Addition of a sequence from α2-antiplasmin transforms human serum albumin into a blood clot component that speeds clot lysis

<p>Abstract</p> <p>Background</p> <p>The plasma protein α₂-antiplasmin (α₂AP) is cross-linked to fibrin in blood clots by the transglutaminase factor XIIIa, and in that location retards clot lysis. Competition for this effect could be clinically useful in patients with thrombosis. We hypothesized that fusion of N-terminal portions of α₂-antiplasmin to human serum albumin (HSA) and production of the chimeric proteins in <it>Pichia pastoris </it>yeast would produce a stable and effective competitor protein.</p> <p>Results</p> <p>Fusion protein α₂AP(13-42)-HSA was efficiently secreted from transformed yeast and purified preparations contained within a mixed population the full-length intact form, while fusions with longer α₂AP moieties were inefficiently secreted and/or degraded. The α₂AP(13-42)-HSA protein, but not recombinant HSA, was cross-linked to both chemical lysine donors and fibrin or fibrinogen by factor XIIIa, although with less rapid kinetics than native α₂AP. Excess α₂AP(13-42)-HSA competed with α₂AP for cross-linking to chemical lysine donors more effectively than a synthetic α₂AP(13-42) peptide, and reduced the α₂AP-dependent resistance to fibrinolysis of plasma clots equally effectively as the peptide. Native α₂AP was found in <it>in vivo </it>clots in rabbits to a greater extent than α₂AP(13-42), however.</p> <p>Conclusion</p> <p>In this first report of transfer of transglutamination substrate status from one plasma protein to another, fusion protein α₂AP(13-42)-HSA was shown to satisfy initial requirements for a long-lasting, well-tolerated competitive inhibitor of α₂-antiplasmin predicted to act in a clot-localized manner.</p

Discovery of Multiply Imaged Galaxies behind the Cluster and Lensed Quasar SDSS J1004+4112

We have identified three multiply imaged galaxies in Hubble Space Telescope images of the redshift z=0.68 cluster responsible for the large-separation quadruply lensed quasar, SDSS J1004+4112. Spectroscopic redshifts have been secured for two of these systems using the Keck I 10m telescope. The most distant lensed galaxy, at z=3.332, forms at least four images, and an Einstein ring encompassing 3.1 times more area than the Einstein ring of the lensed QSO images at z=1.74, due to the greater source distance. For a second multiply imaged galaxy, we identify Ly_alpha emission at a redshift of z=2.74. The cluster mass profile can be constrained from near the center of the brightest cluster galaxy, where we observe both a radial arc and the fifth image of the lensed quasar, to the Einstein radius of the highest redshift galaxy, ~110 kpc. Our preliminary modeling indicates that the mass approximates an elliptical body, with an average projected logarithmic gradient of ~-0.5. The system is potentially useful for a direct measurement of world models in a previously untested redshift range.Comment: 5 pages, 3 figures. Accepted by ApJL. High resolution version of the paper can be found at: http://wise-obs.tau.ac.il/~kerens/papers.htm

Qualitative Assessment of Participant Utilization and Satisfaction With the Seattle Senior Farmers’ Market Nutrition Pilot Program

INTRODUCTION: The Seattle Senior Farmers' Market Nutrition Pilot Program delivered fresh fruits and vegetables to homebound seniors in King County, Washington, from June through October 2001. A primary objective of the program was to increase participants' intake of fruits and vegetables. A qualitative study was conducted to examine the impact of the program on participating homebound seniors. METHODS: Semi-structured interviews were performed with 27 participants in their homes to identify benefits and barriers they encountered and to measure their use and sense of satisfaction with the program. RESULTS: Analysis of the transcribed interviews revealed several common themes:  Participants appreciated the variety and quality of the fresh fruits and vegetables. . Some participants would not have had access to fresh fruits and vegetables without the program. . Home-delivered baskets of fresh fruits and vegetables brought participants joy, stimulated interest in healthy foods, and improved quality of life. . The program newsletter supported consumption of fresh produce. CONCLUSION: Program success was rooted in the multiple ways the program addressed potential barriers and reinforced behavioral intent

Complete analysis of the B-cell response to a protein antigen, from in vivo germinal centre formation to 3-D modelling of affinity maturation

Somatic hypermutation of immunoglobulin variable region genes occurs within germinal centres (GCs) and is the process responsible for affinity maturation of antibodies during an immune response. Previous studies have focused almost exclusively on the immune response to haptens, which may be unrepresentative of epitopes on protein antigens. In this study, we have exploited a model system that uses transgenic B and CD4&lt;sup&gt;+&lt;/sup&gt; T cells specific for hen egg lysozyme (HEL) and a chicken ovalbumin peptide, respectively, to investigate a tightly synchronized immune response to protein antigens of widely differing affinities, thus allowing us to track many facets of the development of an antibody response at the antigen-specific B cell level in an integrated system &lt;i&gt;in&lt;/i&gt; &lt;i&gt;vivo&lt;/i&gt;. Somatic hypermutation of immunoglobulin variable genes was analysed in clones of transgenic B cells proliferating in individual GCs in response to HEL or the cross-reactive low-affinity antigen, duck egg lysozyme (DEL). Molecular modelling of the antibody–antigen interface demonstrates that recurring mutations in the antigen-binding site, selected in GCs, enhance interactions of the antibody with DEL. The effects of these mutations on affinity maturation are demonstrated by a shift of transgenic serum antibodies towards higher affinity for DEL in DEL-cOVA immunized mice. The results show that B cells with high affinity antigen receptors can revise their specificity by somatic hypermutation and antigen selection in response to a low-affinity, cross-reactive antigen. These observations shed further light on the nature of the immune response to pathogens and autoimmunity and demonstrate the utility of this novel model for studies of the mechanisms of somatic hypermutation

First Observation of 15Be

The neutron-unbound nucleus 15Be was observed for the first time. It was populated using neutron transfer from a deuterated polyethylene target with a 59 MeV/u 14Be beam. Neutrons were measured in coincidence with outgoing 14Be particles and the reconstructed decay energy spectrum exhibits a resonance at 1.8(1) MeV. This corresponds to 15Be being unbound by 0.45 MeV more then 16Be thus significantly hindering the sequential two-neutron decay of 16Be to 14Be through this state

In Vivo Clearance of Alpha-1 Acid Glycoprotein Is Influenced by the Extent of Its N-Linked Glycosylation and by Its Interaction with the Vessel Wall

Alpha-1 acid glycoprotein (AGP) is a highly glycosylated plasma protein that exerts vasoprotective effects. We hypothesized that AGP's N-linked glycans govern its rate of clearance from the circulation, and followed the disappearance of different forms of radiolabeled human AGP from the plasma of rabbits and mice. Enzymatic deglycosylation of human plasma-derived AGP (pdAGP) by Peptide: N-Glycosidase F yielded a mixture of differentially deglycosylated forms (PNGase-AGP), while the introduction of five Asn to Gln mutations in recombinant Pichia pastoris-derived AGP (rAGP-N(5)Q) eliminated N-linked glycosylation. PNGase-AGP was cleared from the rabbit circulation 9-fold, and rAGP-N(5)Q, 46-fold more rapidly than pdAGP, primarily via a renal route. Pichia pastoris-derived wild-type rAGP differed from pdAGP in expressing mannose-terminated glycans, and, like neuraminidase-treated pdAGP, was more rapidly removed from the rabbit circulation than rAGP-N(5)Q. Systemic hyaluronidase treatment of mice transiently decreased pdAGP clearance. AGP administration to mice reduced vascular binding of hyaluronic acid binding protein in the liver microcirculation and increased its plasma levels. Our results support a critical role of N-linked glycosylation of AGP in regulating its in vivo clearance and an influence of a hyaluronidase-sensitive component of the vessel wall on its transendothelial passage