Artificial reproduction of wild and cultured barbel (Barbus barbus, Cyprinidae) under controlled conditions

The aim of this work was to compare the effects of controlled reproduction of cultured and wild common barbel, Barbus barbus (L.). Preparations containing different GnRH analogues and dopamine receptor antagonists (Ovopel, Ovaprim) as well as human chorionic gonadotropin (hCG) (in the case of cultured fish) were applied and their influence on ovulation, spermiation and quality of gametes obtained was determined. No differences in the qualitative or quantitative parameters of semen were found between fish stimulated with different hormonal preparations and those not receiving hormonal stimulation. The high suitability of Ovaprim for ovulation induction in (cultured and wild) barbel was confirmed. The highest synchronisation of ovulation was obtained after the application of Ovopel (18 ± 3 h), but the best results of controlled reproduction (expressed as the percentage of ovulations and survival of embryos) were obtained by applying Ovaprim (83.2 ± 4.1). A significantly higher percentage of ovulation was obtained in cultured fish (80-90%) than in wild fish (< 25%).     

Interaction between zinc deficiency and boron toxicity on growth and mineral nutrition of sour orange seedlings

Sour orange (Citrus aurantium L.) seedlings were grown for 3 months in diethylenetriamine pentaacetate (DTPA)‐buffered nutrient solutions to study the effect of Zn stress on the plants’ sensitivity to high boron concentration in the root environment. There were three zinc treatments: 21 μM Zn (LOW Zn‐DTPA), 69 μM Zn (NORMAL Zn‐DTPA) in the nutrient solution, or 12 weekly foliar sprays with ZnSO₄ (FOLIAR‐Zn). In the FOLIAR‐Zn treatment, the nutrient solution contained 21 μM Zn. Zn activities calculated with a chemical equilibrium model, Geochem PC, and expressed as pZn=‐log(Zn⁺²), were 10.2 and 9.7 in the LOW Zn‐DTPA and NORMAL Zn‐DTPA nutrient solutions, respectively. One half of the plants in each Zn treatment were grown in 51 μM B (NORMAL‐B) and the other half in 200 μM B (HIGH‐B) nutrient solution. Seedlings grown in LOW Zn‐DTPA/NORMAL‐B nutrient solution developed Zn deficiency symptoms such as: reduced shoot growth, small and chlorotic leaves, and white roots with visibly shorter and thicker laterals than in Zn sufficient plants. The HIGH‐B treatment decreased shoot growth, leaf and stem dry weight, leaf area, and induced severe leaf B toxicity on seedlings grown in the LOW Zn‐DTPA nutrient solution but the effect was either absent or less pronounced in the NORMAL Zn‐DTPA or FOLIAR‐Zn treatments. Seedlings in the LOW Zn‐DTPA FOLIAR‐Zn treatments but they had lower B concentration on a whole plant basis indicating less B uptake per unit of dry weight. The FOLIAR‐Zn and NORMAL Zn‐DTPA treatments were equally effective in alleviating leaf B toxicity symptoms. The FOLIAR‐Zn treatment, however, was less effective than the NORMAL Zn‐DTPA treatment in alleviating the deleterious effect of high B on leaf dry weight even though the B concentrations in leaves, stems, and roots of the foliar‐sprayed seedlings were similar to the NORMAL Zn‐DTPA seedlings. Leaf concentrations of phosphorus, potassium, magnesium, iron, mangenese, and copper were within the optimal range for citrus with the exception of Ca which was low. Although B and particularly Zn treatments modified the concentration of some of these elements in leaves and roots, these changes were too small to explain the observed growth responses. The observation that B toxicity symptoms in Zn‐deficient citrus could be mitigated with Zn applications is of potential practical importance as B toxicity and Zn deficiency are simultaneously encountered in some soils of semiarid zones.     

The Impact of Fertilization Methods on the Nutrient Balance and Changes of Soil Chemical Features in Crop Rotation

The effect of various fertilization methods on the simplified nutrients balance and changes of soil chemical features in crop rotation was estimated on the basis of 7-year (1994-2000) studies carried out on loess-based, sedimentary-gley, light-clay soil. Different fertilization methods were used: A - mineral fertilization; B - mineral and manure fertilization; C - straw, green manure and mineral fertilization; D - ecological compost and biodynamic preparation fertilization. Particular fertilization methods caused a significant differentiation of the nutrients balance. A positive phosphorus balance was found out in all the fertilization systems. It has been observed that the nitrogen and potassium carry-in with fertilizers was higher than the carry-out with crops in system B and C. A favorable calcium balance was achieved in systems C and D while a favorable magnesium balance - only in the ecologically fertilized systems (D). The changes of soil chemical features were only insignificantly dependent on the nutrient balance. After seven years of research, the contents of humus, phosphorus, potassium, magnesium and calcium increased while the soil reaction decreased in the soil of all the fertilized systems except the ecologically fertilized system.     

Maternal HMB treatment affects bone and hyaline cartilage development in their weaned piglets via the leptin/osteoprotegerin system

It has been demonstrated in animal studies that prenatal administration of β‐hydroxy‐β‐methylbutyrate (HMB, metabolite of leucine) influences general growth and mechanical endurance of long bones in newborn offspring in sex‐dependent manner. The present experiment was conducted to evaluate the effect of HMB treatment of pregnant sows on bone development in offspring at weaning. From 70th day until the 90th day of gestation, sows received either a basal diet (n = 12) or the same diet supplemented with HMB (n = 12) at the dose of 0.2 g/kg of body weight/day. Femora obtained from six males and females in each group weaned at the age of 35 days were examined. Maternal HMB treatment significantly enhanced body weight and changed bone morphology increasing femur mechanical strength in both sexes. Maternal HMB supplementation also elevated bone micro‐ and macroelement concentrations and enhanced content of proteoglycans in articular cartilage. Based on the obtained results, it can be concluded that maternal HMB supplementation in the mid‐gestation period significantly accelerated bone development in both sexes by upregulation of a multifactorial system including leptin and osteoprotegerin. However, the sex (irrespective of the HMB treatment) was the factor which influenced the collagen structure in cartilages and trabecular bone, as demonstrated both by the Picrosirius red staining and performed analysis of thermal stability of collagenous tissues. The structural differences in collagen between males and females were presumably related to a different collagen maturity. No studies conducted so far provided a detailed morphological analysis of bone, articular cartilage, growth plate and the activities of the somatotropic and pituitary–gonadal axes, as well as leptin/osteoprotegerin system in weaned offspring prenatally treated with HMB. This study showed also the relationship between the maternal HMB treatment and bone osteometric and mechanical traits, hormones, and growth and bone turnover markers such as leptin, osteoprotegerin and insulin‐like growth factor‐1.     

The preliminary studies on protein profile in retained and not retained foetal membranes in heavy draft mares

Protein profile of the placenta expresses its function and maintenance. Any alterations can be reflected in qualitative and quantitative changes in this profile. The aim of the present study was the evaluation of protein profile in the placenta of mares suffering from the retention of foetal membranes (FMR) by two separation methods and the comparison with physiologically released tissues. Placentas from 14 healthy, heavy draft mares were collected immediately after the expulsion of newborn. Tissues after homogenization and staining with fluorescent dyes were subjected to electrophoretic as well as chromatographic separation. Electrophoretic gels were statistically analysed for the presence and abundance of examined proteins, while some proteins were identified in chromatographic fractions. Out of 248 spots detected in endometrium, 38 differed significantly between FMR and control animals, while in allantochorion, respective values reached 241 and 27 spots (p < .05). Among identified proteins that expressed higher abundance in endometrium of FMR mares than control animals were prostaglandin reductase, dehydrogenase/reductase SDR family, and placental growth factor. These proteins are involved in regulation of parturition. Additionally, the following proteins responsible for physiological activity of a cell—guanine methyl transferase, aspartyl/asparaginyl beta‐hydroxylase and GTP‐binding protein, were identified. These proteins expressed higher abundance in allantochorion of FMR mares than in controls. This preliminary study confirmed the disturbances in protein pattern between foetal membranes in FMR and healthy mares. Further qualitative and quantitative experiments are necessary to deepen our knowledge on the mechanisms of the retention of foetal membranes in mares.     

Effect of conjugated linoleic acids on the activity and mRNA expression of 5- and 15-lipoxygenases in human macrophages

Lipoxygenases are a family of non-heme enzyme dioxygenases. The role of lipoxygenases is synthesis of hydroperoxides of fatty acids, which perform signaling functions in the body. Studies on conjugated linoleic acids (CLAs) as fatty acids with a potential anti-atherosclerotic function have recently been initiated. The aim of the study was to test the effect of CLAs and linoleic acid on 5- and 15-lipoxygenase (5-LO, 15-LO-1) enzyme activity, their mRNA expression, and concentration in the cells. It was also desired to determine whether the CLAs are substrates for the enzymes. For the experiments monocytic cell line (THP-1) and monocytes obtained from human venous blood were used. Monocytes were differentiated to macrophages: THP-1 (CD14+) by PMA administration (100 nM for 24 h) and monocytes from blood (CD14+) by 7-day cultivation with the autologous serum (10%). After differentiation, macrophages were cultured with 30 microM CLAs or linoleic acid for 48 h. The 15- and 5-lipoxygenase products were measured by HPLC method. mRNA expression and protein content were analyzed by real-time PCR and Western blot analysis. The in vitro studies proved that both CLA isomers are not substrates for 15-LO-1; in ex vivo studies hydroxydecadienoic acid (HODE) concentration was significantly reduced (p = 0.019). The trans-10,cis-12 CLA isomer reduced HODE concentration by 28% (p = 0.046) and the cis-9,trans-11 CLA isomer by 35% (p = 0.028). In macrophages obtained from THP-1 fatty acids did not change significantly mRNA expression of the majority of the investigated genes. CLAs did not change the content of 5-LO and 15-LO-1 proteins in macrophages obtained from peripheral blood. Linoleic acid induced 15-LO-1 expression (2.6 times, p < 0.05). CLAs may perform the function of an inhibitor of lipoxygenase 15-LO-1 activity in macrophages.     

Phyto- and endogenous estrogens differently activate intracellular calcium ion mobilization in bovine endometrial cells

The main purpose of this study was to check whether phyto- and endogenous estrogens influence calcium ion mobilization [Ca(2+)](i) in bovine endometrial cells and whether this action is connected with biological effects i.e. prostaglandin (PG)F(2alpha) production. In our study we used two calcium measurement methods by comparing the microscopic method with widely used quantitative - spectrofluorometric method of [Ca(2+)](i) measurement. We also wanted to confirm whether visualization of calcium ion [Ca(2+)](i) in cells using microscopic method supported by micro image analysis (Micro Image Olympus system) reflects real, qualitative changes in the ion concentration. In both methods a cell-permeable form of fluorescent [Ca(2+)](i) indicator Fura-2 was used. Cultured bovine endometrial epithelial and stromal cells influenced by phorbol-2-myristate-13-acetate (PMA; positive control), estradiol 17-beta (E(2); endogenous estrogen) and active metabolites of phytoestrogens (environmental estrogens) were used as a model to study PGF(2alpha) secretion and [Ca(2+)](i) mobilization in the cells. Equol and para-ethyl-phenol in doses of 10(-8)-10(-6) M increased PGF(2alpha) concentration both in epithelial and stromal cells (P<0.05). In both methods, equol and para-ethyl-phenol did not cause intracellular [Ca(2+)](i) mobilization in epithelial and stromal cells (P>0.05). Both methods revealed that only E(2) and PMA induced intracellular [Ca(2+)](i) mobilization in epithelial and stromal cells (P<0.05). The results of both methods were highly correlated (P<0.001; r=0.82 for epithelial cells and r=0.89 for stromal cells). In conclusion, both methods gave approximately the same results and showed that phytoestrogens, in contrast to PMA and E(2), did not cause intracellular [Ca(2+)](i) mobilization in endometrial cells. The obtained results proved that the [Ca(2+)](i) visualization method supported by micro image analysis can produce similar results to the spectrofluorometric method.