Coronary plaque injury triggers neutrophil activation in patients with coronary artery disease

Activation of leukocytes, in particular polymorphonuclear neutrophils (PMN), is considered an early event in unstable coronary disease. Upon activation PMN liberate myeloperoxidase (MPO), an enzyme which binds to the vessel wall and depletes vascular NO bioavailability. Using coronary balloon angioplasty as a trigger to provoke coronary plaque injury, we assessed the time course of neutrophil activation, local and peripheral levels of myeloperoxidase, and systemic vascular NO bioavailability in patients with stable coronary artery disease. Twenty-four patients with stable CAD were enrolled prior to undergoing percutaneous interventions (PCI, n=14) and diagnostic coronary angiography (n=10), respectively. Following angioplasty arterial MPO plasma levels increased (231.5+/-67.6 to 273.8+/-80.4 pg/mg protein; P<0.01) whereas MPO levels in the coronary sinus decreased (240.8+/-74.4 vs 205.4+/-60.1 pg/mg protein; P<0.01) in the absence of elevated serum markers for myocardial necrosis. Following PCI, patients revealed impaired vascular NO bioavailability as reflected by reduced brachial flow-mediated dilation (FMD; 6.25+/-3.03 to 4.90+/-2.70%; P<0.01), whereas FMD increased in the angiography group. Coronary plaque injury provokes rapid activation of PMN in the absence of myocardial necrosis; the coronary circulation emerges as a primary site for deposition of MPO following injury of the coronary vessel wall. Activation of PMN with release of MPO is not only restricted to the target site, but can be assessed systemically and may represent a critical mechanistic link for impaired systemic vascular NO bioavailability in patients suffering unstable coronary disease.     

Vinculin E29R mutation changes cellular mechanics

We investigated the effect of the point mutation E29R on vinculin under cell mechanical aspects. MEFvcl KO cells were transfected with intact eGFP-vinculin (rescue) or mutant E29R vinculin. Cellular stiffness and adhesion strength of mutant E29R vinculin were considerably higher compared to rescue and MEFvcl KO cells. 2D traction microscopy also indicated markedly higher strain energy in E29R mutant cells compared to rescue and MEFvcl KO cells. Fluorescence recovery after photobleaching showed that the recovery time for mutant E29R cells was drastically slower than for MEFvcl rescue cells and that the mobile fraction was larger for rescue compared to E29R mutant cells. These results indicate that E29R mutation might prime the vinculin head for F-actin binding, which results in higher cell stiffness, contractile force, and strengthening of focal adhesions.     

Phosphorylation of filamin (ABP-280) regulates the binding to the lipid membrane, integrin, and actin

Actin-binding protein (ABP-280; filamin) is a phosphoprotein present in the periphery of the cytoplasm, where it can cross-link actin filaments, associate with lipid membranes, and bind to membrane surface receptors. Given its function and localization in the cell, the hypothesis that it serves as a substrate for p56lck, a lymphocyte-specific member of the src family of protein tyrosine kinases associated with cell surface glycoproteins is considered. The results suggest conformationally-induced regulation of filamin (ABP-280).     

Epidemiological Characteristics of Classical Scrapie Outbreaks in 30 Sheep Flocks in the United Kingdom

Background

Most previous analyses of scrapie outbreaks have focused on flocks run by research institutes, which may not reflect the field situation. Within this study, we attempt to rectify this deficit by describing the epidemiological characteristics of 30 sheep flocks naturally-infected with classical scrapie, and by exploring possible underlying causes of variation in the characteristics between flocks, including flock-level prion protein (PrP) genotype profile. In total, the study involved PrP genotype data for nearly 8600 animals and over 400 scrapie cases.

Methodology/Principal Findings

We found that most scrapie cases were restricted to just two PrP genotypes (ARQ/VRQ and VRQ/VRQ), though two flocks had markedly different affected genotypes, despite having similar underlying genotype profiles to other flocks of the same breed; we identified differences amongst flocks in the age of cases of certain PrP genotypes; we found that the age-at-onset of clinical signs depended on peak incidence and flock type; we found evidence that purchasing infected animals is an important means of introducing scrapie to a flock; we found some evidence that flock-level PrP genotype profile and flock size account for variation in outbreak characteristics; identified seasonality in cases associated with lambing time in certain flocks; and we identified one case that was homozygous for phenylalanine at codon 141, a polymorphism associated with a very high risk of atypical scrapie, and 28 cases that were heterozygous at this codon.

Conclusions/Significance

This paper presents the largest study to date on commercially-run sheep flocks naturally-infected with classical scrapie, involving 30 study flocks, more than 400 scrapie cases and over 8500 PrP genotypes. We show that some of the observed variation in epidemiological characteristics between farms is related to differences in their PrP genotype profile; although much remains unexplained and may instead be attributed to the stochastic nature of scrapie dynamics.     

Specificity and localization of the hepatitis B virus-associated protein kinase.

The nature of the protein kinase (PK) which phosphorylates the core protein of hepatitis B virus in vitro was studied. The PK copurified with the core particles during rate zonal centrifugation and gel chromatography. It showed the same size heterogeneity as the core particles, which consisted of a main fraction of 28-nm particles and a subfraction of 22- to 26-nm particles. DNA-containing heavy core particles with a density of 1.33 to 1.35 g/ml and less endogenous PK than did the light cores. The phosphorylation reaction had a rapid initial phase (several minutes) and a slow but long-lasting second phase (many hours). The PK had a high affinity for ATP (KM = 0.5 mumol/liter). Only few of the several hundred P21.9 subunits in one core particle were phosphorylated in vitro. The only amino acid which was phosphorylated in vitro was serine. The resistance of the introduced phospho group against alkaline phosphatase showed that the PK acceptor, and probably the enzyme itself, was located inside the core particle.