Wim Terpstra

Elevated procoagulant microparticles expressing endothelial and platelet markers in essential thrombocythemia

Essential thrombocythemia is a myeloproliferative neoplasm characterized by an increased risk of both arterial and venous thrombosis. The findings of this study show that patients with this disorder have elevated levels of platelet-and endothelium-derived microparticles, which may support thrombin generation and play a role in the pathophysiology of thromboembolic complications. Background Most cell types, including blood - and vascular cells, produce microparticles upon activation. Since cellular microparticles are known to be elevated in thromboembolic diseases, we hypothesized a role for microparticles in the pathogenesis of thrombosis in essential thrombocythemia. Design and Methods In plasma samples from 21 patients with essential thrombocythemia and ten healthy subjects, the levels and the cellular origin of microparticles were determined by flowcytometric analysis, while the microparticle-associated procoagulant activity was measured using a thrombin generation assay. Results Patients with essential thrombocythemia had significantly higher numbers of circulating annexin V-positive microparticles than controls (median 4500 vs. 2500×106 events/L; p=0.039), including significantly higher numbers of microparticles positive for the platelet marker CD61 (p=0.043), the endothelial markers CD62E (p=0.009) and CD144 (p=0.021), and for tissue factor (p=0.036). CD62E was co-expressed with the platelet marker CD41 on microparticles, suggesting a bilineage origin of such microparticles, which were observed only in patients with risk factors for thrombosis. Patients with essential thrombocythemia had higher plasma levels of mature von Willebrand factor (p=0.045) but similar propeptide levels compared to controls. In thrombin generation analyses, microparticle-rich plasma from patients with essential thrombocythemia had a shorter lag time (p=0.001) and higher peak height (p=0.038) than plasma from controls. Peak height correlated significantly with the total number of microparticles (R=0.634, p<0.001). Conclusions Patients with essential thrombocythemia had higher number of circulating microparticles with platelet and endothelial markers, suggesting ongoing platelet and endothelial activation. This was confirmed by an increased level of mature von Willebrand factor, an abnormal mature von Willebrand factor/propeptide ratio, and a hypercoagulable state reflected in thrombin generation. These findings suggest a role for microparticles in thrombosis in essential thrombocythemia.

Procoagulant myeloblast-derived microparticles in AML patients: changes in numbers and thrombin generation potential during chemotherapy

M. C . V AN AALDEREN ,* 1 M. C . T RAPP ENB URG,* 1 M. VA N SCHILF GA ARDE , P . J . MOLENAAR , H. TEN C ATE , W. E . TERPSTRA* and A . LEY TE *Department of Internal Medicine, and Department of Clinical Chemistry, Onze Lieve Vrouwe Gasthuis, Amsterdam; and Department of Internal Medicine and Cardiovascular Research Institute Maastricht, Maastricht University Medical Center, Maastricht, the Netherlands

Chronic renal failure is accompanied by endothelial activation and a large increase in microparticle numbers with reduced procoagulant capacity

BACKGROUND In patients with chronic renal failure (CRF), cardiovascular disease is the leading cause of increased morbidity and mortality. We hypothesized a role for endothelial activation and microparticle (MP) numbers and procoagulant activity in the pre-thrombotic state of these patients. METHODS We analysed blood samples of 27 patients with CRF [8 chronic kidney disease Stage 4 (CKD4), 9 peritoneal dialysis (PD) and 10 haemodialysis (HD), samples taken before and after HD] and 10 controls. Degree and nature of endothelial activation were assessed by measuring mature von Willebrand factor (vWF) and vWF propeptide levels. Cellular MPs were characterized by flow cytometry and MP-specific thrombin generation (TG) measurements. RESULTS CRF was accompanied by chronic (CKD4 and PD) or acute (HD) endothelial activation. Patients with CRF had substantially higher MP numbers than controls (median 9400 versus 4350×10(6)/L, P=0.001), without significant differences between the treatment subgroups or between pre- and post-HD. The vast majority of MPs were platelet derived. Of the minor populations, endothelial MPs and tissue factor-bearing MPs were more abundant in CRF. MPs were procoagulant, but the increase in numbers was not reflected in a proportional increase in MP-specific TG. CONCLUSION Renal failure is accompanied by endothelial activation of a different nature in CKD4 and PD patients compared to HD patients, and results in all subgroups in an increase of mainly platelet-derived MPs that appear to be less procoagulant than in other disease states, possibly because of the uraemic functional defect of their cellular source.

Elevated numbers and altered subsets of procoagulant microparticles in breast cancer patients using endocrine therapy.

INTRODUCTION Microparticles (MP) can be elevated in cancer and thromboembolic disease. We hypothesized a role for MP in the hypercoagulable state in breast cancer patients using endocrine therapy, in whom both cancer and the use of endocrine therapy are independent risk factors for the development of thrombosis. DESIGN AND METHODS Plasma samples were collected from 40 breast cancer patients using endocrine therapy (20 patients without metastases receiving adjuvant therapy and 20 patients with metastatic disease treated in a palliative setting) and from 20 female healthy controls. The endocrine therapy used was either an anti-estrogen or an aromatase inhibitor. Numbers and cellular origin of MP subsets were analyzed by flowcytometry. MP-associated procoagulant activity was measured using a thrombin generation assay using conditions that allow analysis of MP induced thrombin generation. RESULTS Breast cancer patients using endocrine therapy had higher levels of MP positive for Annexin V (median 10000 vs 6500×10E6/l), P-selectin (330 vs 200×10E6/l), tissue factor (33 vs 15×10E6/l), and of MP derived from platelets (CD41) and leukocytes (CD45). Thrombin generation in plasma was dependent on the presence of MP and thrombin generation performed after addition of isolated MP to normal plasma showed a higher endogenous thrombin potential (1105 vs 1029 nM.min) in breast cancer patients. No differences were observed in MP levels and thrombin generation parameters between the metastatic and adjuvant group. CONCLUSION Breast cancer patients using endocrine therapy have an increased MP number and a higher MP-dependent thrombin generation, irrespective of the presence of metastatic disease. Altered MP subset characteristics in these patients, especially the higher number of (activated) platelet derived MP and leukocyte derived MP, may in part explain a heightened procoagulant state in breast cancer patients using endocrine therapy.

Haemolytic anaemia after nitrofurantoin treatment in a pregnant woman with G6PD deficiency

We present a normotensive, pregnant woman with severe haemolytic anaemia in the third trimester of pregnancy. Owing to normal platelet count diagnoses other than HELLP syndrome were considered and investigated. The patient was treated with nitrofurantoin 3 weeks before presentation and she turned out to have a deficiency of glucose-6-phosphate dehydrogenase. After treatment with blood transfusion, vitamin B12 and folic acid the patient recovered completely. Caesarean delivery was performed because of maternal hypertension and fetal distress at 33 weeks’ gestation.