ALBERT

Description: Abstract 1141 Lacunar stroke accounts for 25% of all ischemic strokes and is characterized by small infarcts located in the deep white matter, basal ganglia or pons resulting from the occlusion of deep branch arteries. In the last years, evidences about the role of endothelial dysfunction in lacunar stroke also named cerebral small vessel disease, have emerged. However platelet activation and aggregation are of major importance in acute thrombosis of atherosclerotic arteries, but their role and that of thrombus formation at sites of arteriolar injury as a precipitating event of lacunar stroke remains unclear. The aim of the present study was to determine platelet and endothelial activation in consecutive patients with recent lacunar stroke in comparison to population-based control subjects matched for age, sex and vascular risk factors. Methods— Platelet activation markers include activated glycoprotein (GP) IIbIIIa, P-selectin expression, platelet microparticles determined by flow cytometry, shear-induced platelet aggregation (SIPA), measured in a Sipagreg device that reproduces rheological conditions of stenotic or small arteries. We also studied endothelial activation markers including von Willebrand factor antigen (vWF), homocystein and high sensitivity C-reactive Protein (hsCRP). All these parameters were measured in 74 consecutive patients with recent lacunar stroke, in whom detectable large artery atherosclerosis or cardiac embolism have been ruled out, and in 74 population-based controls with no stroke history, matched for age, sex, hypertension, and diabetes. All patients were treated by anti-aggregant therapy, 15 (20%) were treated by clopidogrel (75 mg/d) and the remaining 59 cases received only acetylsalicylic acid (75 to 160 mg/d). Repeated blood samples were collected at one- and three-months after symptom onset. Results— One month after symptom onset, patients had similar levels of platelet activation compared to matched controls (p〉0.40 for all-comparisons), including platelet membrane P-selectin with a median value of 1.3% and an interquartile range (IQR) (0.7–2.8) in patients and 1.1% (0.6–2.2) in controls, p=0.37), platelet activated GPIIbIIIa reached 2.9% (0.9–6.7) in patients and 2.8% (0.7–10.2) in controls, p=0.94. Platelet-derived microparticles were 5067/μL of blood (3720–7490) and 5035 (3530–8150) p=0.63l, in patients and controls, respectively. SIPA values at 4000 sec-1 were in the normal range in patients and controls, with no difference between the two groups: the median value of platelet aggregation was 22% (12–37%) in cases and 23% (7–45%) in controls (p=0.96), respectively. Endothelial activation parameters were increased in patients in comparison to controls (vWF p=0.002 and homocystein/creatinemia p=0.025) The median value of vWF antigen was 142% (114–182%) in cases and 122% (97–150%) in controls (p=0.002). The median value of ratio homocysteinemia/creatinemia was 0.151 (0.119–0.187) in cases and 0.134 (0.111–0.165) in controls (p=0.025). The median value of hsCRP was 2.7 mmol/l (1.5–6.0) in cases and 2.0 mmol/l (1.0–3.4) in controls (p=0.059). Level of hsCRP was slightly increased in patients compared to controls (p=0.059). At 3 months, significant decrease in vWF and hsCRP levels (median change in vWF=10%, p=0.004; median change in hsCRP=0.4 mg/L, p=0.02) was detected in patients. Homocystein level and all platelet parameters remained unchanged at this time compared to 1 month. Conclusions— Our results confirm that endothelial dysfunction is a major feature of lacunar stroke. One explanation could be an endothelial response to an acute systemic aggression by an unidentified agent. In contrast, chronic platelet activation, when compared to controls matched for age, sex and vascular risk factors, did not seem to play a central role in the pathophysiology of lacunar stroke. Disclosures: No relevant conflicts of interest to declare.

Description: Myeloproliferative neoplasms (MPN) are associated with an increased risk of arterial and venous thrombosis. Besides clinical factors (previous history of thrombosis, age 〉 60 years, cardiovascular risk factors), procoagulant phenotype, proadhesive properties of the endothelium and secretion of inflammatory cytokines, contribute to the pathogenesis of thrombosis in MPN. Pegylated interferon alpha (IFN) has obviously a great therapeutic efficacy on hematopoietic cell proliferation but its other biological effects particularly on hemostasis and inflammation have not been determined. The aim of the study was to determine whether IFN impacts the biological profile including endothelial and platelet markers compared to MPN patients treated by hydroxyurea (HU) and non-treated (NT) patients. Patients with polycythemia vera (PV) or essential thrombocythemia (ET) treated with IFN or HU or without any cytostatic drug were included. Platelet membrane glycoproteins and platelet activation were measured by flow cytometry. We evaluated shear-induced platelet aggregation (SIPA) at 4000 sec-1 that mimics stenotic arteries. We measured coagulation proteins by their activity and endothelial parameters (von Willebrand factor (VWF) antigen and activity, soluble thrombomodulin). Thrombin generation was measured in platelet poor plasma and platelet rich plasma. Platelet accumulation on immobilized collagen was measured using whole blood perfused at 1500 sec-1 in flow chambers. Statistical analysis was performed by ANOVA to compare the 3 groups of patients (p

Description: Abstract 1087 Heparin-induced thrombocytopenia (HIT) is a severe complication of heparin treatments associated with a high risk of venous and arterial thrombosis. HIT is often difficult to diagnose in clinical practice since most patients present several potential causes of thrombocytopenia. A scoring system named 4T's and based on four criteria (i.e. Thrombocytopenia, Timing of platelet count fall, Thrombosis or other sequelae, oTher cause of thrombocytopenia) is currently used to evaluate the probability of HIT before laboratory testing. Recently, a rapid lateral flow immunoassay (LFIA) based on the use of PF4/polyanion complexes linked to biotin as antigens and of gold nanoparticles coated with antibodies specific to biotin has been developed (Stic Expert HIT®, Stago Asnières France). This test is IgG-specific since gold particles are immobilized on the nitrocellulose strip by anti-human IgG and become visible as a colored line when IgG antibodies to PF4/heparin complexes are present in the patient sample. The aim of our study was to evaluate the performances of this rapid assay in a large prospective cohort of patients with suspected HIT. In addition, we compared the results obtained with serum and plasma samples and evaluated the inter-reader reproducibility of the assay. Patients and methods: Two hundred and sixteen consecutive patients were enrolled from February to June 2012 in 10 French centers. The pretest probability of HIT was evaluated using the 4T's score blind to antibody test results. The Stic Expert HIT® was performed in each center on plasma and serum. IgG-specific ELISA (Asserachrom HPIA IgG®) and serotonin release assay were also performed in the coordinating center (Tours). HIT was confirmed only when both SRA and IgG specific H/PF4 ELISA were positive. Results: Definite HIT was diagnosed in 24 patients and the incidence of HIT therefore equaled 11.1% in our cohort. The risk of HIT was evaluated as low (LR), intermediate (IR) or high (HR) by the 4T's in 27.8%, 63.2% and 9.0% of patients enrolled, respectively. The negative predictive value of (NPV) of the 4T's was 96.6% since definite HIT was diagnosed in 2 of the 59 LR patients. Although interpretation of LFIA results is visual, the inter-reader reproducibility was excellent (Kappa test ratio higher than 0.9) whether the test was performed with plasma or serum. When performed on plasma samples, LFIA was negative in 159 patients without HIT (NPV 99.4%,) and the negative likelihood ratio (LR-) was 0.05. Results obtained with serum samples were similar with NPV and LR- values of 100% and less than 0.01, respectively. On the other hand, LFIA was positive in 54 patients, including 23 with definite HIT (positive predictive value = 42.6%), with a positive LR value of 5.87. The pre-test probability of HIT was 3.4%, 9.7%, and 36.8% in the patients classified with the 4Ts score as having low, intermediate and high risk of HIT, respectively. The post-test probability of HIT was then calculated according to the Bayes theorem and using the LR values of LFIA obtained. The probability of HIT decreased dramatically to 0.5% in IR patients when the test result was negative. Alternatively, this probability increased to 38% only when LFIA was positive. Discussion and Conclusion: One major advantage of the LFIA Stic Expert HIT® is the possibility of obtaining a result in less than 40 minutes and a strategy based on results of both this assay and the 4Ts score can therefore be proposed for the management of patients with suspected HIT in emergency conditions. The Stic Expert HIT® can be performed both on serum and plasma and a negative result is able to confidently rule out the diagnosis of HIT since NPV and LR- values are excellent (〉99% and 〈 0.1, respectively). These performances are particularly useful in LR or IR patients, for whom heparin treatment can be continued safely if the Stic Expert HIT® is negative. Disclosures: No relevant conflicts of interest to declare.

Description: Background: Myeloproliferative neoplasms (MPN) are associated with an increased risk of arterial and venous thrombosis with an annual incidence varying from 1.1% to 6.6% (Patrono, Blood 2013). Comparable to that recommended in non-MPN high-risk patients, low-dose aspirin (81-100 mg/day) is recommended in primary and secondary prophylaxis of thrombosis in polycythemia vera and essential thrombocytemia (Tefferi, Am J Hematol 2012). However, the daily dose of aspirin to optimize efficacy and safety for MPN patients has never been challenged. Aim of the study: Determine the optimal dose of aspirin to achieve biological efficacy in Philadelphia-negative MPN patients. Methods: Patients with Philadelphia-negative MPN who were treated with low-dose aspirin 75 or 100 mg/day were enrolled in this observational study. Patients without any cytostatic drug and patients treated with either pegylated interferon alpha or hydroxyurea were eligible. Major exclusion criteria included inability to adhere to aspirin therapy and chronic oral anticoagulation. Biological efficacy to aspirin was evaluated by platelet aggregation induced by arachidonic acid 1.33 mM on platelet rich plasma and tested at trough level and 24 hours after last aspirin intake. Resistance to aspirin was defined as a maximal platelet aggregation over 20%. According to the results of platelet aggregation, aspirin dose and dosing regimen were modified as follows: from 100 mg/day to 160 mg/day or 75 mg x2/day. Patients enrolled in this have a median follow up of at least 6 months after the analysis. Results: Between January 2012 and February 2014, 77 patients with Philadelphia-negative MPN were included. 53 were treated with aspirin 75 mg/day (69%) and 24 with aspirin 100 mg/day (31%). Out of the 53 patients treated with aspirin 75 mg/day, 12 patients (23%) were resistant to aspirin. Resistance to aspirin was not correlated to sex, age, presentation, JAK2 status, treatment, history of thrombotic or bleeding and hematologic values (see following table). Table 1:The MPN Grade 1 Fibrosis PhenotypePMF (N:33)PV/ET (N:58)Total (N:91)Median age63 yrs55 yrs58 yrsSex1:31:11:2JAK V617F +16/33: (49%)40/58: (69%)56/91: (62%)Median Hgb(g/dL)11.9 (range 7.9-16.4)12.5 (range 8.0-19.8)12.2 (range 7.9-19.8)Median WBC(X10 (9))26.9 (range 1.3-188)8.9 (range 3.5-51.3)10.8 (range 1.3-188 )Median Platelet(X10 (9))179 (range 18.0-1194)505 (range 67-2286)370 (range 18-2286)Leukoerythroblastic Blood Smear15/33 (45%)PV: 17 ET: 5 Total: 22/58 (38%)37/91 (41%)Splenomegaly(cm below costal margin)18/33: (55%) Median: 1025/58: (43%) Median: 643/91(48%) Median: 4Transfusion dependence6/33: (18%)2/58: (4%)8/91: (9%)Presence Of ³ 1 symptom 17/33: (52%)33/58: (57%)50/91: (55%)DIPSS risk intermediate 2 or higher13/33: (39%)17/58: (29%)30/91: (33%)2 or more prior therapies9/33: (27%)36/58: (62%)45/91: (49%)Vital Status (Alive)25/33: (76%)53/58: (91%)78/90: (86%)Median follow up time (yrs)1.9 (range 0.1-9.8)5.7 (range 0-34.5)3.1 (range 0-34.5) An increased dose of 100 mg/day for at least 7 days overcame this biological resistance in 8 out of 8 re-tested patients. The 2 remaining 75mg resistant patients received an increased dose of aspirin but were not retested. Out of the 24 patients under 100 mg/day, only two (8%) were resistant to aspirin. In these patients, increasing the dose to 160 mg/day did not modify their biological response. However a 75 mgx2 /day was effective to overcome biological resistance. No thrombotic or bleeding event was observed during the 6-month follow-up regardless of the aspirin dose. Conclusions: This is the first study to measure in standardized conditions the biological resistance of aspirin in 77 well-characterized Philadelphia-negative MPN patients. Although this does not reach statistical significance a higher proportion of patients treated with aspirin 75mg/day was resistant to anti-platelet therapy compared to patients treated with aspirin 100 mg/day. Increasing the dose of aspirin from 75 to 100 mg once daily overcomes this biological resistance without increasing bleeding side effects and seems the best compromise. Interestingly in rare cases of extreme resistance to 100mg doubling the dose twice daily (75mgX2) was better than an increase of 160mg in one take. This pilot study on a small number of patients with a limited 6-month follow up compared to the low annual incidence of thromboses in these MPNs will be followed by a prospective study on a larger number of patients with an extended follow-up period to determine if biological resistance to aspirin is correlated to the occurrence of thrombotic events. Disclosures No relevant conflicts of interest to declare.