Alterations in the expression of the Atp7a gene in the early postnatal development of the mosaic mutant mice (Atp7amo-ms) – An animal model for Menkes disease

Copper is a trace element that is essential for the normal growth and development of all living organisms. In mammals, the ATP7A Cu-transporting ATPase is a key protein that is required for the maintenance of copper homeostasis. In both humans and mice, the ATP7A protein is coded by the X-linked ATP7A/Atp7a gene. Disturbances in copper metabolism caused by mutations in the ATP7A/Atp7a gene lead to severe metabolic syndromes Menkes disease in humans and the lethal mottled phenotype in mice. Mosaic is one of numerous mottled mutations and may serve as a model for a severe Menkes disease variant. In Menkes patients, mutations in the ATP7A gene often result in a decreased level of the normal ATP7A protein. The aim of this study was to analyse the expression of the Atp7a gene in mosaic mutants in early postnatal development, a critical period for starting copper supplementation therapy in both Menkes patients and mutant mice. Using real-time quantitative RT-PCR, we analysed the expression of the Atp7a gene in the brain, kidney and liver of newborn (P0.5) and suckling (P14) mice. Our results indicate that in mosaic P0.5 mutants, the Atp7a mRNA level is decreased in all analysed organs in comparison with wild-type animals. In two week-old mutants, a significant decrease was observed only in the kidney. In contrast, their hepatic level of Atp7a tended to be higher than in wild-type mice. We speculate that disturbance in the expression of the Atp7a gene and, consequently, change in the copper concentration of the organs, may contribute to the early fatal outcome of mosaic males.     

Photosynthetic net O<Subscript>2</Subscript> evolution enhancement as a sign of acclimation to phosphorus deficiency in bean (<Emphasis Type="Italic">Phaseolus vulgaris</Emphasis> L.) leaves

Primary leaves of bean (Phaseolus vulgaris L.) seedlings cultivated for 14 days in a growth chamber on complete (control) and phosphate deficient (−P) Knop liquid medium were used for measurements. The −P leaves were smaller and showed an increased specific leaf area (SLA). Their inorganic phosphate (P_i) concentration was considerably lowered. They did not show any significant changes in chlorophyll (Chl) (a + b) concentration and in their net CO₂ assimilation rate when it was estimated under the conditions close to those of the seedlings growth. Light response curves of photosynthetic net O₂ evolution (P _NO₂) of the leaves for the irradiation range up to 500 μmol(photon) m⁻² s⁻¹ were determined, using the leaf-disc Clark oxygen electrode. The measurements were taken under high CO₂ concentration of about 1 % and O₂ concentrations of 21 % or lowered to about 3 % at the beginning of measurement. The results obtained at 21 % O₂ and the irradiations close to or higher than those used during the seedlings growth revealed the phosphorus stress suppressive effect on the leaf net O₂ evolution, however, no such effect was observed at lower irradiations. Other estimated parameters of P _NO₂ such as: apparent quantum requirement (QR_A) and light compensation point (LCP) for the control and −P leaves were similar. However, with a high irradiation and lowered O₂ concentration the rate of P _NO₂ for the −P leaves was markedly higher than that for the control, in relation to both the leaf area and leaf fresh mass. This difference also disappeared at low irradiations, but the estimated reduced QR_A values indicate, under those conditions, the increased yield of photosynthetic light reaction, especially in the −P leaves. The presented results confirm the suggestion that during the initial phase of insufficient phosphate feeding the acclimations in the light phase of photosynthesis, both structural and functional appear. They correspond, probably, to the increased energy costs of carbon assimilation under phosphorus stress, e.g. connected with raised difficulties in phosphate uptake and turnover and enhanced photorespiration. Under the experimental conditions especially advantageous for the dark phase of photosynthesis (saturating CO₂ and PAR, low O₂ concentration), those acclimations may be manifested as an enhancement of photosynthetic net O₂ evolution.     

Ecological causes of morphological evolution in the three‐spined stickleback

The central assumption of evolutionary theory is that natural selection drives the adaptation of populations to local environmental conditions, resulting in the evolution of adaptive phenotypes. The three‐spined stickleback (Gasterosteus aculeatus) displays remarkable phenotypic variation, offering an unusually tractable model for understanding the ecological mechanisms underpinning adaptive evolutionary change. Using populations on North Uist, Scotland we investigated the role of predation pressure and calcium limitation on the adaptive evolution of stickleback morphology and behavior. Dissolved calcium was a significant predictor of plate and spine morph, while predator abundance was not. Stickleback latency to emerge from a refuge varied with morph, with populations with highly reduced plates and spines and high predation risk less bold. Our findings support strong directional selection in three‐spined stickleback evolution, driven by multiple selective agents.     

Haptoglobin glycoforms in a case of carbohydrate-deficient glycoprotein syndrome

Alterations in haptoglobin (Hp) glycosylation were examined in the plasma of the first patient with carbohydrate-deficient glycoprotein syndrome (CDGS) who was described in Poland. Hp concentration in the CDGS patient plasma was low (240mg/l) and the Hp phenotype was shown to be 2-2. Three glycoforms of the Hp subunit were observed in SDS-PAGE in CDGS. The densitometric analysis and molecular weight determinations suggested that 50% of glycoforms were fully glycosylated; 30% contained three out of four and 20% only two out of four glycan units compared to those that are present in Hp derived from healthy people. Results with lectins (concanavalin A and Sambucus nigra, Maackia amurensis and Alleuria aurantia agglutinins) indicate that all three glycoforms of subunit of CDGS-Hp contained biantennary complex glycans terminated with 2,6 bound sialic acid, but without fucose or 2,3 linked sialic acid. Hp glycosylation abnormalities described in this work suggest that this case was a type I carbohydrate-deficient glycoprotein syndrome.     

Ein Beitrag zur Flora Galiziens und der Bukowina

Ohne Zusammenfassung     

Interaction of serum amyloid A with human cystatin C—assessment of amino acid residues crucial for hCC–SAA formation (part II)

Secondary amyloid A (AA) amyloidosis is an important complication of some chronic inflammatory diseases, primarily rheumatoid arthritis (RA). It is a serious, potentially life‐threatening disorder caused by the deposition of AA fibrils, which are derived from the circulatory, acute‐phase‐reactant, serum amyloid A protein (SAA). Recently, a specific interaction between SAA and the ubiquitous inhibitor of cysteine proteases—human cystatin C (hCC)—has been proved. Using a combination of selective proteolytic excision and high‐resolution mass spectrometry, the binding sites in the SAA and hCC sequences were assessed as SAA(86–104) and hCC(96–102), respectively. Here, we report further details concerning the hCC–SAA interaction. With the use of affinity tests and florescent ELISA‐like assays, the amino acid residues crucial for the protein interaction were determined. It was shown that all amino acid residues in the SAA sequence, essential for the formation of the protein complex, are basic ones, which suggests an electrostatic interaction character. The idea is corroborated by the fact that the most important residues in the hCC sequence are Ser‐98 and Tyr‐102; these residues are able to form hydrogen bonds via their hydroxyl groups. The molecular details of hCC–SAA complex formation might be helpful for the design of new compounds modulating the biological role of both proteins. Copyright © 2013 John Wiley & Sons, Ltd.     

Antifungal activity of biflavones from Taxus baccata and Ginkgo biloba

Bilobetin and 4'-O-methylamentoflavone were isolated and identified in the needles of Taxus baccata, for the first time in this species. The antifungal activity of biflavones from T. baccata and Ginkgo biloba, namely amentoflavone, 7-O-methylamentoflavone, bilobetin, ginkgetin, sciadopitysin and 2,3-dihydrosciadopitysin towards the fungi Alternaria alternata, Fusarium culmorum, Cladosporium oxysporum was determined employing computer-aided image analysis couplet to a microscope. Bilobetin exhibited a significant antifungal activity with values of ED50 14, 11 and 17 microM respectively. This compound completely inhibited the growth of germinating tubes of Cladosporium oxysporum and Fusarium culmorum at a concentration 100 microM. Activity of ginkgetin and 7-O-methylamentoflavone towards Alternaria alternata was stronger than that of bilobetin. Moreover, slight structural changes in the cell wall of Alternaria alternata exposed to ginkgetin at concentration of 200 microM were observed.     

Combined effect of low-power laser irradiation and anthraquinone anticancer drug aclarubicin on survival of immortalized cells: Comparison with mitoxantrone

The photodynamic response of the anthraquinone anticancer drug aclarubicin (ACL) was investigated in vitro and compared with that of mitoxantrone (MTX). Cultured immortalized rodent B14 and NIH 3T3 cells were used in the experiments as a model for cells with neoplastic phenotype. Long-term cytotoxicity and inhibition of cell proliferation assayed by the clonal growth and MTT-tetrazolium methods were estimated to compare the efficacy of aclarubicin and mitoxantrone in photosensitizing cells and their death after non-thermal exposure to monochromatic laser light. Green He-Ne (543.5 nm) or red semiconductor (670 nm) low-power laser (LPL) irradiations were applied. Different dose-responses of both cell lines to aclarubicin and mitoxantrone were found so that the cytotoxicity of MTX was considerably greater than the cytotoxicity of ACL. Phototherapy response (P < 0.0001) was observed only for B14 cells after sensitisation with aclarubicin. Under the same conditions no significant effect of red light irradiation (semiconductor 670 nm laser) on survival of both cell lines treated with mitoxantrone was found.