Figure 1.
Pharmacological interventions targeting PI3K signaling affect EV-A71 replication in SF268 neural cells.
The effect of PI3K (LY294002 and wortmannin), P38 MAPK (SB203580) (A) and GSK3β (AR-A014418 and LiCl) (B) inhibitors as pharmacological interventions on the virus yields of EV-A71-infected cells was assessed. SF268 cells were infected with EV-A71 (moi 0.5), and various concentrations of compounds were added to the infected cells after 1 h viral adsorption. At 48 h post-infection, the culture supernatants and cell lysates were collected for virus titration using plaque assays. Data are displayed as mean ± s.e.m. from at least two independent experiments performed in duplicates.
Figure 2.
LiCl inhibits EV-A71 replication in SF268 cells (A), (B) and SH-SY5Y cells (C).
Cells were infected with EV-A71 (moi 0.5), and various concentrations of LiCl were added to the infected cells after viral adsorption time. Lysates collected At 8 h (A) and 72 h (B, C) post-infection were analyzed with immunoblotting. Rupintrivir was employed as a positive control because it can effectively inhibit EV-A71 replication. Measurements were made in three independent experiments.
Figure 3.
LiCl suppresses pro-inflammatory IL-6 and IL-1β mRNA expression (A) and IL-6 protein expression (B) in EV-A71-infected SF268 cells.
SF268 cells were infected with EV-A71 (moi 0.5), and various concentrations of LiCl were added to the infected cells after viral adsorption time. At 48 h post-infection, the cell lysates were collected for quantification of the IL-6 and IL-1β mRNA. In a parallel experiment, the IL-6 protein levels in cell culture supernatant were measured using enzyme-linked immunosorbent assay. Data are the mean ± s.e.m. from at least three parallel measurements per experiment.
Figure 4.
The active metabolite of Leflunomide, A771726, suppress virus production (A) and the expression of the pro-inflammatory cytokine IL-6 (B) in EV-A71-infected SF268 cells.
SF268 cells were infected with EV-A71 (moi 0.5), and various concentrations of A771726 were added to the infected cells after viral adsorption time. At 48 h post-infection, the culture supernatants and cell lysates were collected for virus titration (A). In a parallel experiment, the IL-6 protein levels in cell culture supernatant were measured using ELISA (B). Each virus stock to be titered is serially diluted in10-fold series (dilution fold 10−4) and added to Vero cells. Data are the mean ± s.e.m. from at least two parallel measurements per experiment.
Table 1.
Effects of A771726 in combination with LiCl on EV71- induced cytopathic effect.