Antagonism of microRNA-122 in mice by systemically administered LNA-antimiR leads to up-regulation of a large set of predicted target mRNAs in the liver

MicroRNA-122 (miR-122) is an abundant liver-specific miRNA, implicated in fatty acid and cholesterol metabolism as well as hepatitis C viral replication. Here, we report that a systemically administered 16-nt, unconjugated LNA (locked nucleic acid)-antimiR oligonucleotide complementary to the 5′ end of miR-122 leads to specific, dose-dependent silencing of miR-122 and shows no hepatotoxicity in mice. Antagonism of miR-122 is due to formation of stable heteroduplexes between the LNA-antimiR and miR-122 as detected by northern analysis. Fluorescence in situ hybridization demonstrated uptake of the LNA-antimiR in mouse liver cells, which was accompanied by markedly reduced hybridization signals for mature miR-122 in treated mice. Functional antagonism of miR-122 was inferred from a low cholesterol phenotype and de-repression within 24 h of 199 liver mRNAs showing significant enrichment for miR-122 seed matches in their 3′ UTRs. Expression profiling extended to 3 weeks after the last LNA-antimiR dose revealed that most of the changes in liver gene expression were normalized to saline control levels coinciding with normalized miR-122 and plasma cholesterol levels. Combined, these data suggest that miRNA antagonists comprised of LNA are valuable tools for identifying miRNA targets in vivo and for studying the biological role of miRNAs and miRNA-associated gene-regulatory networks in a physiological context.     

A normal rabbit serum containing Golgi-specific autoatibodies identifies a novel 74-kDa trans-Golgi resident protein

A normal rabbit serum has been identified which contains Golgi-specific autoantibodies. In indirect immunofluorescence experiments the serum was found to stain the juxtanuclear Golgi complex in a variety of cell lines, including human skin fibroblasts, rat osteoblasts, rat myoblasts (L6), baby hamster kidney epithelial cells, and human embryonic kidney cells (293). Thus, the antigen(s) recognized by this serum seems to be well conserved and universally expressed in various mammalian cell types. Immunoelectron microscopy revealed that the epitope resides in the luminal side of the Golgi membranes, and that the antigen is concentrated in the trans-face of the Golgi stacks. In agreement with these results, brefeldin A treatment did not release the antigen from the membranes, but caused its redistribution partly into the endoplasmic reticulum but also into the juxtanuclear area, similarly as with other proteins known to be present in the trans-Golgi cisternae or trans-Golgi network. Our immunoprecipitation studies in human skin fibroblasts demonstrated that the serum recognizes specifically only a single protein with a molecular size of 74 kDa. This protein also cosedimented with a known trans-Golgi-specific marker protein, galactosyltransferase, after fractionation of subcellular organelles by Nycodenz gradient centrifugation. The widespread and polarized expression of this 74-kDa trans-Golgi resident protein suggests that it is required for the late Golgi functions in different mammalian cell types.     

Dominance relations in captive groups of adult and juvenile arctic blue foxes (Alopex lagopus)

Dominance relationships were studied in captive Arctic blue fox (Alopex lagopus) groups comprising adults (four males, five females) and juveniles (four males, five females). The results showed that Arctic blue foxes easily formed a social organization with an observable hierarchy, in which adults typically dominated over juveniles. Within the same age group, males usually dominated over females. Dominance correlated most significantly with body weight in autumn, but later that correlation decreased. Urine marking activity was very low during autumn and early winter, but increased significantly prior to and during the breeding season when aggressive encounters were also most pronounced. In addition to several adults, the social status of some juveniles was high at breeding time. Altogether 7 out of 11 females (63.6%) whelped, but the survival rate of litters was low and kits of only two adult females survived (18.2%). It can be concluded that hierarchical development and reproduction in Arctic blue fox groups are markedly influenced by dominance relationships.     

Adipocytes as a Link Between Gut Microbiota-Derived Flagellin and Hepatocyte Fat Accumulation

While the role of both elevated levels of circulating bacterial cell wall components and adipose tissue in hepatic fat accumulation has been recognized, it has not been considered that the bacterial components-recognizing adipose tissue receptors contribute to the hepatic fat content. In this study we found that the expression of adipose tissue bacterial flagellin (FLG)-recognizing Toll-like receptor (TLR) 5 associated with liver fat content (r = 0.699, p = 0.003) and insulin sensitivity (r = -0.529, p = 0.016) in humans (n = 23). No such associations were found for lipopolysaccharides (LPS)-recognizing TLR4. To study the underlying molecular mechanisms of these associations, human HepG2 hepatoma cells were exposed in vitro to the conditioned culture media derived from FLG or LPS-challenged human adipocytes. The adipocyte-mediated effects were also compared to the effects of direct HepG2 exposure to FLG and LPS. We found that the media derived from FLG-treated adipocytes stimulated fat accumulation in HepG2 cells, whereas either media derived from LPS-treated adipocytes or direct FLG or LPS exposure did not. This is likely due to that FLG-treatment of adipocytes increased lipolysis and secretion of glycerol, which is known to serve a substrate for triglyceride synthesis in hepatocytes. Similarly, only FLG-media significantly decreased insulin signaling-related Akt phosphorylation, IRS1 expression and mitochondrial respiratory chain ATP5A. In conclusion, our results suggest that the FLG-induced TLR5 activation in adipocytes increases glycerol secretion from adipocytes and decreases insulin signaling and mitochondrial functions, and increases fat accumulation in hepatocytes. These mechanisms could, at least partly, explain the adipose tissue TLR5 expression associated with liver fat content in humans.     

Dental findings in osteogenesis imperfecta: II. Dysplastic and other developmental defects

Orthopantomograms of 49 patients with osteogenesis imperfecta (OI) classified according to Sillence were examined for dysplastic dentin defects and other developmental abnormalities of the teeth. Thistle-tube-shaped pulps in the permanent teeth were observed in five patients. Four of them had pulp stones in several teeth. Apically extended pulp chambers were present in 3/49 patients, and the structure of the mandible was cystic in 1/49. These defects occurred in patients with type I or unclassifiable OI and were mainly not associated with type I dentinogenesis imperfecta (DI). The prevalence rates of invaginations (10.2%) and hypodontia of permanent teeth (18.4%) exceeded those in the normal population. The frequent developmental disturbances of the teeth in OI may be secondary to the connective tissue defect. The relation of the dysplastic defects other than type I DI to OI remains to be clarified.     

A Concept of Bayesian Regulation in Fisheries Management

Stochastic variability of biological processes and uncertainty of stock properties compel fisheries managers to look for tools to improve control over the stock. Inspired by animals exploiting hidden prey, we have taken a biomimetic approach combining catch and effort in a concept of Bayesian regulation (BR). The BR provides a real-time Bayesian stock estimate, and can operate without separate stock assessment. We compared the performance of BR with catch-only regulation (CR), alternatively operating with N-target (the stock size giving maximum sustainable yield, MSY) and F-target (the fishing mortality giving MSY) on a stock model of Baltic Sea herring. N-targeted BR gave 3% higher yields than F-targeted BR and CR, and 7% higher yields than N-targeted CR. The BRs reduced coefficient of variance (CV) in fishing mortality compared to CR by 99.6% (from 25.2 to 0.1) when operated with F-target, and by about 80% (from 158.4 to 68.4/70.1 depending on how the prior is set) in stock size when operated with N-target. Even though F-targeted fishery reduced CV in pre-harvest stock size by 19–22%, it increased the dominant period length of population fluctuations from 20 to 60–80 years. In contrast, N-targeted BR made the periodic variation more similar to white noise. We discuss the conditions when BRs can be suitable tools to achieve sustainable yields while minimizing undesirable fluctuations in stock size or fishing effort.     

Acquisition of complement factor H is important for pathogenesis of Streptococcus pyogenes infections: evidence from bacterial in vitro survival and human genetic association

Streptococcus pyogenes (or group A streptococcus [GAS]) is a major human pathogen causing infections, such as tonsillitis, erysipelas, and sepsis. Several GAS strains bind host complement regulator factor H (CFH) via its domain 7 and, thereby, evade complement attack and C3b-mediated opsonophagocytosis. Importance of CFH binding for survival of GAS has been poorly studied because removal of CFH from plasma or blood causes vigorous complement activation, and specific inhibitors of the interaction have not been available. In this study, we found that activation of human complement by different GAS strains (n = 38) correlated negatively with binding of CFH via its domains 5-7. The importance of acquisition of host CFH for survival of GAS in vitro was studied next by blocking the binding with recombinant CFH5-7 lacking the regulatory domains 1-4. Using this fragment in full human blood resulted in death or radically reduced multiplication of all of the studied CFH-binding GAS strains. To study the importance of CFH binding in vivo (i.e., for pathogenesis of streptococcal infections), we used our recent finding that GAS binding to CFH is diminished in vitro by polymorphism 402H, which is also associated with age-related macular degeneration. We showed that allele 402H is suggested to be associated with protection from erysipelas (n = 278) and streptococcal tonsillitis (n = 209) compared with controls (n = 455) (p < 0.05). Taken together, the bacterial in vitro survival data and human genetic association revealed that binding of CFH is important for pathogenesis of GAS infections and suggested that inhibition of CFH binding can be a novel therapeutic approach in GAS infections.     

Structure of complement factor H carboxyl-terminus reveals molecular basis of atypical haemolytic uremic syndrome

Factor H (FH) is the key regulator of the alternative pathway of complement. The carboxyl-terminal domains 19-20 of FH interact with the major opsonin C3b, glycosaminoglycans, and endothelial cells. Mutations within this area are associated with atypical haemolytic uremic syndrome (aHUS), a disease characterized by damage to endothelial cells, erythrocytes, and kidney glomeruli. The structure of recombinant FH19-20, solved at 1.8 A by X-ray crystallography, reveals that the short consensus repeat domain 20 contains, unusually, a short alpha-helix, and a patch of basic residues at its base. Most aHUS-associated mutations either destabilize the structure or cluster in a unique region on the surface of FH20. This region is close to, but distinct from, the primary heparin-binding patch of basic residues. By mutating five residues in this region, we show that it is involved, not in heparin, but in C3b binding. Therefore, the majority of the aHUS-associated mutations on the surface of FH19-20 interfere with the interaction between FH and C3b. This obviously leads to impaired control of complement attack on plasma-exposed cell surfaces in aHUS.