Expression profiles of human interferon-alpha and interferon-lambda subtypes are ligand- and cell-dependent

Recent genome-wide association studies suggest distinct roles for 12 human interferon-alpha (IFN-α) and 3 IFN-λ subtypes that may be elucidated by defining the expression patterns of these sets of genes. To overcome the impediment of high homology among each of the sets, we designed a quantitative real-time PCR assay that incorporates the use of molecular beacon and locked nucleic acid (LNA) probes, and in some instances, LNA oligonucleotide inhibitors. We then measured IFN subtype expression by human peripheral blood mononuclear cells and by purified monocytes, myeloid dendritic cells (mDC), plasmacytoid dendritic cells (pDC), and monocyte-derived macrophages (MDM), and -dendritic cells (MDDC) in response to poly I:C, lipopolysaccharide (LPS), imiquimod and CpG oligonucleotides. We found that in response to poly I:C and LPS, monocytes, MDM and MDDC express a subtype pattern restricted primarily to IFN-β and IFN-λ1. In addition, while CpG elicited expression of all type I IFN subtypes by pDC, imiquimod did not. Furthermore, MDM and mDC highly express IFN-λ, and the subtypes of IFN-λ are expressed hierarchically in the order IFN-λ1 followed by IFN-λ2, and then IFN-λ3. These data support a model of coordinated cell- and ligand-specific expression of types I and III IFN. Defining IFN subtype expression profiles in a variety of contexts may elucidate specific roles for IFN subtypes as protective, therapeutic or pathogenic mediators.     

Similarities in Restriction Fragment Patterns of Mitochondrial DNAs of PhytophthoraSpecies Strongly Depend on the Restriction Enzyme Used Due to Heterogeneous Base Distribution and Sequence Conservation

Mitochondrial DNAs of six morphologically different Phytophthora species were digested with 15 restriction enzymes. The numbers of restriction fragments obtained differed considerably from those theoretically expected for random base distribution. Enzymes with relatively many G and C in their recognition sequences produced significantly larger numbers of fragments. Moreover, fragments generated by most of these enzymes were more often shared by two or more species than those from enzymes with more A and T in their recognition sequence. It is concluded that base distribution in mitochondrial DNA of Phytophthora is heterogeneous,AT-rich stretches occurring scattered over the mitochondrial genome and GC-rich regions present in conserved sequences, presumably genes. A practical consequence for taxonomic RFLP studies is that optimal enzymes can be selected, depending on the desired level of resolution.     

Tetrapod phylogeny inferred from 18S and 28S ribosomal RNA sequences and a review of the evidence for amniote relationships [published erratum appears in Mol Biol Evol 1991 May;8(3):398]

The 18S ribosomal RNAs of 21 tetrapods were sequenced and aligned with five published tetrapod sequences. When the coelacanth was used as an outgroup, Lissamphibia (living amphibians) and Amniota (amniotes) were found to be statistically significant monophyletic groups. Although little resolution was obtained among the lissamphibian taxa, the amniote sequences support a sister-group relationship between birds and mammals. Portions of the 28S ribosomal RNA (rRNA) molecule in 11 tetrapods also were sequenced, although the phylogenetic results were inconclusive. In contrast to previous studies, deletion or down- weighting of base-paired sites were found to have little effect on phylogenetic relationships. Molecular evidence for amniote relationships is reviewed, showing that three genes (beta-hemoglobin, myoglobin, and 18S rRNA) unambiguously support a bird-mammal relationship, compared with one gene (histone H2B) that favors a bird- crocodilian clade. Separate analyses of four other genes (alpha- crystallin A, alpha-hemoglobin, insulin, and 28S rRNA) and a combined analysis of all sequence data are inconclusive, in that different groups are defined in different analyses and none are strongly supported. It is suggested that until sequences become available from a broader array of taxa, the molecular evidence is best evaluated at the level of individual genes, with emphasis placed on those studies with the greatest number of taxa and sites. When this is done, a bird-mammal relationship is most strongly supported. When regarded in combination with the morphological evidence for this association, it must be considered at least as plausible as a bird-crocodilian relationship.      

Mycobacterium tuberculosis triggers host type I IFN signaling to regulate IL-1β production in human macrophages

Mycobacterium tuberculosis is a virulent intracellular pathogen that survives in macrophages even in the presence of an intact adaptive immune response. Type I IFNs have been shown to exacerbate tuberculosis in mice and to be associated with disease progression in infected humans. Nevertheless, the mechanisms by which type I IFNs regulate the host response to M. tuberculosis infection are poorly understood. In this study, we show that M. tuberculosis induces an IFN-related gene expression signature in infected primary human macrophages, which is dependent on host type I IFN signaling as well as the mycobacterial virulence factor, region of difference-1. We further demonstrate that type I IFNs selectively limit the production of IL-1β, a critical mediator of immunity to M. tuberculosis. This regulation occurs at the level of IL1B mRNA expression, rather than caspase-1 activation or autocrine IL-1 amplification and appears to be preferentially used by virulent mycobacteria since avirulent M. bovis bacillus Calmette-Guérin (BCG) fails to trigger significant expression of type I IFNs or release of mature IL-1β protein. The latter property is associated with decreased caspase-1-dependent IL-1β maturation in the BCG-infected macrophages. Interestingly, human monocytes in contrast to macrophages produce comparable levels of IL-1β in response to either M. tuberculosis or BCG. Taken together, these findings demonstrate that virulent and avirulent mycobacteria employ distinct pathways for regulating IL-1β production in human macrophages and reveal that in the case of M. tuberculosis infection the induction of type I IFNs is a major mechanism used for this purpose.     

PHYLOGENETIC ANALYSIS OF LEARNING-RELATED NEUROMODULATION IN MOLLUSCAN MECHANOSENSORY NEURONS

In spite of significant advances in our understanding of mechanisms of learning and memory in a variety of organisms, little is known about how such mechanisms evolve. Even mechanisms of simple forms of learning, such as habituation and sensitization, have not been studied phylogenetically. Here we begin an evolutionary analysis of learning-related neuromodulation in species related to the well-studied opisthobranch gastropod, Aplysia californica. In Aplysia, increased spike duration and excitability in mechanosensory neurons contribute to several forms of learning-related changes to defensive withdrawal reflexes. The modulatory transmitter serotonin (5-hydroxytryptamine, or 5-HT), is thought to play a critical role in producing these firing property changes. In the present study, we tested mechanosensory homologs of the tail-withdrawal reflex in species related to Aplysia for 5-HT-mediated increases in spike duration and excitability. Criteria used to identify homologous tail-sensory neurons included position, relative size, resting electrical properties, expression of a sensory neuron-specific protein, neuroanatomy, and receptive field. The four ingroup species studied (Aplysia californica, Dolabella auricularia, Bursatella leachii, and Dolabrifera dolabrifera) belong to two clades (two species each) within the family Aplysiidae. In the first clade (Aplysia/Dolabella), we found that the tail-sensory neurons of A. californica and tail-sensory homologs of a closely related species, D. auricularia, responded to bath-applied serotonin in essentially similar fashion: significant increases in spike duration as well as excitability. In the other clade (Dolabrifera/Bursatella), more distantly related to Aplysia, one species (B. leachii) showed spike broadening and increased excitability. However, the other species (D. dolabrifera) showed neither spike broadening nor increased excitability. The firing properties of tail-sensory homologs of D. dolabrifera were insensitive to 5-HT over a wide range of concentrations. We also performed experiments on two outgroup species (Akera bullata and Bulla gouldiana) and found that spike duration was unaffected by 5-HT, whereas excitability was increased. This study suggests that 5-HT-induced spike broadening arose more recently in opisthobranch evolution, whereas 5-HT-induced excitability increase is a more ancestral trait that may have been expressed in the earliest opisthobranchs. Both traits are absent in the aplysiid species D. dolabrifera, demonstrating that a lineage can lose learning-related mechanisms. The phylogenetic variation observed in the present study presents the opportunity to test general models about learning mechanisms and their evolution in unique ways.     

Evaluation of the <Emphasis Type="Italic">OPTC</Emphasis> gene in primary open angle glaucoma: functional significance of a silent change

Background  

We investigated the molecular basis of primary open-angle glaucoma (POAG) using Opticin (OPTC) as a candidate gene on the basis of its expression in the trabecular meshwork cells involved in the disease pathogenesis. Two hundred POAG patients and 100 controls were enrolled in this study. The coding sequence of OPTC was amplified by PCR from genomic DNA of POAG patients, followed by SSCP, DHPLC and DNA sequencing. Subsequent bioinformatic analysis, site-directed mutagenesis, quantitative RT-PCR and western blot experiments were performed to address the functional significance of a 'silent' change in the OPTC coding region while screening for mutations in POAG patients.     

Untersuchungen zur Tiergesundheit,Leistung und zum Rückstandsverhalten beim Schwein bei gleichzeitiger Aufnahme der Mykotoxine Ochratoxin A,Deoxynivalenol, Zearalenon über das Futter im 90-Tage-Test

The effect of the administration of the mycotoxins OTA, ZEA and DON alone resp. in combination on animal health and the residue behavior of pigs from 50 – 60 kg living weight over 90 days was investigated in 4 separate studies. Due to its fast metabolisation the administration of 1000 µg DON resp. 250 µg ZEA per kg feed alone or in combination with other mycotoxins does not lead to detectable residues of these mycotoxins in organs and tissues. Therefore these mycotoxins should not be relevant to the consumer.There is an effect of the simultaneous administration of ZEA resp. DON on the metabolisation resp. secretion of OTA. OTA is of relevance from the point of view of residue toxicology.