Protein expression by Streptococcus uberis in co-culture with bovine mammary epithelial cells

Three strains of Streptococcus uberis isolated from dairy cows with mastitis were co-cultured with a bovine mammary epithelial cell line (MAC-T) in Dulbecco's modified Eagle's medium without fetal bovine serum. Protein profiles from culture supernatants and bacterial pellets among different treatments were compared by electrophoresis. There were proteins induced or having increased expression in both supernatant and surface-associated samples from S. uberis co-cultured with MAC-T cells. Some of these proteins were recognized by antibodies in serum obtained from a cow infected by S. uberis . In supernatant samples, there were two distinct protein bands at 35 and 36.8 kDa for all three strains of S. uberis co-cultured with MAC-T cells. These two bands were absent when bacterial protein synthesis was inhibited by chloramphenicol. This study clearly indicates that bacterial protein expression was regulated in response to co-culture with mammary epithelial cells.     

In vitro culture of human chondrocytes (1): A novel enhancement action of ferrous sulfate on the differentiation of human chondrocytes

Chondrogenic differentiation of mesenchymal cells is generally thought to be initiated by the inductive action of specific growth factors and depends on intimate cell-cell interactions. The aim of our investigation was to characterize the influences of basic fibroblast growth factor (bFGF) and ferroussulfate (FeSO₄) on proliferation and differentiation of human articular chondrocytes (HAC). This is the first report of the effects of FeSO₄ on chondrogenesis of HAC. Multiplied chondrocytes of hip and shoulder joints were cultured in chondrocyte growth medium supplemented with bFGF, FeSO₄, or both bFGF + FeSO₄ for4weeks. A 20 μl aliquot of a cell suspension containing2 × 10⁷ cells ml⁻¹ was delivered onto each well of 24-well tissue culture plates. Cells cultured with the growth medium only was used as a control. Alamar blue and alcian blue staining were done to determine the chondrocyte proliferation and differentiation, respectively, after 4 weeks. The samples exposed to bFGF, FeSO₄, and combination of both indicated sufficient cell proliferation similar to the control level. Differentiations of the HAC exposed to bFGF, FeSO₄,and bFGF + FeSO₄ were 1.2-, 2.0-, and 2.2-fold of the control, respectively. Therefore, chondrocyte differentiation was significantly enhanced by the addition of FeSO₄ andbFGF + FeSO₄. The combined effects of bFGF and FeSO₄ were additive, rather than synergistic. These results suggest that treatment with ferrous sulfate alone or in combination with basic fibroblast growth factor etc, is a powerful tool to promote the differentiation of HAC for the clinical application. This revised version was published online in August 2006 with corrections to the Cover Date.     

The role of nutrition on epigenetic modifications and their implications on health

Nutrition plays a key role in many aspects of health and dietary imbalances are major determinants of chronic diseases including cardiovascular disease, obesity, diabetes and cancer. Adequate nutrition is particularly essential during critical periods in early life (both pre- and postnatal). In this regard, there is extensive epidemiologic and experimental data showing that early sub-optimal nutrition can have health consequences several decades later.     

The Last Month of Szent-Györgyi in Groningen

Albert (von) Szent-Györgyi started his studies on biological oxidation processes – which also resulted in the discovery of vitamin C, for which he received the Nobel Price in 1937 – in the Laboratory of Physiology of the University in Groningen in 1922–1926. These studies were later continued in Cambridge (UK) and Szeged (Hungary). When he had already received the invitation as well as the financial means to come and work in Cambridge, he still did experiments in Groningen to find out whether the adrenal extract, isolated by him and later found to be a major source of vitamin C, contained the hormone essential for the survival of cats whose adrenals were removed. He was rather upset by the negative results of this experiment, judging by the recollections of a former student of his. This history constitutes an interesting example of the difference between serendipitous discovery and planned invention.     

Scale effects and constraints for sound production in katydids (Orthoptera: Tettigoniidae): correlated evolution between morphology and signal parameters

Male katydids (Orthoptera: Tettigoniidae) produce mating calls by rubbing the wings together, using specialized structures in their forewings (stridulatory file, scraper and mirror). A large proportion of species (ca. 66%) reported in the literature produces ultrasonic signals as principal output. Relationships among body size, generator structures and the acoustic parameters carrier frequency (f_c) and pulse duration (p_d), were studied in 58 tropical species that use pure‐tone signals. A comparative analysis, based on the only available katydid phylogeny, shows how changes in sound generator form are related to changes in f_c and p_d. Anatomical changes of the sound generator that might have been selected via f_c and p_d are mirror size, file length and number of file teeth. Selection for structures of the stridulatory apparatus that enhance wing mechanics via file‐teeth and scraper morphology was crucial in the evolution of ultrasonic signals in the family Tettigoniidae.     

Similarities and differences in the properties of multiple isoforms of maize endosperm casein kinase I (CK-I)

Two novel type I casein kinases named CK-1B and CK-1C have been purified from maize endosperm (three weeks after anthesis) by a six step procedure involving ammonium sulfate precipitation, DEAE-cellulose, Sephadex G-75, Heparin-sepharose, and ATP-agarose chromatography. The catalytic subunits of both enzymes were identified as a 35-37 kDa polypeptide doublet by in situ phosphorylation after SDS/PAGE in active casein gel. Both enzymes required 5-10 mmol · L⁻¹ Mg²⁺ for maximal activity, could utilize only ATP as phosphate donor, were insensitive to heparin, were not autophosphorylated, had a pH optimum at pH 7 to 8.5, and exclusively phosphorylated acidic proteins (casein, phosvitin). Regarding the enzyme differences, their properties were as follows: a) CK-1B could bind on ATP-agarose affinity column, while CK-1C could not; b) the activity of CK-1C was strongly stimulated at low concentrations (1 mmol/L) of spermine, while that of CK-1B was inhibited; c) CK-1B and CK-1C Km values for ATP were 11 μmol · L⁻¹ and 26 μmol · L⁻¹, respectively; d) Mg²⁺ could substituted by Mn²⁺ in the CK-1B catalytic activity (by about 80 percnt;); e) CK-1B phosphorylated serine, while CK-1C both serine and threonine on casein. The combination of these results with those from Babatsikos and Yupsanis (2000) brings the number of investigated maize endosperm CK-I isoforms to three (CK-1B, CK-1C, and CK-1E). This is the first biochemical approach demonstrating that multiple isoforms of CK-I casein kinases are present in the same plant tissue.     

Biological rhythms in birds — development, insights and perspectives

The aim of this review is to show that probably the internal clock of precocial birds is imprinted in the prenatal period by exogenous factors (zeitgeber). The activity of organ functions occurs early during embryonic development, before this function is ultimately necessary to ensure the survival of the embryo. Prenatal activation of some functional systems may have a training effect on the postnatal efficiency.The development of physiological control systems is influenced by endogenous and exogenous factors during the late prenatal and early postnatal period: epigenetic adaptation processes play an important role in the development of animals; they have acquired characteristics which are innated but not genetically fixed. As a rule, the actual value during the determination period has a very strong influence on the set-point of the system. This will be explained using the example of thermoregulation.It is shown in detail that it seems to be possible to imprint the prenatal development of circadian rhythms by periodic changes of the light-dark cycle but not by rhythmic influence of acoustic signals.Altogether, there are more questions open than solved concerning the perinatal genesis of circadian rhythms in birds. Topics are given for the future research.     

Multigenic polymorphism as a source of adaptively restricted population variability

Among 400 Drosophila melanogaster flies individually analysed for nine gene–enzyme systems, a total of 160 different genotypes were found: 78 were repeatedly observed in two to 22 individuals, and 82 appeared only once. An increase in the frequency of rare alleles could be observed in such groups of genotypes that were less frequent. Among 24 most frequent genotypes (in 189 individuals) only four different combinations of five third-chromosomal genes are present, and 12 different combinations of three second-chromosomal genes. Among 82 unique genotypes these combinations were much more versatile: 29 at the third, 22 at the second, and three at the first chromosomes. The proportion of heterozygotes was increasing from most frequent toward unique genotypes (1.5–2.1 ± 0.1), primarily due to heterozygosity in five third-chromosomal loci (0.4–1.1 ± 0.06). When the number of genotypes in 100, 200, 300 and 400 sampled flies was extrapolated to larger samples, it became clear that this increase has an asymptotic character. It must be assumed that for our nine-loci model a total of approximately 200–220 different genotypes may exist in a population of a few thousands individuals, which means that adaptive variation for such a complex gene–enzyme system must be very much limited.     

The distribution of benthic marine algae in relation to the temperature regulation of their life histories

Mainly on the basis of the distribution patterns of 42 species of the recently revised genus Cladopkora (Chlorophyceae) in the north Atlantic Ocean, it appeared possible to distinguish 10 phytogeographic distribution groups of wide applicability. Experimentally determined critical temperatures limiting essential events in the life histories of 17 benthic algal species were used to infer possible phytogeographic boundaries; these appeared to fit closely the phytogeographic boundaries derived from field-distribution data. For a temperate species, at least six different boundaries can be postulated and should be checked in the northern hemisphere: (1) the ‘northern lethal boundary’ (corresponding to the lowest winter temperature which a species can survive); (2) the ‘northern growth boundary’ (corresponding to the lowest summer temperature which, over a period of several months, permits sufficient growth); (3) the ‘northern reproductive boundary’ (corresponding to the lowest summer temperature permitting reproduction over a period of several months); (4–6) the corresponding southern boundaries. Photoperiodic responses may influence the temperature responses. Many phytogeographic boundaries appear to be of a composite nature. For instance, the southern boundary of Laminaria digitata follows the European 10°C February isotherm (which corresponds to the highest winter temperature permitting fertility in the female gametophyte, i.e. to the ‘southern reproductive boundary’), and the American 19°C summer isotherm (corresponding to the ‘southern lethal boundary’). Thus, experimental evidence supports the validity of eight of the following 10 distribution groups (for distribution groups 2 and 6, such evidence could not be found): (1) the amphiatlantic tropical-to-warm temperate group, with a north-eastern extension (examples: Gracilaria foliifera and Centroceras clavulalum); (2) the amphiatlantic tropical-to-warm temperate group, with a north-western extension (example: Hypnea musciformis); (3) the amphiatlantic tropical-to-temperate group (example: Sphacelaria rigidula =furcigera); (4) the amphiatlantic temperate group: the Cladophora rupestris type (examples: Callithamnion hookeri, Dumontia contorta; Laminaria saccharina is transitional to type 10, I., digitata to types 5 and 10); (5) the amphiatlantic temperate group: the Cl. albida type (examples: Scytosiphon lomentaria, Petalonia fascia); (6) the tropical western Atlantic group; (7) the north-east American tropical-to-temperate group (example: Gracilaria tikvahiae); (8) the north-east American temperate group and the corresponding Japanese temperate group (examples: Campylaephora hypneoides and Sargassum muticum); (9) the warm-temperate Mediterranean-Atlantic group, and the corresponding warm-temperate Californian group (examples: Saccorhiza polyschides, Laminaria hyperborea, I., ockroleuca, Macrocystis pyrifera, Hedophyllum sessile); (10) the Arctic group (examples: Saccorhiza dermatodea and Sphacelaria arctica). Distribution groups 6, 9 and 10 have comparatively narrow temperature ranges with a span of 18 22°C between their lethal boundaries and of 5 12°C between their reproductive or growth boundaries. These narrow temperature ranges limit the species in these groups to the tropics; the temperate coasts on the eastern sides of the north Pacific and north Atlantic Oceans and in the southern hemisphere; and to the Arctic, respectively. The narrow temperature ranges of group 9 make the species in this group unfit for life on the western temperate coasts of the north Pacific and north Atlantic Oceans, where algae must cope with annual temperature fluctuations of more than 20°C. Conversely, algae in group 8 (containing the numerous Japanese endemic species) are characterized by wide temperature spans (e.g. 29°C between ‘lethal boundaries’, 12–19°C between ‘growth and/or reproductive boundaries’) and must be potentially capable of occupying wide latitudinal belts on temperate coasts along the east sides of the north Pacific and north Atlantic Oceans. Algae ‘escaped’ from Japan, such as Sargassum muticum, conform to this picture. Apparently Japanese algae do not have the capacity for long distance dispersal. The corresponding east American coasts (30–45 N) harbour very few endemic species, probably as a result of the adverse nature of these sediment coasts for benthic macroalgae and their functioning as a barrier to latitudinal displacements of the flora during glaciations. The remaining distribution groups (1,2,3,4,5,7) are characterized by wide temperature spans and wide distributions, often in both the Atlantic and Pacific Oceans and in both hemispheres. Six temperate species (in distribution groups 4, 5 and 9) with an amphiaequatorial distribution have similar winter-temperature maxima permitting reproduction and corresponding with winter isotherms of 15–17°C; their upper lethal temperatures are more dissimilar and correspond with summer isotherms of 20–30°C. Their amphiaequatorial distribution can be explained by assuming glacial temperature drops along east Pacific and east Atlantic equatorial coasts in narrow belts of intensified upwelling during the presumably intensified glacial circulation of the ocean gyres.