Phosphonic analogues of morphactins. Part III: Peptides containing p-terminal 9-aminofluoren-9-ylphosphonic acid

Peptides of 9-aminofluoren-9-ylphosphonic or 1-aminocyclohexylphosphonic acids were synthesised and found to have typical morphactin activity when tested on Lepidium sativum and Triticum vulgare. The observed effects included root elongation with simultaneous stem shortening. The results obtained indicate that these phosphonic acid derivatives of cyclohexylamine and fluoren-9-ylamine may have a similar mechanism of action.     

Herbicidal activity of derivatives of 9-aminofluoren-9-ylphosphonic acid

A series of derivatives of 9-aminofluoren-9-ylphosphonic acid (phosphonic analogues of morphactins) were synthesized and screened for herbicidal activity against Lepidium sativum, Cucumis sativus and Lycopersicon esculentum. Ethyl 9-(N-alkylamino)fluoren-9-yl(phenyl)phosphinates appeared to be equipotent with glyphosate and thus may be recognized as new lead compounds for further structural modifications.     

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.     

Effects of prostaglandin F(2alpha) and nitric oxide on the secretory function of bovine luteal cells

The objective of the present study was to investigate the influence of prostaglandin F(2alpha) (PGF (2alpha)) and nitric oxide (NO) on production of steroids and PGs by culturing bovine luteal cells obtained from ovaries on days 8-12 of the estrous cycle with a nitric oxide (NO) donor (Spermine NONOate), and a NO synthase inhibitor (N(G)-nitro-L-arginine methyl ester dihydrochloride: L-NAME). When the cells were exposed for 24 h to PGF(2alpha) (10(-7)-10(-5) M), production of progesterone (P(4)) increased significantly at all doses used (P<0.05). Moreover, PGF(2alpha) stimulated PGF(2alpha) production (P<0.01), depressed testosterone (T) production (P<0.05), but did not affect synthesis of prostaglandin E(2) (PGE(2)). Spermine NONOate decreased P(4) production to 66%, 47% and 34% of the control concentration after treatment with 10(-5) M, 10(-4) M and 10(-3) M, respectively, but did not affect T production, and increased PGF(2alpha) synthesis (P<0.05) and PGE(2) (P<0.01) at all doses used. L-NAME increased production of P(4) (P<0.01) but did not affect (P>0.05) secretion of T, PGF(2alpha) and PGE(2). Estradiol-17beta (E(2)) was detectable on the level of sensitivity of assay and was not significantly altered by any treatments. The overall results suggest that PGF(2alpha) and NO produced locally in bovine CL play roles in the regulation of the secretory function of the bovine CL as auto/paracrine factors.     

Forecasting the milk yield of cows on farms equipped with automatic milking system with the use of decision trees

The purpose of this paper was to utilize the decision trees technique to determine the factors responsible for high monthly milk yield in Polish Holstein‐Friesian cows from 27 herds equipped with milking robots. The applied statistical method—the decision tree technique—showed that the most important factors responsible for monthly milk yield of dairy cows using robots were, in descending order of importance: milking frequency, lactation number, month of milking, and type of lying stall. At the same time, it has been ascertained that the highest monthly milk yield (47.24 kg) can be expected from multiparous cows kept in barns with a deep bedding that were milked more frequently than three times per day. On the other hand, the lowest milk production (13.56 kg) was observed among dairy cows milked less frequently than two times a day, with an average number of milked quarters lower than 3.97. The application of the decision trees technique allows a breeder to select appropriate levels of environmental factors and parameters that will help to ensure maximized milk production.     

Domestication affects spawning of the ide (Leuciscus idus)—preliminary study

Ide, Leuciscus idus, breeders in Polish fish farm are cultured or are collected from the wild. The data about differences in reproduction of different ide forms in captivity are limited. In the present paper domesticated and wild stock are reproduced artificially. Cultured fish do not require application of spawning agents. For obtaining good-quality oocytes only stimulation with temperature and photoperiod is necessary.     

Progesterone stimulation by LH involves the phospholipase-C pathway in bovine luteal cells

Luteinizing hormone (LH)-stimulated steroidogenesis in luteal cells is known to be mediated through the activation of cyclic AMP (cAMP)-dependent protein kinase, and to be also modulated by calcium-dependent mechanisms. In the present study, we tested the hypothesis that LH stimulates progesterone (P4) production in bovine luteal cells through activation of phospholipase (PL) C by using a cell culture system. Bovine mid-luteal cells (Days 8-12 of the estrous cycle) were cultured for 24 h and then exposed to a PLC inhibitor (U-73122; 10 microM) with or without LH (10 ng/ml) for 4 h. U-73122 blocked LH-stimulated P4 production without affecting cAMP accumulation. Moreover, exposure of luteal cells to PLC increased P4 production in a dose-dependent manner. These results support the hypothesis that the luteotropic action of LH in bovine luteal cells is mediated not only by activation of adenylate cyclase but also by activation of PLC.     

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.