Changes In Platelet Activation Research Articles

Reduced platelet activation and thrombus formation in male transgenic model mice of Alzheimer's disease suggests early sex-specific differences in platelet pathophysiology

Alzheimer's disease (AD) is the most common form of dementia and characterized by extracellular amyloid-β (Aβ) plaques, intracellular neurofibrillary tau tangles and neurodegeneration. Over 80 % of AD patients also exhibit cerebral amyloid angiopathy (CAA). CAA is a cerebrovascular disease caused by deposition of Aβ in the walls of cerebral blood vessels leading to vessel damage and impairment of normal blood flow. To date, different studies suggest that platelet function, including activation, adhesion and aggregation, is altered in AD due to vascular Aβ deposition. For example, the transgenic AD model mice APP23 mice that exhibit CAA and parenchymal Aβ plaques, show pre-activated platelets in the blood circulation and increased platelet integrin activation leading to a pro-thrombotic phenotype in these mice late stages of AD. However, it is still an open question whether or not platelets exhibit changes in their activation profile before they are exposed to vascular Aβ deposits. Therefore, the present study examined platelets from middle-aged transgenic APP23 mice at the age of 8–10 months. At this age, APP23 mice show amyloid plaques in the brain parenchyma but not in the vasculature. Our analyses show that these APP23 mice have unaltered platelet numbers and size, and unaltered surface expression of glycoproteins. However, the number of dense granules in transgenic platelets was increased while the release was unaltered. Male, but not female APP23 mice, exhibited reduced platelet activation after stimulation of the thrombin receptor PAR4 and decreased thrombus stability on collagen under flow conditions ex vivo compared to control mice. In an arterial thrombosis model in vivo, male APP23 mice showed attenuated occlusion of the injured artery compared to controls. These findings provide clear evidence for early changes in platelet activation and thrombus formation in male mice before development of overt CAA. Furthermore, reduced platelet activation and thrombus formation suggest sex-specific differences in platelet physiology in AD that has to be considered in future studies of platelets and their role in AD.

Decreased platelet activation predicts hepatic decompensation and mortality in patients with cirrhosis.

Patients with cirrhosis show alterations in primary haemostasis, yet prognostic implications of changes in platelet activation remain controversial and assay validity is often limited by thrombocytopenia. We aimed to study the prognostic role of platelet activation in cirrhosis, focusing on bleeding/thromboembolic events, decompensation and mortality. We prospectively included 107 cirrhotic patients undergoing a same-day hepatic venous pressure gradient (HVPG) and platelet activation measurement. Platelet activation was assessed using flow cytometry after protease-activated receptor (PAR)-1, PAR-4, or epinephrine stimulation. Over a follow-up of 25.3 (IQR:15.7-31.2) months, first/further decompensation occurred in 29 patients and 17 died. More pronounced platelet activation was associated with an improved prognosis, even after adjusting for systemic inflammation, HVPG and disease severity. Specifically, higher PAR-4-inducible platelet activation was independently linked to a lower decompensation risk (aHR per 100 MFI [median fluorescence intensity]:0.95 [95%CI:0.90-0.99]; p=0.036) and higher PAR-1-inducible platelet activation was independently linked to longer survival (aHR per 100 MFI:0.93 [95%CI:0.87-0.99]; p=0.040). Thromboembolic events occurred in 8 patients (75% non-tumoral portal vein thrombosis [PVT]). Higher epinephrine-inducible platelet activation was associated with an increased risk of thrombosis (HR per 10 MFI:1.07 [95%CI:1.02-1.12]; p=0.007) and PVT (HR per 10 MFI:1.08 [95%CI:1.02-1.14]; p=0.004). In contrast, of the 11 major bleedings that occurred, 9 were portal hypertension-related, and HVPG thus emerged as the primary risk factor. Preserved PAR-1- and PAR-4-inducible platelet activation was linked to a lower risk of decompensation and death. In contrast, higher epinephrine-inducible platelet activation was a risk factor for thromboembolism and PVT.

Using an agonist panel and aggregation to assess antiplatelet therapy in mice

Abstract Platelets play a crucial role in hemostasis. For cardiovascular and stroke prophylaxis, patients are commonly placed on either single or dual antiplatelet drugs, such as aspirin and/or clopidogrel. Here we outline a mouse model examining antiplatelet therapy and the resulting changes in platelet activation and aggregation. Using commercially available transgenic mice with platelets expressing green fluorescent protein (GFP) or red fluorescent protein (RFP), endogenous platelets were inhibited with a single, dual, or control antiplatelet treatment. We used 100µg aspirin/g mouse or 50µg clopidogrel/g mouse for a single dose; dual antiplatelet therapy was a combined dose of 100µg aspirin/g mouse and 50µg clopidogrel/g; control treatment was DMSO alone. One hour after treatment, whole blood (WB) was collected by aseptic cardiac puncture and mixed in the following groups: aspirin treated RFP and GFP WB, clopidogrel treated RFP and GFP WB, aspirin and clopidogrel treated RFP and GFP WB, control treated RFP and GFP WB. Each combined sample was then treated with a panel of agonists: no agonist, 0.5U/mL thrombin, 20µM ADP, and 500µg/mL arachidonic acid. Thrombin samples were activated for 2 minutes at room temperature while ADP and arachidonic acid samples were activated for 10 minutes at 37°C. Platelets were labeled with anti-CD41a to label all platelets and anti-CD62P to label activated platelets. Aggregation is measured as double-colored GFP and RFP platelet events. Compared to control mice, mice treated with antiplatelet agents demonstrated statistically significant decreases in activation following thrombin administration (ranging from 90% to 83% activated); there was no significant difference in platelet aggregation following thrombin among the samples from different antiplatelet therapies. Following ADP activation, clopidogrel treatment decreased platelet activation (1% activation) compared to control (3% activation) and aspirin-treated (5% activation) mice with no additional decrease in dual treatment (1% activation); unexpectedly, aspirin treatment alone or dual therapy decreased platelet aggregation with ADP (15% and 17% aggregated, respectively) compared to control and clopidogrel (23% and 27% aggregated, respectively). Finally, after arachidonic acid activation, aspirin treatment significantly decreased activation (7% activated) compared to control (20% activated), while clopidogrel decreased activation more than that of aspirin (3% activated), with no additional decrease from dual therapy (3% activated). Though not significant, aspirin treatment (1.4% aggregation) and dual therapy (1.2%) demonstrated a trend with decreased platelet aggregation following arachidonic acid activation compared to control (1.8% aggregation) and clopidogrel-treated platelets (3.2%). These data presented here show a novel application of platelet activation and aggregation of GFP and RFP platelet populations in a mouse model following aspirin and/or clopidogrel treatment. This model can be applied generally to other drugs and disease models for platelet dysfunction using additional markers for platelet activation. Additionally, these experiments can be used to further investigate the function of transfused murine platelets.

Effect of evolocumab on platelet function in patients with acute coronary syndromes. An analysis of the randomized, double-blind, placebo-controlled EVOPACS Trial

Abstract Background Proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibitors substantially lower levels of low-density lipoprotein cholesterol (LDL-C) and favourably impact clinical outcomes in patients with acute coronary syndromes (ACS). Although PCSK9 inhibitors have been suggested in ex vivo studies to suppress platelet activation, the impact of evolocumab on platelet function in ACS patients receiving potent antiplatelet therapy remains largely unknown. Methods The EVOPACS trial was a randomized, double-blind, placebo-controlled trial assessing the use of evolocumab initiated in the acute phase of ACS. Patients presenting with ST-elevation myocardial infarction (STEMI) or Non-ST-elevation ACS (NSTE-ACS) were randomly allocated in a 1:1 ratio to either evolocumab 420mg SC or matching placebo at baseline (during hospitalization for index ACS event) and after 4 weeks. All patients received atorvastatin 40mg daily. Dual antiplatelet therapy consisting of aspirin and a potent P2Y12 inhibitor was initiated according to STEMI / NSTE-ACS guidelines, unless a less potent treatment was indicated (e.g. due to concomitant use of oral anticoagulation). Platelet function was measured at baseline (during index hospitalization) and after 8 weeks, using the Multiplate analyser. The main outcome for this analysis was the change from baseline to 8 weeks in adenosine diphosphate (ADP)-induced platelet aggregation. Results Serial platelet function measurements were available in 260 of all 308 enrolled patients, 124 in the evolocumab and 136 in the placebo group. Mean age was 60.2±11.0 years and 83% were men. Baseline clinical characteristics were well balanced between the two treatment groups. At discharge, the proportion of patients treated with ticagrelor, prasugrel, and clopidogrel was 73%, 11%, and 9% respectively, without differences between groups. Evolocumab significantly reduced LDL-C compared to placebo at 8 weeks (difference in mean percentage change -40.75%, 95% CI -45.39 to -36.11; p<0.001). Change in ADP-induced platelet aggregation between baseline and 8-week follow-up was -5.6±23.5 U (from 24.1±19.8 to 29.7±25.6 U) in the evolocumab group vs. -1.7±198.7 U (from 23.0±14.7 U to 24.7±19.1 U) in the placebo group (p=0.13). Similar results were found in a subgroup of 189 patients (86 in the evolocumab and 103 in the placebo group) treated with ticagrelor at discharge, showing a change in ADP-induced platelet aggregation of -2.7±19.5 U with evolocumab vs. -2.4±19.9 U with placebo (p=0.80). Conclusions In this randomized trial of ACS patients largely treated with a potent P2Y12 inhibitor, evolocumab initiated during index hospitalization on top of high-intensity statin therapy had no measurable effect on ADP-induced platelet aggregation. These findings suggest that the early therapeutic benefit of PCSK9 inhibitors in ACS patients is likely related to plaque regression/stabilization rather than changes in platelet activity.

Sex-related differences in the response of anti-platelet drug therapies targeting purinergic signaling pathways in sepsis.

Sepsis, a complex clinical syndrome resulting from a serious infection, is a major healthcare problem associated with high mortality. Sex-related differences in the immune response to sepsis have been proposed but the mechanism is still unknown. Purinergic signaling is a sex-specific regulatory mechanism in immune cell physiology. Our studies have shown that blocking the ADP-receptor P2Y12 but not P2Y1 receptor was protective in male mice during sepsis, but not female. We now hypothesize that there are sex-related differences in modulating P2Y12 or P2Y1 signaling pathways during sepsis. Male and female wild-type (WT), P2Y12 knock-out (KO), and P2Y1 KO mice underwent sham surgery or cecal ligation and puncture (CLP) to induce sepsis. The P2Y12 antagonist ticagrelor or the P2Y1 antagonist MRS2279 were administered intra-peritoneally after surgery to septic male and female mice. Blood, lungs and kidneys were collected 24 hours post-surgery. Sepsis-induced changes in platelet activation, secretion and platelet interaction with immune cells were measured by flow cytometry. Neutrophil infiltration in the lung and kidney was determined by a myeloperoxidase (MPO) colorimetric assay kit. Sepsis-induced platelet activation, secretion and aggregate formation were reduced in male CLP P2Y12 KO and in female CLP P2Y1 KO mice compared with their CLP WT counterpart. Sepsis-induced MPO activity was reduced in male CLP P2Y12 KO and CLP P2Y1 KO female mice. CLP males treated with ticagrelor or MRS2279 showed a decrease in sepsis-induced MPO levels in lung and kidneys, aggregate formation, and platelet activation as compared to untreated male CLP mice. There were no differences in platelet activation, aggregate formation, and neutrophil infiltration in lung and kidney between female CLP mice and female CLP mice treated with ticagrelor or MRS2279. In human T lymphocytes, blocking P2Y1 or P2Y12 alters cell growth and secretion in vitro in a sex-dependent manner, supporting the data obtained in mice. In conclusion, targeting purinergic signaling represents a promising therapy for sepsis but drug targeting purinergic signaling is sex-specific and needs to be investigated to determine sex-related targeted therapies in sepsis.

Flow Cytometric Assessment of Changes in Platelet Reactivity after Acute Coronary Syndrome: A Systematic Review.

Increased platelet activity is an important predictor for recurrent cardiovascular events in patients with acute coronary syndromes (ACS). Flow cytometry is an advanced method for evaluation of platelet activity. We aimed to summarize the current literature on dynamic changes in platelet activity analyzed by flow cytometry in patients with ACS. Employing the guidelines of Preferred Report Items for Systematic Reviews and Meta-Analyses (PRISMA), we searched PubMed and Embase on October 26, 2021, and identified studies measuring platelet activity with flow cytometry in ACS patients in the acute phase (baseline) and at follow-up in a more stable phase. In the 12 included studies, fibrinogen receptor, α-granule secretion, platelet reactivity index, monocyte-platelet aggregates, neutrophil-platelet aggregates, and reticulated platelets were measured. The fibrinogen receptor and α-granule secretion were either unchanged or lower during follow-up measurements than in the acute phase. Platelet reactivity index showed inconsistent results. Values of monocyte-platelet aggregates and neutrophil-platelet aggregates were lower at follow-up than at baseline (p-values <0.05). Reticulated platelets were either unchanged (p-value >0.64) or lower at 1 to 2 months follow-up (p-value 0.04), and also lower at 5 months to 1-year follow-up (p-value >0.005) compared with baseline. Overall, flow cytometric analyses of platelet function in ACS patients showed that platelet activity was lower at follow-up than at baseline. However, in some patients, platelet activity remained unchanged from baseline to follow-up, possibly indicating a sustained high platelet activity that may increase the risk of recurrent cardiovascular events.

Platelet Activation Is Not Always Associated With Platelet-Related Plasma microRNA Abundance - Results From a Randomized Controlled Trial of Periodontal Patients.

Platelets are involved in a variety of diseases, making their adequate functional assessment is essential. However, due to their easily activatable nature this has some methodological pitfalls. Therefore, the availability of stable, easily measurable surrogate markers would be beneficial. In this regard, some evidence suggests that certain microRNAs (miRNAs) circulating in plasma might be useful. We aimed to corroborate their suitability by analyzing plasma samples obtained in a randomized controlled trial, which assessed the effects of periodontal treatment on platelet function. We hypothesized that miRNA levels mirror changes of platelet activation and -function. Both platelet function and miRNA abundance were quantified using state-of-the-art flow cytometry and qPCR methods. The following miRNAs were quantified: 223-3p, 150-5p, 197-3p, 23a-3p, 126-3p, 24-3p, 21-5p, 27b-3p, 33a-5p, 320a, 191-5p, 28-3p, 451a, 29b-3p, and 1-3p. However, periodontal treatment did not affect the abundance of any investigated miRNAs to a relevant extent. Platelet activation and reactivity indices did neither correlate with any tested miRNA at baseline, nor after the treatment period. In addition, there was no evidence that investigated miRNAs were released by platelets, as suggested previously. In conclusion, our data suggest that in patients suffering from periodontal disease the investigated miRNAs are unlikely to be suitable biomarkers for platelet function. Our data aim to raise awareness that previously determined platelet activation dependent circulating miRNAs are not suitable as platelet biomarkers in all cohorts.

Exposure to fine particulate matter promotes platelet activation and thrombosis via obesity-related inflammation

Short-term exposure to fine particulate matter (PM2.5) increases thrombotic risk particularly in obese individuals, but the underlying mechanisms remain unclear. This study aims to compare the effects of PM2.5 on inflammation and platelet activation in obese versus normal-weight adults, and investigate potential causal pathways. We conducted a panel study measuring blood markers in 44 obese and 53 normal-weight adults on 3 separate occasions in 2017–2018. Associations between PM2.5/black carbon (BC) and biomarkers were estimated using mixed-effect models. An interaction analysis compared PM2.5/BC-related effects between subgroups. Biomarker combinations and mediation analysis were performed to elucidate the biological pathways. There was a significant “low-high-low” trend of PM2.5 levels across the 3 study periods. Increases in pro-inflammatory cytokines and changes of platelet activation and aggregation markers were associated with PM2.5/BC in obese subgroup only. Among obese subjects, the combination of pro-inflammatory cytokines and that of platelet markers increased 26.8% (95% CI: 16.0%, 37.9%) and 14.7% (95% CI: 1.9%, 27.0%) per IQR increase in PM2.5 over 5-day and 7-day averages. Inflammation mediated 24.5% of the pathways through which PM2.5 promoted platelet activation. This study suggested obese people are susceptible to pro-thrombotic impacts of PM2.5 exposures. PM2.5 may aggravate thrombosis through obesity-related inflammation.

Post-TBI splenectomy may exacerbate coagulopathy and platelet activation in a murine model

IntroductionTraumatic brain injury (TBI) induces acute hypocoagulability, subacute hypercoagulability, and persistently elevated risk for thromboembolic events. Splenectomy is associated with increased mortality in patients with moderate or severe TBI. We hypothesized that the adverse effects of splenectomy in TBI patients may be secondary to the exacerbation of pathologic coagulation and platelet activation changes. MethodsAn established murine weight-drop TBI model was utilized and a splenectomy was performed immediately following TBI. Sham as well as TBI and splenectomy alone mice were used as injury controls. Mice were sacrificed for blood draws at 1, 6, and 24 h, as well as 7 days post-TBI. Viscoelastic coagulation parameters were assessed by rotational thromboelastometry (ROTEM) and platelet activation was measured by expression of P-selectin via flow cytometry analysis of platelet rich plasma samples. ResultsAt 6 h following injury, TBI/splenectomy demonstrated hypocoagulability with prolonged clot formation time (CFT) compared to TBI alone. By 24 h following injury, TBI/splenectomy and splenectomy mice were hypercoagulable with a shorter CFT, a higher alpha angle, and increased MCF, despite a lower percentage of platelet contribution to clot compared to TBI alone. However, only the TBI/splenectomy continued to display this hypercoagulable state at 7 days. While TBI/splenectomy had greater P-selectin expression at 1-h post-injury, TBI alone had significantly greater P-selectin expression at 24 h post-injury compared to TBI/splenectomy. Interestingly, P-selectin expression remained elevated only in TBI/splenectomy at 7 days post-injury. ConclusionSplenectomy following TBI exacerbates changes in the post-injury coagulation state. The combination of TBI and splenectomy induces an acute hypocoagulable state that could contribute to an increase in intracranial bleeding. Subacutely, the addition of splenectomy to TBI exacerbates post-injury hypercoagulability and induces persistent platelet activation. These polytrauma effects on coagulation may contribute to the increased mortality observed in patients with combined brain and splenic injuries.

PEAR1 polymorphisms as a prognostic factor in hemostasis and cardiovascular diseases.

Platelet Endothelial Aggregation Receptor (PEAR1), as a platelet receptor, plays a vital role in hemostasis. This receptor, by its extracellular part, causes platelet adhesion and consequently initiates platelet aggregation. Dysfunction of PEAR1 can disrupt platelet aggregation in patients with cardiovascular diseases (CVDs). The content used in this paper has been taken from English language articles (2005-2020) retrieved from Pubmed database and Google scholar search engine using "Cardiovascular Disease", "PEAR1", "Polymorphism", and "Platelet Aggregation" keywords. Some PEAR1 polymorphisms can disrupt homeostasis and interfere with the function mechanism of cardiac drugs. Since polymorphisms in this gene affect platelet function and the platelet aggregation process, PEAR1 could be further studied in the future as an essential factor in controlling the treatment process of patients with cardiovascular diseases. PEAR1 polymorphisms through disruption of the platelet aggregation process can be a risk factor in patients with CVDs. Therefore, controlling patients through genetic testing and the evaluation of PEAR1 polymorphisms can help improve the treatment process of patients. According to the studies on the PEAR1 gene and the effect of different polymorphisms on some crucial issues in CVDs patients (changes in platelet activity), it is clear that if there is a significant relationship between polymorphisms and CVDs, they can be used as prognostic and diagnostic markers. This study aims to evaluate the prognosis and drug treatment of the PEAR1 gene in CVDs patients.

PCSK9 Regulates Nox2-Mediated Platelet Activation via CD36 Receptor in Patients with Atrial Fibrillation

Background: High levels of proprotein convertase subtilisin/kexin 9 (PCSK9) is predictive of cardiovascular events (CVEs) in atrial fibrillation (AF). We hypothesized that PCSK9 may directly induce platelet activation (PA). Methods: We measured platelet aggregation, recruitment, Thromboxane B2 (TxB2) formation and soluble P-selectin levels as markers of PA and soluble Nox2-derived peptide (sNox2-dp), H2O2, isoprostanes and oxidized Low-Density-Lipoprotein (oxLDL) to analyze oxidative stress (OS) in 88 patients having PCSK9 values < (n = 44) or > (n = 44) 1.2 ng/mL, balanced for age, sex and cardiovascular risk factors. Furthermore, we investigated if normal (n = 5) platelets incubated with PCSK9 (1.0–2.0 ng/mL) alone or with LDL (50 µg/mL) displayed changes of PA, OS and down-stream signaling. Results: PA and OS markers were significantly higher in patients with PCSK9 levels > 1.2 ng/mL compared to those with values < 1.2 ng/mL (p < 0.001). Levels of PCSK9 significantly correlated with markers of PA and OS. Platelets incubation with PCSK9 increased PA, OS and p38, p47 and Phospholipase A2 (PLA2) phosphorylation. These changes were amplified by adding LDL and blunted by CD36 or Nox2 inhibitors. Co-immunoprecipitation analysis revealed an immune complex of PCSK9 with CD36. Conclusions: We provide the first evidence that PCSK9, at concentration found in the circulation of AF patients, directly interacts with platelets via CD36 receptor and activating Nox2: this effect is amplified in presence of LDL.

Platelet-Leukocyte Interplay in Cancer Development and Progression.

Beyond their crucial role in hemostasis, platelets are increasingly recognized as regulators of inflammation. Via modulation of the immune system by direct and indirect interactions with leukocytes, platelets regulate several aspects of tumor-associated pathology. They influence inflammatory processes in cancer at various stages: platelets alter the activation status of the endothelium, recruit leukocytes to tumor sites and attune the inflammatory milieu at sites of primary and metastatic tumors. Patients with cancer show systemic changes of platelet activation. Tumor-associated platelet activation facilitates initiation of the coagulation cascade and constitutes a significant risk for thrombosis. Tumor-activated platelets further contribute to cancer progression by promoting critical processes such as angiogenesis and metastasis. Platelets modulate innate leukocyte effector functions such as antigen presentation by dendritic cells, monocyte recruitment and differentiation or neutrophil extracellular trap formation, which sculpture immune responses but also promote thrombosis and metastasis. On the other hand, responses of the adaptive immune system are also regulated by platelets. They are also involved in T-helper cell 17 differentiation, which represents a double-edged sword in cancer progression, as these cells propagate angiogenesis and immunosuppressive activities but are also involved in recruiting immune cells into tumors and stimulating effector CD8+ T cells. Moreover, platelets fine-tune tumor surveillance processes by modulating natural killer cell-mediated cancer cell recognition and effector functions. This review aims at summarizing the role of platelet-leukocyte interactions in the development and progression of cancer and puts its focus on cancer-related alterations of platelet and leukocyte functions and their impact on cancer pathology.

Functional properties of platelets in piglets when changing methods of nutrition

A functionally significant element in ensuring homeostasis of the internal environment of an animal organism is platelets. The state of their activity greatly influences the rheology of blood in small vessels and, thus, the metabolism in tissues. It becomes clear that the process of growth and development of piglets at any age substantially depends on the level of their activity. In this regard, the assessment of agerelated changes in platelet activity in piglets during their early ontogenesis is of great importance. In the work performed, it was found that in piglets during the dairy and vegetable nutrition phase there is an increase in the adhesive, aggregation and secretory properties of platelets. The leading cause of these changes can be the enhancement of receptor processes and activation of the work of post-receptor mechanisms of information transfer in platelets. This is noted in piglets during the milk-plant nutrition phase simultaneously for both strong and weak aggregation inducers. The growth of platelet activity in piglets found during the observation period apparently was due to the intensification of thromboxane generation in platelets as a result of the increased activity of their cyclooxygenase and thromboxane synthetase, as well as an increase in the secretion of ADP molecules from them. The increase in the severity of the hemostatic properties of platelets in piglets during the milk-plant nutrition phase seems to be a serious regulator of their microcirculation and metabolism processes in any environmental conditions.

P1786 Impact of exercise on platelet activity in patients with significant ischemic mitral regurgitation qualified for cardiosurgery treatment

Abstract Purpose Platelet activity is believed to play an important role in patients (pts)after myocardial infarction (MI). This study sought to examine a change in platelet activity after 2D echocardiography semi-supine exercise test (ExE) in pts with at least moderate ischemic mitral regurgitation (IMR) qualified for cardiosurgical treatment - coronary artery bypass grafting alone (CABG) or CABG with mitral reconstruction (CABGmr). Methods We collected venous blood samples from 70 patients (M 41, 66 ± 9 years) with at least moderate IMR, 3-24 weeks after MI. Platelet activity was estimated with the use of rapid, point-of-care platelet analyzer as the time (in seconds) for flowing whole blood to occlude a collagen-epinephrine ring, with shorter closure times, indicating greater activity (CEPI-CT- for assessing the thromboxane A2 –dependent activation pathway). CEPI-CT was evaluated before (CEPI-CT rest) and 20 min after ExE (CEPI-CT ExE). All the patients received 75-100 mg of aspirin daily (without clopidogrel treatment). Effective regurgitation orifice (ERO) was used for quantitative IMR assessment (moderate ≥10-20, severe ≥ 20 mm²). All the pts were referred for CABG (multivessel coronary disease, ejection fraction (EF) 42,6 ± 10,1%, wall motion score index (WMSI) 1.65 ± 0.36). Results We observed a significant decrease in CEPI–CT’s in all analyzed group (rest vs exe: 202 sec vs 124 sec, respectively; p &amp;lt; 0,0001) and in pts with moderate IMR as well as in pts with severe IMR, despite aspirin treatment. The results of the analysis of CEPI-CT and IMR size at rest and after ExE in both groups of pts are shown in Table 1. A negative correlation was found between CEPI-CT ExE and ERO ExE (r=-0,752, p = 0,0042). Conclusions 1. Significant increase in platelet activity after exercise was observed in patients after MI with IMR qualified for cardiosurgical treatment. 2. The increase in platelet activity after exercise is correlated with exercise changes of IMR size. 3. Aspirin may have a limited antithrombotic effect after physical exercise in patients after MI with significant IMR. Table 1. CEPI-CT rest CEPI-CT ExE p ERO rest ERO exe p All 202 124 &amp;lt; 0,0001 17 22 &amp;lt;0,001 Moderate IMR 201 124 0,0001 15 20 &amp;lt; 0,0001 Severe IMR 235,5 123,5 &amp;lt; 0,0001 25 31 &amp;lt; 0,0001

Functional features of hemostasis in physically weakened newborn piglets and calves

The normal state of hematological parameters in mammals at the beginning of ontogenesis is rightfully considered to be functionally very significant for the development of early ontogenesis in them. Hemostasis is considered to be very significant for vital functions and vulnerable blood systems. Its condition affects trophism due to the effect on the state of microcirculation, which is very important at the beginning of ontogenesis. In the neonatal phase, calves and piglets develop the formation of hemostasis and therefore, during these periods it can be easily disrupted. The appearance of hemostatic dysfunctions in the body of young animals, despite its importance, is still very poorly studied. This circumstance was the motivation for the study of the activity of the hemostatic system in calves and piglets that were born physically weakened. It was found that immediately after birth in weakened calves and piglets, there are physiologically unfavorable changes in platelet activity, vascular hemostasis and blood coagulation. This provides them with the appearance of excessive platelet activity and blood clotting with a significant weakening of vascular hemostasis. All these disorders lead to negative changes in their internal organs and create their serious dysfunctions. The development of disorders in the mechanisms of hemostasis in physically weakened calves and piglets during the neonatal phase is a serious mechanism for weakening their adaptation to the environment and slowing the growth and development of animals.

Prognostic Importance of Thrombocyte Indices in Children with Carbon Monoxide Poisoning

\n Prognostic importance of thrombocyte indices in children with carbon monoxide poisoning \n \n Abstract \n \n Objective: Carbon monoxide (CO) poisoning is still being a major cause of morbidity and mortality all over the world. Although the role of platelet functions in CO poisoning is not clear, increased platelet activation and aggregation had been reported previously. Increased thrombotic tendency due to endothelial damage, increased platelet stickiness, and alterations in the fibrinolytic pathway occurs in CO poisoning. The aim of our study was to determine whether platelet indices provide clinical benefit or not in children with CO poisoning. \n \n Materials and Methods: We retrospectively reviwed the records of children with the diagnosis of CO poisoning who admitted to the pediatric emergency departments of Konya Beyhekim State Hospital and Necmettin Erbakan University Meram Medical Faculty. A total of 92 children with CO poisoning and 62 age- and gender-matched healthy controls were included in the study. \n \n Results: While mean platelet volume (MPV) and platelet distribution width (PDW) levels were significantly higher (9,34±0,55 vs 9,78±0,97fL, p=0.001 ; 11,46±2,64 vs 10,57±1,41, retrospectively, p=0.007), platelet count and plateletcrit (PCT) (324,05±82,07 vs 357,27±89,70 x109 p=0.015 ; 0,31±0,06 vs 0,33±0,07, retrospectively, p=0.039) were significantly lower in patients with CO poisoning. MPV levels were also significantly higher in patients with a carboxy hemoglobin (COHb) level higher than 20, when compared with COHb levels between 10-20 (9,40±0,84 vs 10,08±1,22 fL, p=0.003). \n \n Conclusion: Our results showed that platelet indices MPV and PDW are markedly elevated in patients with CO poisoning while platelet count and PCT were decreased. MPV and PDW levels, which reflect the changes in platelet activation and function, may predict the development of especially thromboembolic complications in the course of CO poisoning. MPV and PDW levels may be useful in prognostic estimation of CO poisoning. \n \n Keywords: carbon monoxide; children; platelet indices; poisoning \n \n \n

Platelet activation in diabetic mice models: the role of vascular endothelial cell-derived protein disulfide isomerase-mediated GP IIb/IIIa receptor activation.

GP IIb/IIIa receptor activation plays an important role in thrombosis. The mechanism of early activation of GP IIb/IIIa receptors in diabetic conditions remains unknown. The purpose of this study was to investigate the release of Endothelial microparticle (EMP)-associated protein disulfide isomerase (PDI) after endothelial cell injury induced in diabetes and the changes in platelet activation. We produced an animal model of type 2 diabetes mellitus using ApoE−/− mice. Normal ApoE−/− and diabetic mice were allocated to four groups (n = 15): normal diet, normal diet plus rutin, diabetic, and diabetes plus rutin. The EMP-PDI content and GP IIb/IIIa expression of mice platelets were determined. In addition, EMPs obtained from the four groups were pretreated with the PDI inhibitor rutin; then, their effects on the platelets of normal C57 mice were characterized. Compared with the normal diet group, the diabetic group had significantly increased plasma EMP-PDI content and accelerated platelet activation by increased GP IIb/IIIa expression. In conclusion, EMP-PDI promotes early platelet activation through glycoprotein (GP) IIb/IIIa receptors present on platelet surface in the diabetic state. However, this process could be partially suppressed by the administration of rutin.

Changes in platelet activity during perioperative period in patients with diabetes mellitus

Objective To evaluate the changes in the platelet activity during the perioperative period in the patients with diabetes mellitus. Methods Twenty patients with type 2 diabetes mellitus (group DM) and 22 patients with no diabetes mellitus (group NDM) of both sexes, with body mass index of 20-30 kg/m2, of American Society of Anesthesiologists physical statusⅠ or Ⅱ, scheduled for elective spinal or hip joint surgery, were enrolled in the study.Blood samples were collected on admission to the operating room (T0), immediately after recovery from general anesthesia (T1) and at 24 h after surgery (T2). The expression of platelet P-selectin (CD62P) was measured by flow cytometry, and the positive rate was calculated.The platelet count (PLT) and mean platelet volume (MPV) were determined using an automatic blood cell analyzer. Results Compared with group NDM, the positive rate of CD62P cells at T0, 1 and PLT at T0 were significantly increased (P 0.05). There was no significant difference in the positive rate of CD62P cells, PLT or MPV between each time point within each group (P>0.05). Conclusion Platelets remain hyperactivity that is not further enhanced by diabetes mellitus during the perioperative period in the patients. Key words: Diabetes mellitus; Intraoperative period; Blood platelets

The impact of vitamin C on the relationship among inflammation, lipid peroxidation and platelet activation during analgesic nephropathy in rats.

Oxidative stress and inflammation are involved in the pathogenesis of paracetamol-induced renal damage. This study examines the relationship between 8-iso-prostaglandin F2α (8-iso-PGF2α) and platelet activation as well as the relative contribution of the pro-inflammatory markers interleukin (IL)-1β and tumor necrosis factor-α (TNF-α) in enhanced 8-iso-PGF2α biosynthesis, as a complementary onset during analgesic nephropathy induced by chronic treatment with paracetamol. The protective effects of vitamin C on the aforementioned settings are also investigated. Analgesic nephropathy was induced in Wistar rats. Renal function markers and the activity of antioxidant enzymes were determined spectrophotometrically. Immunoassays were used to measure the pro-inflammatory markers and the markers of lipid peroxidation and platelet activation. The chronic treatment with paracetamol led to renal dysfunction, represented by the elevation of plasma urea and creatinine and the decline in the enzymatic antioxidant status, but did not cause a significant increase in TNF-α and IL-1β. The paracetamol-induced lipid peroxidation and enhanced production of 8-iso-PGF2α was not sufficient to cause changes in platelet activation represented by the level of 11-dehydro thromboxane B2. Our results suggest that oxidative stress cannot circumvent the need of stimulation by circulatory cytokines in order to induce inflammatory response and changes in platelet activation during analgesic nephropathy. Vitamin C proved to be beneficial in restoring the renal function markers to normal, increasing the renal enzymatic antioxidant potential, inhibiting lipid peroxidation, and lowering cytokine production and 11-dehydro thromboxane B2 excretion. The observed effects of vitamin C offer support for its potential use as protective treatment in cases of chronic paracetamol overdose.

Abstract 150: Hypoxia and Ischemia Promote a Maladaptive Platelet Phenotype

Background: An understudied area of platelet biology is that ischemic diseases such as peripheral vascular disease (PVD) can alter the platelet phenotype, and this may lead to unpredictable effects of anti-platelet agents. Methods: Isolated murine and human platelets were exposed to a reduced oxygen environment (hypoxia chamber, 5% O 2 ). Platelet activation was assessed as mean fluorescence intensity (MFI) of surface p-selectin by flow cytometry, and morphologic changes by confocal microscopy. A murine model of critical limb ischemia (CLI) was used to assess changes in platelet activity in wild-type (WT) and in platelet ERK5 -/- mice. Limb blood flow was assessed by laser Doppler imaging. Hypoxia was additionally examined in mice by unilateral pneumonectomy. Western blotting was used to assess protein expression. Results: In human platelets ex vivo at 5% O 2 , platelet activation increased to 5076 MFI±409 from 3548 MFI±187 at 20% O 2 (P=0.005). With the ERK5 inhibitor XMD892, platelet activation at 5% O 2 decreased from 5677 MFI±312 to 3175 MFI±247 (p=0.001). In a murine model of PVD, enhanced platelet activation was sustained for weeks vs. sham, and limb blood flow in platelet ERK5 -/- mice one week after ischemia was augmented by 26%: 0.61±0.03 vs 0.87±0.03 vs, WT mice (p=0.0002), with 50% less platelet activity. In mice with unilateral pneumonectomy vs. sham surgery, observed hypoxia was accompanied by increased platelet ERK5 activation coincident with enhanced platelet activation. In patients with CLI, augmented platelet activation in spite of aspirin treatment was observed via the thromboxane and the PAR1 receptors, and especially via P2Y 12 receptor. Conclusions: Hypoxia and ischemic tissue injury, such as with critical limb ischemia, change the phenotype of the platelet, promoting a pro-thrombotic state that is partly dependent on platelet ERK5. Platelet phenotype and function should be better characterized in ischemic diseases.