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Lupus nephropathy is a severe and frequent complication of systemic lupus erythematosus. Here, we assessed the biomarkers of oxidative stress, inflammation and disease activity in patients with lupus nephritis. Thirty-four patients with active lupus nephritis, 31 patients with inactive lupus nephritis and 20 lupus patients without renal damage (non-lupus nephritis) were studied. Oxidative stress biomarkers malonyldialdehyde, oxidized-to-total glutathione, catalase, superoxide dismutase and total antioxidant status were assessed, as well as inflammation biomarkers CRP, interleukin 6 and monocyte chemoattractant protein 1. Renal tubular disease biomarkers neutrophil gelatinase-associated lipocalin and β2-microglobulin were assessed, together with the classic disease activity biomarkers urinary protein/creatinine ratio, anti-dsDNA, anti-C1q antibody and complement proteins C3 and C4. Significant differences were found between active lupus nephritis and inactive lupus nephritis patients and between active lupus nephritis and non-lupus nephritis patients for all the assessed biomarkers ( P < 0.05), except for catalase, superoxide dismutase and interleukin 6. There is an imbalance in the redox status in active lupus nephritis patients that would be involved in lipid peroxidation of the glomerular basal membrane that would alter its integrity and could also affect renal tubular function in these patients.

Ethical approval: The study was approved by the Bioethics Comitee of Facultad de Ciencias Bioquímicas y Farmacéuticas (Universidad Nacional de Rosario). Res Nº 076/2008. All participants signed the corresponding informed consent. Guarantor: SMMA Contributorship: HFP performed most assays and analysis of results, and wrote the paper. EP and CB performed some assays and collaborated in the analysis of results. JM performed patients selection for the study and aided in medical matters. MB, MJS and SMD performed sample collection and conditioning, as well as some laboratory determinations. HB performed statistical analysis. SMMA coordinated and supervised the whole study. Abstract word count: 251. Actual word count (excluding abstract and references): 2664 AbstractBackground: Antiphospholipid syndrome (APS) is an autoimmune disease characterized by thrombosis, fetal losses and thrombocytopenia associated to antiphospholipid (APL) antibodies (Abs). They are directed to phospholipids, such as cardiolipins (a-cardiolipin, ACA) and lupus anticoagulant (LA), or to complexes formed by phospholipids and protein cofactors, such as β2 glycoprotein 1 (a-β2GP1) and annexin V (a-annexin V). These auto Abs may be considered as a family of Abs involved in thrombotic events and APL activity. On the other hand, some proangiogenic factors are involved in the normal development of placental vasculature, such as the vascular endothelial growth factor (VEGF). Overexpression of VEGF receptor in its soluble form (sVEGFR-1) has been associated to a higher antiangiogenic activity. Our aim was to analyze the association between ACA, LA, a-β2GP1, a-annexin V and sVEGFR-1 with recurrent miscarriage before week 10 of gestation in women with APS. Methods: We studied 24 women (primary or secondary APS), who were divided into two groups: women with recurrent miscarriage before week 10 of gestation [M; n=12] and women with no history of fetal loss [NM; n=12]. ACA, a-β2GP1, aannexin V and sVEGF-R1 levels were assessed by ELISA, while LA was assessed by screening and confirmatory tests. Results: A significant association was observed between the number of positive biomarkers and the belonging group (p<0.05). Besides, a positive result for LA and a-β2GP1 was found to be significantly associated to the M group (p<0.05). Conclusions: LA and a-β2GP1 may be implicated in pregnancies complicated by recurrent miscarriage in women with APS.

Foi desenvolvido e validado um ELISA tipo sanduíche "in house" para a quantificação de anticorpos (Ac) anti-anexina V em plasma. Os parâmetros de validação estudados foram: (i) precisão, expressado como coeficiente de variação (CV) inter-e intra-ensaio, (ii) exatidão, expressado como porcentagem de desvio entre o valor obtido e o valor real, (iii) limite de detecção (LOD), avaliado a partir do branco de reagentes e (iv) robusteza, obtida através da introdução propositada de pequenas variações em diferentes parâmetros. Além disso, a técnica "in house" foi comparada com um método comercial. Encontrou-se que ambos CV foram < 20%, a exatidão foi de 100 ± 20%, o limite de detecção foi menor que 1 U mL -1 e as pequenas variações na técnica não produziram variações significativas nos resultados. A comparação com o método comercial mostrou uma correlação aceitável. Concluiu-se que o método desenvolvido cumpre satisfatoriamente com os parâmetros de padronização e validação para imuno-análise.An "in house" sandwich ELISA for the quantification of plasma anti-annexin V antibodies was developed and validated. The validation parameters studied were: (i) precision, expressed either as the intra-or the inter-assay coefficient of variation (CV), (ii) exactitude, expressed as the percentage deviation between the obtained value and the real value, (iii) limit of detection (LOD), evaluated from the reagents blank and (iv) robustness, obtained by deliberately introducing slight variations in different parameters. Also, a comparison between the "in house" technique and a commercial method was performed. The research revealed that both CV were < 20%, exactitude was within the 100 ± 20% range, limit of detection was below 1 U mL -1 and that slight variations in the technique did not produce any significant variations in the results. Comparison with the commercial method showed an acceptable correlation. It was concluded that the method developed here satisfactorily accomplishes the parameters of standardization and validation for an immunoassay.Keywords: ELISA, annexin V, antibodies, anti-phospholipid syndrome, foetal losses IntroductionAnnexins belong to a family of proteins that are able to bind to negatively charged phospholipids and membrane bilayers through calcium dependent interactions. Though their fine structure has been well described, their functions have not been clearly identified yet.1 Like others, annexins constitute a group of ubiquitous cytoplasmic proteins involved in signal transduction. 2Annexin V is a 320-amino acid-residue, 36-kDa-protein that is folded into a planar cyclic arrangement of four repeats with each repeat composed of five alpha-helical segments. 3,4 It is expressed in various cell types, including placental trophoblasts and vascular endothelial cells. This protein is highly expressed in an apparently constitutive manner by placental trophoblasts and is displayed Quantification of Autoantibodies to Annexin V in Plasma by an "In House" Sandwich ELISA J. Braz. Chem. Soc. 454 on the apical membrane o...