Abstract
Extracellular vesicles (EV) can modulate the responses of cells to toll-like receptor (TLR) ligation; conversely, TLR ligands such as double-stranded RNA (dsRNA) can enhance the release of EV and influence of the composition and functions of EV cargos. Inflamed synovial joints in rheumatoid arthritis (RA) are rich in EV and extracellular RNA; besides, RNA released from necrotic synovial fluid cells can activate the TLR3 signaling in synovial fibroblasts (SFs) from patients with RA. Since EV occur prominently in synovial joints in RA and may contribute to the pathogenesis, we questioned whether EV can interact with dsRNA, a TLR3 ligand, and modify its actions in arthritis. We have used as model the effects on RA SFs, of EV released from monocyte U937 cells and peripheral blood mononuclear cells upon stimulation with Poly(I:C), a synthetic analog of dsRNA. We show that EV released from unstimulated cells and Poly(I:C)-stimulated U937 cells [Poly(I:C) EV] differ in size but bind similar amounts of Annexin V and express comparable levels of MAC-1, the receptor for dsRNA, on the vesicular membranes. Specifically, Poly(I:C) EV contain or associate with Poly(I:C) and at least partially protect Poly(I:C) from RNAse III degradation. Poly(I:C) EV shuttle Poly(I:C) to SFs and reproduce the proinflammatory and antiviral gene responses of SFs to direct stimulation with Poly(I:C). Poly(I:C) EV, however, halt the death receptor-induced apoptosis in SFs, thereby inverting the proapoptotic nature of Poly(I:C). These prosurvival effects sharply contrast with the high toxicity of cationic liposome-delivered Poly(I:C) and may reflect the route of Poly(I:C) delivery via EV or the fine-tuning of Poly(I:C) actions by molecular cargo in EV. The demonstration that EV may safeguard extracellular dsRNA and allow dsRNA to exert antiapoptotic effects on SFs highlights the potential of EV to amplify the pathogenicity of dsRNA in arthritis beyond inflammation (by concurrently enhancing the expansion of the invasive synovial stroma).
Highlights
Extracellular vesicles (EV) are a heterogeneous group of vesicles that are secreted from cells to enter the extracellular space where they can exhibit a variety of immunological properties
Particles with sizes of microvesicles predominated in the EV preparations from unstimulated U937 cells (Con EV) [Figure 1A, median (10th, 90th percentile): 218 nm (114, 371 nm) and 227 (131, 368 nm), n = 2] and Poly(I:C)-stimulated U937 cells [Poly(I:C) EV] [Figure 1B, 346 nm (217, 504 nm) and 287 (187, 413 nm), n = 2]
These results suggested that control EV (Con EV) and Poly(I:C) EV might differ in composition or origin
Summary
Extracellular vesicles (EV) are a heterogeneous group of vesicles that are secreted from cells to enter the extracellular space where they can exhibit a variety of immunological properties. Synovial fluid from patients with RA contains increased amounts of EV derived from platelets, monocytes, lymphocytes and neutrophils [5,6,7,8]. These EV can promote the matrix-degrading and/or proinflammatory properties in synovial fibroblasts (SFs) and/or neutrophils [5, 6, 9, 10], thereby aggravating the arthritis; EV, can have chondroprotective [7] and proresolving [11] functions, which could be exploited therapeutically [4]. EV contain or can associate with proinflammatory cytokines [5], damage-associated molecular patterns (DAMPs) [11], or citrullinated autoantigens [6, 12] and can modulate their proinflammatory actions [6, 13] in inflammatory arthritis
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