Platelet Function tests: A Comparative Review

In physiological hemostasis a prompt recruitment of platelets on the vessel damage prevents the bleeding by the rapid formation of a platelet plug. Qualitative and/or quantitative platelet defects promote bleeding, whereas the high residual reactivity of platelets in patients on antiplatelet therapies moves forward thromboembolic complications. The biochemical mechanisms of the different phases of platelet activation – adhesion, shape change, release reaction, and aggregation – have been well delineated, whereas their complete translation into laboratory assays has not been so fulfilled. Laboratory tests of platelet function, such as bleeding time, light transmission platelet aggregation, lumiaggregometry, impedance aggregometry on whole blood, and platelet activation investigated by flow cytometry, are traditionally utilized for diagnosing hemostatic disorders and managing patients with platelet and hemostatic defects, but their use is still limited to specialized laboratories. To date, a point-of-care testing (POCT) dedicated to platelet function, using pertinent devices much simpler to use, has now become available (ie, PFA-100, VerifyNow System, Multiplate Electrode Aggregometry [MEA]). POCT includes new methodologies which may be used in critical clinical settings and also in general laboratories because they are rapid and easy to use, employing whole blood without the necessity of sample processing. Actually, these different platelet methodologies for the evaluation of inherited and acquired bleeding disorders and/or for monitoring antiplatelet therapies are spreading and the study of platelet function is strengthening. In this review, well-tried and innovative platelet function tests and their methodological features and clinical applications are considered

Assessment of fibrinolytic activity by measuring the lysis time of a tissue-factor-induced clot: a feasibility evaluation.

A clot lysis time assay in which a tissue factor—induced fibrin clot is lysed by exogenously added tissue plasminogen activator has been recently reported. We evaluated the feasibility of clot lysis time in a routine hemostasis laboratory, and its correlation with thrombin activatable fibrinolysis inhibitor and plasminogen activator inhibitor-1 levels and changes with aging in 185 healthy participants. Clot lysis time was assessed by monitoring changes in turbidity during clot formation and subsequent lysis using a computerized kinetic spectrophotometric microtiter plate. After preliminary experiments, 100 and 160 ng/mL tissue plasminogen activator concentrations were chosen for the study. Clot lysis time was calculated by a new mathematical analysis of the lysis curve based on discrete derivative. Clot lysis time, thrombin activatable fibrinolysis inhibitor, and plasminogen activator inhibitor-1 plasma levels showed a normal distribution. For both concentrations of tissue plasminogen activator, clot lysis time progressively increased with increase in age (P < .0001) and was significantly correlated with thrombin activatable fibrinolysis inhibitor antigen, thrombin activatable fibrinolysis inhibitor activity, and plasminogen activator inhibitor-1 antigen (at least P < .01). During linear regression analysis, thrombin activatable fibrinolysis inhibitor and plasminogen activator inhibitor-1 antigen were found to significantly influence clot lysis time (at least P < .01). Clot lysis time determination has a good laboratory performance. Our new method of calculation is independent of the time of reading and allows a more accurate and consistent detection of both short and prolonged lysis times. Our data suggest the feasibility of the use of this test in the work of routine hemostasis laboratory

The Impact of a Single Apheretic Procedure on Endothelial Function Assessed by Peripheral Arterial Tonometry and Endothelial Progenitor Cells

Introduction: Endothelial progenitor cells (EPC) are involved in vascular repair and proliferation, contributing to the long-term outcomes of apheretic treatment. Aim of this study was to investigate the relationships between endothelial function, assessed by levels of bone marrow-derived progenitor cells and endothelial response to hyperaemia, and clinical and biohumoral parameters in high vascular risk patients before, immediately after, 24-hours and 72 hours after a single lipid apheresis procedure. Material and Methods: We evaluated lipid profile, endothelial function and endothelial progenitor cells before (T0), immediately after (T1), 24h after (T2) and 72h after (T3) a lipoprotein apheresis procedure, in 8 consecutive patients [Sex: 62.5% M; Age; 63.29(12), mean, (range) years] with a personal history of acute coronary syndrome, symptomatic peripheral arterial disease and elevated plasma levels of lipoprotein (a) [Lp(a)]. Patients were on regularly weekly or biweekly lipoprotein apheresis, and they were treated with the FDA-approved Heparin-induced Extracorporeal LDL Precipitation (H.E.L.P.) (Plasmat Futura, B.Braun, Melsungen, Germany) technique. PAT values were expressed as the natural logarithm (Ln-RHI, normal values&ge;0.4) of the reactive hyperaemia index (RHI), which is the parameter automatically calculated by the device. Results: We found a reduction in the natural logarithm of reactive hyperaemia index (Ln-RHI), assessed immediately after the procedure (0.57&plusmn;0.21 vs 0.72&plusmn; 0.29); difference between T2 and T0 was statistically significant (0.43&plusmn;0.24 vs 0.72&plusmn;0.29; p=0.006). Reduction in Ln-RHI values was documented in all patients, two subjects showing a Ln-RHI&lt;0.4 at T1, and four at T2. At T3, PAT values were increased significantly (0.91&plusmn;0.18) in comparison to T1 and T2, showing a median value higher than at T0. Cd34+/Kdr+ and Cd133+/Kdr+ showed a minimum increase in median values at T1, and a higher increase at T2, in comparison to baseline. Differences in Cd34+/133+/Kdr+ values at different times were not statistically significant. A significant reduction in circulating endothelial cells (CEC) count at T2 in comparison to T0 was found (12.00&plusmn;8.85 vs 23.86&plusmn;12.39; p=0.024). Discussion: At 24h and 72h after procedures, we found an improvement in endothelial function, expressed by an increase in PAT values and EPC levels, and by a reduction in CEC

Erythrocyte membrane fluidity alterations in sudden sensorineural hearing loss patients: The role of oxidative stress

Introduction Sudden sensorineural hearing loss (SSNHL) involves an acute unexplained hearing loss, nearly always unilateral, that occurs over less than a 72-hour period. SSNHL pathogenesis is not yet fully understood. Cochlear vascular occlusion has been proposed as a potential mechanism of hearing damage and cochlear ischaemia has been related to alterations of cochlear microvessels. In addition, some researchers have focused their attention on the rheological alterations and blood hyperviscosity. Erythrocyte deformability plays a key role in determining blood viscosity, and it is critical to cochlear perfusion. It has been shown that oxidative stressinduced erythrocyte membrane fluidity alterations are linked to the progression of cardiovascular diseases. Methods To determine whether erythrocytes from SSNHL patients show signs of oxidative stress, and whether this condition can modify the haemorheologic profile in these patients, we analysed haemorheologic profile and erythrocyte oxidative stress in 35 SSNHL patients and 35 healthy subjects, matched for age and sex. Fluorescence anisotropy was used to evaluate the fluidity of erythrocyte membranes. Results Our results show a significant structural and functional involvement of erythrocyte membrane alterations in SSNHL, as well as elevated levels of membrane lipid peroxidation and intracellular reactive oxygen species (ROS) production. In addition, erythrocyte-derived ROS and erythrocyte lipid peroxidation positively correlated with whole blood viscosity and erythrocyte deformability. Moreover, in vitro experiments demonstrated that ROS display a key role in erythrocyte membrane fluidity. Conclusion These findings indicate that erythrocyte oxidative stress plays a key role in the pathogenesis of SSNHL and pave the way to new therapeutic interventions