Relevance of non-criteria antiphospholipid antibodies titers among patients with antiphospholipid syndrome and patients with systemic lupus erythematosus.

BACKGROUND: There is an increasing interest in the study of non-criteria antiphospholipid antibodies (aPL) including antibodies targeting domain 1 of the B2 glycoprotein 1 (anti-D1 B2GP1) and antibodies anti phosphatidylserine/ prothrombin (PS/PT). OBJECTIVES: Our aim was to analyze a panel of conventional and non-criteria aPL in a cohort of patients with systemic lupus erythematosus (SLE) and primary antiphospholipid syndrome (APS), to describe if there are differences in aPL titers among groups, to evaluate clinical associations including risk of recurrent events of novel aPL. METHODS: Observational study that evaluated at baseline antibodies against anti-D1 B2GP1 and anti PS/PT. Anti-D1 B2GP1 antibodies were tested using a chemiluminescent immunoassay. IgG and IgM anti PS/PT, aCL and anti B2GP1 by ELISA techniques. Therefore, patients were followed in order to identify new thrombotic events. RESULTS: 133 patients with SLE and 23 with primary APS patients were included. Main APS manifestations were DVT (27%), obstetric morbidity (22%) and arterial thrombosis (10.1%). IgM anti PS/PT antibodies levels were (20.6 - 127) vs 21.9 (11.2 - 39.2) U/ml, p<0.001 in primary APS vs SLE with APS, respectively. Anti-D1 B2GP1, IgG and IgM anti PS/PT were associated with thrombotic and non-thrombotic manifestations. During follow-up, IgG B2GP1 were related with a significant cumulative risk of thrombosis. CONCLUSIONS: We found significant differences in serum titers of non-criteria aPL among patients with primary APS vs SLE with APS. Whether non-criteria aPL antibodies titers are useful to differentiate patients with primary and secondary APS requires further analysis in other populations.

Extracellular vesicles released upon stimulation with antiphospholipid antibodies: An actual direct procoagulant mechanism or a new factor in the lupus anticoagulant paradox?

Antiphospholipid antibodies (aPL) lead to a hypercoagulable state in vivo. Paradoxically, some of these autoantibodies perform as inhibitors of the coagulation cascade in vitro (a phenomenon referred to as "lupus anticoagulant"). The presence of lupus anticoagulant has been related to an increased quantity of plasma extracellular vesicles, which may constitute a direct procoagulant mechanism in antiphospholipid syndrome. This study investigates whether or not endothelial cell-derived extracellular vesicles released upon stimulation with aPL (aPL-EDEVs) are related to a higher direct coagulation activity. Using an in vitro model of endothelium, flow cytometry and a recalcified plasma-based assay, we found that the coagulation activity of aPL-EDEVs is mainly conditioned by the lupus anticoagulant-like activity of autoantibodies. Nevertheless, in the presence of ß2 glycoprotein I, a cofactor of aPL during the stimulation of endothelial cells, the coagulation activity of EDEVs is restored in a mitogen-activated protein kinase kinases 1 and 2 (MEK1/2)-dependent manner. This phenomenon was especially evident when using immunoglobulins G from patients with vascular and obstetric primary antiphospholipid syndrome who manifest refractoriness to treatment. Our findings suggest that the role of aPL-EDEVs in the antiphospholipid syndrome-related hypercoagulable state may not rely on their capacity to enhance clotting directly. While ß2 glycoprotein I performs as a procoagulant cofactor and restores the coagulation activity of extracellular vesicles via MEK1/2 pathway, proportionally, autoantibodies interact with aPL-EDEVs and exhaust their coagulation properties. Further analysis is required to establish whether lupus anticoagulant-like autoantibodies opsonise extracellular vesicles and whether opsonised vesicles may lead to thrombosis by indirect means.

Antiphospholipid Antibodies From Women With Pregnancy Morbidity and Vascular Thrombosis Induce Endothelial Mitochondrial Dysfunction, mTOR Activation, and Autophagy.

Antiphospholipid syndrome (APS) is an autoimmune disease characterized by thrombosis and pregnancy morbidity (PM) obstetric events together with persistent high titers of circulating antiphospholipid antibodies (aPL). Several mechanisms that explain the development of thrombosis and PM in APS include the association of aPL with alterations in the coagulation cascade and inflammatory events. Other mechanisms disturbing cellular homeostases, such as mitochondrial dysfunction, autophagy, and cell proliferation, have been described in other autoimmune diseases. Therefore, the objective of this study was to investigate the impact of aPL from different patient populations on endothelial cell mitochondrial function, activation of the mammalian target of rapamycin (mTOR) and autophagy pathways, and cellular growth. Using an in vitro model, human umbilical vein endothelial cells (HUVECs) were treated with polyclonal immunoglobulin G (IgG) purified from the serum of women with both PM and vascular thrombosis (PM/VT), with VT only (VT), or with PM and non-criteria aPL (seronegative-obstetric APS, SN-OAPS). We included IgG from women with PM without aPL (PM/aPL-) and healthy women with previous uncomplicated pregnancies (normal human serum, NHS) as control groups. Mitochondrial function, mTOR activation, autophagy, and cell proliferation were evaluated by Western blotting, flow cytometry, and functional assays. IgG from women with PM/VT increased HUVEC mitochondrial hyperpolarization and activation of the mTOR and autophagic pathways, while IgG from patients with VT induced endothelial autophagy and cell proliferation in the absence of elevated mTOR activity or mitochondrial dysfunction. IgG from the SN-OAPS patient group had no effect on any of these HUVEC responses. In conclusion, aPL from women with PM and vascular events induce cellular stress evidenced by mitochondrial hyperpolarization and increased activation of the mTOR and autophagic pathways which may play a role in the pathogenesis of obstetric APS.

Frequency of Aspirin Resistance in Ischemic Stroke Patients and Healthy Controls from Colombia.

OBJECTIVE: To evaluate the aspirin resistance prevalence in patients with previous ischemic cerebrovascular disease undergoing aspirin therapy for secondary prevention. MATERIALS AND METHODS: Three hundred fifty patients presenting ischemic strokes and 100 healthy controls under aspirin treatment were evaluated using the optic platelet aggregation test. RESULTS: Aspirin resistance was found in 7.4% of the patients with ischemic stroke and 4% of controls. Aspirin resistance was associated with stroke recurrence in univariate analysis (p = 0.004). Aspirin resistance was not associated with smoking, diabetes, or hypercholesterolemia. CONCLUSION: Aspirin resistance is present in Colombian patients with ischemic stroke as well as in healthy controls.

Microparticles: An Alternative Explanation to the Behavior of Vascular Antiphospholipid Syndrome.

Antiphospholipid syndrome is an autoimmune disease characterized by the persistent presence of antiphospholipid antibodies, along with occurrence of vascular thrombosis and pregnancy morbidity. The variety of antiphospholipid antibodies and their related mechanisms, as well as the behavior of disease in wide groups of patients, have led some authors to propose a differentiation of this syndrome into two independent entities: vascular and obstetric antiphospholipid syndrome. Thus, previous studies have discussed whether specific autoantibodies may be responsible for this differentiation or, in contrast, how the same antibodies are able to generate two different clinical presentations. This discussion is yet to be settled. The capability of serum IgG from patients with vascular thrombosis to trigger the biogenesis of endothelial cell-derived microparticles in vitro is one of the previously discussed differences between the clinical entities of antiphospholipid syndrome. These vesicles constitute a prothrombotic mechanism as they can directly lead to clot activation in murine models and recalcified human plasma. Nevertheless, other indirect mechanisms by which microparticles can spread a procoagulant phenotype could be critical to understanding their role in antiphospholipid syndrome. For this reason, questions regarding the cargo of microparticles, and the signaling pathways involved in their biogenesis, are of interest in attempting to explain the behavior of this autoimmune disease.

Bioseguridad en los equipos de aerosolterapia por cuidadores en atención domiciliaria / Biosecurity in aerosolterapy equipament by caregivers in home care

Objetivo: Identificar el proceso de bioseguridad en los equipos de aerosolterapia por parte de cuidadores en atención domiciliaria en la ciudad de Cali, Colombia, en el año 2016. Método: Se realizó un estudio observacional, descriptivo y de corte transversal. La población de estudio correspondió a 54 adultos, cuidadores primarios (n=16) y profesionales (n=38) pertenecientes al programa de terapia respiratoria de la IPS SISANAR. Se diseñó un instrumento de recolección de datos, tipo encuesta. Fue utilizado el paquete estadístico Stata 10 para el análisis de los datos, los cuales fueron revisados y validados. Las frecuencias y porcentajes se calcularon, así como los intervalos de confianza al 95%. Resultados: Se encontró que el 96.3% de los participantes realiza el lavado de manos clínico antes de ejecutar la terapia nebulizada, de los cuales el 55.8% usa tapa- bocas para el proceso de lavado de los micronebulizadores, el 28.8% usa guantes y el 15.4% no uti- liza ninguno. La desinfección de los micronebulizadores la realizó el 59.3% de los participantes. Esta desinfección es realizada principalmente con hipoclorito y gel antibacterial. Discusión: Es importante formalizar la educación oportuna por medio de protocolos, dirigidos a los pacientes y cuidadores primarios frente a los procesos de limpieza y bioseguridad en el mantenimiento de los equipos de aerosolterapia en atención domiciliaria, lo cual permitirá la disminución de sobreinfecciones asociadas con microorganismos presentes en los equipos.

Objective: To identify the biosecurity process in aerosol therapy equipment used by caregivers and professionals in Respiratory Therapy in a home-based healthcare program in the city of Cali, Colombia, in 2016. Methods: An observational, descriptive and cross-sectional study was performed. The studied population was composed of 54 adults, caregivers (n = 16) and profes- sionals (n = 38) that belonged to the respiratory therapy program of the IPS SISANAR. A sur- vey-type data collection instrument was designed. The statistical package Stata 10 was used for data analysis, which was reviewed and validated. Frequencies and percentages were calculated as well as 95% confidence intervals. Results: It was found that 96.3% of the participants performed clinical handwashing before performing nebulized therapy, where 55.8% used masks to wash the micronebulizers, 28.8% used gloves and 15.4% did not use any. The micronebulizers' disinfection was performed by 59.3% of the participants. This disinfection is mainly done with hypochlorite and hand sanitizer. Discussion: It is important to formalize a proper education through proto- cols for patients and caregivers, to teach them about the cleaning and biosafety processes for the maintenance of the aerosol therapy equipment in home-based healthcare. This will allow a reduction of superinfections associated with microorganisms present in the equipment

An acidic phospholipase A2 with antibacterial activity from Porthidium nasutum snake venom.

Snake venoms are complex mixtures of proteins among which both basic and acidic phospholipases A(2) (PLA(2)s) can be found. Basic PLA(2)s are usually responsible for major toxic effects induced by snake venoms, while acidic PLA(2)s tend to have a lower toxicity. A novel PLA(2), here named PnPLA(2), was purified from the venom of Porthidium nasutum by means of RP-HPLC on a C18 column. PnPLA(2) is an acidic protein with a pI of 4.6, which migrates as a single band under both non-reducing and reducing conditions in SDS-PAGE. PnPLA(2) had a molecular mass of 15,802.6 Da, determined by ESI-MS. Three tryptic peptides of this protein were characterized by HPLC-nESI-MS/MS, and N-terminal sequencing by direct Edman degradation showing homology to other acidic PLA(2)s from viperid venoms. PnPLA(2) displayed indirect hemolytic activity in agarose erythrocyte-egg yolk gels and bactericidal activity against Staphylococcus aureus in a dose-dependent manner, with a MIC and MBC of 32 µg/mL. In addition, PnPLA(2) showed a potent inhibitory effect on platelet aggregation with doses up to 40 µg/mL. This acidic PLA(2), in contrast to basic enzymes isolated from other viperid snake venoms, was not cytotoxic to murine skeletal muscle myoblasts C(2)C(12). This is the first report on a bactericidal protein of Porthidium nasutum venom.