Estrés oxidativo en células endoteliales inducido por el suero de mujeres con diferentes manifestaciones clínicas del síndrome antifosfolípido

Abstract

Introducción. El síndrome antifosfolípido se caracteriza por la presencia persistente de anticuerpos antifosfolípidos y manifestaciones clínicas de trombosis o morbilidad gestacional, las cuales se asocian con estrés oxidativo y disfunción endotelial.Objetivo. Evaluar los marcadores de estrés oxidativo en células endoteliales, inducidos por el suero de mujeres con diferentes manifestaciones clínicas del síndrome antifosfolípido y analizar la capacidad antioxidante de los sueros.Materiales y métodos. Se incluyeron 48 mujeres que fueron clasificadas así: presencia de anticuerpos antifosfolípidos y criterios clínicos de morbilidad gestacional, trombosis vascular o ambas. Como grupos control se incluyeron mujeres negativas para anticuerpos antifosfolípidos. En un modelo in vitro de células endoteliales estimuladas con los sueros de las mujeres del estudio, se determinaron algunos marcadores de estrés oxidativo por citometría de flujo. También, se analizó la capacidad antioxidante de los sueros incluidos.Resultados. Los sueros de los grupos de mujeres con síndrome antifosfolípido que presentaban trombosis, con morbilidad gestacional o sin ella, generaron un incremento significativo (p<0,05 y p<0,001) en los marcadores de estrés oxidativo endotelial, en contraste con el control de suero humano normal. No se observaron diferencias en el efecto de los sueros de los diferentes grupos de estudio sobre la lipoperoxidación endotelial. Tampoco se encontró diferencia en la actividad antioxidante de los sueros.Conclusión. El estrés oxidativo mitocondrial en el endotelio se asocia con la presencia de trombosis. Sin embargo, cuando esta se asocia con morbilidad gestacional, también se genera estrés oxidativo intracelular

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

“…Since both depolarization and hyperpolarization involve dissociation of the electron transport chain and an increase of oxygen consumption, these conditions can be harmful to the cell through the induction of oxidative or reductive stress ( Guo et al, 2011 ; Oaks et al, 2016 ; Zorova et al, 2018 ). We previously reported that serum from women with PM/VT increased the intracellular and mitochondrial production of ROS in HUVECs ( Velásquez et al, 2019 ), the same group of patients whose IgGs induced the high MMP (hyperpolarization) in this study. Taken together, these findings suggest a mechanism by which aPL from women with PM/VT induced HUVEC oxidative stress.…”

“…Human umbilical vein endothelial cells (HUVEC) were isolated from umbilical cords obtained from uncomplicated pregnancies based on a modified protocol by Jaffe et al (1973) and as previously described ( Velásquez et al, 2019 ; Gil-Villa et al, 2020 ). In brief, umbilical veins were perfused with 100 μg/ml type I collagenase (Invitrogen, Waltham, MA, United States) and incubated for 20 min at 37°C.…”

“…Dilute Russell’s viper venom time (dRVVT) screen and dRVVT confirmation (Instrumentation Laboratory) were used to detect LA. In addition, other non-criteria aPL were detected using an in-house ELISA standardized by the reproduction group based on the technique published by Kwak et al (1992) and as previously described ( Velásquez et al, 2019 ). In brief, U-bottom 96-well polystyrene microplates (Maxisorp Nunc TM , Thermo Fisher Scientific) were covered with 30 μl of 50 μg/ml of the following phospholipids suspended in methanol: phosphatidylglycerol, phosphatidic acid, phosphatidylserine, phosphatidylethanolamine, and phosphatidylinositol (Sigma-Aldrich, Saint Louis, MO, United States).…”

“…Human umbilical vein endothelial cells (HUVEC) were isolated from umbilical cords obtained from uncomplicated pregnancies based on a modified protocol by Jaffe et al (1973) and as previously described (Velásquez et al, 2019;Gil-Villa et al, 2020). In brief, umbilical veins were perfused with 100 µg/ml type I collagenase (Invitrogen, Waltham, MA, United States) and incubated for 20 min at 37 • C. Cells were recovered, and after centrifugation (50 g for 5 min), they were seeded in the endothelial cell growth medium (Promocell, Heidelberg, Germany) supplemented with 2% fetal bovine serum (FBS, Gibco, Waltham, MA, United States), 100 U/ml penicillin (Sigma Aldrich, Missouri, United States), 50 µg/ml gentamicin (Genfar, Bogotá, Colombia), and 0.25 µg/ml amphotericin B (Vitalis, Bogotá, Colombia).…”

Obstetric antiphospholipid syndrome: An approach from glycans of the immunoglobulin G

“…The clinical and laboratory features of women included in this study are outlined in Table 1 . Women in this study were classified as Pregnancy Morbidity/Vascular Thrombosis (PM/VT), women that fulfilled the clinical and laboratory revised Sapporo classification criteria for APS (n=7); Seronegative-Obstetric APS (SN-OAPS), women who present clinical features consistent with a diagnosis of obstetric APS but tested persistently negative for conventional anti-cardiolipin (aCL), anti-β2glicoprotein-I (aβ2GPI) and lupus anticoagulant (LA) tests[ 4 ], these women were positive for the non-criteria aPL[ 5 ] (n=8), and Pregnancy Morbidity/aPL- (PM/aPL-), women with a history of gestational morbidity, without autoimmune or chronic diseases, negative for both non-criteria aPL and conventional tests (n=10). These women were compared with a Normal Human Serum (NHS) group, healthy women with previous uncomplicated pregnancies (n= 7).…”