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In addition to its classical receptor, CD40, it is now well established that CD154 also binds αIIbβ3, α5β1, and αMβ2 integrins. Although these integrins are all members of the same family, they bind CD154 differently. The current investigation aims to analyze the interaction of CD154 with α5β1 and αMβ2 and investigate its role in bidirectional signals in various human cell lines. Results obtained herein indicate that the CD154 residues involved in the interaction with α5β1 are N151 and Q166, whereas those involved in αMβ2 binding are common to residues required for CD40, namely Y145 and R203. Soluble CD40/CD154 or αMβ2/CD154 complexes do not interfere with the binding of CD154 to α5β1-positive cells, but inhibit the binding of CD154 to CD40- or αMβ2-positive cells, respectively. Ligation of CD154 on CD154-positive cells with soluble CD40, αIIbβ3, α5β1, or αMβ2 stimulates intracellular signaling, including MAPK phosphorylation. Given that CD154 exists as a trimer, our data strongly suggest that CD154 may bind concomitantly to two receptors of the same or different family, and biologically activate cells expressing both receptors. The characterization of CD154/receptor interactions helps the identification of new therapeutic targets for the prevention and/or treatment of CD154-associated autoimmune and inflammatory diseases.

Thromboelastography uses cups and pins made of Cyrolite ® plastic to analyze the rate of fibrin clot formation in blood samples. In this study, TEG cups and pins were modified by 4 distinct coating types using plasma-enhanced chemical vapor deposition (PECVD): carboxylated, amine-rich, hydrophobic, SiO 2 , and analyzed for surface chemistry and wettability. We tested the hypothesis that the coagulation kinetics of recalcified citrated blood plasma is controlled by surface chemistry, in the absence of clot activator. Only carboxylated surfaces became negatively charged upon wetting, and accelerated clot formation in a highly reproducible manner, whereas Cyrolite ® and the other coatings had delayed and unpredictable clotting times. These data are consistent with a model whereby carboxylated surfaces selectively adsorb and activate factor XII while repelling other more abundant anionic blood proteins, resulting in reproducible clot kinetics.

Vascular PET grafts (Dacron) have shown good performance in large vessels (≥ 6 mm) applications. To address the urgent unmet need for small-diameter (2-6 mm) vascular grafts, proprietary high-compliance nonwoven PET fiber structures were modified with various PEG concentrations using PVA as a cross-linking agent, to fabricate non-thrombogenic mechanically compliant vascular grafts. The blood compatibility assays measured through platelet adhesion (SEM and mepacrine dye) and platelet activation (morphological changes, P-selectin secretion, and TXB2 production) demonstrate that functionalization using a 10% PEG solution was sufficient to significantly reduce platelet adhesion/activation close to optimal literature-reported levels observed on carbon-coated ePTFE.

Excessive distension of venous grafts due to arterial pressure enhances the convective water transport (filtration flow) through the vessel wall, and thus might affect the infiltration of macromolecules such as lipoproteins. In this paired experimental study, filtration velocities were measured at 100 mm Hg for canine jugular veins with or without external supports of expanded polytetrafluoroethylene (ePTFE) arterial prostheses. In addition, to assess the effect of filtration velocity on lipid infiltration or uptake, canine jugular veins were wrapped over half of their lengths with ePTFE arterial prostheses and perfused with dog serum containing 3H-cholesterol at a pressure of 100 mm Hg. At 100 mm Hg, the average filtration velocity of the wrapped jugular veins was 7.9 +/- 1.3 x 10(-6) cm/s whereas the average filtration velocity of the unwrapped veins was 27.3 +/- 2.7 x 10(-6) cm/s (p < 0.005). Moreover, the unwrapped veins had a significantly higher uptake rate of labeled cholesterol than the wrapped veins (10.9 +/- 7.3 x 10(-4) cm/h and 5.0 +/- 1.6 x 10(-4) cm/h, respectively, p < 0.005). In conclusion, under arterial pressure, veins experience excessive distention, which leads to significant increases in both filtration flow and cholesterol uptake. An external wrap or support of ePTFE material protects veins from excessive distension and thus may prevent atherosclerosis in venous grafts by reducing cholesterol uptake.

Hydroxyapatite (HA) and HA-alumina (HA/Al2O3) composites, with Al2O3 contents of 5, 10, 20, and 30%, were synthesized using a wet precipitation method and sintered at 900 and 1300°C. We investigated the effect of sintering temperature and relative concentration of HA and Al2O3 on the chemical composition, surface morphology, and cytotoxicity of the composite powders. The XRD results show that in the 1300°C composites, HA partially decomposed into CaO which combined with Al2O3 to form different calcium aluminates. For the 900°C composites the CaO phase was not detected, though a Ca/P ratio larger than 1.67 measured by XPS suggests that CaO was present in trace amounts. SEM-EDX analysis indicated that the HA microstructure was affected by the sintering temperature, and this HA is present on the surface of Al2O3 particles. The cytotoxicity of the composites was assessed indirectly using the MTT assay. The short-term effect of leachables was quantified by exposing a L929 mouse fibroblast cell line to the degradation products released by the composites after immersion in the cell culture medium. Degradation products were less toxic to L-929 at lower extract concentrations (10, 50%) than at 100% concentration. Cell viability was also influenced by leachable size. © 2007 Wiley Periodicals, Inc. J Biomed Mater Res, 2007

1 P-selectin is involved, with P-selectin glycoprotein (GP)-ligand-1 (PSGL-1), in platelet/leukocyte interactions during thrombo-inflammatory reactions; it also stabilizes platelet aggregates. Its antagonism accelerates thrombolysis and enhances the anti-aggregatory effects of GPIIb–IIIa inhibitors. This study was designed to investigate the mechanisms of P-selectin-mediated platelet aggregation. 2 In freshly isolated human platelets, P-selectin translocation after thrombin stimulation increased rapidly to 48, 72, and 86% positive platelets after 60, 120, and 300 s, respectively. Platelet aggregation at 60 s post-stimulation averaged 46.7±1.9% and its extent followed closely the kinetics of P-selectin translocation. 3 Pre-treatment of platelets with P-selectin antagonists, a recombinant PSGL-1 (rPSGL-Ig) or a blocking monoclonal antibody, significantly delayed platelet aggregation in a dose-dependent manner. At 100 μg ml−1 of rPSGL-Ig, platelet aggregation was completely inhibited up to 60 s post-stimulation and increased thereafter to reach maximal aggregation at 5 min. The second phase of platelet aggregation, in the presence of rPSGL-Ig, was completely prevented by the addition of a GPIIb–IIIa antagonist (Reopro) at 60 s, whereas its addition in the absence of rPSGL-Ig was without any significant effect. 4 Combination of rPSGL-Ig with Reopro or with an inhibitor of Pi3K (LY294002), which reduces GPIIb–IIIa activation, showed to be more effective in inhibiting platelet aggregation, in comparison to the effects observed individually. 5 rPSGL-Ig blocks P-selectin, whereas Reopro and LY294002 block GPIIb–IIIa and its activation, respectively, without a major effect on the percentage of platelets expressing P-selectin. 6 In summary, platelet P-selectin participates with GPIIb–IIIa in the initiation of platelet aggregation. Its inhibition, with rPSGL-Ig, delays the aggregation process and increases the anti-aggregatory potency of Reopro. Thus, combination of P-selectin and GPIIb–IIIa antagonism may constitute a promising therapeutic option in the management of thrombotic disorders. British Journal of Pharmacology (2006) 148, 299–305. doi:10.1038/sj.bjp.0706734

Membrane-covered devices could help treat disease of the vasculature such as aneurysm, rupture, and fistulas. They are also investigated to reduce embolic complication associated with revascularization of saphenous vein graft. The aim of this study is to design a clinically applicable biodegradable membrane-covered stent based on the natural polysaccharide chitosan, which has been developed. The mechanical properties of the membrane is optimized through blending with polyethylene oxide (70:30% Wt CH:PEO). The membrane was able to sustain the mechanical deformation of the supporting self-expandable metallic stents during its deployment. The membrane was demonstrated to resist physiological transmural pressure (burst pressure resistance >500 mm Hg) and presented a high-water permeation resistance (1 mL/cm(2) min(-1) at 120 mmHg). The CH-PEO membrane showed a good hemocompatibility in an ex vivo assay. Heparin and hyaluronan surface complexation with the membrane further reduced platelet adhesion by 50.1 and 63% (p = 0.05). The ability of the membrane-covered devices to be used as a drug reservoir was investigated using the nitric oxide donor sodium nitroprusside (SNP). SNP-loaded membranes displayed significantly reduced platelet adhesion.

Leukotrienes constitute a class of potent bioactive mediators known to play a pivotal role in inflammation. Since their biosynthesis has been shown to be enhanced by platelet-neutrophil interactions, leukotrienes may be involved in these interactions and the arterial response to injury. Therefore, we investigated the effects of the selective leukotriene biosynthesis inhibitor 3-[1-(4-chlorobenzyl)-3-t-butyl-thio-5-isopropylindol-2-yl]-2,2-dimethylpropanoic acid (MK-886) on the acute thrombotic and vasomotor responses after arterial injury by angioplasty. Carotid arterial injury was produced by balloon dilatation in control (molecusol vehicle; n=10) and treated (MK-886, 10 mg kg−1, i.v.; n=9) pigs. The acute thrombotic reaction following deep arterial wall injury was quantified with 51Cr labelled platelets and 111In labelled neutrophils, and the vasomotor response was determined angiographically. Platelet deposition at the site of deep arterial wall injury averaged 56.4±11.0×106 platelets cm−2 in the control group, and was significantly reduced to 18.2±3.8×106 platelets cm−2 (P