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Figure 6 | Virology Journal

Figure 6

From: Inhibition of cyclophilins alters lipid trafficking and blocks hepatitis C virus secretion

Figure 6

Enlarged lipid droplets correlate with inhibition of HCV replication using siRNA knockdown. A) sg-1b replicon cells were transiently transfected with siRNA to target cypA, cypB and cyp40. A RISC-free siRNA construct was used as a control. Whole cell lysates were analyzed for target protein expression by Western blot at 72h and 96h post-transfection. B) Total RNA from siRNA transfected cells was extracted and used for quantitative real-time RT-PCR to measure copies of HCV. Replication in RISC-free control cells was set to 100% for each time point. Results are the average of 4 independent experiments, ± standard deviation. CypA and cyp40 siRNA result in a statistically significant decrease in replicon activity, p < 0.01 for cypA and p < 0.05 for cyp40 compared to RISC-free siRNA by Students paired t-test. C) At 96h post-transfection, lipid droplets were visualized as described. Representative images for each siRNA are shown. D) 20X images were acquired and quantified by the Cellomics ArrayScan VTI HCS reader. The mean fluorescence intensities for BODIPY 493/503 were normalized to the RISC free siRNA control. Those siRNA constructs that resulted in inhibition of HCV [17] also caused the lipid droplets to become enlarged, similar to what is seen with NIM811 treatment.

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