Circulating platelet-derived microparticles in systemic lupus erythematosus

2006 ◽  
Vol 95 (01) ◽  
pp. 94-99 ◽  
Author(s):  
Gino Alfaro ◽  
Manuela Goycoolea ◽  
Teresa Quiroga ◽  
Mauricio Ocqueteau ◽  
Loreto Massardo ◽  
...  
Keyword(s):  
Systemic Lupus Erythematosus ◽  
Lupus Erythematosus ◽  
Thrombin Generation ◽  
Thrombus Formation ◽  
Annexin V ◽  
Coagulation System ◽  
Systemic Lupus ◽  
Double Labeling ◽  
Cellular Source ◽  
Risk For Thrombosis

SummaryThe risk for thrombosis is significantly increased in systemic lupus erythematosus (SLE), affecting both venous and arterial vessels. Activated platelets are known to participate in thrombus formation and growth. A general feature of activated cells is the shedding of microparticles (MP) which support coagulation by exposure of negatively charged phospholipids and possibly tissue factor (TF). In this work we characterized circulating MP in patients with SLE and their relationship with a procoagulant state. Thirty patients with SLE (aged 15–72 years, mean age 38 years) and 20 healthy controls (aged 22–54 years, mean age 34 years) were studied; patients fulfilled 4 revised criteria for SLE. The number and cellular source of circulating MP were determined by flow cytometry using double labeling with specific monoclonal antibodies and annexin V. Thrombin generation was measured as the endogenous thrombin potential (ETP) without the addition of exogenous phospholipids and TF; under these conditions the generation of thrombin depended directly on the number of MP present in plasma. Thrombin anti-thrombin (TAT) and plasmin-antiplasmin (PAP) complexes were measured by ELISA. Compared to the controls, circulating MP were significantly elevated in the patient group (1218 ± 136 vs 653 ± 74 x103/ml plasma, p: 0.0007). In both groups, most of these MP were of platelet origin (927± 131 vs 517 ± 72 x103/ml plasma, p:0.009 ). ETP was higher among patients as compared to the controls (804± 64 vs 631 ± 37 nM thrombin, p: 0.025). Plasma levels of TAT in patients and controls were 3.4 ± 0.8 and 1.4 ± 0.5 µg/L, respectively (p:0.04), and of PAP complexes were 62.5 ± 14 and 24.05± 2.5 µg/ml, respectively (p: 0.014).The number of platelet-derived MP correlated significantly with thrombin generation (r: 0.42; p: 0.038) and TAT levels (r: 0.40; p: 0.035).We did not find an association of circulating MP with disease activity nor with the presence of antiphospholipid antibodies. The increased number of circulating platelet-derived microparticles and their association with high ETP and activation of the coagulation system suggest that these microparticles play an important role in the pathogenesis of the prothrombotic state in SLE patients.

scholarly journals Characterization of the thrombin generation profile in systemic lupus erythematosus

2017 ◽  
Vol 104 (1) ◽  
pp. 35-41 ◽  
Author(s):  
A Kern ◽  
E Barabás ◽  
A Balog ◽  
Sz Burcsár ◽  
M Kiszelák ◽  
...  
Keyword(s):  
Systemic Lupus Erythematosus ◽  
Lupus Erythematosus ◽  
Thrombin Generation ◽  
Area Under The Curve ◽  
Autoimmune Disorder ◽  
Systemic Lupus ◽  
Time To Peak ◽  
Cat Assay ◽  
Coagulation Tests ◽  
Healthy Control

Systemic lupus erythematosus (SLE) is a multisystemic inflammatory autoimmune disorder. Thrombotic events occur at a higher incidence among SLE patients. The investigation of thrombin generation (TG) with calibrated automated thrombogram (CAT) test as a global hemostasis assay is applicable for the overall functional assessment of the hemostasis. The aim of this study was to characterize the hemostatic alterations observed in SLE by CAT assay. In this study, CAT parameters and basic coagulation parameters of SLE patients (n = 22) and healthy control subjects (n = 34) were compared. CAT area under the curve (i.e., endogenous thrombin potential) was lower than normal in SLE (807 vs. 1,159 nM*min, respectively), whereas other CAT parameters (peak, lag time, time to peak, and velocity index) and the basic coagulation tests were within the normal range. The presence of anti-phospholipid antibodies and the applied therapy was not associated with hemostasis parameters in SLE. We concluded that the reported high risk of thrombosis is not related to TG potential.

scholarly journals Thrombin Generation Related to Netosis in Patients with Systemic Lupus Erythematosus

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 10-11
Author(s):  
Elena Monzón Manzano ◽  
Ihosvany Fernandez-Bello ◽  
Raul Justo Sanz ◽  
Ángel Robles Marhuenda ◽  
Paula Acuña ◽  
...  
Keyword(s):  
Systemic Lupus Erythematosus ◽  
Fluorescence Microscopy ◽  
Lupus Erythematosus ◽  
Antiphospholipid Antibodies ◽  
Research Funding ◽  
Thrombin Generation ◽  
Healthy Controls ◽  
Systemic Lupus ◽  
Blood Samples ◽  
History Of

NETosis is a process suffered by neutrophils that consists in the loss of their function and the release of their nuclear material as large web-like structure called neutrophil extracelular traps (NETs). Many authors demonstrated that NETs participate in the pathogenesis of autoimmune diseases, such as systemic lupus erythematosus (SLE), because the release of autoantigens amplifies inflammatory responses, perpetuating the exacerbation of autoimmunity. On the other hand, NETs may play a prominent role in thrombosis because they serve as a negative charge scaffold to trap platelets and coagulation factors, promoting blood clot formation. Objetive: to determine participation of NETs in the hypercoagulable state of patients with SLE. Methods: 32 patients with SLE without antiphospholipid antibodies and without history of thrombotic events were included after signing informed consent; 88 sex- and age-matched healthy controls were also recruited. Blood samples were drawn in citrate tubes (3.2%). Neutrophils were isolated by centrifugation of whole blood with a Percoll gradient at 500 g, 25 min, 5ºC. To induce NETs formation, 2.5x105 isolated neutrophils were incubated in RPMI-1640 medium with or without 100 nM phorbol 12-myristate 13-acetate (PMA) for 45 min, 37ºC. To verify NETs formation, neutrophils were seeded on cover glasses pretreated with poly-L-lysine in RPMI-1640 medium with or without 100 nM PMA for 45 min, 37ºC. Samples were fixed and later incubated first, with an anti-human myeloperoxidase and then, with Alexa Fluor 488 goat anti-rabbit IgG. Finally, samples were embedded in mounting medium with DAPI and were observed by fluorescence microscopy with a Nikon Eclipse 90i microscope. Cell free DNA (cfDNA) was determined in poor platelet plasma obtained by centrifugation of whole blood (2500 g for 15 min), using the Quant-iT™ Pico Green dsDNA assay (Thermo Fisher Scientific, Waltham, MA, USA) according to the manufacturer's instructions. To assess thrombin generation associated to NETs, 2.5x105 neutrophils from patients with SLE or from controls were incubated with either buffer or 100 nM PMA during 45 min. Then they were centrifuged at 5000g, 3 min and resuspended in 40-μL of rich platelet rich plasma (PRP) from healthy controls adjusted to 106 platelets/µL obtained from blood samples drawn either in citrate or citrate plus corn trypsin inhibitor (CTI) tubes. CTI is an inhibitor of FXIIa. Calibrated automated thrombogram (CAT) was performed without addition of any trigger. Results: We observed that plasma from patients with SLE had increased free nucleic acids (cfDNA in fluorescence units, controls: 94.90±21.29, SLE patients: 112.4±26.59; P=0.0211). In accordance with this observation, analyses by fluorescence microscopy showed that neutrophils from SLE patients, but not from controls, had NETs even in basal conditions. Moreover, neutrophils from these patients generated more NETs in presence of 100 nM PMA (Figure 1). To evaluate whether the increment of NETs observed in patients with SLE had consequences on the hemostasis of these patients, we tested thrombin generation of neutrophils from either patients with SLE or controls in the presence of platelets from healthy controls. Neutrophils from patients with SLE produced more thrombin than those from healthy controls under basal conditions and after stimulation with 100 nM PMA. These increments were avoided when PRP was collected from blood samples drawn with CTI (Figure 2). Conclusions: Neutrophils from SLE patients without antiphospholipid antibodies and with no history of thrombotic seemed more prone to form NETs than those from healthy controls. NETs might be considered as a key element in the prothrombotic profile of patients with SLE and their analyses by thrombin generation test might be useful to detect risk of occurrence of thrombotic events in these patients and to prevent its occurrence by therapeutic management. This work was supported by grants from FIS-FONDOS FEDER (PI19/00772). EMM holds a predoctoral fellowship from Fundación Española de Trombosis y Hemostasia (FETH-SETH). Disclosures Fernandez-Bello: Stago: Speakers Bureau; Pfizer: Speakers Bureau; SOBI,: Research Funding; Roche: Speakers Bureau; Novartis: Speakers Bureau; Takeda: Research Funding, Speakers Bureau; NovoNordisk: Current Employment, Research Funding, Speakers Bureau. Justo Sanz:Takeda: Current Employment. Alvarez Román:Bayer: Consultancy; Grifols: Research Funding; Pfizer,: Research Funding, Speakers Bureau; SOBI,: Consultancy, Research Funding, Speakers Bureau; Takeda: Research Funding, Speakers Bureau; NovoNordisk,: Research Funding, Speakers Bureau; Roche: Speakers Bureau; Novartis: Speakers Bureau. García Barcenilla:Novartis: Speakers Bureau; Roche: Speakers Bureau; Pfizer,: Speakers Bureau; NovoNordisk: Research Funding, Speakers Bureau; Takeda: Research Funding, Speakers Bureau; Bayer: Speakers Bureau. Canales:Sandoz: Speakers Bureau; Roche: Honoraria; Sandoz: Honoraria; Karyopharm: Honoraria; Roche: Speakers Bureau; Takeda: Speakers Bureau; Roche: Honoraria; Takeda: Speakers Bureau; Novartis: Honoraria; Sandoz: Speakers Bureau; Karyopharm: Honoraria; Roche: Speakers Bureau; Janssen: Honoraria; Janssen: Speakers Bureau; iQone: Honoraria; Sandoz: Honoraria; Gilead: Honoraria; Janssen: Speakers Bureau; Celgene: Honoraria; Janssen: Honoraria; Novartis: Honoraria. Jimenez-Yuste:F. Hoffman-La Roche Ltd, Novo Nordisk, Takeda, Sobi, Pfizer: Consultancy; F. Hoffman-La Roche Ltd, Novo Nordisk, Takeda, Sobi, Pfizer, Grifols, Octapharma, CSL Behring, Bayer: Honoraria; Grifols, Novo Nordisk, Takeda, Sobi, Pfizer: Research Funding. Butta:Novartis: Speakers Bureau; NovoNordisk: Speakers Bureau; Takeda: Research Funding, Speakers Bureau; SOBI: Speakers Bureau; Grifols: Research Funding; ROCHE: Research Funding, Speakers Bureau; Pfizer: Speakers Bureau.

Prevalence and Clinical Correlation of Anti-Phospholipid–Binding Protein Antibodies in Anticardiolipin-Negative Patients With Systemic Lupus Erythematosus and Women With Unexplained Recurrent Miscarriages

2005 ◽  
Vol 129 (1) ◽  
pp. 61-68
Author(s):  
Nicola Bizzaro ◽  
Elio Tonutti ◽  
Danilo Villalta ◽  
Marilina Tampoia ◽  
Renato Tozzoli
Keyword(s):  
Systemic Lupus Erythematosus ◽  
Lupus Erythematosus ◽  
Pregnancy Loss ◽  
Recurrent Pregnancy Loss ◽  
Annexin V ◽  
Control Group ◽  
Systemic Lupus ◽  
Serum Samples ◽  
Phospholipid Binding ◽  
Anionic Phospholipids

Abstract Context.—Anti-phospholipid antibodies (aPL) are a heterogeneous group of autoantibodies, the presence of which is associated with thrombotic events and miscarriage. Objective.—To establish whether antibodies directed against phospholipid-binding plasma proteins such as β2-glycoprotein I (β2GPI), prothrombin (PT), and annexin V (Anx V) constitute a risk factor for thromboembolism in patients with systemic lupus erythematosus (SLE) and for miscarriage in women with recurrent pregnancy loss (RPL), independently of the presence of the classic anticardiolipin (aCL) antibodies, and whether their determination together with that of aCL would help to increase the diagnostic sensitivity of aPL tests. Design.—The prevalence of various antibodies directed toward phospholipids (CL and other anionic phospholipids [APL]) and phospholipid-binding proteins (β2GPI, PT, and Anx V) was determined by immunoenzymatic methods in 311 serum samples. Patients.—Twenty-five patients with aCL-positive primary anti-phospholipid syndrome (pAPS); 89 patients with SLE, 23 of whom had thrombotic complications (SLE/APS) and 66 of whom had no thrombosis; and 77 women with unexplained recurrent pregnancy loss comprised our study group. One hundred twenty healthy subjects matched for age and sex were studied as the control group. Results.—Immunoglobulin (Ig) G and/or IgM aAPL, anti-β2GPI, anti-PT, and IgG anti-Anx V antibodies were detected in 25 (100%), 20 (80%), 15 (60%), and 6 (24%), respectively, of the 25 aCL-positive pAPS patients; IgG and/or IgM aCL, aAPL, anti-β2GPI, anti-PT, and IgG anti-Anx V antibodies were detected in 33 (37%), 42 (47%), 31 (35%), 40 (45%), and 12 (13%) of the 89 SLE patients, respectively. Of the 56 SLE patients who proved to be aCL negative, anti-β2GPI was present in 3 patients (5%), anti-PT in 13 (23%) patients, and anti-Anx V in 5 (9%) patients. In the subset of 23 SLE/APS patients, IgG anti-PT prevalence was higher than that of the other autoantibodies (87% vs 70% aCL, 66% aAPL, 57% anti-β2GPI, and 4% anti-Anx V), and in 26% of cases, IgG anti-PT was the only antibody present. Anti-PT had a slightly lower specificity than aCL (46% vs 49%); however, the occurrence of both antibodies brought the specificity to 92.4%. The highest risk for thrombosis in SLE patients was associated with the presence of IgG anti-PT antibody (odds ratio [OR] 15.3, P < .001, vs 6.5 aCL, 3.5 aAPL, 3.4 anti-β2GPI, 0.2 anti-Anx V). Fifty-one of the 77 women with recurrent pregnancy loss were negative for all antibodies investigated; the prevalence of IgG and/or IgM aCL, aAPL, anti-β2GPI, anti-PT, and IgG anti-Anx V antibodies was 6% (5), 12% (9), 6% (5), 16% (12), and 17% (13), respectively. Of the 67 aCL-negative women, none had anti-β2GPI antibodies, 7 (11%) were anti-PT positive, and 13 (19%) were anti-Anx V positive. In the subgroup of 26 recurrent pregnancy loss patients who had at least one antibody, anti-Anx V was present in 50% of cases (in 42% as the sole antibody) and was the only antibody significantly associated with miscarriage (P = .02). Conclusions.—The results of this study indicate that it is useful to measure anti-PT antibodies in addition to the more widely used aCL and anti-β2GPI antibodies in the prognostic evaluation of SLE patients for the risk of thrombosis, and the results also confirm that anti-Anx V antibodies may play an important role in recurrent pregnancy loss.

scholarly journals Insights into the Procoagulant Profile of Patients with Systemic Lupus Erythematosus without Antiphospholipid Antibodies

2020 ◽  
Vol 9 (10) ◽  
pp. 3297
Author(s):  
Elena Monzón Manzano ◽  
Ihosvany Fernández-Bello ◽  
Raúl Justo Sanz ◽  
Ángel Robles Marhuenda ◽  
Francisco Javier López-Longo ◽  
...  
Keyword(s):  
Systemic Lupus Erythematosus ◽  
Lupus Erythematosus ◽  
Antiphospholipid Antibodies ◽  
Thrombin Generation ◽  
Thrombotic Event ◽  
Systemic Lupus ◽  
Activation Markers ◽  
Global Tests ◽  
Platelet Aggregates ◽  
Pai 1

We aimed to identify the key players in the prothrombotic profile of patients with systemic lupus erythematosus (SLE) not mediated by antiphospholipid antibodies, as well as the potential utility of global coagulation tests to characterize hemostasis in these patients. Patients with SLE without antiphospholipid antibodies and without signs of thrombosis were included. The kinetics of clot formation were determined by ROTEM®. Platelet activation markers were determined by flow cytometry. Thrombin generation associated with Neutrophil Extracellular Traps (NETs) and microparticles (MPs) was measured by calibrated automated thrombogram (CAT). The plasma levels of PAI-1 were also determined. ROTEM® showed a procoagulant profile in SLE patients. SLE patients had activated platelets and more leukocyte/platelet aggregates at basal conditions. The plasma PAI-1 and platelet aggregates correlated with several ROTEM® parameters. The thrombin generation associated withthe tissue factor (TF) content of MPs and with NETs was increased. Our results suggest the utility of global tests for studying hemostasis in SLE patients because they detect their procoagulant profile, despite having had neither antiphospholipid antibodies nor any previous thrombotic event. A global appraisal of hemostasis should, if possible, be incorporated into clinical practice to detect the risk of a thrombotic event in patients with SLE and to consequently act to prevent its occurrence.

Clinical significance of anti-annexin V antibodies in patients with systemic lupus erythematosus

1997 ◽  
Vol 54 (3) ◽  
pp. 209-213 ◽  
Author(s):  
Junichi Kaburaki ◽  
Masataka Kuwana ◽  
Mihoko Yamamoto ◽  
Shinichi Kawai ◽  
Yasuo Ikeda
Keyword(s):  
Systemic Lupus Erythematosus ◽  
Clinical Significance ◽  
Lupus Erythematosus ◽  
Annexin V ◽  
Systemic Lupus

Thromboelastometry Shows a Prothrombotic State in Systemic Lupus Erythematosus Related to Diminished Fibrinolysis and Microparticle-Derived Tissue Factor

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 2317-2317
Author(s):  
Ihosvany Fernandez Bello ◽  
Francisco Javier López Longo ◽  
Victor Jiménez Yuste ◽  
Miguel Canales ◽  
Juan Ovalles ◽  
...  
Keyword(s):  
Systemic Lupus Erythematosus ◽  
Tissue Factor ◽  
Lupus Erythematosus ◽  
Thrombin Generation ◽  
Alpha Angle ◽  
Endothelial Damage ◽  
Hypercoagulable State ◽  
Prothrombotic State ◽  
Systemic Lupus ◽  
Pai 1

Abstract Background: Systemic Lupus Erythematosus (SLE) is an autoimmune disorder of unknown origin with a high mortality pattern due to the development of a premature cardiovascular disease. The presence in these patients of a dysfunctional endothelium together with a hypercoagulable milieu may contribute to an increased incidence of thrombotic events. Increased thrombin generation, elevated levels of circulating microparticles and plasmatic levels of PAI-1 may contribute to the prothrombotic phenotype of the disease but data are scarce. Objectives: 1-To characterize the prothrombotic state in SLE by rotational thromboelastometry (ROTEM) and thrombin generation linked to tissue factor bearing microparticles. 2- To evaluate endothelial damage in patients with SLE and its relationship with the prothrombotic state of the disease. 3- To evaluate the influence of PAI-1 in the prothrombotic state of SLE. Material and methods: 39 patients with SLE and 25 sex and age matched healthy subjects were included. Whole blood was drawn in standard BD sodium citrate tubes (3.2%). ROTEM was performed in naTEM condition. Clotting time (CT, time from start of measurement until initiation of clotting [in seconds], alpha angle, which reflects the rate of fibrin polymerisation (tangent to the curve at 2-mm amplitude [in degrees]), maximum clot firmness, which reflects the maximum tensile strength of the thrombus (MCF, [in mm]), the time that clot takes to increase from 2mm above baseline to 20mm above baseline (CFT) and A5, amplitude at 5 min, were recorded. To evaluate the presence of tissue factor bearing microparticles, thrombin generation was determined by Calibrated Automated Thrombogram (CAT) in presence of 4 mM phospholipid (MP-Reagent, Diagnostica Stago, Spain). The endogenous thrombin potential (ETP, the total amount of thrombin generated over time); the lag time (the time to the beginning of the explosive burst of thrombin generation); the peak height of the curve (the maximum thrombin concentration produced); and the time to the peak were evaluated. Antigenic levels of E-selectin and PAI-1 were determined by ELISA (R&D Systems, MN, USA and Affymetrix eBioscience, Vienna, Austria) respectively. Results: ROTEM parameters showed a hypercoagulable profile in LES patients. CT and CFT were shorter (p<0.001 in both cases), and MCF, alpha angle and A5 were higher (p<0.001, p<0.02 and p<0.001 respectively) when compared to healthy controls. On the other hand, CAT parameters did not show differences between both groups. Nevertheless, in LES patients but not in controls, CAT parameters significantly correlated with ROTEM ones (Table 1). Table 1.Correlations in the LES patients group between CAT and ROTEM parameters. Analyses were performed with Spearman test. N: number of determinations; p* denotes significance.CTCFTMCFMCF-talphaA5A10ETP(MP)r-,301-,375*,369*-,368*,378*,364*,393*p,0840,028*0,031*0,032*0,027*0,034*0,021*N34343434343434Peak(MP)r-,353*-,396*,372*-,345*,396*,393*,402*p0,040*0,020*0,030*0,045*0,020*0,021*0,018*N34343434343434ttPeak(MP)r,240,222-,242,217-,223-,209-,210p,171,207,168,217,205,236,232N34343434343434 In order to evaluate endothelial damage, plasma E-selectin and PAI-1 were determined. No differences were found in E-selectin level whereas increased PAI-1 levels were seen in LES group (p<0.006). PAI-1 did not correlate to ROTEM parameters. Conclusions: ROTEM can detect a hypercoagulable state in patients with SLE. The hypercoagulable state may be linked to increased tissue factor bearing microparticles and increased PAI-1 plasma levels. Disclosures No relevant conflicts of interest to declare.

scholarly journals Increased thrombin generation and activity in patients with systemic lupus erythematosus and anticardiolipin antibodies: evidence for a prothrombotic state [see comments]

Blood ◽  
1993 ◽  
Vol 81 (11) ◽  
pp. 2958-2963
Author(s):  
JS Ginsberg ◽  
C Demers ◽  
P Brill-Edwards ◽  
M Johnston ◽  
R Bona ◽  
...  
Keyword(s):  
Systemic Lupus Erythematosus ◽  
Lupus Erythematosus ◽  
Thrombin Generation ◽  
Anticardiolipin Antibodies ◽  
Laboratory Evaluation ◽  
Prothrombotic State ◽  
Systemic Lupus ◽  
Cross Sectional ◽  
Significant Difference ◽  
The Mean

The objective of this study is to determine whether patients with systemic lupus erythematosus (SLE) and anticardiolipin antibodies (ACA) have biochemical evidence of an ongoing prothrombotic state. Using a cross-sectional analysis of a cohort design in an outpatient SLE clinic setting, 43 consecutive patients with SLE participated. Patients underwent clinical and laboratory evaluations on two separate occasions at least 3 months apart. As part of the clinical evaluation, the following were ascertained: (1) the ongoing use of warfarin therapy; (2) the presence of prior venous and arterial thromboembolic disease by history, critical review of objective tests, and examination for reflux in the deep veins of the legs as an indicator of venous thrombosis; and (3) disease-related activity by performing a lupus activity criteria count (LACC). As part of the laboratory evaluation, blood was taken on both occasions and assayed for prothrombin fragments (F1 + 2) and fibrinopeptide A (FPA), as indices of thrombin generation and activity, respectively, and ACA. For the analyses, patients were classified as ACA+ if the assay was abnormal on both occasions and ACA- if the assay was negative on both occasions or negative on one occasion and positive on the other. ACA+ patients had: (1) a significantly higher mean level of F1 + 2 (1.07 nmol/L) than ACA- patients (0.79 nmol/L; P = .02) and patients receiving warfarin (0.47 nmol/L; P = .009) and (2) a significantly higher mean level of FPA (1.01 nmol/L) than ACA- patients (0.45 nmol/L; P = .02). When patients with prior thromboembolism were excluded from the analysis, significant differences in the mean levels of F1 + 2 and FPA between ACA+ and ACA- patients were still seen, whereas when patients with prior thromboembolism and/or active disease were excluded from the analysis, a significant difference in the mean level of FPA and a nonsignificant trend in the mean level of F1 + 2 were seen. The results of this study support the hypothesis that the presence of ACA in SLE patients is associated with an ongoing prothrombotic state.

scholarly journals Prothrombotic State, Platelet Activation and Netosis in Systemic Lupus Erythematosus

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 1141-1141
Author(s):  
Elena Monzón Manzano ◽  
Ihosvany Fernandez-Bello ◽  
Raul Justo Sanz ◽  
Larissa Valor ◽  
Francisco Javier López-Longo ◽  
...  
Keyword(s):  
Systemic Lupus Erythematosus ◽  
Platelet Activation ◽  
Lupus Erythematosus ◽  
Research Funding ◽  
Thrombin Generation ◽  
Hypercoagulable State ◽  
Healthy Controls ◽  
Prothrombotic State ◽  
Systemic Lupus ◽  
Pai 1

Introduction: Systemic lupus erythematosus (SLE) is a chronic autoimmune disease of unknown origin characterized by a hypercoagulable state and a high mortality rate. Mechanisms that cause the accelerated deterioration of cardiovascular health in SLE are unknown. Objectives: to characterize the prothrombotic state in SLE patients by global coagulation assays and the contribution of platelets, endothelial damage, microparticles and neutrophil extracellular traps (NETs) in their prothombotic profile. Material and methods: 72 patients and 90 healthy controls were recruited. Patients were classified according to clinical characteristics in: 32 with lupus (SLE group), 29 with SLE and antiphospholipid antibodies (aFL, SLE+aFL group) and 12 who met the criteria for SLE and antiphospholipid syndrome (APS, SLE+APS group). Experimental protocol was approved by La Paz University Hospital Ethics Committee. Venous blood collected in BD sodium citrate tubes (3.2%) was centrifuged at 150 g for 20 min at 23ºC to obtain platelet-rich plasma (PRP). PPP was obtained by centrifugation at 1500 g for 15 min at 23ºC. To obtain neutrophils, whole blood was centrifuged to 1600 rpm 25 min using a Ficoll gradient and red cells were lysed. Rotational thromboelastometry (ROTEM®) was performed in naTEM condition. Clotting time (CT, time from start of measurement until initiation of clotting [in seconds]); alpha angle (tangent to the curve at 2-mm amplitude [in degrees]), Ax (clot firmness at time x, [in mm]) and maximum clot firmness (MCF, [in mm]) were recorded. Procoagulant activity associated to microparticle's content of tissue factor was determined in PPP by Calibrated Automated Thrombogram (CAT) using MP-reagent (4 mM phospholipids, Diagnostica Stago, Spain). We evaluated the endogenous thrombin potential (ETP, the total amount of thrombin generated over time); the lag time (the time to the beginning of the explosive burst of thrombin generation); the peak height of the curve (the maximum thrombin concentration produced) and the time to the peak. Thrombin generation associated to NETs was also measured by CAT. Neutrophils from healthy controls or from LES patients were stimulated with 100 nM PMA in RPMI medium during 45 min at 37º and then cocultivated with PRP adjusted to 105 platelets/µL. NETs formation was verified by fluorescent microscopy performed with DAPI and an anti-myeloperoxidase antibody. Plasma levels of LDL-ox, E-Selectin and PAI-1 were determined by Elisa (R&D Systems, MN, USA and Affymetrix eBioscience, Vienna, Austria, respectively). Platelet activation was analysed by flow cytometry (FCM, FACScan, BD Biosciences). Fibrinogen receptor activation was evaluated through PAC1-FITC binding and release of granule's content was assessed with monoclonal antibodies (mAbs) anti-CD63 and anti P-selectin in quiescent and 100 µM TRAP and 10 µM ADP stimulated platelets. Data were analysed with Graphpad prism and p ≤0.05 was stablished as statistical significance. Results: PAI-1 plasma level was increased in all patient's groups, whereas LDL-ox and E-selectin showed no differences with control cohort (Fig.1). ROTEM demonstrated a procoagulant profile in SLE and SLE+aPL but not in SLE+APS group (Fig. 2). PAI-1 levels correlated with several ROTEM parameters (Table 1). SLE patients and SLE+aFL showed a basal platelet activation. Moreover, SLE group exposed more P-selectin and CD63 than controls (Fig.3). Regarding thrombin generation associated to tissue-factor content of microparticles, no differences were observed between SLE patients and healthy controls. On the other hand, SLE patients had an increased peak of thrombin generation related to NETs formation (control group: 170.3± 58.0, SLE patients: 230.6±39.3, p=0.019). Conclusions: ROTEM® detected a hypercoagulable state in SLE and SLE+aPL patients. The hypercoagulable state might be linked to increased PAI-1 plasma levels and basal platelet activation in SLE and SLE+aPL groups. Moreover, neutrophils from SLE patients seemed to present a basal activation that induced a NETs-related procoagulant state in these patients. SLE+APS patients did not show a hypercoagulable state perhaps because of the presence of lupus anticoagulant and/or to therapeutic treatment of these patients. This work was supported by grants from the FIS-FONDOS FEDER (PI15/01457, NB). NVB holds a Miguel Servet tenure track grant from FIS-FONDOS FEDER (CP14/00024). Disclosures Fernandez-Bello: Novartis, Pfizer, ROCHE, Stago: Speakers Bureau. Robles:ABBVIE, SANDOZ FARMACEUTICA: Speakers Bureau. Álvarez Roman:Sobi: Consultancy, Speakers Bureau; CSL Behring: Consultancy, Speakers Bureau; Roche: Consultancy, Speakers Bureau; Pfizer: Consultancy, Speakers Bureau; Bayer: Consultancy, Speakers Bureau; Novartis: Consultancy, Speakers Bureau; Amgen: Consultancy, Speakers Bureau; Takeda: Research Funding; NovoNordisk: Consultancy, Speakers Bureau. Canales:Celgene: Honoraria; Gilead: Honoraria; Novartis: Honoraria; Janssen: Honoraria, Speakers Bureau; Sandoz: Honoraria; iQone: Honoraria; Takeda: Speakers Bureau; SOBI: Research Funding; Karyopharm: Honoraria; F. Hoffmann-La Roche Ltd: Honoraria, Speakers Bureau. Jimenez-Yuste:Bayer, CSL Behring, Grifols, Novo Nordisk, Octapharma, Pfizer, Roche, Sobi, Shire: Consultancy, Honoraria, Other: reimbursement for attending symposia/congresses , Research Funding, Speakers Bureau. Butta:Novartis: Consultancy; Roche, Pfizer: Speakers Bureau.

Inhibition of Lupus Anticoagulant Activity by Hexagonal Phase Phosphatidylethanolamine in the Presence of Prothrombin

1998 ◽  
Vol 80 (12) ◽  
pp. 936-941 ◽  
Author(s):  
Marion Tannenbaum ◽  
Carolyn Neville ◽  
Paul Fortin ◽  
Joyce Rauch
Keyword(s):  
Systemic Lupus Erythematosus ◽  
Lupus Erythematosus ◽  
Lupus Anticoagulant ◽  
Plasma Proteins ◽  
Anticoagulant Activity ◽  
Hexagonal Phase ◽  
Annexin V ◽  
Systemic Lupus ◽  
Glycoprotein I ◽  
No Inhibition

SummaryWe have previously demonstrated that lupus anticoagulant antibodies from patients with systemic lupus erythematosus (SLE) specifically recognize hexagonal (II) phase phosphatidylethanolamine (PE), but not bilayer PE (Thromb Haemost 1989; 62: 892). In those studies, the involvement of proteins in this recognition was not evaluated. To address this issue, we have isolated IgG lupus anticoagulant antibodies from the plasma of SLE patients and evaluated the inhibition of lupus anticoagulant activity by hexagonal (II) phase PE in the presence and absence of purified plasma proteins. All six of the IgG lupus anticoagulant antibodies tested were inhibited by hexagonal (II) phase PE in the presence, but not the absence, of human prothrombin. In contrast, little or no inhibition was observed with prothrombin alone or with PE in combination with either β2-glycoprotein I or annexin V. These data indicate that, for certain lupus anticoagulant antibodies, inhibition by hexagonal (II) phase PE is dependent on prothrombin, suggesting that these antibodies recognize a complex of PE and prothrombin.

Sign in / Sign up

Export Citation Format

Share Document