Comparative proteomic analysis of PAI‐1 and TNF‐alpha‐derived endothelial microparticles

Abstract

Endothelium-derived microparticles (EMPs) are small vesicles released from endothelial cells in response to cell injury, apoptosis, or activation. Elevated concentrations of EMPs have been associated with many inflammatory and vascular diseases. EMPs also mediate long range signaling and alter downstream cell function. Unfortunately, the molecular and cellular basis of microparticle production and downstream cell function is poorly understood. We hypothesize that EMPs generated by different agonists will produce distinct populations of EMPs with unique protein compositions. To test this hypothesis, different EMP populations were generated from human umbilical vein endothelial cells by stimulation with plasminogen activator inhibitor type 1 (PAI-1) or tumor necrosis factor-alpha (TNF-α) and subjected to proteomic analysis by LC/MS. We identified 432 common proteins in all EMP populations studied. Also identified were 231 proteins unique to control EMPs, 104 proteins unique to PAI-1 EMPs and 70 proteins unique to TNF-α EMPs. Interestingly, variations in protein abundance were found among many of the common EMP proteins, suggesting that differences exist between EMPs on a relative scale. Finally, gene ontology (GO) and KEGG pathway analysis revealed many functional similarities and few differences between the EMP populations studied. In summary, our results clearly indicate that EMPs generated by PAI-1 and TNF-α produce EMPs with overlapping but distinct protein compositions. These observations provide fundamental insight into the mechanisms regulating the production of these particles and their physiological role in numerous diseases.

Endothelial cell-derived microparticles induce plasmacytoid dendritic cell maturation: potential implications in inflammatory diseases

“…A recent proteomic study comparing EMP generated by different agonists identified 432 common proteins in quiescent EMP, plasminogen activator inhibitor type 1-induced EMP and TNF-induced EMP. 5 Variations in protein abundance in these different EMP were found. 5 Our data are rather in favor of an increase of EMP in inflammatory conditions (as observed here after TNF-α stimulation of endothelial cells) being sufficient to induce PDC maturation.…”

“…1 All microparticles, whatever their cell origin, have negatively charged phospholipids -such as phosphatidylserine -in their outer membrane leaflet, accounting for their procoagulant properties. [1][2][3] They also express proteins, characteristic of their cellular origin, on their surface and carry proteins packaged from numerous cellular compartments, 4,5 as well as mRNA. 6 Microparticles are different from exosomes, since these latter are smaller (30-90 nm) and derived from endocytic compartments leading to an enrichment of tetraspanin molecules.…”

Robust Label-free, Quantitative Profiling of Circulating Plasma Microparticle (MP) Associated Proteins

“…There is increasing evidence that MP are part of a sophisticated intercellular communication network and are involved in repair of damaged tissue (8,9). Protein, lipid, and RNA composition is dependent on the stimuli and the cell type (8,10,11). Therefore, MP can be considered as a repository of physiological processes taking place in the vascular system, and are potentially useful biomarkers for the assessment of severity, treatment, and prognosis of vascular diseases.…”

Endothelial microparticles activate endothelial cells to facilitate the inflammatory response

“…Despite a relationship between EMP protein composition and their parental ECs having been demonstrated (13), to the best of our knowledge, the effect of EMPs on their parental ECs remains unknown. Since the major pathological changes during sepsis occur in the microvascular and pulmonary microvascular endothelial cells, human pulmonary microvascular endothelial cells (HPMECs) were used for the present study.…”

“…EMPs may also activate target cells, including monocytic cells, and amplify harmful responses including inflammation, thrombosis and vascular dysfunction, but research in this area is lacking. A direct correlation between the proteins that compose EMPs and tumor necrosis factor-α (TNF-α)-stimulated ECs has been previously demonstrated (13). The endothelial proteins transferred by EMPs may be involved in the interaction between EMPs and their target cells, which may result in endothelial dysfunction (14).…”