Binding and orientation of conantokins in PL vesicles and aligned PL multilayers.

The association of a ligand with its cognate cell surface receptor can be facilitated by interactions between the ligand and the lipid phase of the cell membrane. With respect to the N-methyl-D-aspartate receptor (NMDAR), we have previously established a low affinity, nonreceptor-mediated interaction of the peptidic conantokins with synaptic membranes in conjunction with a high affinity binding to the NMDARs present therein [Klein, R. C.

Transcriptional profiling of the transition from normal intestinal epithelia to adenomas and carcinomas in the APCMin/+ mouse.

Mutations in the adenomatous polyposis coli (APC) gene that result in excessive beta-catenin-induced cell signaling are implicated in the risk of colon cancer. Although the mechanism of APC-mediated tumorigenesis is known, the pathways that translate beta-catenin signaling into tumor growth in vivo are undefined. To address this, gene expression profiles of normal intestinal epithelial cells were compared with those from adenomas and carcinomas from APC(Min/+) mice, a model of APC-related colorectal cancer.

The fibrinolytic system in dissemination and matrix protein deposition during a mycobacterium infection.

The fibrinolytic system is known to play an important role in the inflammatory response to bacterial infections. In the present study, relationships between protein components of the fibrinolytic system and infectivity by Mycobacterium avium were analyzed. Infections were initiated through noninvasive intratracheal administration of M.

In utero transplantation of wild-type fetal liver cells rescues factor X-deficient mice from fatal neonatal bleeding diatheses.

Factor X (FX)-deficient embryos suffer partial embryonic lethality with approximately 30% of the embryos arresting at midgestation. The remaining animals survive to term but die perinatally mainly from abdominal or intracranial hemorrhage. We have rescued FX-deficient mice by transplanting fetal liver cells from FX+/+, Rosa26 fetuses into midgestation embryos derived from FX+/- heterozygous crosses.

Endogenous plasmin converts Glu-plasminogen to Lys-plasminogen on the monocytoid cell surface.

Recently, we showed that localization of Glu-plasminogen on cell surfaces enhances its conversion to Lys-plasminogen by exogenous plasmin. This leads to stimulation of plasminogen activation because Lys-plasminogen is the preferred substrate on cell surfaces. Here, we show that Glu-plasminogen was converted to Lys-plasminogen on monocytoid cells in the absence of exogenous plasmin.

Direct binding properties of conantokins to native N-methyl-d-aspartate receptors.

Conantokin-G (con-G) is a small, gamma-carboxyglutamic acid (Gla)-containing peptide that functions neurophysiologically by inhibiting the N-methyl-d-aspartate receptor (NMDAR). In the current study, the receptor binding properties of an alanine-rich, Gla-deficient con-G variant, Ala-con-G, were assessed following tracer radioiodination with 125I. Direct binding experiments with [125I]Ala-con-G yielded a single site defined by a Kd value of 516 +/- 120 nm.

Structure-function analysis of the streptokinase amino terminus (residues 1-59).

Streptokinase (SK) binds to plasminogen (Pg) to form a complex that converts substrate Pg to plasmin. Residues 1-59 of SK regulate its capacity to induce an active site in bound Pg by a nonproteolytic mechanism and to activate substrate Pg in a fibrin-independent manner. We analyzed 24 SK mutants to better define the functional properties of SK-(1-59).

A fast-acting, modular-structured staphylokinase fusion with Kringle-1 from human plasminogen as the fibrin-targeting domain offers improved clot lysis efficacy.

To develop a fast-acting clot dissolving agent, a clot-targeting domain derived from the Kringle-1 domain in human plasminogen was fused to the C-terminal end of staphylokinase with a linker sequence in between. Production of this fusion protein in Bacillus subtilis and Pichia pastoris was examined. The Kringle domain in the fusion protein produced from B.

Thrombospondin 1--a regulator of adenoma growth and carcinoma progression in the APC(Min/+) mouse model.

Thrombospondin 1 (TSP-1) is a multifunctional extracellular matrix protein that is an endogenous regulator of tumor angiogenesis. The effects of TSP-1 on adenoma formation and development into cancerous lesions has been evaluated in the Min(/+) (multiple intestinal neoplasia) mouse model. These mice develop multiple adenomas in the small intestine due to a mutation in the homologous APC (adenomatous polyposis coli) gene.

Activated protein C blocks p53-mediated apoptosis in ischemic human brain endothelium and is neuroprotective.

Activated protein C (APC) is a systemic anti-coagulant and anti-inflammatory factor. It reduces organ damage in animal models of sepsis, ischemic injury and stroke and substantially reduces mortality in patients with severe sepsis. It was not known whether APC acts as a direct cell survival factor or whether its neuroprotective effect is secondary to its anti-coagulant and anti-inflammatory effects.

Critical role for conversion of glu-plasminogen to Lys-plasminogen for optimal stimulation of plasminogen activation on cell surfaces.

When Glu-plasminogen, the native circulating form of the zymogen, is bound to cell surfaces, its activation is markedly enhanced compared with the reaction in solution. This results in localization of the broad-spectrum proteolytic activity of plasmin on cell surfaces. The cell-associated plasmin plays a key role in fibrinolysis, cell migration, and prohormone processing.

Tissue factor deficiency causes cardiac fibrosis and left ventricular dysfunction.

Exposure of blood to tissue factor (TF) activates the extrinsic (TF:FVIIa) and intrinsic (FVIIIa:FIXa) pathways of coagulation. In this study, we found that mice expressing low levels of human TF ( approximately 1% of wild-type levels) in an mTF(-/-) background had significantly shorter lifespans than wild-type mice, in part, because of spontaneous fatal hemorrhages. All low-TF mice exhibited a selective heart defect that consisted of hemosiderin deposition and fibrosis.

Mice with a severe deficiency of the endothelial protein C receptor gene develop, survive, and reproduce normally, and do not present with enhanced arterial thrombosis after challenge.

The endothelial cell Protein C receptor (EPCR) functions to enhance activation of anticoagulant Protein C (PC) by the thrombin/ thrombomodulin (Tm) complex on the surface of the endothelium. This overall system functions in anticoagulation, profibrinolytic, and antiinflammatory responses. Mice with a severe targeted deficiency of this receptor have been generated by integration of exogenous DNA elements into the 5'-untranslated region of the EPCR gene.

The X-ray crystallographic structure of the angiogenesis inhibitor angiostatin.

Angiogenesis inhibitors have gained much public attention recently as anti-cancer agents and several are currently in clinical trials, including angiostatin (Phase I, Thomas Jefferson University Hospital, Philadelphia, PA). We report here the bowl-shaped structure of angiostatin kringles 1-3, the first multi-kringle structure to be determined. All three kringle lysine-binding sites contain a bound bicine molecule of crystallization while the former of kringle 2 and kringle 3 are cofacial.

Review article: medical treatment of severe ulcerative colitis.

Approximately 15% of patients with ulcerative colitis have a severe attack requiring hospitalization at some time during their illness. This treatment leads to a remission in 60-80% of patients and non-responders may require a total colectomy. Mortality in severe episodes of ulcerative colitis decreased from 31-61% in the 1950s to 5-9% in the 1960s thanks to the introduction of steroids and to a policy of early colectomy.

Fibrinogen stabilizes placental-maternal attachment during embryonic development in the mouse.

In humans, maternal fibrinogen (Fg) is required to support pregnancies by maintaining hemostatic balance and stabilizing uteroplacental attachment at the fibrinoid layer found at the fetal-maternal junction. To examine relationships between low Fg levels and early fetal loss, a genetic model of afibrinogenemia was developed. Pregnant mice homozygous for a deletion of the Fg-gamma chain, which results in a total Fg deficiency state (FG(-/-)), aborted the fetuses at the equivalent gestational stage seen in humans.

Gene targeting of components of the fibrinolytic system.

A number of in vitro and in vivo observations have implicated components of the fibrinolytic system in events associated with diverse physiological and pathophysiological processes, ranging from embryo implantation to cancer. Advances in gene targeting technology have led to the generation of mice deficient for components of the fibrinolytic system. Remarkably, these animals survive to adulthood with few spontaneous life threatening events.

Detection of anti-Saccharomyces cerevisiae antibodies in Crohn's disease: is it a reliable diagnostic and prognostic marker?

Background: In the past few years, serologic markers have been proposed in inflammatory bowel disease. Anti-Saccharomyces cerevisiae antibodies showed high specificity for Crohn's disease. A prognostic role for serology has also been hypothesised.

Domain interactions between streptokinase and human plasminogen.

Plasmin (Pm), the main fibrinolytic protease in the plasma, is derived from its zymogen plasminogen (Plg) by cleavage of a peptide bond at Arg(561)-Val(562). Streptokinase (SK), a widely used thrombolytic agent, is an efficient activator of human Plg. Both are multiple-domain proteins that form a tight 1:1 complex.

Accelerated skin wound healing in plasminogen activator inhibitor-1-deficient mice.

Components of the fibrinolytic system have been implicated in cell migratory events associated with tissue remodeling. Studies in plasminogen-deficient mice (PG(-/-)) indicated that skin wound healing is impaired, but is resolved with an additional fibrinogen deficiency. Plasminogen activator inhibitor-1 (PAI-1) expression by keratinocytes has been identified shortly after wound injury.

Kinetic and mechanistic characterization of NMDA receptor antagonism by replacement and truncation variants of the conantokin peptides.

The characterization of conantokin-T (con-T), conantokin-R (con-R), and variants thereof, using the whole-cell patch clamp technique, was undertaken to evaluate the contribution of various residues towards the onset and recovery of N-methyl-D-aspartate (NMDA) receptor inhibition in cultured embryonic murine hippocampal neurons. The results obtained indicate that the two most C-terminal gamma-carboxyglutamic acid (Gla) residues of the conantokins, while not essential for activity, provided for more tenacious binding to the receptor. Specifically, con-T[gamma10K/gamma14K] and con-R[gamma11A/gamma15A] displayed 5.