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Proteomics in biomarker discovery for clinical purposes

Publication . Penque, Deborah

Clinical Proteomics Dedicated to the study of the PROTEOME PROFILE associated with the HEALTHY AND DISEASE STATE, in the search for DIAGNOSTIC / PROGNOSTIC / MONITORING BIOMARKERS or as TARGETS for the development of new therapeutic approaches

2015-09-18Lecture

Open access

Obstructive Sleep Apnea: New insights into antioxidant activity and cellular response to stress

Publication . Vaz, Fátima; Feliciano, Amélia; Silva, Ana Rita; Coelho, Cristina; Torres, Vukosava Millic; Bozanic, Vesna; Malhotra, Atul; Pinto, Paula; Bárbara, Cristina; Penque, Deborah

Obstructive sleep apnea (OSA) is a sleep¬related breathing disorder characterized by recurrent episodes of apnea/and hypopnea during sleep with resultant intermittent hypoxia and sleep fragmentation, leading to cardiometabolic diseases. OSA affects 3.7% to 26% of adult population, but frequently is underdiagnosed. By 2DIGE-proteomics approach, we have investigated red blood cells (RBC) in OSA to uncover new insights into putative chronic stress-induced RBC dysfunction that lead to inflammation and metabolic syndrome associated with OSA. A number of proteins as potential candidate biomarkers for OSA, such as the cytosolic antioxidant regulators, catalase (CAT) and peroxiredoxin2 (PRX2) was identified, needing additional biochemical research validation.

2015-09-28Conference object

Open access

Proteomics of Red Blood Cells from Patients with Obstructive Sleep Apnea

Publication . Feliciano, Amelia; Bozanic, Vesna; Torres, Vukosava Milic; Matthiesen, Rune; Carvalho, Ana S.; Almeida, Andreia; Alexandre, Bruno; Vaz, Fátima; Malhotra, Atul; Pinto, Paula; Bárbara, Cristina; Penque, Deborah

Obstructive sleep apnea (OSA) is a common public health concern causing metabolic and cardiovascular consequences. Although OSA is a systemic disease, the molecular mechanisms and specific genes/proteins associated with such processes remain poorly defined.To identify dysregulated proteins that could be useful as candidate biomarkers of diagnosis/prognosis of OSA., is the aim of this study

2015-09-28Conference object

Open access

Obstructive Sleep Apnea: a proteomics study of the effects of positive airway pressure therapy

Publication . Valentim-Coelho, Cristina; Vaz, Fátima; Martins, Inês L.; Feliciano, Amélia; Pinto, Paula; Cristina, Bárbara; Penque, Deborah

Obstructive Sleep Apnea (OSA) syndrome is a common public health concern characterized by recurrent episodes of apneas and hypopneas during sleep. These obstructive events result in recurrent intermittent hypoxia and sleep fragmentation that can lead to metabolic and cardiovascular diseases. We recently demonstrated that OSA can cause alterations in the red blood cells (RBC) proteome that may be associated with OSA outcomes1,2. Here we intend to investigate whether the first-line therapy for OSA, the positive airway pressure (PAP) can revert or modulate these proteome alterations. RBCs from Snorers and patients with severe OSA before/after 6 months of PAP treatment (n=10/condition) were depleted of hemoglobin, analyzed by 2D-DIGE using Progenesis SameSpotsv4.5. The differentially abundant proteins were identified by MALDI-MS/MS and protein annotations acquired by DAVIDv6.8. Western blotting (WB) validation was performed for Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and (overoxidized) GAPDHSO3 on a new Cohort (n=59). SPSS software was used to correlation studies with peroxiredoxin-2 (PRDX2) redox-oligomeric forms and several clinical parameters. Ten protein spots showed significant differences (Anova p<0.05) among groups and were associated with cell death, protein oligomerization and response to stress. Three proteoforms of GAPDH were identified decreased in OSA RBC (Anova p<0.05) and 6 months of PAP treatment increased these GAPDH proteoforms to the control levels. By WB, we confirmed these data by showing that the decreased GAPDH monomeric/tetrameric forms in OSA were increased by PAP treatment. PAP also increased GAPDHSO3 tetramers. In OSA, GAPDH monomers and GAPDHSO3 tetramers correlated positively with the respiratory disturbance index or triglycerides and adrenalin, respectively. After PAP, GAPDHSO3 tetramers correlated positively with PAP-induced PRDX2SO2/3 decameric forms, described having chaperone activity in cell protection. OSA induces alterations in the redox/oligomeric state of GAPDH and PRDX2 that can be reverted/modulated by PAP therapy. The clinical significant of these findings needs further validation and investigation.

2019-07-08Conference object

Restricted access

Shotgun proteomics of red blood cells from obstructive sleep apnea patients under positive airway pressure (PAP) treatment

Publication . Coelho, Cristina Valentim; Osório, Hugo; Vaz, Fatima; Neves, Sofia; Pinto, Paula; Barbara, Cristina; Penque, Deborah

Obstructive Sleep Apnea (OSA) syndrome is characterized by recurrent episodes of apneas and hypopneas during sleep, leading to recurrent intermittent hypoxia and sleep fragmentation. No treated OSA can result in metabolic and cardiovascular diseases. By 2D gel-based proteomics approach we have demonstrated that OSA can cause alterations in the red blood cells (RBC) proteome that may be associated with OSA outcomes. OSA induces alterations in the redox/oligomeric states of RBC proteins such as gyceraldehyde-3-phosphate dehydrogenase (GAPDH) and peroxiredoxin-2 (PRDX2) that can be reverted or modulated by PAP treatment. In this study, we applied a shotgun proteomics strategy to further investigate the RBC proteome from patients with OSA before and after PAP treatment to better understand the regulation of RBC homeostasis in the context of OSA and/or under effect of PAP treatment. As a first approach, RBCs samples, corresponding to Snorers patients as control (n=23) and patients with OSA before and after six months of PAP treatment (n=33/condition) were selected from our biobank1. Samples were randomly pooled (n=3 per group/condition) and lysed 1:6 with 5mM sodium phosphate buffer containing 100 mM of N-ethylmaleimide, a reagent that alkylates free sulfhydryl groups, before haemoglobin depletion by using HemovoidTM system. Depleted samples were alkylated, reduced and digested with trypsin and chymotrypsin. The resulting peptides were cleaned with C18 columns and analysed in triplicate by a Nano High Performance Liquid Chromatography (nanoHPLC) on-line coupled to a high-resolution accurate-mass Orbitrap mass spectrometer (Q Exactive, Thermo Scientific) with a nano electrospray ionization source (nanoESI). The acquired mass spectrometry data were analysed by MaxQuant v1.5.8.3 and Perseus v2.0.3.1 software. The preliminary results corroborated our previous findings by showing that proteins associated with stress response and antioxidant regulatory system were the most changed in OSA RBC compared with Snorers ones. The active catalytic cysteine (Cys 51) in the PRDX2 was identified trioxidized –SO3H almost exclusively in OSA RBC before PAP treatment. Further analyses and validation of these data are in progress, which will certainly provide a better understanding of RBC molecular mechanisms and their proteins/PTMs associated with OSA pathology and/or response to PAP therapy.

2022-05Conference object

Open access