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. 2017 Mar 28;7:45375. doi: 10.1038/srep45375

Figure 2. Nedd4 knockdown inhibits JEV infection.

Figure 2

SK-N-SH cells were transfected with Nedd4-specific siRNA (12.5, 25, 50, 100 nM) or the Non-Target (NT) siRNA (100 nM) for 72 h. (A) The reduction of Nedd4 after treatment with different concentrations of siRNA was measured by RT-qPCR. An MTT assay was used to assess the siNedd4-induced cytotoxic effect. (B) The level of Nedd4 protein was detected by Western blot, quantitated by densitometric analysis using ImageJ software and normalized to GAPDH. (C) Cells transfected with Nedd4 siRNA or the NT siRNA (50 nM) were infected with JEV (MOI = 1.0). JEV infection was assessed by monitoring intracellular JEV RNA at 24 h p.i. by RT-qPCR. (D) SK-N-SH cells were fixed at 24 h p.i. and were subjected to immunofluorescence to detect JEV E (green). The nuclei were stained with DAPI (blue). Images were obtained using a confocal microscope and quantitated using ImageJ software (Scale bar = 50 μm). The data represent the mean ± SD of three independent experiments. *, P < 0.05; **, P < 0.01; ***, P < 0.001.