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. 2006 May;80(9):4510–4520. doi: 10.1128/JVI.80.9.4510-4520.2006

FIG. 2.

FIG. 2.

CsA suppressed the replication of HCV genome, irrespective of the presence of the structural proteins. (A) Detection of HCV proteins from NNC#2 (NN/1b/FL) genome-length replicon. Core (a), E2 (b), NS3 (c), NS5A (d), NS5B (e), and tubulin (f) in Huh-7, NNC#2 (NN/1b/FL), and MH-14 (NN/1b/SG) cells analyzed by immunoblot analysis are shown. (B) HCV RNA in Huh-7, NNC#2 (NN/1b/FL), and MH-14 (NN/1b/SG) cells quantified by real-time RT-PCR analysis. The data represent the means of three independent experiments. (C) CsA decreased the production of HCV proteins in NNC#2 (NN/1b/FL), as well as in MH-14 (NN/1b/SG) cells. After treatment with 1-μg/ml CsA (+) for 5 days or without treatment (−), total-cell lysates of NNC#2 (NN/1b/FL) and MH-14 (NN/1b/SG) cells, together with Huh-7 cells as a negative control, were recovered to examine the production of HCV NS5A (top), NS5B (middle), and tubulin as an internal control (bottom) by immunoblot analysis. The same result was obtained at day 7 after treatment. (D) The sensitivity to CsA of HCV genome-length replicon was almost the same as that of the subgenomic replicon. HCV RNA was quantified by real-time RT-PCR analysis using total RNA from NNC#2 (NN/1b/FL), SN1A#2 (Con1/1b/FL), and SNC#7 (Con1/1b/FL) cells treated with various concentrations of CsA for 7 days. The relative amount of HCV RNA was plotted against the concentration of CsA (in micrograms per milliliter). (E) Effect of CsA on cell proliferation. NNC#2 (NN/1b/FL) cells were treated with various amount of CsA for 7 days. Cell numbers were counted, and cell numbers relative to those of cells without treatment were plotted against the concentration of CsA.