In vivo metabolism of a mutant apolipoprotein, apoA-IIowa, associated with hypoalphalipoproteinemia and hereditary systemic amyloidosis.

Apolipoprotein (apo) A-I is the major protein constituent of plasma high density lipoproteins (HDL). A kindred has been identified in which a glycine to arginine mutation at residue 26 in apoA-I is associated with hypoalphalipoproteinemia and hereditary systemic amyloidosis. We isolated the mutant protein, termed apoA-IIowa, from the plasma of an affected subject and studied its in vivo metabolism compared to that of normal apoA-I in two heterozygous apoA-IIowa subjects and two normal controls. Normal and mutant apoA-I were radioiodinated with 131I and 125I, respectively, reassociated with autologous plasma lipoproteins, and simultaneously injected into all subjects. Kinetic analysis of the plasma radioactivity curves demonstrated that the mutant apoA-IIowa was rapidly cleared from plasma (mean fractional catabolic rate [FCR] 0.559 day-1) compared with normal apoA-I (mean FCR 0.244 day-1) in all four subjects. The FCR of normal apoA-I was also substantially faster in the heterozygous apoA-IIowa subjects (mean FCR 0.281 days-1) than in the normal controls (mean FCR 0.203 days-1). Despite the rapid removal from plasma of apoA-IIowa, the cumulative urinary excretion of its associated radioactivity after 2 weeks (44%) of the injected dose) was substantially less than that associated with normal apoA-I (78% of injected dose), indicating extravascular sequestration of radiolabeled apoA-IIowa.(ABSTRACT TRUNCATED AT 250 WORDS)     

Long-term cathode performance and the microbial communities that develop in microbial fuel cells fed different fermentation endproducts

To better understand how cathode performance and substrates affected communities that evolved in these reactors over long periods of time, microbial fuel cells were operated for more than 1 year with individual endproducts of lignocellulose fermentation (acetic acid, formic acid, lactic acid, succinic acid, or ethanol). Large variations in reactor performance were primarily due to the specific substrates, with power densities ranging from 835 ± 21 to 62 ± 1 mW/m³. Cathodes performance degraded over time, as shown by an increase in power of up to 26% when the cathode biofilm was removed, and 118% using new cathodes. Communities that developed on the anodes included exoelectrogenic families, such as Rhodobacteraceae, Geobacteraceae, and Peptococcaceae, with the Deltaproteobacteria dominating most reactors. Pelobacter propionicus was the predominant member in reactors fed acetic acid, and it was abundant in several other MFCs. These results provide valuable insights into the effects of long-term MFC operation on reactor performance.     

New targets and emerging therapies for reducing LDL cholesterol

PURPOSE OF REVIEW: Effective therapies for lowering LDL-cholesterol reduce the incidence of cardiovascular disease and provide associated decreases in morbidity and mortality. Progress in our understanding of metabolism and innovations in drug design have jointly identified promising new drug targets and alternative approaches to old targets. This review focuses on the mechanism, safety and efficacy of emerging LDL-cholesterol lowering therapies. RECENT FINDINGS: Decreasing apolipoprotein B expression or preventing the formation of a stable lipoprotein structure by inhibiting microsomal triglyceride transfer protein attenuates the secretion of atherogenic lipoproteins containing apolipoprotein B into the plasma. Increases in LDL receptor-mediated cholesterol clearance occur when hepatic cholesterol stores are reduced secondary to inhibition of squalene synthase or LDL receptor degradation is disrupted by reduced activity of proprotein convertase subtilisin kexin type 9. Each of these developing therapies demonstrably reduces LDL-cholesterol levels. SUMMARY: The emergence of modalities that act in series and in parallel with available agents may allow more effective LDL-cholesterol lowering in those patients intolerant of current therapy, and may permit decremental reductions in LDL-cholesterol for those unable to achieve aggressive LDL-cholesterol goals using existing agents.     

Implications for the domain arrangement of axonin-1 derived from the mapping of its NgCAM binding site.

The neuronal cell adhesion molecule axonin-1 is composed of six immunoglobulin and four fibronectin type III domains. Axonin-1 promotes neurite outgrowth, when presented as a substratum for neurons in vitro, via a neuronal receptor that has been identified as the neuron-glia cell adhesion molecule, NgCAM, based on the blocking effect of polyclonal antibodies directed to NgCAM. Here we report the identification of axonin-1 domains involved in NgCAM binding. NgCAM-conjugated microspheres were tested for binding to COS cells expressing domain deletion mutants of axonin-1. In addition, monoclonal antibodies directed to axonin-1 were assessed for their ability to block the axonin-1-NgCAM interaction, and their epitopes were mapped using the domain deletion mutants. The results suggest that the four amino-terminal immunoglobulin domains of axonin-1 form a domain conglomerate which is necessary and sufficient for NgCAM binding. Surprisingly, NgCAM binding to membrane-bound axonin-1 was increased strongly by deletion of the fifth or sixth immunoglobulin domains of axonin-1. Based on these results and on negative staining electron microscopy, we propose a horseshoe-shaped domain arrangement of axonin-1 that obscures the NgCAM binding site. Neurite outgrowth studies with truncated forms of axonin-1 show that axonin-1 is a neurite outgrowth-promoting substratum in the absence of the NgCAM binding site.     

The effect of controlled growth factor delivery on embryonic stem cell differentiation inside fibrin scaffolds

The goal of this project was to develop 3-D biomaterial scaffolds that present cues to direct the differentiation of embryonic stem (ES) cell-derived neural progenitor cells, seeded inside the scaffolds, into mature neural phenotypes, specifically neurons and oligodendrocytes. Release studies were performed to determine the appropriate conditions for retention of neurotrophin-3 (NT-3), sonic hedgehog, and platelet-derived growth factor (PDGF) by an affinity-based delivery system incorporated into fibrin scaffolds. Embryoid bodies containing neural progenitors were formed from mouse ES cells, using a 4−/4+ retinoic acid treatment protocol, and then seeded inside fibrin scaffolds containing the drug delivery system. This delivery system was used to deliver various growth factor doses and combinations to the cells seeded inside the scaffolds. Controlled delivery of NT-3 and PDGF simultaneously increased the fraction of neural progenitors, neurons, and oligodendrocytes while decreasing the fraction of astrocytes obtained compared to control cultures seeded inside unmodified fibrin scaffolds with no growth factors present in the medium. These results demonstrate that such a strategy can be used to generate an engineered tissue for the potential treatment of spinal cord injury and could be extended to the study of differentiation in other tissues.     

Neurite Fasciculation Mediated by Complexes of Axonin-1 and Ng Cell Adhesion Molecule

Neural cell adhesion molecules composed of immunoglobulin and fibronectin type III-like domains have been implicated in cell adhesion, neurite outgrowth, and fasciculation. Axonin-1 and Ng cell adhesion molecule (NgCAM), two molecules with predominantly axonal expression exhibit homophilic interactions across the extracellular space (axonin- 1/axonin-1 and NgCAM/NgCAM) and a heterophilic interaction (axonin-1–NgCAM) that occurs exclusively in the plane of the same membrane (cis-interaction). Using domain deletion mutants we localized the NgCAM homophilic binding in the Ig domains 1-4 whereas heterophilic binding to axonin-1 was localized in the Ig domains 2-4 and the third FnIII domain. The NgCAM–NgCAM interaction could be established simultaneously with the axonin-1–NgCAM interaction. In contrast, the axonin-1–NgCAM interaction excluded axonin-1/axonin-1 binding. These results and the examination of the coclustering of axonin-1 and NgCAM at cell contacts, suggest that intercellular contact is mediated by a symmetric axonin-1₂/NgCAM₂ tetramer, in which homophilic NgCAM binding across the extracellular space occurs simultaneously with a cis-heterophilic interaction of axonin-1 and NgCAM. The enhanced neurite fasciculation after overexpression of NgCAM by adenoviral vectors indicates that NgCAM is the limiting component for the formation of the axonin-1₂/NgCAM₂ complexes and, thus, neurite fasciculation in DRG neurons.