Cell-derived vesicles exposing coagulant tissue factor in saliva

On vascular damage, coagulation is initiated by extravascular tissue factor (TF). Intravascular TF, which is present on circulating cell-derived vesicles, is noncoagulant under physiologic conditions but prothrombotic under pathologic conditions. Human saliva triggers coagulation, but the mechanism and physiologic relevance are unknown. Because saliva is known to contain TF, we hypothesized that this TF may also be associated with cell-derived vesicles to facilitate coagulation when saliva directly contacts blood. The saliva-induced shortening of the clotting time of autologous plasma and whole blood from healthy subjects (n = 10) proved TF-dependent. This TF was associated with various types of cell-derived vesicles, including microparticles and exosomes. The physiologic function was shown by adding saliva to human pericardial wound blood collected from patients undergoing cardiac surgery. Addition of saliva shortened the clotting time from 300 ± 96 to 186 ± 24 seconds (P = .03). Our results show that saliva triggers coagulation, thereby reducing blood loss and the risk of pathogens entering the blood. We postulate that our reflex to lick a wound may be a mechanism to enable TF-exposing vesicles, present in saliva, to aid in the coagulation process and thus protect the organism from entering pathogens. This unique compartmentalization may be highly conserved because also animals lick their wounds.     

Increases in myeloperoxidase levels after exercise in myocardial perfusion scintigraphy are not induced by myocardial ischemia

BACKGROUND: Increased systemic levels of myeloperoxidase (MPO) have been reported in patients with acute myocardial ischemia. We studied the association between exercise-induced myocardial ischemia measured by myocardial perfusion scintigraphy (MPS) and the magnitude and time course of changes in MPO levels in humans. METHODS: One hundred and twenty six patients underwent symptom limited exercise MPS. Myocardial ischemia was assessed semi-quantitatively. Plasma samples were taken before the start of exercise (baseline), at maximum exercise and every hour up to 6 h after maximum exercise. RESULTS: Myocardial ischemia was present in 42 (33%) patients. MPO levels rapidly increased during exercise in patients with and without ischemia (to 131% (106-172%) and 145% (121-199%) of baseline, respectively). MPO levels and absolute changes in MPO did not differ between patients with and without ischemia at any time point. None of the patient characteristics, including presence of ischemia, was independently predictive of the absolute increase in MPO levels during exercise. CONCLUSIONS: Exercise related immediate increases in MPO levels do not reflect myocardial ischemia.     

P-selectin- and CD63-exposing platelet microparticles reflect platelet activation in peripheral arterial disease and myocardial infarction

BACKGROUND: Platelet-derived microparticles (PMPs) are generally considered a marker of platelet activation in cardiovascular disease. We studied the extent to which PMP subpopulations parallel platelet activation in vitro and in vivo. METHODS: Using flow cytometry, we analyzed PMP subpopulations from resting and activated platelets in vitro (n = 6) as well as from plasma samples of patients with stable angina, peripheral arterial disease, or myocardial infarction [non-ST-elevation (NSTEMI) and ST-elevation (STEMI)] and from older, age- and sex-matched and young healthy individuals [n = 10 for all groups except NSTEMI (n = 11)]. Coagulation markers prothrombin fragment F(1 + 2) and thrombin-antithrombin complexes were determined by ELISA. The PMP-associated fraction of soluble (s)P-selectin was estimated by ELISA. RESULTS: In vitro, stimulation of platelets with thrombin receptor-activating peptide (15 micromol/L) or the calcium ionophore A23187 (2.5 micromol/L) increased fractions of both platelets and PMPs exposing P-selectin or CD63 (P <0.001 for all). Whereas the number of PMPs released by A23187-stimulated platelets increased significantly (P <0.001), the number of PMPs released from thrombin receptor-activating peptide-stimulated platelets remained constant (P >0.05). Ex vivo, numbers of circulating PMPs were comparable in all groups. Compared with young persons, P-selectin-exposing PMPs were increased in older persons (P = 0.02) and were further increased in patients with NSTEMI (P = 0.007) and STEMI (P = 0.045). CD63-exposing PMPs were increased in patients with peripheral arterial disease (P = 0.041), NSTEMI (P = 0.001), and STEMI (P = 0.049). Subpopulations exposing P-selectin or CD63 correlated with each other (r = 0.581; P <0.001), but neither correlated with the plasma concentrations of F(1 + 2) or thrombin-antithrombin complexes. The PMP-associated fraction of sP-selectin constituted only 2.2 (4.7)% [mean (SD)] of total sP-selectin. CONCLUSIONS: PMP subpopulations reflect platelet activation status better than the total number of PMPs. Increased concentrations of circulating PMP subpopulations are found in aging, and further increases are encountered in peripheral arterial disease and myocardial infarction.     

Expression of inflammation-related genes in endothelial cells is not directly affected by microparticles from preeclamptic patients

BACKGROUND: Inflammation and endothelial dysfunction are prominent in preeclampsia. Microparticles (MPs) may link these processes, as MPs induce the production of pro-inflammatory cytokines by endothelial cells and cause endothelial dysfunction. AIM: To study changes in expression of inflammation-related genes in human endothelial cells in response to MPs from preeclamptic patients. METHODS: Human umbilical vein endothelial cells (HUVECs) were incubated for various time intervals in the absence or presence of isolated MP fractions from preeclamptic patients (n = 3), normotensive pregnant women (n = 3), non-pregnant controls (n = 3), and interleukin (IL)-1alpha as a positive control. Total RNA was isolated and used for multiplex ligation-dependent probe amplification (MLPA) and real-time polymerase chain reaction (PCR). RESULTS: IL-1alpha enhanced the expression of IL-1alpha, IL-2, IL-6, and IL-8; nuclear factor of kappa light chain enhancer in B-cells (NFkappaB)-1, NFkappaB-2, and NFkappaB-inhibitor; cyclin-dependent kinase inhibitor and monocyte chemotactic protein-1; and transiently increased tissue factor expression. RNA expression of inflammation-related genes and genes encoding adhesion receptors, however, were unaffected by any of the MP fractions tested. CONCLUSION: MLPA is a suitable assay to test the inflammatory status of endothelial cells, because incubation with IL-1alpha triggered substantial changes in RNA expression in endothelial cells. Taken together, it seems unlikely that MPs from preeclamptic patients induce endothelial dysfunction by directly affecting the expression of inflammation-related genes in these cells.