Microvesicles released from activated CD4+ T cells alter microvascular endothelial cell function

Abstract

Background: Microvesicles are vesicles shed by plasma membranes following cell activation and apoptosis. The role of lymphocyte-derived microvesicles in endothelial function remains poorly understood.

Methods: CD4⁺ T cells isolated from peripheral blood of healthy human donors were stimulated using anti-CD3/anti-CD28-coated beads. Proteomic profiling of microvesicles was performed using linear discriminant analysis (LDA) from activated T cells (MV.Act) and nonactivated T cells (MV.NAct). In addition, data processing analysis was performed using MaxQUANT workflow. Differentially expressed proteins found in MV.Act or MV.NAct samples with identification frequency = 100%, which were selected by both LDA (p < .01) and MaxQUANT (p < .01) workflows, were defined as "high-confidence" differentially expressed proteins. Functional effects of MV.Act on human primary microvascular endothelial cells were analysed.

Results: T cells released large amounts of microvesicles upon stimulation. Proteomic profiling of microvesicles using LDA identified 2279 proteins (n = 2110 and n = 851 proteins in MV.Act and MV.NAct, respectively). Protein-protein interaction network models reconstructed from both differentially expressed proteins (n = 594; LDA p ≤ .01) and "high-confidence" differentially expressed proteins (n = 98; p ≤ .01) revealed that MV.Act were enriched with proteins related to immune responses, protein translation, cytoskeleton organisation and TNFα-induced apoptosis. For instance, MV.Act were highly enriched with IFN-γ, a key proinflammatory pathway related to effector CD4⁺ T cells. Endothelial cell incubation with MV.Act induced superoxide generation, apoptosis, endothelial wound healing impairment and endothelial monolayer barrier disruption.

Conclusions: T cell receptor-mediated activation of CD4⁺ T cells stimulates the release of microvesicles enriched with proteins involved in immune responses, inflammation and apoptosis. T cell-derived microvesicles alter microvascular endothelial function and barrier permeability, potentially promoting tissue inflammation.

Keywords: T cells; endothelial cell; extracellular vesicle; microvesicle; proteomics.