunning can only be presumed. Increased mRNA stability can be confered via adenosine which is produced during ischemia by ATP-breakdown. The demasking of unknown genes - via differential display reverse transcription polymerase chain reaction (DDRT-PCR) - should provide a more comprehensive view of the mechanisms underlying both processes.
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  • 68
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    Molecular and cellular biochemistry 201 (1999), S. 111-123 
    ISSN: 1573-4919
    Keywords: complement factor I ; TPA ; protein kinase C ; gene expression ; Hep G2 cells
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract This study examined the role of the protein kinase C (PKC) signalling pathway in the regulation of expression of human complement factor I (CFI) gene. The production of CFI by Hep G2 cells was enhanced in a dose- and time-dependent fashion by 12-O-tetradecanoyl-1,2-phorbol 13-acetate (TPA), a potent PKC activator. 4α-phorbol didecanoate, an inactive phorbol ester, had no effect on CFI synthesis. The TPA-dependent increase in CFI secretion was correlated with an increase in CFI mRNA levels. Forskolin, a cAMP-inducing agent, augmented the TPA response. W7, an inhibitor of protein kinase A and genistein, an inhibitor of protein tyrosine kinase(s) both did not prevent the increase in CFI expression mediated by TPA. However, calphostin C, a specific inhibitor of PKC, abolished the TPA-induced increase in CFI mRNA levels. Down regulation of intracellular PKC levels by prior exposure of Hep G2 cells to a high concentration of TPA also blocked the increase in CFI mRNA levels induced by TPA suggesting that the TPA effects were mediated via activation of PKC. mRNA decay studies indicated that the half-life of CFI mRNA in TPA-induced cells was not significantly different from control. Nuclear run-on transcriptional assays on the other hand demonstrated that whereas the CFI gene is transcribed under basal conditions in Hep G2 cells, TPA induced a 3-4 fold increase in the transcription rate of CFI gene in 24 h. The transcription rate of GAPDH gene did not change, indicating that the effects were not general on gene transcription. Transient transfections of Hep G2 cells with chloramphenicol acetyltransferase reporter gene (CAT) constructs containing a series of sequential 5′ deletions of the CFI promoter and CAT assays showed that the sequence between -136 and -130, containing an AP-1 consensus sequence (TGAGTCA) was required for the TPA response. This observation was substantiated by the finding that mutation of this AP-1 site to TttaTCA or TtAtcCA abolished the TPA responsiveness. The enhancement of the activity of transfected chimeric CAT constructs by TPA was abrogated by calphostin C and by pyrrolidine dithiocarbamate (an inhibitor of NF-κB and AP-1 transactivation). These results indicate that TPA regulation of CFI gene requires PKC signalling and is mediated by via a TPA response element (TRE) in the CFI promoter region located at -136/-130 and involves the transactivation of AP-1 and NF-κB transcription factors. We suggest that PKC may be one of the intracellular pathways that control CFI gene expression and that cellular processes (involving growth factors, hormones, cytokines etc.) that activate PKC may upregulate the expression of the CFI gene.
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  • 69
    ISSN: 1573-4919
    Keywords: regucalcin ; calcium-binding protein ; gene expression ; Ca2+-ATPase ; brain microsomes ; aging
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    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The expression of calcium-binding protein regucalcin and its effect on the microsomal Ca2+-ATPase activity in rat brain tissues was investigated. The expression of regucalcin mRNA was demonstrated by reverse transcription-polymerase chain reaction (RT-PCR) analysis in brain tissues using rat regucalcin-specific primers. Regucalcin concentration in the brain tissues was about 5 × 10-9 M as measured using enzyme-linked immunoadsorbent assay (ELISA), and this level was lowered with increasing age (50 weeks old). The presence of regucalcin (10-9 to 10-7 M) in the enzyme reaction mixture caused a significant decrease in Ca2+-ATPase activity in the brain microsomes of young rats (5 weeks old). Meanwhile, the enzyme activity was not significantly altered by the addition of calmodulin (1 or 50 μg/ml), calbindin (1 or 10 μg/ml), and S-100 A protein (5 or 25 μg/ml), which are other Ca2+-binding proteins in rat brain. The effect of regucalcin to inhibit microsomal Ca2+-ATPase activity was weakened in the brain of rats with increasing age (50 weeks old). The present study demonstrates that regucalcin is expressed in the brain, and that it can uniquely inhibit Ca2+-ATPase activity in the brain microsomes of rats. The findings suggest that regucalcin plays a role in the regulation of microsomal Ca2+-ATPase activity in rat brain tissues.
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  • 70
    ISSN: 1573-1561
    Keywords: Acutilol diterpenes ; biogeography ; chemical defenses ; Dictyota ; diffuse coevolution ; fish ; plant–herbivore interactions ; sea urchin ; secondary metabolites
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Because herbivory is more intense in the tropics, tropical seaweeds may be better defended against herbivory than are temperate seaweeds. A “diffuse” coevolutionary corollary to this hypothesis is that tropical herbivores should be more resistant to seaweed defenses than temperate herbivores because tropical herbivores more commonly encounter heavily defended seaweeds. We begin to test the latter prediction using three newly discovered diterpenoid secondary metabolites from the tropical brown alga Dictyota acutiloba. We tested the feeding deterrent properties of these compounds against common herbivorous fishes and sea urchins from warm-temperate North Carolina versus tropical Guam using standardized laboratory feeding assays. The temperate herbivores were deterred by lower concentrations of secondary metabolites than the tropical herbivores. In no case was a tropical herbivore more deterred by a compound than a temperate herbivore, suggesting that temperate herbivores may be more strongly affected by seaweed chemical defenses. Feeding by the temperate pinfish Lagodon rhomboides was significantly reduced by two of the three diterpenes at a concentration that was only 13–18% of the natural concentration found in the alga. Feeding by four species of tropical fishes (two parrotfishes and two surgeonfishes) was unaffected by metabolite concentrations that deterred the temperate fish. At 100% of natural concentrations, only one of the three compounds deterred the two parrotfishes, and none of the three compounds deterred the surgeonfishes. Contrasts between the temperate sea urchin Arbacia punctulata and the tropical sea urchin Diadema savignyi showed a similar pattern; low concentrations of acutilol A acetate strongly deterred the temperate, but not the tropical, urchin. Tropical herbivores appear more resistant than temperate herbivores to seaweed chemical defenses.
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  • 71
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    Water, air & soil pollution 85 (1995), S. 823-828 
    ISSN: 1573-2932
    Keywords: fish ; mercury ; lake chemistry ; biomagnification
    Source: Springer Online Journal Archives 1860-2000
    Topics: Energy, Environment Protection, Nuclear Power Engineering
    Notes: Abstract We determined mercury in fish (perch Perca fluviatilis) from 26 Russian lakes in three regions over four years. The lakes ranged in size from 2 to 395,000 ha, in pH from 4.5 to 10.0, and in color from 3 to 190 hazen. Sixteen lakes were drainage lakes, with permanent outlets, and 10 were seepage lakes, with no permanent inlets or outlets. The lakes were generally located in forested regions with little or no human habitation in the watershed. The three regions were geologically distinct: Precambrian Shield granitic bedrock covered with thin soil; Triassic bedrock covered with thick glacial tills; and Triassic bedrock covered with thin sediments. At each lake water samples were collected and analyzed for pH, add neutralizing capacity (ANC), major cations, and anions. Dissolved mercury species were estimated with a thermodynamic equilibrium model (MINTEQA2). Mercury content of dorsal muscle varied from 0.04 to 1.0 μg/g wet weight, and was linearly related to calculated HgCH3Cl (r20.68, p〈0.001). Lake HgCH3Cl, in turn, was related to lake pH (r2=0.86, p〈0.001). Stepwise multiple regression selected lake HgCH3Cl and color as the factors most highly related to fish mercury content, with the model accounting for 75% of the variation.
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  • 72
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    Water, air & soil pollution 85 (1995), S. 51-62 
    ISSN: 1573-2932
    Keywords: fish ; invertebrates ; osmoregulation ; ionoregulation ; aluminum ; calcium ; organic acids ; low pH ; acid lakes ; refugia ; mixing zones
    Source: Springer Online Journal Archives 1860-2000
    Topics: Energy, Environment Protection, Nuclear Power Engineering
    Notes: Abstract This paper presents an overview of the response to acidification of aquatic fauna with special emphasis on Zooplankton, benthos, and fishes. Changes in behavior, body chemistry, reproduction, and species diversity are presented based on laboratory experiments and field studies in both Europe and North America. Differences in species sensitivity are discussed as they relate, not only to acidification but also to low calcium concentrations in the water, elevated aluminum concentrations, and presence of naturally occurring organic acids. The mechanisms—behavioral, physiological and ecological—enabling aquatic fauna to survive in acidified waters are discussed.
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  • 73
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    Water, air & soil pollution 85 (1995), S. 419-424 
    ISSN: 1573-2932
    Keywords: acidification ; acid episodes ; critical load ; mountain ; streams ; benthic fauna ; fish
    Source: Springer Online Journal Archives 1860-2000
    Topics: Energy, Environment Protection, Nuclear Power Engineering
    Notes: Abstract Investigations in the southern part of the Scandinavian mountain range have shown a direct correlation between snowpack pH and the lowest pH in small streams. In streams with catchments 〈100 km2 a snowpack pH below 4.8 resulted in a stream pH below 5.5. As the snowpack pH in the southern mountain range is 4.0–4.6, lower values to the south and at high altitudes, large areas are affected by acid deposition. The acidity of the snowpack is released almost directly into the streams during thaw, due to the large snowpack, rapid thaw, steep terrain and thin soils. The acidification of the snow has lead to an extensive fauna depletion, especially in smaller streams without upstream lakes. The abundance of benthos in acidified streams in the municipality of Härjedalen (11.000 km2) is today only one tenth of the abundance before acidification. Fish populations have declined to the same extent, and several acid-sensitive species have been lost. It is concluded that mountain streams and fauna are extremely sensitive to acidification, and that even large reductions of emissions will be insufficient.
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  • 74
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    Water, air & soil pollution 85 (1995), S. 2419-2424 
    ISSN: 1573-2932
    Keywords: critical load ; acidity ; water acidification ; soil acidification ; fish ; empirical models ; sulphur deposition ; acid episodes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Energy, Environment Protection, Nuclear Power Engineering
    Notes: Abstract The critical load of acidity for surface waters is based on the concept that the inputs of acids to a catchment do not exceed the weathering less a given amount of ANC. The Steady State Water Chemistry (SSWC) Method is used to calculate critical loads, using present water chemistry. To ensure no damage to biological indicators such as fish species a value for ANClimit of 20 μeq/l has been used to date for calculating critical loads. The SSWC-method is sensitive to the choice of the ANClimit. In areas with little acid deposition the probability of acid episodes leading to fish kills is small even if the ANClimit is set to zero, while in areas with high acidic deposition fish kills may occur at this value. Thus, the ANClimit can be a function of the acidifying deposition to the lake, nearing zero at low deposition and increasing to higher values at higher deposition. A formulation for such an ANClimit has been worked out, and we have tested the effect of the ANClimit as a linear function of the deposition, assuming ANClimit = 0 at zero deposition with a linear increase to 50 ueq/l at a deposition of 200 meq.m−2.yr−1. For areas with high deposition the effect of a variable ANClimit is small, while in areas with low deposition the effect is significant. For Norway the exceeded area decreases from 36 to 30% using a variable ANClimit instead of a fixed value of 20 μeq/l.
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  • 75
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    Water, air & soil pollution 85 (1995), S. 1461-1466 
    ISSN: 1573-2932
    Keywords: ozone ; wheat ; Triticum aestivum ; growth ; senescence ; biomass partitioning ; photosynthesis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Energy, Environment Protection, Nuclear Power Engineering
    Notes: Abstract In closed-chamber fumigation experiments dry matter partitioning and chlorophyll fluorescence of wheat were studied, analysing the effects of ozone during different stages of plant development. Ozone causes enhanced leaf senescence, leading to a loss of green leaf area and, consequently to a decreased supply of assimilates, affecting (in increasing order of severeness) stem, ear and grain productivity because of reduced storage pools for translocation. Leaves of plants before shooting stage were most sensitive but the lack of green leaf area after ear emergence had the most pronounced effects on grain yield. Measurements of photochemical capacity showed that evidence for negative ozone effects could be found in changes of chlorophyll fluorescence parameters in leaf sections not yet showing visible ozone injury. Negative effects on photosynthesis were more distinct with increasing accumulated ozone dose, with increasing age of leaf tissue and with increasing ozone sensitivity of the cultivar. The changes in chlorophyll fluorescence are most likely to be explained by a decreased pool size of plastoquinones caused by ozone.
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  • 76
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    Water, air & soil pollution 90 (1996), S. 557-562 
    ISSN: 1573-2932
    Keywords: heavy metal ; accumulation ; fish
    Source: Springer Online Journal Archives 1860-2000
    Topics: Energy, Environment Protection, Nuclear Power Engineering
    Notes: Abstract Levels of cadmium, copper, zinc, lead and iron were determined seasonally in the liver, spleen, kidney, gill and muscle tissues of Mullus barbatus and Sparus aurata from the Iskenderun Gulf, East Mediterranean coast of Turkey. Wet digested tissues were analysed by atomic absorption spectroscopy. Metal levels were higher in liver, spleen and kidney compared with the gill and muscle tissues in both species; the levels of all metals in a given tissue were always higher in Mullus barbatus than in Sparus aurata.
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  • 77
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    Water, air & soil pollution 94 (1997), S. 163-180 
    ISSN: 1573-2932
    Keywords: acid deposition ; liming ; catchment ; lake ; water chemistry ; fish ; soil ; model
    Source: Springer Online Journal Archives 1860-2000
    Topics: Energy, Environment Protection, Nuclear Power Engineering
    Notes: Abstract In June 1983 a whole-catchment liming experiment was conducted at Tjønnstrond, southernmost Norway, to test the utility of terrestrial liming as a technique to restore fish populations in remote lakes with short water-retention times. Tjønnstrond consists of 2 small ponds of 3.0 and 1.5 ha in area which drain a 25-ha catchment. The area is located at about 650–700 meters above sea-level in sparse and unproductive forests of spruce, pine and birch with abundant peatlands. A dose of 3 ton/ha of powdered limestone were spread by helicopter to the terrestrial area. No limestone was added to the ponds themselves. The ponds were subsequently stocked with brown and brook trout. Liming caused large and immediate changes in surface water chemistry; pH increased from 4.5 to 7.0, Ca increased from 40 to 200 μeq/L, ANC increased from –30 to +70 μeq/L, and reactive-Al decreased from about 10 to 3 μmol/L. During the subsequent 11 years the chemical composition of runoff has decreased gradually back towards the acidic pre-treatment situation. The major trends in concentrations of runoff Ca, ANC, pH, Al and NO3 in runoff are all well simulated by the acidification model MAGIC. Neither the measured data nor the MAGIC simulations indicate significant changes in any other major ion as a result of liming. The soils at Tjønnstrond in 1992 contained significantly higher amounts of exchangeable Ca relative to those at the untreated reference catchment Storgama. In 1992 about 75% of the added Ca remains in the soil as exchangeable Ca, 15% has been lost in runoff, and 10% is unaccounted for. The whole-catchment liming experiment at Tjønnstrond clearly demonstrates that this liming technique produces a long-term stable and favourable water quality for fish. Brown trout in both ponds in 1994 have good condition factors, which indicate that the fish are not stressed by marginal water quality due to re-acidification. The water quality is still adequate after 11 years and 〉20 water renewals. Concentrations of H+ and inorganic Al have gradually increased and approach levels toxic to trout, but the toxicity of these are offset by the continued elevated Ca concentrations. Reduced sulphate deposition during the last 4 years (1990–94) has also helped to slow and even reverse the rate of reacidification. The experiment at Tjønnstrond demonstrates that for this type of upland, remote terrain typical of large areas of southern Norway, terrestrial liming offers a suitable mitigation technique for treating acidified surface waters with short retention times.
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  • 78
    ISSN: 1573-3017
    Keywords: cortisol ; BKME ; fish ; interrenal ; pituitary ; morphol ogy ; acute stress ; endocrine dysfunction
    Source: Springer Online Journal Archives 1860-2000
    Topics: Energy, Environment Protection, Nuclear Power Engineering
    Notes: Abstract The effects of bleached kraft mill effluent (BKME) on blood cortisol levels and the morphology of the pituitary-interrenal axis were investigated in two species of teleost fish, the northern pike, Esox lucius, and the yellow perch, Perca flavescens, sampled upstream and downstream from a pulp and paper mill on the St Maurice River, Quebec. Fish were acutely stressed by a standardized capture and sampling protocol at both sites, and their ability to elevate blood cortisol levels in response to the capture stress was compared. Blood cortisol levels in fish from the upstream site (〉100 ng/ml plasma) were higher than the levels in fish from the BKME site, and the pituitary corticotropes and the interrenal steroidogenic cells of the upstream fish were larger and had larger nuclei compared with cells from the downstream fish. The low blood cortisol levels in fish exposed to BKME were correlated to cellular atrophy within the hypothalamo-pituitary-interrenal (HPI) axis. The reduced ability to elevate blood cortisol in response to an acute stress may be an endocrine dysfunction occuring in fish chronically exposed to chemical stressors in their environment
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  • 79
    ISSN: 1573-3017
    Keywords: fish ; mercury ; natural selection ; allozyme ; population
    Source: Springer Online Journal Archives 1860-2000
    Topics: Energy, Environment Protection, Nuclear Power Engineering
    Notes: Abstract Measurements of the differential tolerance between enzyme genotypes and shifts in allozyme frequencies in populations from contaminated habitats have prompted the use of allozymes as markers of population-level toxicant effects. However, such studies often do not consider other factors that influence allele frequencies, including natural clines, migration, the intensity and specificity of selection and toxicant-induced genetic bottlenecks. In addition, selection components other than survival are not included. Consequently, the associated conclusions remain speculative. To assess this approach rigorously, a simulation study was conducted with the mosquitofish (Gambusia holbrooki) GPI-2 locus. Laboratory studies have shown the GPI-238/38 homozygote at this locus to be less tolerant than other genotypes during acute exposure to mercury. The GPI-2100/100 genotype has also been shown to have a reproductive disadvantage at lower mercury concentrations. Simple and then more complex models were used to quantify the relative effects of viability selection, random genetic drift and migration on the GPI-238 allele frequency. Simulations were also performed to assess the contribution of sexual and fecundity selection. A simple population model suggested that viability selection plays a greater role than does mortality-driven, genetic drift in the decrease of the sensitive allele under the conditions of this study. A more complex, stochastic model indicated that no significant mortality-driven drift was taking place in this system. In both models, migration mitigated the effect of selection. Sexual and fecundity selection had little effect on the allele frequencies in these simulations. We conclude that, provided the system under study is clearly understood, shifts in allele frequency can indicate the population-level effects of pollutants.
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  • 80
    ISSN: 1573-3017
    Keywords: genotoxicity ; Chernobyl ; fish ; flow cytometry ; coefficient of variation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Energy, Environment Protection, Nuclear Power Engineering
    Notes: Abstract Flow cytometric (FCM) analysis was used to assess the potential impact of chronic radionuclide exposure in fish populations inhabiting contaminated sites in the vicinity of the Chernobyl nuclear accident. Four species of fish, channel catfish (Ictalurus punctatus), crucian carp (Carassius carassius), carp (Cyprinus carpio) and tench (Tinca tinca), were collected within a 10 km radius of the Chernobyl Nuclear Power Plant and compared with 'control' populations from two uncontaminated locations far removed from the plant. Assays of whole blood, as well as separate erythrocyte and leukocyte components, revealed aneuploid-like patterns in the DNA histograms of some fish, as well as widened G0/G1 peaks. None of the fish collected from the uncontaminated sites demonstrated these kinds of changes in their DNA histograms. Increases in the coefficient of variation (CV) of the G0/G1 peak, indicating abnormal DNA distributions, were observed in several of the fish from Chernobyl relative to the control populations. Cell cycle perturbation in fish from the contaminated sites was also detected, with a higher percentage of cells in G2/M phase relative to the controls. Leukocytes proved more sensitive than erythrocytes, as they displayed a larger number of abnormal DNA histograms. Variations in the cellular DNA content similar to those reported here have been shown for other vertebrate species exposed to radiation and other genotoxic agents in laboratory and field settings.
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  • 81
    ISSN: 1573-3017
    Keywords: contamination risks ; fish ; Mississippi River ; ecological factors ; taxonomy
    Source: Springer Online Journal Archives 1860-2000
    Topics: Energy, Environment Protection, Nuclear Power Engineering
    Notes: Abstract We compared contamination levels in fish from contaminated and uncontaminated floodplain swamps of the lower Mississippi River to assess differences in contamination risks between swamps, across different taxonomic and ecological groupings of fishes within and between swamps, and with seasonality in river stage. Fish tissue levels of inorganic contaminants were substantially lower than environmental levels in both swamps, suggesting either that fish were not uptaking these contaminants, or they were effectively eliminating the contaminants from their bodies. Tissue levels of organic contaminants were high relative to environmental levels, suggesting that these contaminants were bioaccumulating. Organic contaminants were significantly higher in fish from the contaminated swamp (Devil's Swamp) than in fish from a reference swamp up river (Tunica Swamp). Because the organic contaminants were largely confined to sediments, we expected bottom-oriented fishes to have higher concentrations than pelagic fishes. Assuming that uptake was primarily through the food chain, we expected top predators to exhibit higher concentrations than low-level consumers. We also expected year- round swamp residents to exhibit higher accumulations than more transitory users of backswamp habitat. However, organic contaminant levels did not differ in the directions expected for any of these groupings. We did observe differences in organic contaminant levels within and between swamps for different taxonomic groupings of fishes (species and genera). Some taxa occupying low to middle positions in the food web (e.g., gizzard shad, Lepomis spp.) exhibited higher concentrations than taxa near the top of the food web. Within Devil's Swamp, organic contaminant levels were significantly higher at low river stage, when fish were confined to the swamp, than at high river stage, when fish were free to move between the river and the swamp. We caught more species and more fish per unit effort in Devil's Swamp than in Tunica Swamp, contrary to expectations if contaminants in the former were negatively impacting population and community structure. Species richness differences between swamps were a consequence of catch differences, with higher catch corresponding to inclusion of more rare species. The lower catch in Tunica Swamp may have resulted from physical modifications of its waterways to support agriculture and hunting. The results of this study underscore the importance in factoring information on the taxonomy and ecology of organisms, and seasonal changes in environmental conditions, into assessments of contamination risks.
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  • 82
    ISSN: 1573-3017
    Keywords: fish ; speciation ; copper ; bioavailability ; bioaccumulation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Energy, Environment Protection, Nuclear Power Engineering
    Notes: Abstract Neon tetras Paracheirodon innesi were exposed to various species of copper during exposures to evaluate the bioavailability of free copper (0 to 2 mg/l), copper apparently complexed to humic substances (0 to 160 mg/l), and copper adsorbed on kaolin clays (0 to 182 mg/l). The results of the experiments demonstrated that free copper is the most bioavailable form. Both humic substances and kaolin clay particulates reduced copper bioavailability to the fish. However, fish accumulated a fraction of copper complexed to humic substances and part of copper adsorbed on kaolin clays.
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  • 83
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    Molecular and cellular biochemistry 151 (1995), S. 55-60 
    ISSN: 1573-4919
    Keywords: regucalcin ; calcium ; gene expression ; kidney damage ; rat kidney cortex
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The alteration of Ca2+-binding protein regucalcin mRNA expression in the kidney cortex of rats administered cisplatin and cephaloridine, which can induce kidney damage, was investigated. Cisplatin (0.25, 0.5 and 1.0 mg/100 g body weight) or cephaloridine (25, 50 and 100 mg/100 g) was intraperitoneally administered in rats, and 1, 2 and 3 days later they were sacrificed. The alteration in serum findings after the administration of cisplatin (1.0 mg/100 g) or cephaloridine (50 and 100 mg/100 g) demonstrated chemically induced kidney damage; blood urea nitrogen (BUN) concentration increased markedly and serum inorganic phosphorus or calcium concentration decreased significantly. Moreover, the administration of cisplatin (1.0 mg/100 g) or cephaloridine (100 mg/100 g) caused a remarkable increase of calcium content in the kidney cortex of rats, indicating kidney damage. The expression of regucalcin mRNA in the kidney cortex was markedly reduced by the administration of cisplatin or cephaloridine in rats, when the mRNA levels were analyzed by Northern blotting using rat liver regucalcin cDNA (0.9 kb). The mRNA decreases were seen with the used lowest dose of cisplatin or cephaloridine. The present study clearly demonstrates that the mRNA expression of Ca2+-binding protein regucalcin in the kidney cortex of rats is decreased by chemically induced kidney damage.
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  • 84
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    Molecular and cellular biochemistry 152 (1995), S. 131-141 
    ISSN: 1573-4919
    Keywords: gene expression ; mRNA ; proto-oncogenes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Polyomavirus large T-antigen transgenic mice develop cardiac hypertrophy characterized by an increase in atrial natriuretic factor and β-myosin heavy chain isoform expression. The aim of this study was to examine changes in proto-oncogene expression in hypertrophied hearts from the transgenic mice. Expression of early growth response-1 (Egr-1) mRNA was detected in hearts from all 15 transgenic mice, but was not detectable in 13 control mice. Reverse transcriptase-polymerase chain reaction experiments usingEgr-1-specific primers confirmed the increase inEgr-1 mRNA in enlarged hearts from the transgenic mice. Expression of c-jun,junD and Ha-ras mRNAs was increased in the transgenic hearts 3, 17 and 2.8-fold, respectively. Western blots showed an increase in c-myc, c-jun and ras protein in hypertrophied transgenic hearts. Immunofluorescence analyses confirmed an increase in Egr-1 and c-jun protein in transgenic cardiomyocytes. Proliferating cell nuclear antigen, Ki-ras and HSP 90 mRNAs were decreased 22, 2.7 and 3-fold, respectively in the transgenic hearts. Not altered in most hypertrophied hearts was expression of c-fos, junB, p53, c-neu, c-myc, HSP70, HSP27, TGF-β or IGF-1 mRNAs. Proto-oncogene and growth factor gene expression in hypertrophy induced by PVLT expression is modulated, with some proto-oncogenes increased and others decreased in expression.
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  • 85
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    Molecular and cellular biochemistry 154 (1996), S. 65-70 
    ISSN: 1573-4919
    Keywords: gene expression ; regulatory elements ; plasmid ; oligonucleotides
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract A potentially powerful pharmacological approach to modulating the expression of specific, disease-related genes involves the inhibition of transcription factor binding to promoter or enhancer elements through oligonucleotide-mediated triple-helix formation. In vivo, the typical target for intermolecular triplex formation would most likely be torsionally-strained rather than relaxed duplex DNA. To determine the effects of strained DNA on triplex formation, we investigated the interactions between a G/T rich oligonucleotide and both supercoiled and relaxed plasmid DNA using a restriction endonuclease protection assay. Both the kinetics of formation and dissociation of purine-motif triplexes were unaffected by the conformational state of the duplex DNA. Similarly, the topological state of the plasmid targets was not affected by triplex formation. Taken together, these observations suggest that stable intermolecular triplexes can form in vivo under conditions of moderate torsional strain.
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  • 86
    ISSN: 1573-4919
    Keywords: estrogen ; apolipoprotein ; gene expression ; mice ; atherosclerosis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Estrogen protects against developing premature coronary artery disease.However, the mechanism of protective effects of estrogen still remainspoorly understood. One mechanism by which estrogen can have protectiveeffects apppears to be through modulation of plasma lipoproteins. We showedthat the mouse can be used as animal model to study estrogen-mediatedsynthesis and secretion of lipoproteins since, unlike the rat, the mousedoes not up-regulate LDL receptors (Srivastava et al. [4]). Since inbredstrains of mice differ in their genetic background and show differingresponsiveness to dietary lipids, we examined how various inbred strains ofmice respond to estradiol administration, and whether some mouse strainsshow responses similar to rats. 17b-estradiol was administered to male micefrom 15 different inbred strains, and the changes in plasma levels oflipids, apoB, apoAI, and apoE were examined. Total cholesterol decreased inall but one strain, apoAI levels decreased in all but 3 strains while apoBlevels and apoB/apoAI ratios increased in all but 2 strains, suggesting thatin contrast to rats, the apoB-containing lipoproteins increased relative toHDL in all strains of mice examined. Basal and estradiol-induced changes intotal cholesterol were significantly correlated with changes in apoAI, butnot apoB, reflecting the predominance of HDL over other lipoproteins inmouse plasma. The effects of estrogen on plasma apoE levels varied amongvarious inbred strains of mice tested. Plasma apoE levels increased in sevenstrains treated with estrogen, and remained unchanged in the rest. Toexamine whether changes of plasma apoproteins are associated with thechanges in the respective hepatic mRNA levels, apoAI, B and E mRNA werequantified by RNase protection assay. Hepatic apoE mRNA did not showcorrelation with either basal or post treatment plasma apoE levels in any ofthe strains. Similarly, most of the mouse strains did not show correlationof plasma apoAI and apoB levels with the corresponding hepatic mRNA levels.These results suggest that estrogen regulates plasma lipoproteinconcentrations primarily by posttranscriptional mechansims, and there werestrain-related differences in the estrogen-mediated regulation oflipoprotein metabolism.
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  • 87
    ISSN: 1573-4919
    Keywords: regucalcin ; Ca2+-binding protein ; insulin ; gene expression ; HepG2 cells ; transformed cells
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The expression of hepatic Ca2+-binding protein regucalcin in the cloned human hepatoma cells (HepG2) was investigated. The change in regucalcin mRNA levels was analyzed by Northern blotting using rat liver regucalcin complementary DNA (0.9 kb of open reading frame). Regucalcin mRNA was expressed in HepG2 cells, although the mRNA was markedly expressed in normal rat liver. Moreover, regucalcin protein in HepG2 cells was detected by Western blot analysis using a polyclonal rabbit anti-regucalcin antibody. Regucalcin mRNA expression in HepG2 cells was clearly stimulated by the culture with insulin (10-8 M) of the effective concentration. Regucalcin protein in HepG2 cells was also increased by the treatment of insulin (10-8 M). The present results demonstrate that regucalcin is expressed in the transformed HepG2 cells, and that the expression is stimulated by insulin.
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  • 88
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    Molecular and cellular biochemistry 177 (1997), S. 1-6 
    ISSN: 1573-4919
    Keywords: gene expression ; mRNA secondary structure ; single tube RT-PCR ; TNF receptor I
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The secondary structure of human tumor necrosis factor receptor I (TNFR-I) mRNA based on its lowest folding energy was predicted. Three combinations of primers selected from open-regions and four combinations of primers from closed-regions of TNFR-I mRNA structure were employed for single-tube reverse transcription-polymerase chain reaction (RT-PCR) for the determination of TNFR-I gene expression in U937 cell. All the primers were designed with the same criteria. However, the different primers generated distinct quantities of RT-PCR products from the same concentration of TNFR-I mRNA, implying that the determination of gene expression by RT-PCR was affected by the mRNA secondary structure. In addition, the sensitivity of the open-region RT-PCR was approximately one hundred-fold higher than that in the closed-regions of TNFR-I mRNA. The low efficiency of the closed-region RT-PCR was not correlated with the G/C content of the TNFR-I mRNA structure. These results suggest that consideration of the influence of intrinsic mRNA structure of a gene is essential prior to the determination of gene expression by quantitative RT-PCR, and this open-region strategy of primer design may yield an efficient primer for in vitro amplification of cDNA by RT-PCR.
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  • 89
    ISSN: 1573-4919
    Keywords: cholesteryl ester ; CETP ; Caco-2 ; polymerase chain reaction ; gene expression ; mRNA ; alternative splicing
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Cholesteryl ester transfer protein (CETP) is a plasma protein involved in the reverse cholesterol transport and expressed in several human tissues and cell lines. We studied CETP expression in Caco-2 cell line, a model of the human enterocyte epithelium. By reverse-transcriptase polymerase chain reaction, we could demonstrate that in basal condition Caco-2 cells have a low rate of expression of active CETP mRNA. Furthermore, we found that even in this cell line CETP mRNA alternative splicing occurs with deletion of exon 9 sequence. Densitometric analysis of the in vitro amplified fragments showed that under basal conditions about 60% of reverse transcribed CETP cDNA corresponds to exon 9-deleted transcripts. After challenge with 50 µM sodium oleate, there is a ∼2 fold increase in the transcription rate of the full-length CETP cDNA, as measured by competitive PCR, which is accompanied to an increased activity measured in the cell-conditioned medium. On the contrary, no significant change is seen in the amount of exon 9-deleted cDNA. Consequently, an inversion in the ratio of full-length and exon 9-deleted CETP cDNA is evident, suggesting that sodium oleate selectively enhances the expression of full-length CETP mRNA.
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  • 90
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    Molecular and cellular biochemistry 178 (1998), S. 283-287 
    ISSN: 1573-4919
    Keywords: regucalcin ; calcium-binding protein ; gene expression ; fetal development ; rat liver
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The expression of hepatic calcium-binding protein regucalcin mRNA in fetal rats was investigated. The alteration in regucalcin mRNA levels was analyzed by Northern blotting using liver regucalcin cDNA (0.9 kb with complete open reading frame). Hepatic regucalcin mRNA levels were progressively increased with fetal development; the mRNA was clearly expressed at 15 and 21 days of pregnancy but only slightly at the 8 days. Meanwhile, β-actin mRNA levels in the fetal liver were remarkable at 8 and 15 days of pregnancy. The fetal liver regucalcin mRNA levels at 15 days of pregnancy were significantly decreased by overnight-fasting of maternal rats. The oral administration of calcium chloride (50 mg Ca/100 g body weight) to maternal rats at 15 days of pregnancy caused a remarkable elevation (about 2 fold) of regucalcin mRNA levels in the fetal liver; this increase was seen 60 and 180 min after the calcium administration. After birth, regucalcin mRNA was increasingly expressed in the livers of newborn and weanling rats, while hepatic β-actin mRNA expression was not appreciably altered with increasing ages. These findings demonstrate that the expression of hepatic regucalcin mRNA is increased with fetal development, and that the gene expression may be stimulated by the ingestion of dietary calcium.
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  • 91
    ISSN: 1573-4919
    Keywords: extremely low frequency magnetic fields ; gene expression ; neuron derived orphan receptor-1 ; signal transduction ; Chinese hamster ovary K1 cells
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Enhanced expression of neuron derived orphan receptor (NOR-1) gene was observed by exposure of Chinese hamster ovary K1 (CHO-K1) cells to an extremely low frequency magnetic field (ELFMF) of 50 Hz at 400 mT, but not at 5 mT. The enhanced expression, reaching the maximum at 6 h, was transient and reduced to the control level after exposure to 400 mT ELFMF for 24 h. The NOR-1 expression induced by treatment with forskolin and TPA was further enhanced by the simultaneous treatment with 400 mT ELFMF, in which the maximum response was at 3 h. The NOR-1 expression by these treatments was induced more earlier than that by 400 mT ELFMF alone. When cells were treated with an inhibitor of the protein kinase C (calphostin C or crocetin) and Ca2+ entry blockers (nifedipin and dantrolen) during the 400 mT ELFMF exposure, the enhanced NOR-1 expression was not observed. Exposure of CHO-K1 cells to the high-density 400 mT ELFMF may affect the signal transduction in the cells, resulting in the enhanced NOR-1 gene expression.
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  • 92
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    Molecular and cellular biochemistry 188 (1998), S. 41-48 
    ISSN: 1573-4919
    Keywords: zinc ; transcription factors ; gene expression ; organogenesis ; Xenopus laevis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Zinc regulates the gene expression machinery. It affects the structure of chromatin, the template function of its DNA, the activity of numerous transcription factors and of RNA polymerases. Hence, it determines both the types of mRNA transcripts synthesized and the rate of transcription itself. Alterations in one or more of these zinc dependent processes have been proposed to account for the proliferative arrest and teratology induced by zinc deficiency. To examine this proposal, studies of zinc during X. laevis development have been initiated. The kinetics of X. laevis oocyte zinc uptake and storage and of zinc utilization during embryogenesis have been examined first. Vitellogenin carries zinc into the oocyte. Ten % of the total zinc (10 ng/egg) remains within the cytosol while 90% (90 ng/egg) is stored in the yolk platelets associated with lipovitellin. The cytosolic pool is the source of the zinc for all newly formed metalloproteins involved in embryo development. The yolk platelet zinc pool is stored for later use during early metamorphosis. It is now possible to examine zinc transfer to molecules, such as e.g. transcription factors, and the role of the metal in their function in development and organogenesis.
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  • 93
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    Molecular and cellular biochemistry 189 (1998), S. 107-111 
    ISSN: 1573-4919
    Keywords: gene expression ; electromagnetic fields ; superinduction ; anisomycin ; immediate early gene
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Rat pheochromocytoma PC12 cells have been treated with nerve growth factor (NGF) at final concentrations of 2, 4, 8, and 16 ng/ml, and then were exposed to 60-Hz, sinusoidal magnetic fields (MF) of 12.5, 25, 50, and 100 μT (rms) for 30 min. Transcript levels for both c-fos and glyceraldehyde-3 -phosphate dehydrogenase were determined by Northern blot analysis using 32P-labeled cDNA probes. No change in c-fos expression was measured at any condition employed. Treatment of PC12 cells with a combination of agents (NGF, forskolin, and tetradecanoylphorbol acetate [TPA]) increased c-fos expression over that detected with NGF alone. MF exposure of cells treated with the three-agent regimen produced two outcomes, either no change or a doubling of c-fos expression. In subsequent experiments, cells were treated with NGF, NGF + forskolin + TPA, or pre-treated with anisomycin and then treated with NGF + forskolin + TPA. It was determined that MF exposure, like superinduction with anisomycin, increased c-fos expression only in cultures which were not yet exhibiting maximal c-fos expression. It is hypothesized that MF exposure, like anisomycin, may alter the activity of key intracellular protein kinases.
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  • 94
    ISSN: 1573-4919
    Keywords: mechanical stretch ; smooth muscle cells ; differential display ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Physical forces induce profound changes in cell phenotype, shape and behavior. These changes can occur in vascular structures as a result of pressure overload and their effects can be seen in atherosclerotic vessels in which smooth muscle cells have undergone hyperplastic and hypertrophic changes. At the molecular level, mechanical stimuli are converted into chemical ones and lead to modulation of gene expression and/or the activation of a new repertoire of genes whose encoded proteins help the cells to adapt to their microenvironment. In this study, we have used a two primer-based mRNA differential display technique to identify candidate mechano-responsive genes in pulmonary artery smooth muscle cells. As compared to the original method described by Liang and Pardee, this technique uses two arbitrary primers instead of an anchored oligo(dt) plus an arbitrary primer in the polymerase chain reaction. The chief advantages of these modifications are an increase in the efficiency of the amplification and in the identification of differentially expressed clones. Using this approach, we compared the pattern of expressed genes in cells cultured under static conditions with those in cells that were mechanically stretched (1 Hz) for 24 h in a well-defined in vitro mechanical system. Three candidate genes that showed reproducible differences were chosen for further characterization and cloning. One clone was under expressed in stretched cells and had a DNA sequence with 90% homology to the human fibronectin gene. Two other clones were highly expressed in stretched cells and had a 92% and a 83% sequence homology with human platelet-activating factor (PAF) receptor and rat insulin-like growth factor-I (IGF-I) genes respectively. Northern blot analysis confirmed low levels of fibronectin mRNA transcripts in stretched cells. In contrast, accumulation of PAF receptor mRNA occurred 30 min after mechanical stretch was initiated whereas IGF-I mRNA levels peaked at 8 h. Both mRNA levels were sustained for up to 24 h of mechanical stretching. These results demonstrate the usefulness of the two primer-based mRNA differential display that enabled us to identify and characterize alterations at the level of gene expression among matrix proteins, G-protein coupled receptors and growth factors, each of whose response to mechanical strain is different. A more complete understanding of these responses will provide further insight into the pathologic processes associated with hypertension and atherosclerosis.
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  • 95
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    Molecular biology reports 24 (1997), S. 221-230 
    ISSN: 1573-4978
    Keywords: gene expression ; ribonucleoprotein ; RNase MRP ; RNase P ; transcription
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We report on the expression of mouse RNase MRP RNA in human embryonic kidney 293 cells upon DNA transfection. Stable cell lines were selected by cotransfection with a neo r gene. Transcription of wild-type and deletion mutants of MRP RNA and ribonucleoprotein formation were assessed by RNase protection and immunoprecipitation experiments. Mouse MRP RNA as expressed in 293 cells readily associates with human proteins to form a chimeric Th ribonucleoprotein. 5' truncated MRP RNAs, however, failed to associate with Th antigen(s) and deletion of the 3' sequences of MRP RNA greatly reduced the expression in stable as well as in transient transfectants. Abbreviations: nt(s) – nucleotide(s); RNP – ribonucleoprotein.
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  • 96
    ISSN: 1573-4978
    Keywords: gene expression ; nuclear matrix proteins ; ocular lens epithelial cells ; transcription factors
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Association of transcription factors with the nuclear matrix represents a mechanism by which nuclear architecture may influence transcriptional control of gene expression. This investigation examines nuclear matrix associated proteins (NMP's) isolated from ocular lens epithelial cells by monitoring DNA binding activities using consensus oligonucleotides recognized by the transcription factors YY1, AML-1, AP-1, SP-1 and ATF. The nuclear matrix fractions tested included an immortilized human lens epithelial cell line containing the SV40 large T-antigen, and two mouse lens epithelial cell lines derived from either a normal mouse or a cataract mouse. A rabbit epidermal epithelial cell line and HeLa cells were also included in this study for comparison. The data from these experiments reveal that ubiquitously represented and tissue restricted regulatory proteins are associated with nuclear matrix of lens epithelial cells. The functional significance of the nuclear matrix association of these transcription factors remains to be determined. However, our findings raise the possibility that the transcription factors associated with the nuclear matrix could have specific roles in gene regulation and eye tissue development.
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  • 97
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    Molecular biology reports 21 (1995), S. 85-86 
    ISSN: 1573-4978
    Keywords: fish ; repetitive DNA ; SINE ; LINE
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A highly repetitive DNA element has been isolated fromOpsariichthys uncirostris. It contains several oligo-dA tracts and potential regions for the secondary structures.
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  • 98
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    Molecular biology reports 22 (1995), S. 33-35 
    ISSN: 1573-4978
    Keywords: fish ; repetitive DNA ; RFLP ; satellite DNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A member of satellite repetitive DNA was isolated and sequenced from a saltwater fishSillago japonica (Percoidei). This sequence consists of several oligo-dA/dT tracts and two inverted repeats which resemble each other. Dot blot hybridization analysis using a satellite DNA clone pSJ2 among the species in the suborder Percoidei revealed that the pSJ2 sequence was amplified at least after the family Sillaginidae had been derived.
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  • 99
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    Molecular biology reports 23 (1996), S. 205-210 
    ISSN: 1573-4978
    Keywords: autoantigen ; cDNA cloning ; gene expression ; ribonucleoprotein Ro ; 5′RACE
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Autoantibodies to SS-A/Ro are among the most common found in sera of patients with systemic rheumatic diseases. These autoimmune diseases can affect various organ systems of the body and are variable in their manifestations and presentation. One of the autoimmune targets is the 60 kDa SS-A/Ro protein known to be associated with small cytoplasmic Y RNAs. To study systematically the expression of the protein, we have cloned the mouse full length 60 kDa SS-A/Ro cDNA using 5′ RACE based on a cDNA sequence reported in the mouse genome project. The recombinant protein derived from the putative full-length construct was shown to react with human prototype anti-SS-A/Ro serum Ge in western blot and immunoprecipitation and comigrated with cellular 60 kDa SS-A/Ro protein in 3T3 cells. Cellular expression, measured by RT-PCR, was highest in mouse brain, followed by lung, muscle, kindney and heart. Lower levels were found in testis, liver and spleen. Like the human 60 kDa SS-A/Ro protein, the deduced mouse homolog has 538 amino acids. Sequence analysis showed 89.9% identity and 95.0% similarity between the mouse and human proteins.
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  • 100
    ISSN: 1573-5028
    Keywords: β-fructofuranosidase ; invertase ; gene expression ; gene structure ; flower buds ; Daucus carota
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Three genomic clones (Inv *Dc1, Inv *Dc2 and Inv *Dc3) were isolated by using the cDNA for carrot cell wall β-fructofuranosidase as a probe. The expression patterns of the three genes differed markedly. High levels of Inv *Dc1 transcripts were found in leaves and roots of young carrot, whereas in plants with developing tap roots no transcripts were detected. A high level of mRNA of Inv *Dc1 was also present in suspension-cultured cells. In developing reproductive organs, only low levels of transcripts of Inv *Dc1 were found in flower buds and flowers and none at later stages of development. In contrast, Inv *Dc2 and Inv *Dc3 were not expressed in vegetative plant organs. Invb1 *Dc1 was exclusively and strongly expressed in flower buds, and Inv *Dc3 at a very low level in suspension-cultured cells.
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