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. 2011 Oct 24;6(10):e25854. doi: 10.1371/journal.pone.0025854

Figure 13. LMB treatment influences virus production.

Figure 13

(A). Cells were treated with 10 ng/ml LMB for various time periods to determine the incubation time required to affect infection. Cells were infected with HCVcc (JFH1) and the drug was added immediately after infection. Cells were grown in the medium containing LMB, for the time indicated and HCV RNA was then extracted and quantified by RT-qPCR. The values are expressed as a percent of the amount of HCV RNA present in cells grown without LMB. (B). Control experiments carried out to demonstrate that the incubation of cells for 2–8 h with 10 ng/ml LMB (shown in (A) had no toxic effect on cell viability. Cell viability after LMB treatments was determined by counting live and dead cells after trypan blue staining, or by measuring cellular ATP present in culture wells as described in Materials and Methods. Untreated cells and cells treated with 10% DMSO (to induce cell death) were used as negative and positive controls, respectively. The results are expressed as a percent of the value obtained for an untreated control. (C). Treatment with LMB early in infection significantly decreases infection levels. Cells were infected with HCVcc 2 h at 37°C in the presence of 10 ng/ml LMB (T0) or infected with HCVcc and then treated with the drug at the indicated time points after infection (2 h, 4 h, or 6 h) and then incubated for a further 8 h. The treatment of cells with LMB 0–6 h post infection significantly decreases infection levels, whereas the same treatment (for 8 h) applied 24 h after infection has no effect on intracellular HCV RNA levels, as shown by comparison with the untreated control. (D) Control experiments for (C) showing that the application of LMB or its solvent (ethanol) at the same concentrations and for the same time period as used for (C) does not influence cell viability, as demonstrated by comparison with an untreated control. DMSO (at a concentration of 10%) was used as a positive control, to decrease cell viability. Values are expressed as a percent of untreated control.