Lactadherin blocks thrombosis and hemostasis in vivo : correlation with platelet phosphatidylserine exposure

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/73556/1/j.1538-7836.2008.03010.x.pd

Leukocyte- and Platelet-Derived Microvesicle Interactions following In Vitro and In Vivo Activation of Toll-Like Receptor 4 by Lipopolysaccharide

BACKGROUND: Pro-coagulant membrane microvesicles (MV) derived from platelets and leukocytes are shed into the circulation following receptor-mediated activation, cell-cell interaction, and apoptosis. Platelets are sentinel markers of toll-like receptor 4 (TLR4) activation. Experiments were designed to evaluate the time course and mechanism of direct interactions between platelets and leukocytes following acute activation of TLR4 by bacterial lipopolysaccharide (LPS). METHODOLOGY/PRINCIPAL FINDINGS: Blood from age-matched male and female wild type (WT) and TLR4 gene deleted (dTLR4) mice was incubated with ultra-pure E. coli LPS (500 ng/ml) for up to one hour. At designated periods, leukocyte antigen positive platelets, platelet antigen positive leukocytes and cell-derived MV were quantified by flow cytometry. Numbers of platelet- or leukocyte-derived MV did not increase within one hour following in vitro exposure of blood to LPS. However, with LPS stimulation numbers of platelets staining positive for both platelet- and leukocyte-specific antigens increased in blood derived from WT but not dTLR4 mice. This effect was blocked by inhibition of TLR4 signaling mediated by My88 and TRIF. Seven days after a single intravenous injection of LPS (500 ng/mouse or 20 ng/gm body wt) to WT mice, none of the platelets stained for leukocyte antigen. However, granulocytes, monocytes and apoptotic bodies stained positive for platelet antigens. CONCLUSIONS/SIGNIFICANCE: Within one hour of exposure to LPS, leukocytes exchange surface antigens with platelets through TLR4 activation. In vivo, leukocyte expression of platelet antigen is retained after a single exposure to LPS following turn over of the platelet pool. Acute expression of leukocyte antigen on platelets within one hour of exposure to LPS and the sustained expression of platelet antigen on leukocytes following a single acute exposure to LPS in vivo explains, in part, associations of platelets and leukocytes in response to bacterial infection and changes in thrombotic propensity of the blood

Gradual alteration of mitochondrial structure and function by β-amyloids: importance of membrane viscosity changes, energy deprivation, ROS production and cytochrome c release.

Intracellular amyloid beta-peptide (Abeta) accumulation is considered to be a key pathogenic factor in sporadic Alzheimer's disease (AD), but the mechanisms by which it triggers neuronal dysfunction remain unclear. We hypothesized that gradual mitochondrial dysfunction could play a central role in both initiation and progression of sporadic AD. Thus, we analyzed changes in mitochondrial structure and function following direct exposure to increasing concentrations of Abeta(1-42) and Abeta(25-35) in order to look more closely at the relationships between mitochondrial membrane viscosity, ATP synthesis, ROS production, and cytochrome c release. Our results show the accumulation of monomeric Abeta within rat brain and muscle mitochondria. Subsequently, we observed four different and additive modes of action of Abeta, which were concentration dependent: (i) an increase in mitochondrial membrane viscosity with a concomitant decrease in ATP/O, (ii) respiratory chain complexes inhibition, (iii) a potentialization of ROS production, and (iv) cytochrome c release

Update on functional and genetic laboratory assays for the detection of platelet microvesicles

Functional and genetic assays for measuring platelet microvesicles (PMVs) are presented and discussed. Functional assays concern two groups of methods: a) homogeneous assays using the cofactor activity of phospholipids (PPLs) contained in PMVs and present in assayed plasmas, and a coagulation or a thrombin generation assay (TGA) as “end points”; b) capture-based assays, in which PMVs bind to an immobilized ligand, such as Annexin V in the presence of calcium, or monoclonal antibodies (MoAbs) specific for membrane proteins. Genetic assays aim to detect micro-RNA (miRNA) present in PMVs: miRNA must be extracted from plasma, and the expression pattern can be determined by various methods such as quantitative real-time PCR, microarray or sequencing. All these technical approaches introduce new exploration tools for measuring or quantitating PMVs or their associated activities, as biomarkers for disease evolution, their diagnosis or prognosis, and for monitoring of some antithrombotic or anti-inflammatory therapies. They offer invaluable analytical tools for research, drug discovery and epidemiological studies and have a strong potential as diagnostic tests