Analysis of the binding of p-chlorophenyl-methoxybenzyl-ketoxime (CPMB-oxime) to mitochondrial cytochrome c reductase.

Cytochrome c reductase is inhibited by p-chlorophenyl-methoxybenzyl-ketoxime (CPMB-oxime). CPMB-oxime induces a red-shift of the reduced spectrum of cytochrome b. The inhibitor blocks the oxidation of ubihydroquinone at the QP center of this enzyme in a non-competitive way. The binding stoichiometry equals one inhibitor molecule per Qp center. The apparent Kd in a red-shift assay was 6.9 +/- 0.6 microM. All binding characteristics analysed in this study were very similar to those of the E-beta-methoxyacrylate inhibitors, although the chemical structure is different from these inhibitors. This result is interpreted as a support for the inhibitory mechanism based on the model of a 'catalytic switch' proposed recently for the E-beta-methoxyacrylate inhibitors (MOA-inhibitors (Brandt and von Jagow, Eur. J. Biochem.     

HISTORICAL BIOGEOGRAPHY OF THE HOLARCTIC: AREA RELATIONSHIPS, ANCESTRAL AREAS, AND DISPERSAL OF NON-MARINE ANIMALS

Abstract — Based on published phylogenies for 73 groups of Holarctic non-marine animals, interrelationships between the four Holarctic infraregions (western and eastern Nearctic, western and eastern Palearctic) are examined. The study includes analysis of resolved area cladograms, ancestral areas and dispersal indicated by cladistic subordinateness. Area relationships reflecting present continental configurations (Nearctic vs. Palearctic) dominate the material to the extent that one might speak of a general Holarctic area pattern. Paleocontinental (western Nearctic+eastern Palearctic, western Palearctic+eastern Nearctic) and disjunct patterns are relatively more frequent among groups of higher taxonomic rank. The western Nearctic seems to have played a bigger role than the other infraregions as a center of origin. Two computer programs for constructing resolved area cladograms, viz., COMPONENT 1.5 and COMPONENT 2.0, are compared. The three standard assumptions for biogeographical analysis are compared and arguments are presented in favour of Assumption 0.     

Shedding of Endogenous Interleukin-6 Receptor (IL-6R) Is Governed by A Disintegrin and Metalloproteinase (ADAM) Proteases while a Full-length IL-6R Isoform Localizes to Circulating Microvesicles

Generation of the soluble interleukin-6 receptor (sIL-6R) is a prerequisite for pathogenic IL-6 trans-signaling, which constitutes a distinct signaling pathway of the pleiotropic cytokine interleukin-6 (IL-6). Although in vitro experiments using ectopically overexpressed IL-6R and candidate proteases revealed major roles for the metalloproteinases ADAM10 and ADAM17 in IL-6R shedding, the identity of the protease(s) cleaving IL-6R in more physiological settings, or even in vivo, remains unknown. By taking advantage of specific pharmacological inhibitors and primary cells from ADAM-deficient mice we established that endogenous IL-6R of both human and murine origin is shed by ADAM17 in an induced manner, whereas constitutive release of endogenous IL-6R is largely mediated by ADAM10. Although circulating IL-6R levels are altered in various diseases, the origin of blood-borne IL-6R is still poorly understood. It has been shown previously that ADAM17 hypomorphic mice exhibit unaltered levels of serum sIL-6R. Here, by quantification of serum sIL-6R in protease-deficient mice as well as human patients we also excluded ADAM10, ADAM8, neutrophil elastase, cathepsin G, and proteinase 3 from contributing to circulating sIL-6R. Furthermore, we ruled out alternative splicing of the IL-6R mRNA as a potential source of circulating sIL-6R in the mouse. Instead, we found full-length IL-6R on circulating microvesicles, establishing microvesicle release as a novel mechanism for sIL-6R generation.     

Genetic diversity and restoration of a recalcitrant clonal sedge (Tetraria capillaris Cyperaceae)

The genetic diversity of a clonal sedge (Tertraria capillaris) was assessed using isozyme analysis of 11 loci. Of 29 enzyme systems tested, eight were selected which gave interpretable bands with consistently good resolution. Though seedlings of the species are rarely observed in nature, isozyme analysis showed that for the study transects containing 100 sample plants, the majority of plants at the site were sexually rather than clonally derived. Young plants generated from embryos grown in vitro from excised seeds showed a high level of genetic diversity which could account for the genetic diversity measured in the parent population. In terms of restoration of the species, 85% of the assessed genetic diversity at the study site could be retained if 25 T. capillaris plants were taken at random. The study illustrates how genetic assessment coupled with tissue culture methods provides a feasible model for the recovery of most of the assessed local genetic diversity of a clonal species.     

Inhibition of palmitate‐induced GADD34 expression augments apoptosis in mouse insulinoma cells (MIN6)

Saturated fatty acids like palmitate induce endoplasmic reticulum (ER) stress in pancreatic beta‐cells, an event linked to apoptotic loss of β‐cells in type 2 diabetes. Sustained activation of the ER stress response leads to expression of growth arrest and DNA damage‐inducible protein 34 (GADD34), a regulatory subunit of protein phosphatase 1. In the present study, we have used small interfering RNA in order to knockdown GADD34 expression in insulin‐producing MIN6 cells prior to induction of ER stress by palmitate and evaluated its consequences on RNA‐activated protein kinase‐like ER‐localized eIF2alpha kinase (PERK) signalling and apoptosis. Salubrinal, a specific inhibitor of eukaryotic initiation factor 2α (eIF2α) dephosphorylation, was used as a comparison. Salubrinal treatment augmented palmitate‐induced ER stress and increased GADD34 levels. Both GADD34 knockdown and salubrinal treatment potentiated the cytotoxic effects of palmitate as evidenced by increased DNA fragmentation and activation of caspase 3, with the fundamental difference that the former did not involve enhanced levels of GADD34. The data from this study suggest that sustained activation of PERK signalling and eIF2α phosphorylation sensitizes insulin‐producing MIN6 cells to lipoapoptosis independently of GADD34 expression levels. Copyright © 2014 John Wiley & Sons, Ltd.     

What is the role of cytokines in human colostrum?

No one has ever doubted that maternal milk, in comparison to formula milk, has a far superior nutritional value. Colostrum has a well acknowledge crucial value for the survival of the animal species that cannot receive immunoglobulins through the placenta. Until recently the presence of cytokines in colostrum was unsuspected but it has been now clarified that normally there are at least four cytokines, namely interleukin 1 and 6, tumor necrosis factor and interferon gamma, that may exert an important immunostimulatory role particularly on the oropharyngeal-associated lymphoid tissue. As a corollary, physiological concentration of cytokines administered per os may exert a useful adjuvant activity in aged or immunodeficient people.     

In vivo priming by DNA injection occurs predominantly by antigen transfer.

DNA vaccines can stimulate both humoral and cytolytic immune responses. Although bone marrow-derived elements present the expressed Ag, the mechanisms for acquiring immunogenic peptides have yet to be fully elucidated. APCs may become directly transfected by plasmid DNA or process extracellular proteins produced by other transfected cells. Using a transactivating plasmid system and bone marrow chimeras, we show that both mechanisms appear to be involved; however, the bulk of the immune response is dependent on expression of Ag by nonlymphoid tissues and transfer to APCs. These in vivo studies are the first to define the role of transfected nonlymphoid cells in generating Ag for presentation by bone marrow-derived APCs after needle injection with plasmid DNA.