Impact of modulators of nitric oxide synthesis on the level of cytokines in bronchoalveolar lavage in experimental antiphospholipid syndrome.

OBJECTIVE: Aim: The aim of the research is to study the cytokine prof i le (IL-1ß, IL 6, TNF-α, IL-4, IL-10) in bronchoalveolar lavage of lungs in experimental APS and its correction with L-arginine and aminoguanidine. PATIENTS AND METHODS: Materials and Methods: Antiphospholipid syndrome was modeled on white female BALB/c mice. L-arginine (25 mg/kg) and aminoguanidine (10 mg/kg) were used for its correction. The concentration of cytokines in bronchoalveolar lavage from the lungs was assessed using the ELISA test. RESULTS: Results: It was established that in cases of APS the concentration of proinf l ammatory cytokines IL-1ß, IL-6 and TNF-a increased in 1.9, 2.3 and 6.6 times, respectively, compare to the control. At the same time a decrease of the IL-4 in 1.7 and IL-10 in 1.8 times was found in the APS group compare to the control. L-arginine reduced the level of proinf l ammatory cytokines IL-1ß by 22%, IL-6 - by 36%, and TNF-α - by 23% compare to the animals with APS. At the same time, the level of anti-inf l ammatory cytokines increased: IL-4 - by 46%, IL-10 - by 57% compare to the APS animal group. Aminoguanidine, a selective iNOS inhibitor, did not cause any signif i cant decrease in pro-inf l ammatory cytokines but the level of anti-inf l ammatory cytokines IL-4 increased by 44% and IL-10 - by 49%. CONCLUSION: Conclusions: The precursor of the NO synthesis L-arginine leads to a decrease in the concentrations of IL-1ß, IL-6, TNF-a and an increase of IL-4 and IL-10 compare to the group of BALB/c mice with APS.

Neutrophil glucose flux as a therapeutic target in antiphospholipid syndrome.

Neutrophil hyperactivity and neutrophil extracellular trap release (NETosis) appear to play important roles in the pathogenesis of the thromboinflammatory autoimmune disease known as antiphospholipid syndrome (APS). The understanding of neutrophil metabolism has advanced tremendously in the past decade, and accumulating evidence suggests that a variety of metabolic pathways guide neutrophil activities in health and disease. Our previous work characterizing the transcriptome of APS neutrophils revealed that genes related to glycolysis, glycogenolysis, and the pentose phosphate pathway (PPP) were significantly upregulated. Here, we found that neutrophils from patients with APS used glycolysis more avidly than neutrophils from people in the healthy control group, especially when the neutrophils were from patients with APS with a history of microvascular disease. In vitro, inhibiting either glycolysis or the PPP tempered phorbol myristate acetate- and APS IgG-induced NETosis, but not NETosis triggered by a calcium ionophore. In mice, inhibiting either glycolysis or the PPP reduced neutrophil reactive oxygen species production and suppressed APS IgG-induced NETosis ex vivo. When APS-associated thrombosis was evaluated in mice, inhibiting either glycolysis or the PPP markedly suppressed thrombosis and circulating NET remnants. In summary, these data identify a potential role for restraining neutrophil glucose flux in the treatment of APS.

Antiphospholipid-exposed trophoblast-derived extracellular vesicles express elevated levels of TLR7/8-activating microRNAs and induce endometrial endothelial activation, in part, through TLR7.

Women with antiphospholipid syndrome (APS) are at high risk for miscarriage and preeclampsia. Unlike pro-thrombotic systemic APS, obstetric APS is associated with insufficient placentation, as well as inflammation and vascular dysfunction at the maternal-fetal interface. Antiphospholipid antibodies (aPL) can target the placental trophoblast and induce inflammation. We reported that aPL trigger trophoblast cells to produce elevated levels of IL-8 through activation of Toll-like receptor 4 (TLR4). Downstream of TLR4, we found this IL-8 response is mediated by a TLR8-activating microRNA (miR), miR-146a-3p, which is also released by the trophoblast via extracellular vesicles (EVs). Since endothelial dysfunction is a feature of obstetric APS, we sought to determine if other miRs that can activate the RNA sensors, TLR7 and/or TLR8, are released by the trophoblast via EVs after exposure to aPL, and if these EVs can activate human endometrial endothelial cells (HEECs). Using a human first trimester extravillous trophoblast cell line we found that aPL elevated their release of small EVs (<150 nm). These extracellular vesicles released from trophoblast cells exposed to aPL expressed elevated levels of TLR7/8-activating miR-21a and miR-29a, in addition to the previously reported miR-146a-3p. Extracellular vesicles from aPL-exposed human trophoblast cells triggered human endometrial endothelial cells to generate an inflammatory IL-8 response, in part through TLR7. This study highlights EVs as a mode of communication between the placenta and the maternal vasculature, as well as a potential role for TLR7/8-activating miRs in contributing to inflammation at the maternal-fetal interface in obstetric APS.

H3K4me3-Mediated FOXJ2/SLAMF8 Axis Aggravates Thrombosis and Inflammation in ß2GPI/Anti-ß2GPI-Treated Monocytes.

Antiphospholipid syndrome (APS) is characterized by thrombus formation, poor pregnancy outcomes, and a proinflammatory response. H3K4me3-related monocytes activation are key regulators of APS pathogenesis. Therefore, H3K4me3 CUT&Tag and ATAC-seq are performed to examine the epigenetic profiles. The results indicate that the H3K4me3 signal and chromatin accessibility at the FOXJ2 promoter are enhanced in an in vitro monocyte model by stimulation with ß2GPI/anti-ß2GPI, which mimics APS, and decreases after OICR-9429 administration. Furthermore, FOXJ2 is highly expressed in patients with primary APS (PAPS) and is the highest in patients with triple-positive antiphospholipid antibodies (aPLs). Mechanistically, FOXJ2 directly binds to the SLAMF8 promoter and activates SLAMF8 transcription. SLAMF8 further interacts with TREM1 to stimulate TLR4/NF-κB signaling and prohibit autophagy. Knockdown of FOXJ2, SLAMF8, or TREM1 blocks TLR4/NF-κB and provokes autophagy, subsequently inhibiting the release of inflammatory and thrombotic indicators. A mouse model of vascular APS is established via ß2GPI intraperitoneal injection, and the results suggest that OICR-9429 administration attenuates the inflammatory response and thrombus formation by inactivating FOXJ2/SLAMF8/TREM1 signaling. These findings highlight the overexpression of H3K4me3-mediated FOXJ2 in APS, which consequently accelerates APS pathogenesis by triggering inflammation and thrombosis via boosting the SLAMF8/TREM1 axis. Therefore, OICR-9429 is a promising candidate drug for APS therapy.

Deep proteomic analysis of obstetric antiphospholipid syndrome by DIA-MS of extracellular vesicle enriched fractions.

Proteins in the plasma/serum mirror an individual's physiology. Circulating extracellular vesicles (EVs) proteins constitute a large portion of the plasma/serum proteome. Thus, deep and unbiased proteomic analysis of circulating plasma/serum extracellular vesicles holds promise for discovering disease biomarkers as well as revealing disease mechanisms. We established a workflow for simple, deep, and reproducible proteome analysis of both serum large and small EVs enriched fractions by ultracentrifugation plus 4D-data-independent acquisition mass spectrometry (4D-DIA-MS). In our cohort study of obstetric antiphospholipid syndrome (OAPS), 4270 and 3328 proteins were identified from large and small EVs enriched fractions respectively. Both of them revealed known or new pathways related to OAPS. Increased levels of von Willebrand factor (VWF) and insulin receptor (INSR) were identified as candidate biomarkers, which shed light on hypercoagulability and abnormal insulin signaling in disease progression. Our workflow will significantly promote our understanding of plasma/serum-based disease mechanisms and generate new biomarkers.

Small-Extracellular-Vesicle-Derived miRNA Profile Identifies miR-483-3p and miR-326 as Regulators in the Pathogenesis of Antiphospholipid Syndrome (APS).

Primary antiphospholipid syndrome (PAPS) is a systemic autoimmune disease associated with recurrent thrombosis and/or obstetric morbidity with persistent antiphospholipid antibodies (aPL). Although these antibodies drive endothelial injury and thrombophilia, the underlying molecular mechanism is still unclear. Small extracellular vesicles (sEVs) contain miRNAs, key players in intercellular communication. To date, the effects of miRNA-derived sEVs in PAPS are not well understood. We characterised the quantity, cellular origin and miRNA profile of sEVs isolated from thrombotic APS patients (PAPS, n = 50), aPL-carrier patients (aPL, n = 30) and healthy donors (HD, n = 30). We found higher circulating sEVs mainly of activated platelet origin in PAPS and aPL patients compared to HD, that were highly engulfed by HUVECs and monocyte. Through miRNA-sequencing analysis, we identified miR-483-3p to be differentially upregulated in sEVs from patients with PAPS and aPL, and miR-326 to be downregulated only in PAPS sEVs. In vitro studies showed that miR-483-3p overexpression in endothelial cells induced an upregulation of the PI3K-AKT pathway that led to endothelial proliferation/dysfunction. MiR-326 downregulation induced NOTCH pathway activation in monocytes with the upregulation of NFKB1, tissue factor and cytokine production. These results provide evidence that miRNA-derived sEVs contribute to APS pathogenesis by producing endothelial cell proliferation, monocyte activation and adhesion/procoagulant factors.

Association between neutrophil extracellular traps (NETs) and thrombosis in antiphospholipid syndrome.

BACKGROUND: The release of neutrophil extracellular traps (NETs) is the basis of immune-mediated thrombosis. Data on the clinical relevance of NETs in antiphospholipid syndrome-related thrombosis are scarce. OBJECTIVE: The aim of this study was to evaluate whether the NET regulator proteins PADI4, ELANE, and MPO are associated with thrombosis in APS. METHODS: A total of 152 thrombotic APS (t-APS) patients and 123 individuals without thrombosis (controls) were included. The following markers of NETs were evaluated: PADI4, ELANE, and MPO gene expression by qPCR and circulating levels of citrullinated histone H3 (H3cit) and myeloperoxidase-DNA complexes (MPO-DNA) by ELISA. RESULTS: The levels of circulating MPO-DNA and MPO mRNA expression and PADI4 mRNA expression were higher in t-APS patients than in controls. The mean differences were 0.05 OD (95% CI 0.01 to 0.09) in MPO-DNA levels, 1.07 AU (95% CI 0.20 to 1.93) for MPO mRNA and 0.20 AU (95% CI 0.03 to 0.36) in PADI4 mRNA fold-change. These differences were more pronounced in triple-positive patients, who had 56% increased levels of MPO-DNA, 44% increased MPO mRNA expression and 69% increased PADI4 mRNA expression compared to controls. Additionally, circulating MPO-DNA levels and MPO mRNA expression were higher in patients with recurrent thrombosis than in patients with incident thrombosis and controls. In recurrent thrombosis, levels of MPO-DNA were 43.8% higher and MPO mRNA expression was 2-fold higher than in controls. Levels of circulating MPO-DNA and PADI4 mRNA expression did not differ substantially between primary and secondary APS. CONCLUSION: Thrombotic APS was associated with increased NET formation, which was more pronounced among patients with poorer prognosis, such as those with triple antiphospholipid positivity and recurrent thrombosis. Our results provide evidence on the association of NETs and the severity of APS-related thrombosis.

Adenosine diphosphate-induced aggregation is enhanced in platelets obtained from patients with thrombotic primary antiphospholipid syndrome (t-PAPS): Role of P2Y12 -cAMP signaling pathway.

BACKGROUND: Thrombotic antiphospholipid syndrome (t-PAPS) is characterized by arterial, venous, or microvascular occlusions, which are explained, in part, by the presence of antiphospholipid (aPL) antibodies. Although there is much evidence indicating that isolated aPL antibodies increase the activity of platelets obtained from healthy volunteers, platelet function in t-PAPS has not been as widely studied. OBJECTIVE: To evaluate platelet reactivity in t-PAPS patients. METHODS: Platelet aggregation, protein expression, and cyclic nucleotide levels were carried out in platelet rich plasma (PRP) or washed platelets (WPs) obtained from t-PAPS or healthy volunteers. RESULTS: ADP-induced aggregation was significantly higher in PRP obtained from t-PAPS than obtained from the control. The protein expression of P2Y12 receptor and Gs alpha was significantly higher and lower, respectively in WPs from t-PAPS patients. In PRP incubated with iloprost or sodium nitroprusside, the residual platelet reactivity induced by ADP was still higher in PRP from t-PAPS than from the control. Lower intracellular levels of cyclic guanosine monophosphate (cGMP) and cyclic adenosine monophosphate (cAMP) were observed in unstimulated PRP from t-PAPS patients. The protein expression of soluble guanylate cyclase subunits and phosphodiesterases types 3 and 5 did not differ. The antiplatelet activity of ticagrelor was similar between the groups and cilostazol significantly potentiated this response. Isolated aPL antibodies obtained from t-PAPS patients potentiated ADP-induced aggregation in healthy platelets but did not affect the inhibitory responses induced by iloprost or sodium nitroprusside. CONCLUSIONS: The overexpression of P2Y12 receptor, accompanied by lower levels of cAMP and cGMP levels produced greater amplitude of ADP aggregation in platelets from t-PAPS patients.

P2RY8 variants in lupus patients uncover a role for the receptor in immunological tolerance.

B cell self-tolerance is maintained through multiple checkpoints, including restraints on intracellular signaling and cell trafficking. P2RY8 is a receptor with established roles in germinal center (GC) B cell migration inhibition and growth regulation. Somatic P2RY8 variants are common in GC-derived B cell lymphomas. Here, we identify germline novel or rare P2RY8 missense variants in lupus kindreds or the related antiphospholipid syndrome, including a "de novo" variant in a child with severe nephritis. All variants decreased protein expression, F-actin abundance, and GPCR-RhoA signaling, and those with stronger effects increased AKT and ERK activity and cell migration. Remarkably, P2RY8 was reduced in B cell subsets from some SLE patients lacking P2RY8 gene variants. Low P2RY8 correlated with lupus nephritis and increased age-associated B cells and plasma cells. By contrast, P2RY8 overexpression in cells and mice restrained plasma cell development and reinforced negative selection of DNA-reactive developing B cells. These findings uncover a role of P2RY8 in immunological tolerance and lupus pathogenesis.

Defibrotide Inhibits Antiphospholipid Antibody-Mediated Neutrophil Extracellular Trap Formation and Venous Thrombosis.

OBJECTIVE: Defibrotide is a heterogenous mixture of polyanionic oligonucleotides currently approved for treatment of transplant-associated venoocclusive disease. While defibrotide has a known role in limiting endothelial cell activation, some studies have also demonstrated anti-leukocyte properties. In a recent study, we found that neutrophil extracellular traps (NETs) play a role in the thrombotic complications of antiphospholipid syndrome (APS). In the present study, we investigated the hypothesis that defibrotide might act to mitigate APS-relevant NET formation in vitro and in mouse models. METHODS: We used in vitro assays and a mouse model to determine the mechanisms by which defibrotide inhibits NET formation and venous thrombosis in APS. RESULTS: At doses ranging from 1 to 10 µg/ml, defibrotide significantly suppressed NET formation from control neutrophils stimulated with IgG isolated from patients with APS. Defibrotide increased levels of intracellular cyclic AMP in neutrophils, and its suppressive effects on NET formation were mitigated by blocking adenosine A2A receptor or by inhibiting the cyclic AMP-dependent kinase protein kinase A. Defibrotide at doses ranging from 15 to 150 mg/kg/day inhibited NET formation and venous thrombosis in a model of antiphospholipid antibody-accelerated thrombosis-an effect that was reduced in adenosine A2A receptor-knockout mice. CONCLUSION: This study is the first to demonstrate mechanisms by which defibrotide counteracts neutrophil-mediated thrombotic inflammation inherent to APS.

Protein Phosphatase 2A Activation Via ApoER2 in Trophoblasts Drives Preeclampsia in a Mouse Model of the Antiphospholipid Syndrome.

[Figure: see text].

Exosome-Contained APOH Associated With Antiphospholipid Syndrome.

Background: Antiphospholipid syndrome (APS) is a systemic autoimmune disease that can lead to thrombosis and/or pregnancy complications. Exosomes, membrane-encapsulated vesicles that are released into the extracellular environment by many types of cells, can carry signals to recipient cells to affect angiogenesis, apoptosis, and inflammation. There is increasing evidence suggesting that exosomes play critical roles in pregnancy. However, the contribution of exosomes to APS is still unknown. Methods: Peripheral plasma was collected from healthy early pregnancy patients (NC-exos) and early pregnancy patients with APS (APS-exos) for exosome extraction and characterization. The effect of exosomes from different sources on pregnancy outcomes was determined by establishing a mouse pregnancy model. Following the coincubation of exosomes and human umbilical vein endothelial cells (HUVECs), functional tests examined the features of APS-exos. The APS-exos and NC-exos were analyzed by quantitative proteomics of whole protein tandem mass tag (TMT) markers to explore the different compositions and identify key proteins. After incubation with HUVECs, functional tests investigated the characteristics of key exosomal proteins. Western blot analysis was used to identify the key pathways. Results: In the mouse model, APS-exos caused an APS-like birth outcome. In vitro experiments showed that APS-exos inhibited the migration and tube formation of HUVECs. Quantitative proteomics analysis identified 27 upregulated proteins and 9 downregulated proteins in APS-exos versus NC-exos. We hypothesized that apolipoprotein H (APOH) may be a core protein, and the analysis of clinical samples was consistent with finding from the proteomic TMT analysis. APOH-exos led to APS-like birth outcomes. APOH-exos directly enter HUVECs and may play a role through the phospho-extracellular signal-regulated kinase pathway. Conclusions: Our study suggests that both APS-exos and APOH-exos impair vascular development and lead to pregnancy complications. APOH-exos may be key actors in the pathogenesis of APS. This study provides new insights into the pathogenesis of APS and potential new targets for therapeutic intervention.

Cold Agglutinin-Mediated Autoimmune Hemolytic Anemia in Association with Antiphospholipid Syndrome.

Antiphospholipid syndrome and cold agglutinin-mediated autoimmune hemolytic anemia are 2 distinct immune-mediated hematologic disorders. While no clear association exists between these 2 entities, complement activation is known to occur in both of them. Herein, we report a unique case of cold agglutinin hemolytic anemia in a patient with a known primary antiphospholipid syndrome.

miR-19b-3p and miR-20a-5p are associated with the levels of antiphospholipid antibodies in patients with antiphospholipid syndrome.

Monocytes play a key role in pathophysiology of antiphospholipid syndrome (APS), nevertheless it is unclear if microRNA expression is associated with particular APS features. Identify whether miR-19b-3p and miR-20a-5p expression in monocytes are associated with hallmarks of the APS. Fifty-seven APS patients and 18 healthy controls were studied. Expression of miR-19b-3p and miR-20a-5p was measured in monocytes by RT-qPCR. Both miR-19b-3p (AUC = 0.835, 95% CI 0.733-0.938; P < 0.001) and miR-20a-5p (AUC = 0.857, 0.757-0.957; P < 0.001) discriminated APS patients from healthy individuals. A cut-off point of 1.98 for miR-19-3p and 2.18 for miR-20a-5p showed that APS patients with low microRNA expression had higher levels of IgM and IgG anticardiolipin antibodies than patients with high microRNA expression. In addition, APS patients with low microRNA expression had higher IgG anti-ß2 glycoprotein I antibody levels than their counterparts with high microRNA expression. Finally, miR-19b-3p and miR-20a-5p expression levels were significantly higher in APS patients using oral anticoagulants. Monocyte expression of miR-19b-3p and miR-20a-5p is low in APS, and patients with the lowest microRNA expression presented the highest levels of antiphospholipid antibodies.

Extracellular Vesicles and Antiphospholipid Syndrome: State-of-the-Art and Future Challenges.

Antiphospholipid syndrome (APS) is a systemic autoimmune disorder characterized by thromboembolism, obstetric complications, and the presence of antiphospholipid antibodies (aPL). Extracellular vesicles (EVs) play a key role in intercellular communication and connectivity and are known to be involved in endothelial and vascular pathologies. Despite well-characterized in vitro and in vivo models of APS pathology, the field of EVs remains largely unexplored. This review recapitulates recent findings on the role of EVs in APS, focusing on their contribution to endothelial dysfunction. Several studies have found that APS patients with a history of thrombotic events have increased levels of EVs, particularly of endothelial origin. In obstetric APS, research on plasma levels of EVs is limited, but it appears that levels of EVs are increased. In general, there is evidence that EVs activate endothelial cells, exhibit proinflammatory and procoagulant effects, interact directly with cell receptors, and transfer biological material. Future studies on EVs in APS may provide new insights into APS pathology and reveal their potential as biomarkers to identify patients at increased risk.

Extracellular Vesicles Tune the Immune System in Renal Disease: A Focus on Systemic Lupus Erythematosus, Antiphospholipid Syndrome, Thrombotic Microangiopathy and ANCA-Vasculitis.

Extracellular vesicles (EV) are microparticles released in biological fluids by different cell types, both in physiological and pathological conditions. Owing to their ability to carry and transfer biomolecules, EV are mediators of cell-to-cell communication and are involved in the pathogenesis of several diseases. The ability of EV to modulate the immune system, the coagulation cascade, the angiogenetic process, and to drive endothelial dysfunction plays a crucial role in the pathophysiology of both autoimmune and renal diseases. Recent studies have demonstrated the involvement of EV in the control of renal homeostasis by acting as intercellular signaling molecules, mediators of inflammation and tissue regeneration. Moreover, circulating EV and urinary EV secreted by renal cells have been investigated as potential early biomarkers of renal injury. In the present review, we discuss the recent findings on the involvement of EV in autoimmunity and in renal intercellular communication. We focused on EV-mediated interaction between the immune system and the kidney in autoimmune diseases displaying common renal damage, such as antiphospholipid syndrome, systemic lupus erythematosus, thrombotic microangiopathy, and vasculitis. Although further studies are needed to extend our knowledge on EV in renal pathology, a deeper investigation of the impact of EV in kidney autoimmune diseases may also provide insight into renal biological processes. Furthermore, EV may represent promising biomarkers of renal diseases with potential future applications as diagnostic and therapeutic tools.

VWF, Platelets and the Antiphospholipid Syndrome.

The antiphospholipid syndrome (APS) is characterized by thrombosis and/or pregnancy morbidity with the persistent presence of antiphospholipid antibodies (aPLs). Laboratory criteria for the classification of APS include the detection of lupus anticoagulant (LAC), anti-cardiolipin (aCL) antibodies and anti-ß2glycoprotein I (aß2GPI) antibodies. Clinical criteria for the classification of thrombotic APS include venous and arterial thrombosis, along with microvascular thrombosis. Several aPLs, including LAC, aß2GPI and anti-phosphatidylserine/prothrombin antibodies (aPS/PT) have been associated with arterial thrombosis. The Von Willebrand Factor (VWF) plays an important role in arterial thrombosis by mediating platelet adhesion and aggregation. Studies have shown that aPLs antibodies present in APS patients are able to increase the risk of arterial thrombosis by upregulating the plasma levels of active VWF and by promoting platelet activation. Inflammatory reactions induced by APS may also provide a suitable condition for arterial thrombosis, mostly ischemic stroke and myocardial infarction. The presence of other cardiovascular risk factors can enhance the effect of aPLs and increase the risk for thrombosis even more. These factors should therefore be taken into account when investigating APS-related arterial thrombosis. Nevertheless, the exact mechanism by which aPLs can cause thrombosis remains to be elucidated.

Micronized / ultramicronized palmitoylethanolamide (PEA) as natural neuroprotector against COVID-19 inflammation.

Coronavirus Disease 2019 (COVID-19) is upsetting the world and innovative therapeutic solutions are needed in an attempt to counter this new pandemic. Great hope lies in vaccines, but drugs to cure the infected patient are just as necessary. In the most severe forms of the disease, a cytokine storm with neuroinflammation occurs, putting the patient's life at serious risk, with sometimes long-lasting sequelae. Palmitoylethanolamide (PEA) is known to possess anti-inflammatory and neuroprotective properties, which make it an ideal candidate to be assumed in the earliest stage of the disease. Here, we provide a mini-review on the topic, pointing out phospholipids consumption in COVID-19, the possible development of an antiphospholipid syndrome secondary to SARS-CoV-2 infection, and reporting our preliminary single-case experience concerning to a 45-year-old COVID-19 female patient recently treated with success by micronized / ultramicronized PEA.

Autoantibodies to Annexin A2 and cerebral thrombosis: Insights from a mouse model.

INTRODUCTION: Antiphospholipid syndrome (APS) is an autoimmune disorder manifested by thromboembolic events, recurrent spontaneous abortions and elevated titers of circulating antiphospholipid antibodies. In addition, the presence of antiphospholipid antibodies seems to confer a fivefold higher risk for stroke or transient ischemic attack. Although the major antigen of APS is ß2 glycoprotein I, it is now well established that antiphospholipid antibodies are heterogeneous and bind to various targets. Recently, antibodies to Annexin A2 (ANXA2) have been reported in APS. This is of special interest since data indicated ANXA2 as a key player in fibrinolysis. Therefore, in the present study we assessed whether anti-ANXA2 antibodies play a pathological role in thrombosis associated disease. MATERIALS AND METHODS: Mice were induced to produce anti-ANXA2 antibodies by immunization with ANXA2 (iANXA2) and control mice were immunized with adjuvant only. A middle cerebral artery occlusion stroke model was applied to the mice. The outcome of stroke severity was assessed and compared between the two groups. RESULTS: Our results indicate that antibodies to ANXA2 lead to a more severe stroke as demonstrated by a significant larger stroke infarct volume (iANXA2 133.9 ± 3.3 mm3 and control 113.7 ± 7.4 mm3; p = 0.017) and a more severe neurological outcome (iANXA2 2.2 ± 0.2, and control 1.5 ± 0.18; p = 0.03). CONCLUSIONS: This study supports the hypothesis that auto-antibodies to ANXA2 are an independent risk factor for cerebral thrombosis. Consequently, we propose screening for anti-ANXA2 antibodies should be more widely used and patients that exhibit the manifestations of APS should be closely monitored by physicians.

Artemisinin analogue SM934 protects against lupus-associated antiphospholipid syndrome via activation of Nrf2 and its targets.

Kidney is a major target organ in both antiphospholipid syndrome (APS) and systemic lupus erythematosus (SLE). The etiology of antiphospholipid syndrome nephropathy associated lupus nephritis (APSN-LN) is intricate and remains largely unrevealed. We proposed in present work, that generation of antiphospholipid antibodies (aPLs), especially those directed towards the oxidized neoepitopes, are largely linked with the redox status along with disease progression. Moreover, we observed that compromised antioxidative capacity coincided with turbulence of inflammatory cytokine profile in the kidney of male NZW×BXSB F1 mice suffered from APSN-LN. SM934 is an artemisinin derivative that has been proved to have potent immunosuppressive properties. In current study, we elaborated the therapeutic benefits of SM934 in male NZW×BXSB F1 mice, a murine model develops syndrome resembled human APS associated with SLE, for the first time. SM934 treatment comprehensively impeded autoantibodies production, inflammatory cytokine accumulation and excessive oxidative stress in kidney. Among others, we interpreted in present work that both anti-inflammatory and antioxidative effects of SM934 is closely correlated with the enhancement of Nrf2 signaling and expression of its targets. Collectively, our finding confirmed that therapeutic strategy simultaneously exerting antioxidant and anti-inflammatory efficacy provide a novel feasible remedy for treating APSN-LN.