[Level of Microparticles in Patients with Essential Thrombocythemia and Its Relation with JAK2V617F Mutation]

Abstract

Objective: To detect the levels of microparticles (MP) in plasma of patients with esseutial thrombo-cythermia(ET) and analyze the relationship between the JAK2V617F mutant and MP in ET patients.

Methods: The numerical values of MPs were analysed by using flow cytometry. Venous blood of 56 ET patients and 28 healthy persons was collected in the morning and anticoagulated with sodium citrate (1∶9). The RMP, PMP, TF⁺MP and EMP were detected by FCM using phycoerythrin (PE)-conjugated monoclonal antibodies to CD235a for red blood cells, CD61 for platelets, CD142 for tissue factor (TF) and CD62E for endothelial cells, respectively. Forward scatter was set in scale using fluorescent microspheres of 0.8 μm. Standard fluorescent microbeads (0-0.8 μm) in diameter were used to set the microparticles gate. Genomic DNA was extracted from mononuclear cells by using a commercial DNA isolation kit and amplified by allele specific polymerase chain reaction (PCR). According to the size of molecular weight, the amplified products were separated by electrophoresis on a 2% agarose gel to screen 26 JAK2V617F mutations.

Results: The detection results showed that the MP levels in ET group were higher than those in normal control group: RMP (157.2±304.9/μl vs 21.3±18.4/μl), PMP (1378.9±2454/μl vs 113.8±97.1/μl), TF⁺MP (123±354.6/μl vs 21±15.7/μl) and EMP (1434.7±2601.9/μl vs 291.7±297.7/μl) (P<0.05). In 8 patients with thrombus, the levels of these four kinds of MP were higher than those of controls without thrombus: RMP (258.2±327.7/μl vs 55.6±63.8/μl), PMP (1853±1874.7/μl vs 444.7±712/μl), TF⁺MP (120.4±112.5/μl vs 60.3±128.3/μl) and EMP (1637.1±1563.5/μl vs 629.4±1000.2/μl) (P<0.05). The levels of those 4 kinds of MP in 17 patients with splenomegaly were significantly higher than those in 11 patients without splenomegaly: RMP (306.8±491.1/μl vs 59.3±51/μl), PMP (2944.8±3767.6/μl vs 334.8±420.2/μl), TF⁺MP (162.9±166.8/μl vs 31.0±28.7/μl) and EMP (3031.8±4048.8/μl vs 701.5±729.2/μl) (P<0.05). The level of other MP showed no difference between MPN patients with JAK2V617F mutation and without JAK2V617F mutation (P>0.05), except TF⁺MP (154.7±516.3/μl vs 100.5±126.6/μl) (P<0.05).

Conclusion: The numerical values of MP detected are more in ET patients than those in healthy controls. The number of MP is higher in patients with thrombus than that without thrombus, so do in patients with splenomegaly and without splenomegaly. Patients with JAK2V617F mutation show higher number of TF⁺MP than that without JAK2V617F mutation. But the other three kinds of MP show no this difference.