Stefano Barelli

Plasma/serum proteomics: pre-analytical issues

High-throughput proteomics technologies tend to provide highly sensitive information about living tissues and biological fluids. Analytes are characterized by intrinsic and extrinsic properties, the latter depending on each phase of their preparation, sometimes adding artifacts with crucial repercussions in result reliability and interpretation. This review aims to address some issues that can be encountered when handling plasma and serum in experimental and clinical proteomic settings.

Oxidation of proteins: Basic principles and perspectives for blood proteomics

Protein oxidation mechanisms result in a wide array of modifications, from backbone cleavage or protein crosslinking to more subtle modifications such as side chain oxidations. Protein oxidation occurs as part of normal regulatory processes, as a defence mechanism against oxidative stress, or as a deleterious processes when antioxidant defences are overcome. Because blood is continually exposed to reactive oxygen and nitrogen species, blood proteomics should inherently adopt redox proteomic strategies. In this review, we recall the biochemical basis of protein oxidation, review the proteomic methodologies applied to analyse redox modifications, and highlight some physiological and in vitro responses to oxidative stress of various blood components.

Hepcidin as a new biomarker for detecting autologous blood transfusion.

Autologous blood transfusion (ABT) is an efficient way to increase sport performance. It is also the most challenging doping method to detect. At present, individual follow‐up of haematological variables via the athlete biological passport (ABP) is used to detect it. Quantification of a novel hepatic peptide called hepcidin may be a new alternative to detect ABT. In this prospective clinical trial, healthy subjects received a saline injection for the control phase, after which they donated blood that was stored and then transfused 36 days later. The impact of ABT on hepcidin as well as haematological parameters, iron metabolism, and inflammation markers was investigated. Blood transfusion had a particularly marked effect on hepcidin concentrations compared to the other biomarkers, which included haematological variables. Hepcidin concentrations increased significantly: 12 hr and 1 day after blood reinfusion, these concentrations rose by seven‐ and fourfold, respectively. No significant change was observed in the control phase. Hepcidin quantification is a cost‐effective strategy that could be used in an “ironomics” strategy to improve the detection of ABT. Am. J. Hematol. 91:467–472, 2016.

Urinary di‐(2‐ethylhexyl) phthalate metabolites for detecting transfusion of autologous blood stored in plasticizer‐free bags

Autologous blood transfusion (ABT) efficiently increases sport performance and is the most challenging doping method to detect. Current methods for detecting this practice center on the plasticizer di(2‐ethlyhexyl) phthalate (DEHP), which enters the stored blood from blood bags. Quantification of this plasticizer and its metabolites in urine can detect the transfusion of autologous blood stored in these bags. However, DEHP‐free blood bags are available on the market, including n‐butyryl‐tri‐(n‐hexyl)‐citrate (BTHC) blood bags. Athletes may shift to using such bags to avoid the detection of urinary DEHP metabolites.

Generation of procoagulant collagen‐ and thrombin‐activated platelets in platelet concentrates derived from buffy coat: the role of processing, pathogen inactivation, and storage

Collagen‐ and thrombin‐activated (COAT) platelets (PLTs), generated by dual‐agonist stimulation with collagen and thrombin (THR), enhance THR generation at the site of vessel wall injury. There is evidence that higher amounts of procoagulant COAT PLTs are associated with stroke, while a decreased ability to generate them is associated with bleeding diathesis. Our aim was to study PLT functions, particularly the ability to generate COAT PLTs, in PLT concentrates (PCs) from buffy coat. Thus, we investigated the effect of processing, pathogen inactivation treatment (amotosalen‐UVA), and PC storage.

The Role of Plasma Transfusion in Massive Bleeding: Protecting the Endothelial Glycocalyx?

Massive hemorrhage is a leading cause of death worldwide. During the last decade several retrospective and some prospective clinical studies have suggested a beneficial effect of early plasma-based resuscitation on survival in trauma patients. The underlying mechanisms are unknown but appear to involve the ability of plasma to preserve the endothelial glycocalyx. In this mini-review, we summarize current knowledge on glycocalyx structure and function, and present data describing the impact of hemorrhagic shock and resuscitation fluids on glycocalyx. Animal studies show that hemorrhagic shock leads to glycocalyx shedding, endothelial inflammatory changes, and vascular hyper-permeability. In these animal models, plasma administration preserves glycocalyx integrity and functions better than resuscitation with crystalloids or colloids. In addition, we briefly present data on the possible plasma components responsible for these effects. The endothelial glycocalyx is increasingly recognized as a critical component for the physiological vasculo-endothelial function, which is destroyed in hemorrhagic shock. Interventions for preserving an intact glycocalyx shall improve survival of trauma patients.

Antiplatelet Therapy and Anticoagulation

Preoperative withdrawal of antiplatelet (AP) therapy in secondary prevention multiplies the risk of myocardial infarction, stent thrombosis, stroke, or death, by five to ten times. Aspirin is a lifelong therapy and should never be interrupted. Dual AP therapy with aspirin plus clopidogrel, prasugrel, or ticagrelor is essential for at least 6 weeks after coronary revascularization using bare-metal stents, 12 months after treatment for acute coronary syndrome, and 6–12 months after implanting drug-eluting stents. These time spans might even be prolonged in high-risk cases. Elective surgery should be postponed until after these periods. Vital or urgent operations undertaken earlier must be performed under continued dual AP therapy.

Acquired Hemostatic Disorders

Acquired hemostatic disorders comprise thrombocytopenia and platelet dysfunction, coagulation factor deficiencies, excessive anticoagulation, and hemorrhagic complications due to antiplatelet drugs, anticoagulation, and thrombolysis. Blood disorders associated with myeloproliferative neoplasms and disseminated intravascular coagulation can cause both bleeding and thrombosis. Heparin-induced thrombocytopenia, antiphospholipid antibody syndrome, and thrombotic microangiopathies are conditions that can cause thrombocytopenia, but they are more frequently responsible for thrombosis than for bleeding.