RR formation increasingly sensitive to Ribavirin when IMPDH protein level is reduced by specific siRNA knockdown.

<p>HeLa cells were either untransfected, transfected with siCTPS1, or siIMPDH2 for 44 h and followed by incubation of Ribavirin at various concentrations for an additional 4 h. (A) IMPDH2 expression was monitored by Western blot using rabbit anti-IMPDH2 antibody. Serial dilutions of untreated HeLa cell lysates (100%, 50%, and 25%) were used to help to comparatively quantitate the degree of protein knockdown. The level of actin was used as a loading control. Relative IMPDH2 protein levels were quantified using image J and normalized to the serial dilution standards. (B) Percentage of cells with RR were quantified using Mayachitra Imago with a range of 131 to 298 cells counted per data point. Cells treated with siIMPDH2 had a significant increase in percentage of cells with RR at 0.2 µM and 0.05 µM of Ribavirin analyzed by Fisher's exact test. (C–D) Representative image of cells at the lowest concentration of Ribavirin transfected with siCTPS1 or siIMPDH2. Arrows, rods; arrowhead, ring. Bar, 5 µm.