Plasma-derived Extracellular Vesicles Research Articles

Plasma-derived Extracellular Vesicles Contain Predictive Biomarkers and Potential Therapeutic Targets for Myocardial Ischemic (MI) Injury

Myocardial infarction (MI) triggers a potent inflammatory response via the release of circulatory mediators, including extracellular vesicles (EVs) by damaged cardiac cells, necessary for myocardial healing. Timely repression of inflammatory response are critical to prevent and minimize cardiac tissue injuries, nonetheless, progression in this aspect remains challenging. The ability of EVs to trigger a functional response upon delivery of carried bioactive cargos, have made them clinically attractive diagnostic biomarkers and vectors for therapeutic interventions. Using label-free quantitative proteomics approach, we compared the protein cargo of plasma EVs between patients with MI and from patients with stable angina (NMI). We report, for the first time, the proteomics profiling on 252 EV proteins that were modulated with >1.2-fold after MI. We identified six up-regulated biomarkers with potential for clinical applications; these reflected post-infarct pathways of complement activation (Complement C1q subcomponent subunit A (C1QA), 3.23-fold change, p = 0.012; Complement C5 (C5), 1.27-fold change, p = 0.087), lipoprotein metabolism (Apoliporotein D (APOD), 1.86-fold change, p = 0.033; Apolipoprotein C-III (APOCC3), 2.63-fold change, p = 0.029) and platelet activation (Platelet glycoprotein Ib alpha chain (GP1BA), 9.18-fold change, p < 0.0001; Platelet basic protein (PPBP), 4.72-fold change, p = 0.027). The data have been deposited to the ProteomeXchange with identifier PXD002950. This novel biomarker panel was validated in 43 patients using antibody-based assays (C1QA (p = 0.005); C5 (p = 0.0047), APOD (p = 0.0267); APOC3 (p = 0.0064); GP1BA (p = 0.0031); PPBP (p = 0.0465)). We further present that EV-derived fibrinogen components were paradoxically down-regulated in MI, suggesting that a compensatory mechanism may suppress post-infarct coagulation pathways, indicating potential for therapeutic targeting of this mechanism in MI. Taken together, these data demonstrated that plasma EVs contain novel diagnostic biomarkers and therapeutic targets that can be further developed for clinical use to benefit patients with coronary artery diseases (CADs).

Abstract A30: Chronic lymphocytic leukemia-derived extracellular vesicles mediate NFkB signaling and pro-inflammatory cytokine release in monocytes

Abstract The pathogenesis of chronic lymphocytic leukemia (CLL) is stringently associated with a tumor-supportive microenvironment and a dysfunctional immune system. Extracellular vesicles (EV) released by CLL cells are taken up by non-malignant cells in the microenvironment and mediate major disease-related changes in recipient cells. In the current study, we characterized CLL EVs, especially focusing on vesicle-incorporated RNA transcripts, and defined the role of CLL EVs in changing the myeloid tumor microenvironment. EVs were isolated from blood plasma of CLL patients and healthy donors as well as from supernatant of the CLL cell line MEC-1 by a serial centrifugation protocol. Characterization of EVs by electron microscopy, Nanoparticle Tracking Analysis (NTA) and Western blotting revealed vesicles 30 to 350 nm in size, which are positive for various EV marker proteins such as Rab5a and Hsp70. Quantification of blood plasma-derived EVs indicated an enrichment of B-cell derived EVs in plasma of CLL patients compared to healthy donors, although absolute EV counts were not altered in CLL. Focusing on RNA analysis, an enrichment of small RNAs in EVs was observed. Subsequent small RNA sequencing revealed a unique microRNA signature of MEC-1 EVs, with CLL-relevant miRNAs such as miR-21, miR-155 and miR-146a being the most abundant miRNAs. Moreover, full length and 5'end fragment forms of Y RNAs, another class of small non-coding RNAs, were enriched in MEC-1 EVs and CLL plasma EVs. Further evaluating the functional relevance of CLL EVs within the tumor microenvironment, a rapid uptake of CLL cell-derived EVs by human monocytes and macrophages was observed. Uptake of CLL EVs in monocytes induced NFkB signaling and the release of multiple pro-inflammatory cytokines such as CCL2, CCL3, IL-6 and IL-8, which are also upregulated in plasma of CLL patients. In conclusion, tumor-derived EVs harbor a distinct set of non-coding RNAs. The uptake of EVs in recipient cells and the concomitant transfer of incorporated RNAs mediate substantial phenotypic changes in target cells. In the current study, this is exemplified for monocytes, which present several disease-relevant alterations upon EV uptake, including cellular activation and secretion of pro-inflammatory cytokines. Citation Format: Franziska Haderk, Laura Llao Cid, Etienne Moussay, Jerome Paggetti, Karolin Willmund, Jana Seiler, Sven Diederichs, Maria Goebel, Jan Duerig, Thorsten Zenz, Stephan Stilgenbauer, Marc Zapatka, Peter Lichter, Martina Seiffert. Chronic lymphocytic leukemia-derived extracellular vesicles mediate NFkB signaling and pro-inflammatory cytokine release in monocytes. [abstract]. In: Proceedings of the AACR Special Conference: Function of Tumor Microenvironment in Cancer Progression; 2016 Jan 7–10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2016;76(15 Suppl):Abstract nr A30.

Erratum: Vesicular Galectin-3 levels decrease with donor age and contribute to the reduced osteo-inductive potential of human plasma derived extracellular vesicles.

Aging | doi:10.18632/aging.100917. Sylvia Weilner, Verena Keider, Melanie Winter, Eva Harreither, Benjamin Salzer, Florian Weiss, Elisabeth Schraml, Paul Messner, Peter Pietschmann, Florian Hildner, Christian Gabriel, Heinz Redl, Regina Grillari-Voglauer, Johannes Grillari

Plasma Proteome Database as a resource for proteomics research: 2014 update

Plasma Proteome Database (PPD; http://www.plasmaproteomedatabase.org/) was initially described in the year 2005 as a part of Human Proteome Organization’s (HUPO’s) pilot initiative on Human Plasma Proteome Project. Since then, improvements in proteomic technologies and increased throughput have led to identification of a large number of novel plasma proteins. To keep up with this increase in data, we have significantly enriched the proteomic information in PPD. This database currently contains information on 10 546 proteins detected in serum/plasma of which 3784 have been reported in two or more studies. The latest version of the database also incorporates mass spectrometry-derived data including experimentally verified proteotypic peptides used for multiple reaction monitoring assays. Other novel features include published plasma/serum concentrations for 1278 proteins along with a separate category of plasma-derived extracellular vesicle proteins. As plasma proteins have become a major thrust in the field of biomarkers, we have enabled a batch-based query designated Plasma Proteome Explorer, which will permit the users in screening a list of proteins or peptides against known plasma proteins to assess novelty of their data set. We believe that PPD will facilitate both clinical and basic research by serving as a comprehensive reference of plasma proteins in humans and accelerate biomarker discovery and translation efforts.