Abstract
Endothelial activation and alteration during dengue virus (DENV) infection are multifactorial events; however, the role of extracellular vesicles (EVs) in these phenomena is not known. In the present study, we characterized the EVs released by DENV-2 infected U937 macrophage cell line and evaluated the changes in the physiology and integrity of the EA.hy926 endothelial cells exposed to them. U937 macrophages were infected, supernatants were collected, and EVs were purified and characterized. Then, polarized endothelial EA.hy926 cells were exposed to the EVs for 24 h, and the transendothelial electrical resistance (TEER), monolayer permeability, and the expression of tight junction and adhesion proteins and cytokines were evaluated. The isolated EVs from infected macrophages corresponded to exosomes and apoptotic bodies, which contained the viral NS3 protein and different miRs, among other products. Exposure of EA.hy926 cells to EVs induced an increase in TEER, as well as changes in the expression of VE-cadherin and ICAM in addition leads to an increase in TNF-α, IP-10, IL-10, RANTES, and MCP-1 secretion. These results suggest that the EVs of infected macrophages transport proteins and miR that induce early changes in the physiology of the endothelium, leading to its activation and eliciting a defense program against damage during first stages of the disease, even in the absence of the virus.
Highlights
Eukaryotic cells secrete extracellular vesicles (EVs) into the bodily fluids, such as blood, saliva, urine, or breast milk [1]
Endothelial cells EA.hy926 activated by EVs of dengue virus (DENV) infected U937 macrophages as CD63, and Apoptotic Bodies (AB) marker Histone-3 (H3, 1:2000, Cell Signaling) were used as primary antibodies
The correlation results showed that regardless of whether endothelial cells are exposed to complete viral particles (IC) or vesicles from infected U937 macrophages (P (+) or EVs (+)4G2), the effect observed in transendothelial electrical resistance (TEER) is very similar, indicating that it is not Endothelial cells EA.hy926 activated by EVs of DENV infected U937 macrophages necessary for the virus to have contact with the endothelial cells to cause alterations in these, being the information carried by the vesicles from cells that did have direct interaction with the virus enough for this, even if the vesicles were treated with the 4G2 antibody
Summary
Eukaryotic cells secrete extracellular vesicles (EVs) into the bodily fluids, such as blood, saliva, urine, or breast milk [1]. These EVs are spherical particles surrounded by a phospholipid bilayer containing proteins or nucleic acids that control a broad range of biological functions when they are sensed by a receptor cell [2]. The composition of EVs varies depending on the type of cell from which they arise, or the cellular status thereof [3, 4]. The sponsors did not play any role in the study design, data collection and analysis, decision to publish or preparation of the manuscript
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