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Plasma QconCATs reveal a gender-specific proteomic signature in apheresis platelet plasma supernatants.

ABSTRACT: Clinical translation of proteomic technologies is often hampered by technical limitations, including inter-laboratory inconsistencies of label-free derived relative quantification, time-consuming analytical approaches and the subsequent challenge of performing proteomic analyses on large cohorts of subjects. Here we introduce plasma QconCAT-based targeted proteomics, an approach that allows the simultaneous absolute quantitation down to the picogram level of hundreds of proteins in a single liquid chromatography-selected reaction monitoring mass spectrometry run. We demonstrate the robustness of the approach by analyzing apheresis platelet concentrate supernatants at storage day 1 and the end of the shelf life for this blood-derived therapeutic, day 5. The targeted approach was repeatable and robust revealing potential gender-specific signatures across a set of three male and female donors. This technical note represents a proof-of-principle of the application of QconCAT-based MRM strategies to transfusion-medicine relevant issues, such as storage and gender-dependent proteomic signatures in blood-derived therapeutics.

Biological significance

Gender differences in the proteome composition of apheresis platelet supernatants have always been postulated, and might underlie a higher risk of adverse reactions when transfusing apheresis products from female donors. Preliminary proteomic studies provided an overview of gender-dependent relative compositional differences in the proteome of apheresis platelet supernatants during routine storage in the blood bank. Here we apply a proteomics approach for absolute quantitation of approximately 100 proteins in apheresis platelet supernatants from male and female donors at storage days 1 and 5. Absolute quantitative proteomic analyses allowed us to confirm and expand on previous observations about gender and storage-dependency of platelet supernatant protein profiles.

SUBMITTER: Dzieciatkowska M

PROVIDER: S-EPMC5794015 | biostudies-literature | 2015 Apr

REPOSITORIES: biostudies-literature

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Plasma QconCATs reveal a gender-specific proteomic signature in apheresis platelet plasma supernatants.

Dzieciatkowska Monika M D'Alessandro Angelo A Hill Ryan C RC Hansen Kirk C KC

Journal of proteomics 20150302

Clinical translation of proteomic technologies is often hampered by technical limitations, including inter-laboratory inconsistencies of label-free derived relative quantification, time-consuming analytical approaches and the subsequent challenge of performing proteomic analyses on large cohorts of subjects. Here we introduce plasma QconCAT-based targeted proteomics, an approach that allows the simultaneous absolute quantitation down to the picogram level of hundreds of proteins in a single liqu ...[more]

PMID: 25743772

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Project description:Proteomics has identified potential pathways involved in platelet storage lesions, which correlate with untoward effects in the recipient, including febrile non-haemolytic reactions. We hypothesize that an additional pathway involves protein mediators that accumulate in the platelet supernatants during routine storage in a donor gender-specific fashion. Apheresis platelet concentrates were collected from 5 healthy males and 5 females and routinely stored. The 14 most abundant plasma proteins were removed and the supernatant proteins from days 1 and 5 were analyzed via 1D-SDS-PAGE/nanoLC-MS/MS, before label-free quantitative proteomics analyses. Findings from a subset of 18 proteins were validated via LC-SRM analyses against stable isotope labeled standards. A total of 503 distinct proteins were detected in the platelet supernatants from the 4 sample groups: female or male donor platelets, either at storage day 1 or 5. Proteomics suggested a storage and gender-dependent impairment of blood coagulation mediators, pro-inflammatory complement components and cytokines, energy and redox metabolic enzymes. The supernatants from female donors demonstrated increased deregulation of structural proteins, extracellular matrix proteins and focal adhesion proteins, possibly indicating storage-dependent platelet activation. Routine storage of platelet concentrates induces changes in the supernatant proteome, which may have effects on the transfused patient, some of which are related to donor gender.Biological significanceThe rationale behind this study is that protein components in platelet releasates have been increasingly observed to play a key role in adverse events and impaired homeostasis in transfused recipients. In this view, proteomics has recently emerged as a functional tool to address the issue of protein composition of platelet releasates from buffy coat-derived platelet concentrates in the blood bank. Despite early encouraging studies on buffy coat-derived platelet concentrates, platelet releasates from apheresis platelets have not been hitherto addressed by means of extensive proteomics technologies. Indeed, apheresis platelets are resuspended in donors' plasma, which hampers detection of less abundant proteins, owing to the overwhelming abundance of albumin (and a handful of other proteins), and the dynamic range of protein concentrations of plasma proteins. In order to cope with these issues, we hereby performed an immuno-affinity column-based depletion of the 14 most abundant plasma proteins. Samples were thus assayed via GeLC-MS, a workflow that allowed us to cover an unprecedented portion of the platelet supernatant proteome, in comparison to previous transfusion medicine-oriented studies in the literature. Finally, we hereby address the issue of biological variability, by considering the donor gender as a key factor influencing the composition of apheresis platelet supernatants. As a result, we could conclude that platelet supernatants from male and female donors are not only different in the first place, but they also store differently. This conclusion has been so far only suggested by classic transfusion medicine studies, but has been hitherto unsupported by actual biochemistry/proteomics investigations. In our opinion, the main strengths of this study are related to the analytical workflow (immunodepletion and GeLC-MS) and proteome coverage, the translational validity of the results (from a transfusion medicine standpoint) and the biological conclusion about the intrinsic (and storage-dependent) gender-related differences of platelet supernatants.

Dataset Information

Plasma QconCATs reveal a gender-specific proteomic signature in apheresis platelet plasma supernatants.

Biological significance

Publications

Plasma QconCATs reveal a gender-specific proteomic signature in apheresis platelet plasma supernatants.

Similar Datasets

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