Figure 2. Effects of endosomal acidification in the replication of PEC or MNV-1.

A. LLC-PK cells were pre-treated with mock (medium) or chloroquine (70 μM) for 1h, and infected with PEC (MOI 50) in the presence of mock or cholorquine. In all cells, GCDCA (100 μM) was present in the media to support virus replication during virus infection. Following virus infection, cells were thoroughly washed with PBS and fresh medium containing mock or chloroquine was added to the cells. Cells were then further incubated and collected at 12 h PI. In a separate experiment, chloroquine was added to the PEC-infected cells at 6 PI and cells were collected at 12 h PI for the determination of viral replication by TCID₅₀ assay. B. RAW267.4 cells were pre-treated with mock (medium) or chloroquine (200 or 100 μM) for 1h then infected with MNV-1 (MOI 10) in the presence of mock or chloroquine at 4 °C for 1 h. Cells were then washed with PBS and fresh medium containing mock or chloroquine was added to the cells for further incubation at 37°C. At 4 h PI, cells were thoroughly washed again with PBS and fresh medium without chloroquine was added to the cells. Cells were collected at 12 h PI for the determination of virus replication by the TCID₅₀ assay. C. Cells were pretreated with chloroquine (50 or 70 μM) for 1 h, and infected with MNV-1 (10 MOI) in the presence of mock or chloroquine at 4°C for 1 h. Cells were then washed with PBS and fresh medium containing chloroquine was added to the cells. Cells were further incubated and collected at 12 h PI for determination of virus replication by the TCID₅₀ assay. (A to C) Asterisk indicates a significant reduction of viral titers in the treatment group compared to the control (P < 0.05).