The knockdown of ApoB increased lipid accumulation and promoted HCV production.
(A) Huh-7 cells were transduced with ApoB shRNAs. The protein level of ApoB was analyzed by western blotting. Actin was used as the loading control. (B) Huh-7 cells were transduced with ApoB shRNA1-shRNA5. Immunostaining was performed with an anti-ApoB antibody. LDs were stained with BODIPY 493/503. Nuclei were stained with DAPI. Fluorescence signals were visualized by laser confocal microscopy. Scale bars, 10 μM. (C and D) The intracellular and secreted TG concentrations in cells in B were analyzed using a TG quantification kit. TG, triglyceride. (E) Huh-7 cells were transduced with ApoB shRNA1 or shRNA2 for 2 days prior to HCV infection (MOI = 1) for 4 days. Cell lysates and culture supernatants were collected to infect Huh-7 cells. In-cell western blot analysis was performed with an anti-core antibody. Cells were further labeled with IRDye 800-conjugated secondary antibodies and scanned with an Odyssey infrared imaging system. (F and G) Viral titration was performed by immunostaining. (H) The intracellular HCV RNA level was analyzed by real-time PCR. GAPDH was used as the internal control. The statistical significance was determined by unpaired two-tailed Student’s t-tests. The data are presented as the means ± SDs of n = 3 biological repeats. n.s., not significant. ** P < 0.01.