Damage measured by Damage Index for Antiphospholipid Syndrome (DIAPS) in antiphospholipid antibody-positive patients included in the APS ACTION registry.

OBJECTIVES: Our primary objective was to quantify damage burden measured by Damage Index for Antiphospholipid Syndrome (DIAPS) in aPL-positive patients with or without a history of thrombosis in an international cohort (the APS ACTION cohort). Secondly, we aimed to identify clinical and laboratory characteristics associated with damage in aPL-positive patients. METHODS: In this cross-sectional study, we analysed the baseline damage in aPL-positive patients with or without APS classification. We excluded patients with other autoimmune diseases. We analysed the demographic, clinical and laboratory characteristics based on two subgroups: (i) thrombotic APS patients with high vs low damage; and (ii) non-thrombotic aPL-positive patients with vs without damage. RESULTS: Of the 826 aPL-positive patients included in the registry as of April 2020, 586 with no other systemic autoimmune diseases were included in the analysis (412 thrombotic and 174 non-thrombotic). In the thrombotic group, hyperlipidaemia (odds ratio [OR] 1.82; 95% CI 1.05, 3.15; adjusted P = 0.032), obesity (OR 2.14; 95% CI 1.23, 3.71; adjusted P = 0.007), aß2GPI high titres (OR 2.33; 95% CI 1.36, 4.02; adjusted P = 0.002) and corticosteroid use (ever) (OR 3.73; 95% CI 1.80, 7.75; adjusted P < 0.001) were independently associated with high damage at baseline. In the non-thrombotic group, hypertension (OR 4.55; 95% CI 1.82, 11.35; adjusted P = 0.001) and hyperlipidaemia (OR 4.32; 95% CI 1.37, 13.65; adjusted P = 0.013) were independent predictors of damage at baseline; conversely, single aPL positivity was inversely correlated with damage (OR 0.24; 95% CI 0.075, 0.77; adjusted P = 0.016). CONCLUSIONS: DIAPS indicates substantial damage in aPL-positive patients in the APS ACTION cohort. Selected traditional cardiovascular risk factors, steroids use and specific aPL profiles may help to identify patients more prone to present with a higher damage burden.

Anti-C1q antibodies in lupus nephritis children with glomerular microthrombosis.

AIM: Glomerular microthrombosis (GMT) was a common vascular lesion in patients with lupus nephritis (LN). The objective of this study was to investigate the relationship between serum anti-beta2-glycoprotein I antibodies (a-ß2GP1) and anti-complement 1q antibodies (a-C1q) antibodies and to investigate the possible mechanism of GMT in children with LN. METHODS: The subjects were 191 children with LN diagnosed by renal biopsy in our hospital from January 2017 to January 2020. The patients were divided into GMT group and non-GMT group. The clinical manifestations, laboratory tests, renal pathology, prognosis of the two groups and the relationship between a-ß2GP1 and a-C1q antibodies were observed. RESULTS: In 191 children with LN, 52 cases (27.23%) presented with GMT. The value of C3, haemoglobin (Hb), estimate glomerular filtration rate (eGFR) and anticardiolipin antibody (ACA) in GMT group were lower than that of non-GMT group (p < .05, p < .01). The value of serum creatinine (Scr), 24 h proteinuria (PRO), urine red blood cells (RBC), N-acetyl-ß-d-glucosidase (NAG) and retinol-binding protein (RBP), a-C1q, a-ß2GP1, Systemic Lupus Erythematosus Disease Activity Index (SLEDAI) and renal histopathological activity index (AI) score in GMT group were higher than that of non-GMT group (p < .05, p < .01). The positive proportions of serum a-C1q and a-ß2GP1 in GMT group were higher than those in non-GMT group (p < .05). According to Spearman correlation analysis, a-C1q was positively correlated with AI score, SLEDAI, a-ß2GP1, GMT, LN-III and LN-IV. Hb, eGFR and a-C1q Ab were associated with the formation of GMT in children with LN. The complete proteinuria remission and renal survival in GMT group were significantly lower than those in non-GMT group (p < .05, p < .01). CONCLUSION: LN children with GMT had more severe clinical manifestations and renal pathologic damages, and poor outcome. Serum a-C1q level was positively correlated with a-ß2GP1, and a-ß2GP1 may be involved in the formation of GMT in children with LN, which might involve in the activation of complement classical pathway.

Anticardiolipin and anti-beta 2-glycoprotein I antibodies in patients with unexplained articular manifestations.

OBJECTIVE: To determine the frequency of antiphospholipid antibodies (aPL) in patients with unexplained articular manifestations. MATERIAL AND METHODS: Three hundred thirteen patients suffering from arthritis or arthralgia without evident cause and 266 healthy blood donors (HBD) were included in the study. Anticardiolipin antibodies (aCL) and anti-beta 2-glycoprotein I antibodies (aß2GPI) were measured by ELISA. RESULT: Out of the 313 patients, 250 were females and 63 were males. The mean age of patients was 49 ± 14 years (17-87 years). One hundred eleven patients have arthralgia and 202 have arthritis. The frequency of aCL and/or aß2 GPI (24.9%) was significantly higher in patients than in HBD (10.9%). The frequency of aß2GPI was 23.6% in patients and 9.4% in the control group (p < 10-3 ). aß2GPI-IgA was significantly more frequent in patients than in the control group (20.4% vs. 7.5%, p < 10-3 ). aß2GPI was most commonly observed than aCL in patients (23.6% vs. 6.4%, p < 10-6 ). IgA isotype of aß2GPI was the most frequent in 20.4% of patients while IgG and IgM were detected in 5.4% and 2.9% respectively. CONCLUSION: This study showed that aPL were common in patients with articular manifestations and were mainly directed against ß2 GPI. The role of these antibodies remains to be specified.

High frequency of antiphospholipid antibodies in primary biliary cirrhosis.

AIM: To evaluate, retrospectively, the frequency of autoantibodies of antiphospholipid syndrome (APLS) in Tunisian patients with primary biliary cirrhosis (PBC). PATIENTS AND METHODS: We analyzed 80 PBC sera and 80 sera from blood donors. ELISA was used to determine the frequency of antibodies against cardiolipin (aCL IgG, IgA, and IgM) and beta 2 glycoprotein I (aß2GPI IgG, IgA, and IgM). RESULTS: The frequency of antiphospholipid antibodies (aCL and/or aß2GPI) was significantly higher in PBC patients than in controls (70 vs. 5%, P < 10(-6)). The frequency of aCL antibodies (IgG, IgA or IgM) was significantly higher in PBC patients than in the control group (23.7 vs. 3.7%, P = 0.0005). The frequencies of aCL IgA and aCL IgM in PBC patients' sera were significantly higher than those in the control group (10 vs. 0%, P = 0.003 and 20 vs. 2.5%, P = 0.001, respectively). Two patients of eighty (2.5%) had aCL IgG, aCL IgA and aCL IgM. The frequency of aß2GPI antibodies (IgG, IgA, or IgM) was significantly higher in PBC patients than in the control group (70 vs. 1.2%, P < 10(-6)). The frequencies of aß2GPI IgG, aß2GPI IgA, and aß2GPI IgM in PBC patients' sera were significantly higher in patients than in the control group (12.5 vs. 0%, P = 0.003; 62.5 vs. 1.2%, P < 10(-6); and 21.2 vs. 0%, P < 10(-4), respectively). CONCLUSION: Autoantibodies related to APLS (aCL and aß2GPI) were present in the majority of patients with PBC, reflecting the ability of these antibodies to engage mediators of damage.

Antiphospholipid Antibody Testing in a Maximum Care Hospital: Method-Dependent Differences.

Background: Antiphospholipid antibody (aPL) testing is critical for the classification of antiphospholipid syndrome. The 2023 ACR/EULAR classification criteria recommend the use of enzyme-linked immunosorbent assays (ELISAs) and specific thresholds for aPL positivity. Since non-ELISA methods are increasingly used, we compared and evaluated ELISA and non-ELISA aPL assays in a real-world maximum care hospital setting. Methods: Between January 2021 and June 2024, anticardiolipin (aCL; IgG and IgM) and anti-beta2 glycoprotein I (aß2GPI; IgG and IgM) antibodies were measured using ELISA (n = 5115) and a chemiluminescence-based automated immunoassay (CLIA) (n = 3820). Results of parallel testing were compared, and associations with clinical and laboratory characteristics were evaluated. Results: A total of 946 samples were tested using ELISA and CLIA in parallel. A total of 136 (14%) specimens were positive for at least one aPL, and 55 (6%) specimens were from patients diagnosed with APS. Among the latter, 47 (85%) and 41 (75%) patients were positive when ELISA- or CLIA-based aPL assays were used, respectively. After applying the >40 units threshold of the new classification criteria, the number of aPL-positive specimens was significantly lower. In the entire cohort, the agreement between ELISA and CLIA aPL assays was acceptable only for aß2GPI IgG; the results from the two methods did not agree for aCL IgG/IgM and aß2GPI IgM. In APS patients, the agreement between ELISA and CLIA aPL assays was acceptable for aß2GPI IgG and IgM but poor for aCL IgG and IgM. Antibody levels in APS patients were significantly higher using CLIA compared to ELISA. Conclusions: The method-dependent discrepancies between ELISA- and CLIA-based aPL assays regarding the quantitative and qualitative results are substantial. Both methods are suitable for APS classification, but the choice of aPL assay may influence the classification, and therefore, aPL results should be interpreted carefully in the clinical context.

Antiphospholipid antibody testing.

Antiphospholipid antibodies (aPL) are a family of autoantibodies targeting phospholipid-binding proteins and are associated with several clinical settings, and most notably define the antiphospholipid syndrome (APS). These antibodies can be identified using a variety of laboratory tests, which include both solid-phase immunological assays and functional clotting assays that detect lupus anticoagulants (LA). aPLs are linked to a range of adverse medical conditions, such as thrombosis and complications affecting the placenta and fetus, potentially leading to morbidity and mortality. The specific aPL identified, along with the pattern of reactivity, correlates with the severity of these conditions. Therefore, laboratory testing for aPL is crucial for evaluating the risk of complications and for fulfilling certain classification criteria for APS, which are also applied as diagnostic markers in medical practice. This review provides an overview of the available laboratory tests currently for measuring aPL and discusses their clinical implications.

Frequency of positive antiphospholipid antibodies in pregnant women with SARS-CoV-2 infection and impact on pregnancy outcome: A single-center prospective study on 151 pregnancies.

Background: At the beginning of the SARS-CoV-2 pandemic, there was a lack of information about the infection's impact on pregnancy and capability to induce de novo autoantibodies. It soon became clear that thrombosis was a manifestation of COVID-19, therefore the possible contribution of de novo antiphospholipid antibodies (aPL) raised research interest. We aimed at screening SARS-CoV-2 positive pregnant patients for aPL. Methods: The study included consecutive pregnant women who were hospitalized in our Obstetric Department between March 2020 and July 2021 for either a symptomatic SARS-CoV-2 infection or for other reasons (obstetric complications, labour, delivery) and found positive at the admission nasopharyngeal swab. All these women underwent the search for aPL by means of Lupus Anticoagulant (LA), IgG/IgM anti-cardiolipin (aCL), IgG/IgM anti-beta2glycoprotein I (aB2GPI). Data about comorbidities, obstetric and neonatal complications were collected. Results: 151 women were included. Sixteen (11%) were positive for aPL, mostly at low titre. Pneumonia was diagnosed in 20 women (5 with positive aPL) and 5 required ICU admission (2 with positive aPL). Obstetric complications occurred in 10/16 (63%) aPL positive and in 36/135 (27%) negative patients. The occurrence of HELLP syndrome and preeclampsia was significantly associated with positive aPL (p=0,004). One case of maternal thrombosis occurred in an aPL negative woman. aPL positivity was checked after at least 12 weeks in 7/16 women (44%): 3 had become negative; 2 were still positive (1 IgG aB2GPI + IgG aCL; 1 IgM aB2GPI); 1 remained positive for IgG aCL but became negative for aB2GPI; 1 became negative for LA but displayed a new positivity for IgG aCL at high titre. Conclusions: The frequency of positive aPL in pregnant women with SARS-CoV-2 infection was low in our cohort and similar to the one described in the general obstetric population. aPL mostly presented as single positive, low titre, transient antibodies. The rate of obstetric complications was higher in aPL positive women as compared to negative ones, particularly hypertensive disorders. Causality cannot be excluded; however, other risk factors, including a full-blown picture of COVID-19, may have elicited the pathogenic potential of aPL and contributed themselves to the development of complications.

Clinical and laboratory characteristics of Brazilian versus non-Brazilian primary antiphospholipid syndrome patients in AntiPhospholipid Syndrome Alliance for Clinical Trials and InternatiOnal Networking (APS ACTION) clinical database and repository.

BACKGROUND: Antiphospholipid syndrome (APS) is characterized by episodes of thrombosis, obstetric morbidity or both, associated with persistently positive antiphospholipid antibodies (aPL). Studying the profile of a rare disease in an admixed population is important as it can provide new insights for understanding an autoimmune disease. In this sense of miscegenation, Brazil is characterized by one of the most heterogeneous populations in the world, which is the result of five centuries of interethnic crosses of people from three continents. The objective of this study was to compare the clinical and laboratory characteristics of Brazilian vs. non-Brazilian primary antiphospholipid syndrome (PAPS) patients. METHODS: We classified PAPS patients into 2 groups: Brazilian PAPS patients (BPAPS) and PAPS patients from other countries (non-BPAPS). They were compared regarding demographic characteristics, criteria and non-criteria APS manifestations, antiphospholipid antibody (aPL) profile, and the adjusted Global Antiphospholipid Syndrome Score (aGAPSS). RESULTS: We included 415 PAPS patients (88 [21%] BPAPS and 327 [79%] non-BPAPS). Brazilian patients were significantly younger, more frequently female, sedentary, obese, non-white, and had a higher frequency of livedo (25% vs. 10%, p < 0.001), cognitive dysfunction (21% vs. 8%, p = 0.001) and seizures (16% vs. 7%, p = 0.007), and a lower frequency of thrombocytopenia (9% vs. 18%, p = 0.037). Additionally, they were more frequently positive for lupus anticoagulant (87.5% vs. 74.6%, p = 0.01), and less frequently positive to anticardiolipin (46.6% vs. 73.7%, p < 0.001) and anti-ß2-glycoprotein-I (13.6% vs. 62.7%, p < 0.001) antibodies. Triple aPL positivity was also less frequent (8% vs. 41.6%, p < 0.001) in Brazilian patients. Median aGAPSS was lower in the Brazilian group (8 vs. 10, p < 0.0001). In the multivariate analysis, BPAPS patients still presented more frequently with livedo, cognitive dysfunction and sedentary lifestyle, and less frequently with thrombocytopenia and triple positivity to aPL. They were also less often white. CONCLUSIONS: Our study suggests a specific profile of PAPS in Brazil with higher frequency of selected non-criteria manifestations and lupus anticoagulant positivity. Lupus anticoagulant (not triple positivity) was the major aPL predictor of a classification criteria event.

Methodology of generation and purification of anti-beta 2 glycoprotein I antibodies.

In this Method Article we are showing the methodology for generation and purification of Anti-Beta 2 Glycoprotein I (ß2GPI) antibodies. First ß2GPI was purified from human plasma, and recognized by Western Blot and anti-ß2GPI antibodies of serum from patients with antiphospholipid syndrome (APS). The C57BL/6 mice were immunized intraperitonealy with 150 µg of protein in adjuvant (ß2GPI or bovine serum albumin) on days 1, 8 and 14. Then the anti-ß2GPI antibodies were purified by affinity chromatography (Affi-Gel protein A sepharose) and affinity column using human ß2GPI coupled to CNBr-activated Sepharose 4B. Titles of anti-ß2GPI antibodies were determined by ELISA assays. •We purified ß2GPI with great efficacy and that is recognized antigenically by serum from patients with SAP or an anti-ß2gpi antibody.•We found that our purified antibody had 13 fold increased activity in ELISA test compared with the control and in Western Blot recognized with ß2GPI (reference and purified).

Antiphospholipid syndrome's genetic and epigenetic aspects.

Studies on last genetic and epigenetic predisposition to APS are summarized. It is well known that genetic predisposition is in HLA system (DR4 and DRw53) and that lupus anticoagulant (LA) and anticardiolipin antibodies (aCL) are both associated with the same HLA antigens. Other genes, outside the MHC, give their contribution to the development of this autoimmune syndrome, such as IRF5, STAT4 and those related to inherited thrombophilia - factor V Leiden and G20210A prothrombin polymorphisms. Finally, post-transcriptional modifications of anti-beta2GPI antibodies could be implicated too. The most important discovery of last years is that altered microRNAs' expression is linked to autoimmunity, thrombosis, early atherosclerosis, and oxidative stress in APS.

Pathogenic Role of Complement in Antiphospholipid Syndrome and Therapeutic Implications.

Antiphospholipid syndrome (APS) is an acquired autoimmune disease characterized by thromboembolic events, pregnancy morbidity, and the presence of antiphospholipid (aPL) antibodies. There is sound evidence that aPL act as pathogenic autoantibodies being responsible for vascular clots and miscarriages. However, the exact mechanisms involved in the clinical manifestations of the syndrome are still a matter of investigation. In particular, while vascular thrombosis is apparently not associated with inflammation, the pathogenesis of miscarriages can be explained only in part by the aPL-mediated hypercoagulable state and additional non-thrombotic effects, including placental inflammation, have been described. Despite this difference, evidence obtained from animal models and studies in APS patients support the conclusion that complement activation is a common denominator in both vascular and obstetric APS. Tissue-bound aPL rather than circulating aPL-beta2 glycoprotein I immune complexes seem to be responsible for the activation of the classical and the alternative complement pathways. The critical role of complement is supported by the finding that complement-deficient animals are protected from the pathogenic effect of passively infused aPL and similar results have been obtained blocking complement activation. Moreover, elevated levels of complement activation products in the absence of abnormalities in regulatory molecules have been found in the plasma of APS patients, strongly suggesting that the activation of complement cascade is the result of aPL binding to the target antigen rather than of a defective regulation. Placental complement deposits represent a further marker of complement activation both in animals and in patients, and there is also some suggestive evidence that complement activation products are deposited in the affected vessels. The aim of this review is to analyze the state of the art of complement involvement in the pathogenesis of APS in order to provide insights into the role of this system as predictive biomarker for the clinical manifestations and as therapeutic target.

Exploring the plasmatic platelet-activating factor acetylhydrolase activity in patients with anti-phospholipid antibodies.

PURPOSE: To explore the role of plasmatic platelet-activating factor acetylhydrolase (PAF-AH), a marker of cardiovascular risk, in patients with anti-phospholipid antibodies (aPL). METHODS: PAF-AH activity was assessed in a series of 167 unselected patients screened for aPL in a context of thrombotic events, risk of thrombosis or obstetric complications and in 77 blood donors. RESULTS: 116/167 patients showed positive results for at least one aPL among IgG/IgM anti-prothrombin/phosphatidylserine (aPS/PT), anti-cardiolipin (aCL), anti-beta2-glycoprotein I (aß2GPI) or lupus anticoagulant (LAC), while 51/167 patients resulted aPL-negative. LAC+ patients disclosed higher PAF-AH than LAC-negative (22.1 ± 6.4 nmol/min/ml vs. 19.5 ± 4.1 nmol/min/ml; p = 0.0032), and aPL-negative patients (p = 0.03). Patients presenting positive IgG aß2GPI disclosed higher PAF-AH than patients with only IgM aß2GPI-positive antibodies (23.1 ± 7.2 nmol/min/ml vs. 20.1 ± 5.3 nmol/min/ml; p = 0.035), as well as than patients showing only isolated LAC, aCL or aPS/PT (16.9 ± 3.8 nmol/min/ml; p = 0.003). CONCLUSIONS: PAF-AH plasmatic activity is particularly up-regulated in LAC+ and in aß2GPI IgG+ patients, possibly representing an alternative prognostic biomarker for the therapeutic management of APS patients.

Clinical and laboratory characteristics of Brazilian versus non-Brazilian primary antiphospholipid syndrome patients in AntiPhospholipid Syndrome Alliance for Clinical Trials and InternatiOnal Networking (APS ACTION) clinical database and repository

Abstract Background: Antiphospholipid syndrome (APS) is characterized by episodes of thrombosis, obstetric morbidity or both, associated with persistently positive antiphospholipid antibodies (aPL). Studying the profile of a rare disease in an admixed population is important as it can provide new insights for understanding an autoimmune disease. In this sense of miscegenation, Brazil is characterized by one of the most heterogeneous populations in the world, which is the result of five centuries of interethnic crosses of people from three continents. The objective of this study was to compare the clinical and laboratory characteristics of Brazilian vs. non-Brazilian primary antiphospholipid syndrome (PAPS) patients. Methods: We classified PAPS patients into 2 groups: Brazilian PAPS patients (BPAPS) and PAPS patients from other countries (non-BPAPS). They were compared regarding demographic characteristics, criteria and non-criteria APS manifestations, antiphospholipid antibody (aPL) profile, and the adjusted Global Antiphospholipid Syndrome Score (aGAPSS). Results: We included 415 PAPS patients (88 [21%] BPAPS and 327 [79%] non-BPAPS). Brazilian patients were significantly younger, more frequently female, sedentary, obese, non-white, and had a higher frequency of livedo (25% vs. 10%, p < 0.001), cognitive dysfunction (21% vs. 8%, p = 0.001) and seizures (16% vs. 7%, p = 0.007), and a lower frequency of thrombocytopenia (9% vs. 18%, p = 0.037). Additionally, they were more frequently positive for lupus anticoagulant (87.5% vs. 74.6%, p = 0.01), and less frequently positive to anticardiolipin (46.6% vs. 73.7%, p < 0.001) and anti-ß2-glycoprotein-I (13.6% vs. 62.7%, p < 0.001) antibodies. Triple aPL positivity was also less frequent (8% vs. 41.6%, p < 0.001) in Brazilian patients. Median aGAPSS was lower in the Brazilian group (8 vs. 10, p < 0.0001). In the multivariate analysis, BPAPS patients still presented more frequently with livedo, cognitive dysfunction and sedentary lifestyle, and less frequently with thrombocytopenia and triple positivity to aPL. They were also less often white. Conclusions: Our study suggests a specific profile of PAPS in Brazil with higher frequency of selected non-criteria manifestations and lupus anticoagulant positivity. Lupus anticoagulant (not triple positivity) was the major aPL predictor of a classification criteria event.

Myocardial Ischaemia, Coronary Atherosclerosis and Pulmonary Pressure Elevation in Antiphospholipid Syndrome Patients.

BACKGROUND: Thrombotic events in antiphospholipid syndrome (APS) involve venous and arterial circulation with the possible involvement of coronary or pulmonary microcirculation. OBJECTIVES: To evaluate the influence of antiphospholipid antibodies (aPL) and on myocardial ischaemia assessed by single-photon emission computerized tomography (SPECT), coronary atherosclerosis assessed by multidetector computerized tomography (MDCT) and pulmonary pressure assessed by transthoracic echocardiography (TTE) in patients with primary antiphospholipid syndrome (PAPS). MATERIAL AND METHODS: TTE, SPECT (Tc 99m sestamibi) and MDCT-based coronary calcium scoring were performed in 26 consecutive PAPS patients (20 females, 6 males, aged 20-61, mean 39.7) without any signs of other autoimmunological disease and without clinical symptoms of heart disease. RESULTS: Out of 26 patients, TEE showed normal left and right ventricle function in 25 (96.2%) and elevated (≥ 30 mm Hg) right ventricle systolic pressure in 7 (26.9%) patients. SPECT revealed myocardial perfusion defects in 15 (57.7%) patients: exercise-induced in 6 (23.1%) and persistent in 11 (42.3%). MDCT revealed coronary calcifications in 4 (15.4%) patients. The number of plaques ranged from 1 to 11 (median 2), volume 3-201.7 mm³ (median 7), calcium scores 1.3-202.6 (median 5.7). In the group with perfusion defects or coronary calcifications (n = 15), all the patients showed elevated aCL IgG. CONCLUSIONS: In most of the relatively young APS patients, without any symptoms of ischemic heart disease, SPECT showed myocardial perfusion defects, and coronary calcifications in 1/6 of them. Right ventricle systolic pressure was elevated in 1/4 of APS patients. These pathologies, well known as cardiovascular risk markers, were associated with elevated levels of the IgG class of both anti-cardiolipin and antiB2 GPI antibodies. Thus, in a high percentage of APS patients, clinically silent myocardial ischaemia, pulmonary pressure elevation and coronary atherosclerosis are present and related to the presence of antiphospholipid antibodies.

Both IgG and IgM anti-beta2 glycoprotein I antibodies assays are clinically useful to the antiphospholipid syndrome diagnosis.

OBJECTIVES: The aim of our study was to evaluate the clinical values of anti-beta2 glycoprotein I antibodies (anti-beta2GPI) IgG and IgM comparing with lupus anticoagulant (LA), anticardiolipin antibodies (aCL) in the two clinical groups of antiphospholipid syndrome (APS), vascular thrombosis (VT) and pregnancy morbidity (PM). METHODS: Eighty patients who fulfilled the APS clinical criteria, VT n = 34; PM n = 40, both VT and PM n = 6 were included. LA, aCL and three anti-beta2GPI ELISA kits were tested. RESULTS: Sensitivities of LA, aCL and anti-beta2GPI assays were found respectively 62, 26 and 41% in VT, and 28, 28 and 30% in PM. The sensitivity for the APS diagnosis could reach to 63% using triple tests. The presence of LA (P<0·01, OR = 4·3) or anti-beta2GPI IgG alone (P<0·05, OR = 8·4) was significantly associated with VT. IgM isotype was found more frequent in PM (92%) than in VT (57%) among all positive anti-beta2GPI cases. CONCLUSION: Both IgG and IgM anti-beta2GPI assays were useful when clinical features of APS presented, even its standardization is ongoing. A decreased by half sensitivity of LA in PM compared with that in VT underlines the importance of adding anti-beta2GPI in PM of APS, especially IgM isotype although recent review questioned its significance.

The introduction of anti-phosphatidylserine/prothrombin autoantibodies in the laboratory diagnostic process of anti-phospholipid antibody syndrome: 6 months of observation.

PURPOSE: To evaluate the impact of the introduction of the anti-phosphatidylserine/prothrombin autoantibodies (aPS/PT) in the laboratory diagnostic process of anti-phospholipid antibody syndrome (APS). METHODS: Four hundred and twenty-one patients (71.5 % females; 53 ± 15 years) presenting a medical prescription for aPS/PT antibodies were consecutively enrolled in the study from March 2013 to August 2013. During the same period, aPS/PT were additionally investigated in a selected series of 62 patients characterized by difficult lupus anticoagulant (LA) tests interpretation and in a retrospective series of 52 LA positive cases with available data about anti-prothrombin (aPT) antibodies. The aPS/PT antibodies, as well as the anti-cardiolipin (aCL), the anti-ß2 glycoprotein I (aß2GPI) and the aPT antibodies were analyzed by ELISA. LA was tested according to the recommended criteria, performing both the screen and the confirm steps. RESULTS: Overall, aPS/PT IgM positive (>30 U/ml) and/or IgG frankly positive (>40 U/ml) antibodies were found in 49/421 (11.6 %) cases. Among the LA positive patients, we found 56.1 % aPS/PT positive versus 31.7 % aCL and/or aß2GPI positive cases, with limited (17.1 %) simultaneous positivity. The PS/PT complex resulted the newly recognized specificity in about 27 % of patients recruited from the subset with difficult LA test interpretation. Compared to aPT antibodies, the aPS/PT antibodies displayed a much higher sensitivity (55.8 versus 15.4 %) in LA positive patients. CONCLUSIONS: The introduction of aPS/PT antibodies in the diagnostic process of APS is highly recommended, since they disclose a notable diagnostic performance and a high correlation with LA activity, such that they can be a viable alternative.

Performance of two new, automated chemiluminescence assay panels for anticardiolipin and anti-beta2-glycoprotein I antibodies in the laboratory diagnosis of the antiphospholipid syndrome.

INTRODUCTION: Anticardiolipin (aCL) and anti-ß2-glycoprotein I (aß2GPI) antibodies are two of the three laboratory criteria for antiphospholipid syndrome (APS). All assays for antiphospholipid antibodies (aPL), coagulation assays as well as ELISAs, show methodological shortcomings, which makes the search for better assays everlasting. The purpose of this study was to investigate the diagnostic performance of two new, fully automated systems (Zenit RA and HemosIL Acustar) applying chemiluminescent technology for aPL detection. METHODS: The study cohort consisted of a patient population presenting with thrombosis. In such patient population, the demonstration of aPL determines whether a patient has APS or not with implications for treatment. One hundred and twenty-four patients with thrombotic complications, of whom 26 were patients with definite APS, were integrated in this study. Besides, aPL titres were compared to the Sapporo standards. RESULTS: Results of both systems agreed well with ELISA and mutually. Analysis of the discrepant results between Zenit and Acustar finally led to one misclassification as APS. CONCLUSION: Diagnostic performances of both Zenit RA and HemosIL Acustar were comparable with odds ratios lower limits of CI of 5 for aß2 GPI IgG for Zenit and Acustar and 6 and 5 for aCl IgG on Zenit and Acustar, respectively. However, even with these new automated systems, titres differed largely between systems, especially for aß2 GPI IgG.

The antiphospholipid syndrome: a large elephant with many parts or an elusive chameleon disguised by many colours?

The antiphospholipid syndrome (APS) is characterized by a range of clinical features (primarily thrombosis and/or obstetric-related), together with the presence of antiphospholipid antibody (aPL) as detected by a diverse range of laboratory tests. APS remains a significant diagnostic and management challenge for clinicians across a wide range of specialties, some 30 years after APS was first described as a discrete clinical entity. This is due to ongoing issues regarding nomenclature, the diagnosis of APS in individual patients, the expanding range of recognized clinical manifestations and of APS-related laboratory tests, and management issues in particular APS patient subgroups (including obstetric and catastrophic APS). In addition to the presence of appropriate clinical features, the diagnosis of APS fundamentally requires the finding of positive aPL test result(s), which is hampered by ongoing problems with assay reproducibility and standardization. This review focuses on ongoing dilemmas and issues related to clinical and laboratory aspects of APS including: (1) diagnostic challenges posed by the protean clinical manifestations of APS; (2) current nomenclature and recent proposals for revision of the 2006 international classification criteria; (3) an overview of some key issues related to aPL testing; (4) potential pitfalls of applying the APS classification criteria as diagnostic criteria; and (5) the controversial subgroups of seronegative APS and non-APS aPL positivity.

Characterization of antiphospholipid antibodies in chronic hepatitis B infection

BACKGROUND: Many infections are associated with antiphospholipid antibodies (aPLs). The purpose of this study was to investigate the prevalence, persistence, clinical significance, and characteristics of aPLs in hepatitis B virus (HBV)-infected patients. METHODS: This study included 143 patients with HBV infection and 32 healthy individuals as controls. The presence of anticardiolipin antibodies (aCL Ab), anti-beta2-glycoprotein I antibodies (beta2GPI Ab), and lupus anticoagulant (LA) was assessed. RESULTS: The total prevalence of aPLs in HBV-infected patients was 12.6% (18 of 143). Of these 18 patients, 15 had low to medium titers of aCL Ab (10 with IgM, 4 with IgG, and 1 with both isotypes). beta2GPI Ab and LA were detected in 3 (2.1%) and 2 (1.4%) patients with HBV infection, respectively. In follow-up specimens from 14 patients with elevated levels of aCL Ab or beta2GPI Ab, 10 (71.4%) showed the persistent presence of aPLs. No clinical manifestations related to aPLs were identified. CONCLUSION: In HBV-infected patients, the most frequently detected antiphospholipid antibodies were IgM aCL Ab, which have a weak association with the clinical manifestations of APS. Unlike the transient presence reported for other infection-associated aPLs, most aPLs were persistently detected over a 12-week period in patients with HBV infection.

Activated protein C resistance and lupus anticoagulant activity induced by plasma and purified monospecific human IgG anti-β2-glycoprotein-I antibodies / Resistencia a la proteína C activada y actividad de anticoagulante lúpico inducidas por plasma y por anticuerpos purificados humanos del IgG anti β2-glicoproteína I

ABSTRACT Introduction. We investigated the activated protein C resistance (APCR) phenotype and the lupus anticoagulant (LA), activity induced by anti-β2-glycoprotein-I (anti-β2GP-I) antibodies. Patients and methods. We studied plasma and sera samples from 29 patients with persistently positive anti-β2GP-I: 22 with thrombosis (12 with primary APS, 10 with APS secondary to SLE) and seven without thrombosis (all with SLE); 25 healthy subjects were studied as controls. We detected anticardiolipin antibodies (ACA); IgG (and its subclasses) and IgM anti-β2GP-I, on irradiated and non-irradiated plates by ELISA. APCR was assessed by the activated partial thromboplastin time (APTT)-based assay and by the modified test. The FV Leiden mutation was studied by PCR. LA determination included screening and confirmatory dRVVT. Serum anti-β2GP-I were affinity purified on sepharose columns and their isotype, subclass, and reactivity against various antigens were studied by ELISA. Results. We found that titers of IgG anti-β2GP-I on irradiated plates were higher than on non-irradiated plates (p = 0.002), IgG2 was the predominant subclass. Fifteen patients (13 with thrombosis) had LA and 15 (also 13 with thrombosis) induced the APCR phenotype. Eleven (all with thrombosis) had both. Two patients were heterozygous for the Leiden mutation. Two purified antibodies, monospecific for β2GP-I, induced an in vitro APCR phenotype and LA activity. Conclusions. Our results seem to indicate that the inhibition of the APC anticoagulant function by IgG2 anti-β2GP-I with LA activity may be one of the responsible mechanisms of thrombophilia in patients with APS.

Introducción. Investigamos la resistencia a la proteína C activada (RPCA) y la actividad de anticoagulante lápico (AL), inducidas por anticuerpos anti-β2-glicoproteína-I (anti-β2GP-I). Pacientes y métodos. Estudiamos los plasmas y sueros persistentemente positivos para anti-β2GP-I de 29 pacientes: 22 tuvieron trombosis (12 con síndrome de antifosfolípidos (SAF) primario y 10 con SAF secundario a lupus erítematoso generalizado (LEG)) y siete sin trombosis (todos con LEG). Como controles estudiamos 25 sueros de personas clínicamente sanas. Detectamos anticuerpos anticardiolipina, anti-β2GP-I IgG (y sus subclases) e IgM por ELISA en placas irradiadas y no irradiadas. Evaluamos la RPCA por medio del tiempo parcial de tromboplastina activada y por la prueba modificada. Estudiamos la mutación FV de Leiden por PCR y el anticoagulante lápico con el método de dRVVT screening y confirmatorio. Después de purificar los anti-β2GP-I séricos con una columna de antígeno unido a sefarosa, analizamos por ELISA sus isotipos, subclases y reactividad contra β2GP-I y algunos fosfolípidos. Resultados. Los títulos de anti-β2GP-I IgG fueron más altos en placas irradiadas que en no irradiadas (p = 0.002), predominó la subclase IgG2. Quince plasmas (13 de pacientes con trombosis) tuvieron AL y 15 (13 también de pacientes con trombosis) indujeron el fenotipo de RPCA. Once plasmas (todos de pacientes con trombosis) indujeron ambas actividades. Dos pacientes fueron heterocigotos para la mutación de Leiden. Dos anticuerpos purificados monoespecíficos para β2GP-I indujeron el fenotipo de la RPCA y la actividad de AL in vitro. Conclusiones. Nuestros resultados sugieren que la RPCA, inducida por los anti-β2GP-I que concomitantemente tienen actividad de AL, puede tener implicaciones patogénicas en la trombofílía del SAF.