A Mouse Model for Infection with Enterovirus A71 in Small Extracellular Vesicles

Figures

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  FIG 1. Characterization of sEVs from EV-A71-infected cells. (A and B) TEM images of empty sEVs (blue arrows) (A) and sEVs with virus-like particles (green arrow) (B). (C) Diameter of sEVs measured by NTA. (D) Immunoblotting of EV-positive markers (CD63 and TSG101), negative markers (albumin and calnexin), and viral capsid proteins (VP1) in sEVs and cell lysis. (E) Detection of viral RNA in EV-A71 and sEVs by qRT-PCR (mean ± SD; 3 independent experiments). (F) Schematic drawing of in vitro BBB model. (G) Comparison of permeabilities between sEVs and EV-A71 (mean ± SD; 3 independent experiments). In all panels: ns, not significant; *, P ≤ 0.05; **, P ≤ 0.01; ***, P ≤ 0.001; ****, P ≤ 0.0001.

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  FIG 2. sEVs showed stronger neurovirulence than EV-A71 in the mouse model. (A and B) Size (A) and weight (B) differences of mice in different treatment groups at 3 dpi. (C) Survival curve of mice after injection. (D) Absolute quantification of EV-A71 RNA in gut and brain was performed by qRT-PCR. The virus RNA copies for various tissues of each infected mouse were shown as mean for three independent experiments. The viral RNA loads of different treatment groups were compared. Statistical analyses were performed using Mann-Whitney U test. (E) Representative H&E images of brain damage. The EV-A71-infected group exhibited mild neuropil vacuolation and neuronal loss (arrows), and the sEV-infected group exhibited numerous inflammatory cells and vesicular degeneration (arrows). A representative result is shown. In all panels: *, P ≤ 0.05; **, P ≤ 0.01; ***, P ≤ 0.001; ****, P ≤ 0.0001. NC, negative control.