Co-expression pattern of dopamine beta-hydroxylase (DβH) and neuropeptide Y (NPY) within sympathetic innervation of ovary and umbilical cord of the European bison (Bison bonasus L.)

Co-expression of dopamine β-hydroxylase (DβH) and neuropeptide Y (NPY) has never been examined in ovary (OV) and umbilical cord (UC) of the European bison (Eb), the endangered wild species. The OV and UC samples were harvested from seasonally eliminated Eb females (45–120 days post coitum). Frozen histological sections were examined by double fluorescent immunohistochemistry (dF-IHC), using the primary mouse anti-DβH monoclonals and rabbit anti-NPY polyclonals and then the immunocomplexes were visualized with FITC and CY3 fluorophores, respectively. Numerous DβH immunoreactive nerve fibers (DβH-IRs) and a little less frequent NPY-IRs were found in the bundle-like structures, innervating mainly perivascular regions of the OV. The NPY-IRs constantly co-expressed DβH, while some DβH-IRs did not express NPY. This specific pattern of innervation was observed both in the stromal and cortical regions of the OV. The simultaneous co-expression of DβH and NPY were also detected in the UC, in which specific single or bundle-like structures ran along the smooth muscles of blood vessels. The spatial-specific co-expression of DβH and NPY in OV and UC, may suggest that these markers are involved in the control of vascularization that regulates nourishing blood circulation required for proper pregnancy maintenance and efficient embryo/fetus development in the Eb.     

Design and synthesis of 1-aryl-5-anilinoindazoles as c-Jun N-terminal kinase inhibitors

Starting from pyrazole HTS hit (1), a series of 1-aryl-1H-indazoles have been synthesized as JNK3 inhibitors with moderate selectivity against JNK1. SAR studies led to the synthesis of 5r as double digital nanomolar JNK3 inhibitor with good in vivo exposure.     

Mitoxantrone changes spectrin-aminophospholipid interactions

Understanding drug-membrane and drug-membrane protein interactions would be a crucial step towards understanding the action and biological properties of anthracyclines, as the cell membrane with its integral and peripheral proteins is the first barrier encountered by these drugs. In this paper, we briefly describe mitoxantrone-monolayer and mitoxantrone-bilayer interactions, focusing on the effect of mitoxantrone on the interactions between erythroid or nonerythroid spectrin with phosphatidylethanolamine-enriched mono- and bilayers. We found that mitoxantrone markedly modifies the interaction of erythroid and nonerythroid spectrins with phosphatidylethanolamine/phosphatitydcholine (PE/PC) monolayers. The change in Δπ induced by spectrins is several-fold larger in the presence of 72?nM mitoxantrone than in its absence: spectrin/mitoxantrone complexes induced a strong compression of the monolayer. Spin-labelling experiments showed that spectrin/mitoxantrone complexes caused significant changes in the order parameter measured using a 5′-doxyl stearate probe in the bilayer, but they practically did not affect the mobility of 16′-doxyl stearate. These results indicate close-to-surface interactions/penetrations without significant effect on the mid-region of the hydrophobic core of the bilayer. The obtained apparent equilibrium dissociation constants indicated relatively similar mitoxantrone-phospholipid and mitoxantrone-spectrin (erythroid and nonerythroid) binding affinities. These results might in part, explain the effect of mitoxantrone on spectrin distribution in the living cells.     

Increased level of fibrinogen chains in the proteome of blood platelets in secondary progressive multiple sclerosis patients

Epidemiological studies indicate a high risk of stroke, heart failure and myocardial infarction in patients with multiple sclerosis, especially in its secondary progressive (SPMS) phase. Some ischaemic events are directly associated with abnormal platelet functions and their prothrombotic activity. Recent reports, including this study, confirm the increased activation of circulating platelets in SPMS, and also show increased platelet reactivity, among other responses, as well as strong aggregation. In this current study, we conducted a comparative analysis of the platelet proteome in SPMS patients and in healthy controls, to demonstrate the quantitative and qualitative differences likely to affect functional changes observed in SPMS. During densitometry evaluation of 2‐D fluorescence difference gel electrophoresis, we observed differences between the electrophoretic patterns of SPMS platelets and the control samples. To determine a detailed characterisation of the proteome changes in the SPMS patients’ blood platelets, in the next stage, we performed mass spectrometry of selected spots and indicated the increased presence of four proteins (fibrinogen, α‐2 macroglobulin, septin‐14 and tubulin β‐1 chain). The most important of these is the increased amount of prothrombotic protein, fibrinogen, which seems to confirm the accuracy of the imaging and potentially explains the increased risk of platelet‐origin thrombotic events. This study provides new knowledge of the potential existence of the molecular mechanisms responsible for the acceleration of the platelet pro‐coagulant function in SPMS. This can help to identify new targets for therapy, which can then be used not only in the second stage of the disease.     

Quantification and purity determination of newly synthesized thioacridines by capillary liquid chromatography

Capillary liquid chromatography (CLC) was applied for quantification and impurity profile determination of ten newly synthesized acridine thioderivatives. A reversed-phase CLC system employing two different stationary phases, Nucleosil C18 and LiChrosorb RP-select B, was used. The mobile phase composition was optimized to get a satisfactory separation of impurities from the main acridine component in a reasonable analysis time. Significant differences in the chromatographic behavior between acridine derivatives containing and lacking amino groups were observed. Optimized separation conditions were used in CLC to measure the calibration curves of the acridine derivatives in a concentration range from 1.0 x 10(-6) to 1.0 x 10(-3) M at two different detector wavelengths (214 and 230 nm). Limits of detection and quantification of all the substances were determined. The detection limits went down to units of microM for most of the derivatives. CLC was also demonstrated to be a suitable method for the purity determination of test batches of the acridine thioderivatives.     

Clinical features,treatment and genetic background of Treacher Collins syndrome

Treacher Collins syndrome (TCS) is an autosomal dominant disorder of craniofacial development. The major features of the disease include midface hypoplasia, micrognathia, microtia, conductive hearing loss and cleft palate. Current procedures of surgical treatment of TCS are discussed and novel findings concerning the genetic background of TCS are described. The TCS locus has been mapped to chromosome 5q31.3-32. The TCOF1 gene contains 26 exons and encodes a 1411 amino acid protein named treacle. In the TCOF1 gene 51 mutations have been identified. Most of these mutations are insertions or deletions, which result in an introduction of a premature termination codon into the reading frame. Mutational spectra support the hypothesis that TCS results from haploinsufficiency of treacle.     

RNase HI Is Essential for Survival of Mycobacterium smegmatis

RNases H are involved in the removal of RNA from RNA/DNA hybrids. Type I RNases H are thought to recognize and cleave the RNA/DNA duplex when at least four ribonucleotides are present. Here we investigated the importance of RNase H type I encoding genes for model organism Mycobacterium smegmatis. By performing gene replacement through homologous recombination, we demonstrate that each of the two presumable RNase H type I encoding genes, rnhA and MSMEG4305, can be removed from M. smegmatis genome without affecting the growth rate of the mutant. Further, we demonstrate that deletion of both RNases H type I encoding genes in M. smegmatis leads to synthetic lethality. Finally, we question the possibility of existence of RNase HI related alternative mode of initiation of DNA replication in M. smegmatis, the process initially discovered in Escherichia coli. We suspect that synthetic lethality of double mutant lacking RNases H type I is caused by formation of R-loops leading to collapse of replication forks. We report Mycobacterium smegmatis as the first bacterial species, where function of RNase H type I has been found essential.     

The development of benzimidazoles as selective rho kinase inhibitors

Rho Kinase (ROCK) is a serine/threonine kinase whose inhibition could prove beneficial in numerous therapeutic areas. We have developed a promising class of ATP-competitive inhibitors based upon a benzimidazole scaffold, which show excellent potency toward ROCK (IC₅₀ <10 nM). This report details the optimization of selectivity for ROCK over other related kinases such as Protein kinase A (PKA).     

Comparative analysis of cis-regulation following stroke and seizures in subspaces of conserved eigensystems

Background  

It is often desirable to separate effects of different regulators on gene expression, or to identify effects of the same regulator across several systems. Here, we focus on the rat brain following stroke or seizures, and demonstrate how the two tasks can be approached simultaneously.