Astrocyte- and hepatocyte-specific expression of genes from the distal serpin subcluster at 14q32.1 associates with tissue-specific chromatin structures

The distal serpin subcluster contains genes encoding alpha1-antichymotrypsin (ACT), protein C inhibitor (PCI), kallistatin (KAL) and the KAL-like protein, which are expressed in hepatocytes, but only the act gene is expressed in astrocytes. We show here that the tissue-specific expression of these genes associates with astrocyte- and hepatocyte-specific chromatin structures. In hepatocytes, we identified 12 Dnase I-hypersensitive sites (DHSs) that were distributed throughout the entire subcluster, with the promoters of expressed genes accessible to restriction enzyme digestion. In astrocytes, only six DHSs were located exclusively in the 5' flanking region of the act gene, with its promoter also accessible to restriction enzyme digestion. The acetylation of histone H3 and H4 was found throughout the subcluster in both cell types but this acetylation did not correlate with the expression pattern of these serpin genes. Analysis of histone modifications at the promoters of the act and pci genes revealed that methylation of histone H3 on lysine 4 correlated with their expression pattern in both cell types. In addition, inhibition of methyltransferase activity resulted in suppression of ACT and PCI mRNA expression. We propose that lysine 4 methylation of histone H3 correlates with the tissue-specific expression pattern of these serpin genes.     

Flow cytometric analysys of Enterococcus faecalis pAD1(-) strains response to cAD1 pheromone

Conjugative plasmids transfer in Enterococcus faecalis is inducted by sex pheromones. The pheromone is excreted by recipient cells and induces expression of aggregation protein AS in donor cells. This protein is involved in formation of matting aggregates. Use of flow cytometry and anti-As monoclonal antibodies allowed collect of interesting data pheromone response. However, according to our knowledge, no study focused on unspecific influence on particular pheromone for plasmid-free recipient strains. Six pAD1 (-) and tree pAD1 (+) Enterococcus faecalis stains were cultivated for 18h in BHI, with and without cAD1 pheromone (Sigma, Germany), respectively. The bacteria were washed, stained with carboksyfluorescein (FCDA, and analyzed by flow cytometry in FACS BD scan cytometr. Relative fluorescence and size of aggregation was used to compare influence on particular strains. Surprisingly, the results shows divergence in fluorescence, size of aggregates and degree of correlation between fluorescence of aggregates and their sizes among pAD1(-) strains, allowing for distinguish of two groups. Three of studied strains have higher fluorescence than pAD (+) stains. Correlation between fluorescence and size of aggregates, significant higher than in pAD1(+) stains, decrease from r = 0.88 to r=0.74 in reaction to cAD1. The strains if other group fluorize with lower intensity than pAD1 (+). Furthermore, 30.4% pAD1 (-) of second group have no detectable fluorescence. In contrast to pAD1 (-) ) strains of the first group and pA1 (+) strains, low (r=0.55) correlation between fluorescence and size of aggregates of group II increase up to r=0.74 after incubation with cAD1 pheromone. Previous study of these pAD1 (-) strains, currently assigned to group II, shown their low frequency of collecting aph2" gene encoded on other conjugative plasmid, pMG. According to these results, such flow cytometric analysis may be used to predict ability of strain to collect unrelated conjugative plasmid.     

Synthesis and biological activity of N-methylated analogs of endomorphin-2

In this paper, we describe the synthesis of a series of endomorphin-2 analogs containing N-methylated amino acids, consecutively in each position. The μ-opioid receptor binding affinities of the new analogs were determined in the displacement experiments. Their in vivo antinociceptive activity was assessed in the hot-plate test in mice after central (icv) and peripheral (ip) administration. [Sar²]endomorphin-2, which had the highest μ-receptor affinity, also showed the strongest analgesic effect when administered centrally and was the only analog that retained activity after peripheral injection.     

Calcium ionophore A23187 action on cardiac myocytes is accompanied by enhanced production of reactive oxygen species

We show that rat neonatal cardiac myocytes exposed to 1 micromol/l of the calcium ionophore A23187 respond with an enhanced production of reactive oxygen species (ROS). This dose is not cytotoxic to the myocytes. A higher concentration (10 micromol/l) evokes less ROS production and is significantly cytotoxic 24 h after exposure, but not immediately after removal of the A23187, when ROS are measured. Both cell death and the decrease in mitochondrial potential are only partially sensitive to MPT inhibitor cyclosporin A. Experiments performed to elucidate the sources of ROS included use of the nitric oxide synthase (NOS) inhibitor L-NAME; NOS involvement was excluded. Experiments with the oxidative phosphorylation uncoupler CCCP revealed that mitochondria are at least partially responsible for the observed effect. Further studies with cyclooxygenase (COX) and lipoxygenase (LOX) inhibitors (indomethacin and MK886, respectively) showed that these enzymes could also be sources of ROS when the calcium level is elevated. Their effect appeared to be independent of phospholipase A(2) inhibition, suggesting that COX and LOX stimulation is not due to elevated substrate (arachidonic acid) concentration but rather to a direct effect of calcium.     

Evaluation of selected alcoholic antiseptics activity against clinical bacterial strains

The aim of this study was to evaluate antimicrobial activity of selected alcoholic antiseptics against clinical strains, which possessed in majority a high level of drug resistance: MRSA (7), MSSA (3), E. coli: (9): strains producing ESBL (4), P. aeruginosa: (4), E. cloacae: (3), K. pneumoniae: (3). These strains were defined by MIC value, using antibiotic agar dilution method according to NCCLS. Fourteen alcoholic antiseptics were used in this study. Beside alcohol, they contained other active substances like iodine, hydrogen peroxide, chlorhexidine. Some additional agents were included for easier application, such as: gelling, moisturizing, aromatic or coloring substances. The objective of this study was also to determine the dependence of bactericidal activity on preparations (concentration). Product undiluted and diluted two and four times in water was analyzed according to prEN 12054 standard; 30 seconds and 1 minute contact time was used. The obtained data indicate that all tested undiluted antiseptics possessed bactericidal activity described by producers. However antiseptics (dilution leads to decrease and even loss of bactericidal activity. Two-times dilution of gel almost completely inactivated the product. Antimicrobial activity after 30 seconds of contact time was not affected by presence of additional agents in the tested antiseptics.     

The effect of aromatase inhibitor, fadrozole, on sGnRHa stimulated LH secretion in goldfish (Carassius auratus) and common carp (Cyprinus carpio)

The aromatase inhibitor, fadrozole, was applied to common carp and goldfish in order to examine its ability to increase the spontaneous and sGnRHa stimulated LH secretion. First, trials in goldfish in 2003 showed fadrozole's moderate ability to potentiate sGnRHa-stimulated LH secretion. However, this ability was much weaker than that obtained with dopamine antagonist, pimozide. There was no ovulation in fadrozole-treated fish. Several experiments on goldfish and carp during the two consecutive years with the different treatment regimes and doses of fadrozole did not confirm the optimistic results obtained in 2003. This shows that fadrozole is unable to replace the antidopaminergic drugs being used in fisheries practice.