Bacteriophage vB_EcoM_FV3: a new member of “rV5-like viruses”

A proposed new genus of the family Myoviridae, “rV5-like viruses”, includes two lytic bacteriophages: Escherichia coli O157: H7-specific bacteriophage rV5 and Salmonella phage PVP-SE1. Here, we present basic properties and genomic characterization of a novel rV5-like phage, vB_EcoM_FV3, which infects E. coli K-12-derived laboratory strains and replicates at high temperature (up to 47 °C). The 136,947-bp genome of vB_EcoM_FV3 contains 218 open reading frames and encodes 5 tRNAs. The genomic content and organization of vB_EcoM_FV3 is more similar to that of rV5 than to PVP-SE1, but all three phages share similar morphological characteristics and form a homogeneous phage group.     

Synthesis of N-arylsulfonyl DL-phenylserine derivatives exhibiting anti-inflammatory activity in experimental studies

Synthesis and evaluation of anti-inflammatory activity in rats with adjuvant arthritis of aryl sulfonyl derivatives of nonproteinogenic aromatic amino acids is reported. The studied compounds were synthesized by introducing residues of benzene-, p-toluene-, and p-bromobenzene sulfonic acids into threo-DL-phenylserine and erythro-DL-p-nitrophenylserine structures. From the set of 12 compounds tested in animal screening, N-(p-bromobenzenesulfonyl)-erythro-DL-p-nitrophenylserine ethyl ester 12 demonstrated the most pronounced anti-inflammatory activity. This compound inhibited inflammation process in polyarthritis phase by 53% (P < 0.001) though it was slightly toxic (LD50 > 6,000 mg kg(-1) for mice).     

Effects of age, period and cohort on stroke mortality among a middle-aged Lithuanian urban population from 1980 to 2004

AIMS: The main purpose of this paper was to assess the effect of age, period, and cohort on stroke mortality rates among a Lithuanian urban population aged 25-64 years (1041 men and 724 women) between 1980 and 2004. METHODS: Routine stroke mortality data were obtained from official Kaunas region mortality register by codes 430-438 and I60-I69 in the 9th and in the 10th revisions of the International Classifications of Diseases (ICD), respectively. Mortality rates per 100,000 persons for men and women were age-adjusted using the age distribution of the European Standard Population. Age-specific mortality rates were analysed by sex, period, and birth cohort in eight 5-year age groups and five 5-year age groups. Goodness of fit of the Poisson regression models were evaluated using Pearson and Freeman-Tukey residuals. The age-period and age-period-cohort models provided a significantly better fit than a model with the factors ;;age' and ;;cohort'. RESULTS: During the study period, mortality rates decreased from 46.8 to 33.0 per 100,000 for men, and from 20.2 to 18.1 per 100,000 for women (average annual decrease of -1.3%, p<0.1 for men, and -1.6%, p<0.03 for women). An age effect was present in both sexes. The definite upward period effect was observed from 1990 to 1994 both among men and women, and was followed by a sharp fall during 2000-4. Cohort and period effects have contained relevant information which partially explained trends in stroke mortality among a 25-64 year-old Lithuanian urban population. CONCLUSIONS: During the period of 1980-2004, the mortality trend declined among women only. The period effect contains relevant information for the explanation of increasing mortality rates during 2000-4 among men and women. The Poisson regression models could be applied for the examination and explanation of the different causes of the population mortality.     

The first case of combined coagulation factor V and coagulation factor VIII deficiency in Poland due to a novel p.Tyr135Asn missense mutation in the MCFD2 gene

Congenital combined coagulation factor V and coagulation factor VIII deficiency (F5F8D) is a rare bleeding disorder due to mutations in the LMAN1 or MCFD2 genes. Here we report the first Polish family with F5F8 deficiency due to a mutation in the MCFD2 gene. The proposita suffered from mild bleeding including epistaxis, menorrhagia, bleeding after dental extraction, and bruising after minor traumas. The F5F8 deficiency was diagnosed due to an excessive postpartum bleeding at the age of 31. Analysis of further family members revealed a second affected individual. Sequencing of the MCFD2 gene and its flanking regions in both patients demonstrated a novel homozygous missense mutation within the second elongation factor hand domain resulting in a substitution of tyrosine by asparagine at amino acid position 135 (p.Tyr135Asn). This variant represents the third missense mutation found in the MCFD2 gene and most likely disrupts the MCFD2-LMAN1 interaction, thus leading to the disease phenotype.     

Subcellular localization of calcium-permeable AMPA receptors in spinal motoneurons

Activation of Ca(2+)-permeable alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptors has been linked to potent effects on survival and dendritic outgrowth of spinal motoneurons. Ca(2+) permeability of AMPA receptors is controlled by the GluR2 subunit. Whole-cell electrophysiological studies have suggested that GluR2-containing and GluR2-lacking AMPA receptors may coexist in individual motoneurons. However, there has not been a direct demonstration of heterogeneity in AMPA receptor subunit composition in single motoneurons, nor of distinct subcellular distributions of GluR2-containing and GluR2-lacking receptors. In the present study, we have used confocal microscopy, immunocytochemistry and Ca(2+) imaging to characterize the subcellular localization of AMPA receptors in cultured rat spinal motoneurons. Immunoreactivity for GluR2 and GluR4 was concentrated in clusters, the vast majority of which were found in dendrites at synapses. Double-labelling for GluR2 and GluR4 revealed variability in relative expression of GluR2 and GluR4 between clusters within individual motoneurons; most AMPA receptor clusters were immunoreactive for both GluR2 and GluR4, but a significant minority of clusters were immunoreactive for GluR2 only or for GluR4 only. The majority of GluR2-immunonegative AMPA receptor clusters was present in dendrites, but the relative proportion of GluR2-immunonegative and GluR2-immunopositive clusters was similar in dendrites and soma. Imaging of [Ca(2+)](i) rises triggered by AMPA receptor activation confirmed Ca(2+) influx in motoneuron dendrites. These findings strongly support a model in which GluR2-containing and GluR2-lacking AMPA receptors coexist in motoneurons, clustered at synapses, and mixed in a relative proportion that varies considerably between cell membrane microdomains.     

Insights into pathological mechanisms of missense mutations in C-terminal domains of von Willebrand factor causing qualitative or quantitative von Willebrand disease

The carboxyl-terminal domains of von Willebrand factor, D4-CK, are cysteine-rich implying that they are structurally important. In this study we characterized the impact of five cysteine missense mutations residing in D4-CK domains on the conformation and biosynthesis of von Willebrand factor. These variants were identified as heterozygous in type 1 (p.Cys2619Tyr and p.Cys2676Phe), type 2A (p.Cys2085Tyr and p.Cys2327Trp) and as compound heterozygous in type 3 (p.Cys2283Arg) von Willebrand disease. Transient expression of human cell lines with wild-type or mutant von Willebrand factor constructs was performed. The mutated and wild-type recombinant von Willebrand factors were quantitatively and qualitatively assessed and compared. Storage of von Willebrand factor in pseudo-Weibel-Palade bodies was studied with confocal microscopy. The structural impact of the mutations was analyzed by homology modeling. Homozygous expressions showed that these mutations caused defects in multimerization, elongation of pseudo-Weibel-Palade bodies and secretion of von Willebrand factor. Co-expressions of wild-type von Willebrand factor and p.Cys2085Tyr, p.Cys2327Trp and p.Cys2283Arg demonstrated defective multimer assembly, suggesting a new pathological mechanism for dominant type 2A von Willebrand disease due to mutations in D4 and B domains. Structural analysis revealed that mutations p.Cys2283Arg, p.Cys2619Tyr and p.Cys2676Phe disrupted intra-domain disulfide bonds, whereas p.Cys2327Trp might affect an inter-domain disulfide bond. The p.Cys2327Trp variant is distinguished from the other mutants by an electrophoretic mobility shift of the multimer bands. The results highlight the importance of cysteine residues within the carboxyl-terminal of von Willebrand factor on structural conformation of the protein and consequently multimerization, storage, and secretion of von Willebrand factor.